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Nitrogen cycling in an integrated "biomass for energy" system

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Title:
Nitrogen cycling in an integrated "biomass for energy" system
Alternate title:
Biomass for energy system
Creator:
Moorhead, Kevin Keith, 1956-
Publication Date:
Language:
English
Physical Description:
x, 141 leaves : ill. ; 28 cm.

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Subjects / Keywords:
Anaerobic digestion ( jstor )
Biomass ( jstor )
Bodies of water ( jstor )
Nitrogen ( jstor )
Nutrients ( jstor )
Plants ( jstor )
Sediments ( jstor )
Sewage sludge ( jstor )
Sludge digestion ( jstor )
Soil science ( jstor )
Dissertations, Academic -- Soil Science -- UF
Nitrogen cycle ( fast )
Soil Science thesis Ph. D
Water hyacinth ( fast )
Lake Apopka ( local )
Genre:
bibliography ( marcgt )
theses ( marcgt )
non-fiction ( marcgt )

Notes

Thesis:
Thesis (Ph. D.)--University of Florida, 1986.
Bibliography:
Includes bibliographical references (leaves 131-140).
Additional Physical Form:
Also available online.
General Note:
Typescript.
General Note:
Vita.
Statement of Responsibility:
by Kevin Keith Moorhead.

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NITROGEN CYCLING IN AN INTEGRATED
"BIOMASS FOR ENERGY" SYSTEM














By

KEVIN KEITH MOORHEAD



























A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY


UNIVERSITY OF FLORIDA 1986

















ACKNOWLEDGEMENTS



This drssertation reports results from a project that contributes to a cooperative program between the Institute of Food and Agricultural Scieieces .IFAS) of the University of Florida and the Gas Research institute (GRI), entitled "Methane from Biomass and Waste." Financial support from the IFAS/GRI cooperative program is gratefully acknowledged.

Dr. D. A. Graetz was chairman and Dr. K. R. Reddy was cochairman of my supervisory committee. Their friendship and guidance will always be cherished. The other members of the committee were Dr. G. E o. is, Dr. J. G. A. Fiskell, and Dr. R. A. Ncrdstedt.

Special thanks go to Bill Pcthiier who ran r.umercus 'N s:fe.s for this research. Other people who provided as sistanca during thiC :d:~ies include Bill Christy, Stephen McCrackn, PeLe-r Kcottje, Terry an, Veronica Campbell, Premila Rao. Ed Hopwood and Dave Cartlii. (iove Linda designed the statistic !l analyses. I appreciate he use. t Dr. John Moore's facilities for fiber analyses. Carolyn Pickles ;i lrenda Clutter typed the majority of this dissertation on a: IBM computer. Finally, this package is dedicated to my parents, sisters and brother. A trip home always gave me a boost to carry on.









ii

















TABLE OF CONTENTS



Page

ACKNOWLEDGEMENTS .... .. ................... ii

LIST OF TABLES ................... .. . . v

LIST OF FIGURES. .................. . . vii

ABSTRACT . . . . . . . . . . . .... ix

INTRODUCTION. ............... ..... . 1

LITERATURE REVIEW . . . . . . .... .. . . 4

Water Hyacinth Biomass Production ............. 4
Anaerobic Digestion . . . ... . . . .... . 13
Waste By-Product Recycling. ........ ... . . 16
Conclusions .. . ... . . . ... . . . . . 21

WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION. .... .... 23

Materials and Methods .......... . . ...... . 24
Results and Discussion... . . . .......... . 26
Conclusions .. ..... . . . . . .. . . 38

EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER
HYACINTH SYSTEMS . . . . . . . . . . 40

Materials and Methods . . . . . . . . . . 42
Results and Discussion. . . . . .. . . . . 44
Conclusions ................... .... . 58

ANAEROBIC DIGESTION OF WATER HYACINTH. .... . .. . . 60

Materials and Methods . . . . . . . . . . 61
Results and Discussion. . . ..... . . . . 63
Conclusions ................ .. .. . . 74

TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTH 76

Materials and Methods ... . . ..... ...... . 78
Results and Discussion. ............... ... . 80
Conclusions ... .......... . . . . . 93


iii









DECOMPOSITION OF FRESH AND ANAEROBICALLY DIGESTED PLANT
BIOMASS IN SOIL. . . . . . . . . . . . 95

Materials and Methods . . . . . . . . . . 96
Results and Discussion .................. 98
Conclusions ....... . ... ...... . . ... ..111

MASS BALANCE OF NITROGEN IN AN INTEGRATED "BIOMASS FOR ENERGY"
SYSTEM . . . . . . . . . . . . . . 112

Nutrient-Enriched Systems . . . . . . . . . 112
Nutrient-Limited Systems . . . . . . . . . 116

CONCLUSIONS . . . . . . . . . . . . 119

Water Hyacinth Productivity and Detritus Production . . 119 Detritus and Nitrogen Transformations . . . . . . 120
Anaerobic Digestion of Water Hyacinth .......... 120
Digester Effluent Recycling ................ 121
Digester Sludge Recycling . . . . . . . . . 122

APPENDICES

A DIGESTER EFFLUENT CHARACTERISTICS DURING WATER
HYACINTH TREATMENT ................... 124

B SOIL CHARACTERISTICS FROM ADDED FRESH AND ANAEROBICALLY
DIGESTED PLANT BIOMASS ....... .......... 126

BIBLIOGRAPHY . . . . .. . .. .. .. . . .. 131

BIOGRAPHICAL SKETCH. ................... . . 141



























iv















LIST OF TABLES


TABLE PAGE

1. Seasonal water hyacinth yield and detritus production .. 30 2. Seasonal water hyacinth shoot and root lengths .. ... 32 3. Seasonal nitrogen uptake by water hyacinth and detritus .. 35 4. Nitrogen balance for the two reservoirs ... ....... 37

5. Total plant N and 15NO3-N assimilation. ..... . . . 53



15
6. Total plant N and NH4-N assimilation .... ........... .54

7. Mass balance of added 15N3-N in sediment-water-plant systems . . . . . . . . . . . . . 56
15 +
8. Mass balance of added NH 4-N in sediment-water-plant systems . . . . . . . . . . . . . 57

9. Characteristics of the inoculum used in the batch digesters . . . . . . . . . . . . 64

10. Gas production during anaerobic digestion of high and
low N water hyacinth plants ............... 65

11. Nitrogen balance for the batch digesters. ... .... 68

12. Nitrogen-15 balance for the batch digesters ...... 69 13. Characteristics of digester effluents before sludge
removal ............ . . . . . . 71

14. Characteristics of screened effluents sludgee removed)
after digestion . . . . . . . . . . . 72

15. Characteristics of fresh and digested biomass residues. . 73 16. Initial characteristics of the digester effluents
and nutrient medium ... . . ............... . 81

17. First-order kinetic descriptions of NH -N loss with time. 86 18. Nitrogen-15 balance for labeled effluents ........ 88


v









TABLE PAGE

19. Distribution of nutrients in water hyacinth shoots and roots in diluted and undiluted effluents of
digested high N plants. ................. 89

20. Net assimilation or loss of plant nutrients in diluted or undiluted effluents from digested high N plants. ... 91 21. Characteristics of the digester effluents and nutrient medium after water hyacinth treatment ........ . 92

22. Characteristics of the fresh and digested plant biomass 99 23. Soil NO -N concentration from added fresh and digested
plant biomass . . . . . . . . . . . .105

24. Carbon and 15N mineralization from added fresh and
digested plant biomass. ................. .106

25. Soil pH (1:2 w/v) from added fresh and digested plant
biomass . . . . . . . . . . . . .109

26. Mehlich I extractable constituents at Day 90 from
added fresh and digested plant biomass. ......... .110

27. Effluent pH during water hyacinth treatment ........ .124 28. Effluent dissolved 02 concentration during water
hyacinth treatment. . . . . . . . . . .125

29. Soil ammonium concentrations from added fresh and
digested plant biomass. ................. .127

30. Mehlich I extractable constituents at Day 0 from
added fresh and digested plant biomass. ......... .128

31. Mehlich I extractable constituents at Day 30 from
added fresh and digested plant biomass. ......... .129

32. Mehlich I extractable constituents at Day 60 from
added fresh and digested plant biomass. ......... .130















vi
















LIST OF FIGURES


FIGURE PAGE

1. Integrated water hyacinth aquaculture system
of biomass production, bioconversion to methane
and digester waste recycling. ............. 2

2. A generalized diagram of a water hyacinth plant ...... 5 3. Nitrogen cycling in a water hyacinth production sytem .. 9 4. Nitrogen cycling during anaerobic digestion ....... 15 5. Nitrogen cycling in soil treated with plant residues. . 18

6. Weekly averages of daily temperatures and solar
radiation . . . . . . . . . . . . 27

7. Monthly averages of daily primary productivity and
detritus production .................. 28

8. Seasonal plant tissue nitrogen content. .......... 33

9. Dissolved O in sediment-water-plant systems with added nitra e . . . . . . . . . . . 45

10. Dissolved 02 in sediment-water-plant systems with added ammonium. .................. ... 46

11. The pH of sediment-water-plant systems with added nitrate . . . . . . . . . . . 48

12. The pH of sediment-water-plant systems with
added ammonium. .................. ... 49

13. Nitrogen loss from sediment-water-plant systems
with added nitrate. .................. 50

14. Nitrogen loss from sediment-water-plant systems
with added ammonium .................. 51

15. Dry weight gains of water hyacinths in digester
effluents and nutrient medium .............. 83



vii









FIGURE PAGE

16. Carbon evolution from soil applied fresh and digested plant biomass. ........... . .. 101

17. Decomposition stages and rate constants of fresh and digested plant biomass added to soil. ........ 103

18. Nitrogen cycling in an integrated system for water hyacinths growing in nutrient-enriched systems. ..... 113 19. Nitrogen cycling in an integrated system for water hyacinths growing in nutrient-limited systems ...... 117















































viii















Abstract of Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosopy


NITROGEN CYCLING IN AN INTEGRATED "BIOMASS FOR ENERGY" SYSTEM

By

Kevin Keith Moorhead

May, 1986

Chairman: Dr. D. A. Graetz
Cochairman: Dr. K. R. Reddy
Major Department: Soil Science

A series of experiments were conducted to evaluate N cycling in three components of an integrated "biomass for energy" system, i.e. water hyacinth production, anaerobic digestion of hyacinth biomass, and recycling of digester effluent and sludge. Plants assimilated 50 to 90% of added N in hyacinth production systems. Up to 28% of the total plant N was contained in hyacinth detritus. Nitrogen loading as plant
-1 -1
detritus into hyacinth ponds was 92 to 148 kg N ha yr

Net mineralization of plant organic 15N during anaerobic digestion was 35 and 70% for water hyacinth plants with low (10 g N kg-1 dry tissue) and high (35 g N kg-1) N content, respectively. Approximately 20% of the N was recovered in the digested sludge while the remaining N was recovered in the effluent.

Water hyacinth growth in digester effluents was affected by

electrical conductivity (0.7 to 6.7 dS m-1 ) and 1NH -N concentration




ix









(23 to 289 mg N L- ). Biomass yields were maximum at electrical
-1 15 +
conductivities of < 2.5 dS m and NH -N concentrations of < 100 mg N
-1

Addition of water hyacinth biomass to soil resulted in

decomposition of 39 to 50% of added C for fresh plant biomass and 19 to


151
23% of added C for digested biomass sludge. Only 8% of added 15N in digested sludges was mineralized to 1NO3-N despite differences in initial N content (27 and 39 g N kg-I dry sludge). In contrast, 3 and 33% of added 15N in fresh biomass with low and high N content, respectively, was recovered as 15NO3-N.

Total 15N recovery after anaerobic digestion ranged from 70 to 100% of the initial plant biomass 15N. Land application of digester sludge resulted in the mineralization of 2% of initial biomass 15N into plant available form. Use of water hyacinth for digester effluent treatment resulted in recycling of 21 to 38% of the initial biomass 15N. Total N recovery by sludge and effluent recycling in the integrated "biomass for energy" system was 48 to 60% of the initial plant biomass 15N. The remaining 15N was lost from the system during anaerobic digestion and effluent recycling.

















x















INTRODUCTION



Several types of aquatic plants are widely distributed in

freshwater lakes and streams. These plants assimilate nutrients and produce biomass, which could potentially be used for beneficial purposes. Water hyacinth (Eichhornia crassipes [Mart] Solms) is one of the dominant aquatic plants distributed throughout the tropical and subtropical regions of the world. This freshwater macrophyte has already been evaluated for use in treating nutrient-enriched waters such as sewage effluent (Cornwell et al., 1977; Wolverton and McDonald, 1979; Reddy et al., 1985), agricultural drainage water (Reddy and Bagnall, 1981; Reddy et al., 1982), anaerobic digester effluent (Hanisak et al., 1980), and fertilized fish ponds (Boyd, 1976). The characteristics that make this plant grow rapidly in polluted waters make it an ideal candidate for large-scale nutrient removal and water purification (Reddy and Sutton, 1984).

An integrated aquaculture system has been developed using water hyacinth for water treatment and for total resource recovery. The components of an integrated aquaculture system are schematically illustrated in Fig. 1. Water hyacinth plants have been used for wastewater treatment while the biomass produced was harvested periodically and processed through anaerobic digestion to produce methane. The process produced a waste by-product which must be disposed of, or preferably utilized, in an environmentally-safe manner.

1









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The waste by-product contains digested biomass sludge (solid) and effluent (liquid). The digested biomass sludge was applied to soil as a nutrient source for plants. The effluent was recycled in water hyacinth ponds for nutrient recovery by plants. This type of integrated system will provide low cost water treatment and total resource recovery. Efficient utilization of by-products could potentially reduce the cost effectiveness of the system.

The overall objective of this study was to assess nitrogen cycling in the three components of an integrated "biomass for energy" system. Nitrogen is often identified as a limiting factor for plant growth and is used to establish loading rates in the disposal of solid and liquid waste. Information on N cycling is limited to studies on the individual components of the integrated system, i.e. the water hyacinth production system (Boyd, 1976; DeBusk et al., 1983; Reddy, 1983), anaerobic digestion (Hashimoto et al., 1980; Field et al., 1984), and effluent and sludge recycling (Ryan et al, 1973; Hanisak et al., 1980; Terry et al., 1981; Atalay and Blanchar, 1984). No attempt has been made to establish N cycling within the entire integrated system.

The specific objectives of this study were 1) to determine growth rate and detritus production of water hyacinth grown in eutrophic lake water; 2) to determine the effect of detritus on N transformations in water hyacinth systems; 3) to evaluate N and C mineralization during anaerobic digestion of water hyacinth biomass; 4) to evaluate the potential of water hyacinth to grow in anaerobic digester effluents for N recovery; and 5) to determine N and C mineralization during decomposition of fresh and digested biomass added to soil.
















LITERATURE REVIEW



The three components of the integrated "biomass for energy" system were 1) the water hyacinth production system; 2) anaerobic digestion of water hyacinth biomass; and 3) recycling of digested biomass sludge and effluent. An integrated approach of wastewater renovation using aquatic macrophytes with utilization of biomass for energy production is economically appealing.



Water Hyacinth Biomass Production

The first component of the integrated "biomass for energy" system was an aquatic system for the production of biomass as well as water quality improvement. Although several aquatic plants naturally grow in polluted waters, one the most productive plants appears to be water hyacinth (Reddy et al., 1983).

Water hyacinth is a mat-forming, free-floating vascular aquatic

plant with wide distribution in sub-tropical and tropical regions. The plant consists of a submerged rooting system and an aerial photosynthetic petiole and leaf (shoot) system (Fig. 2). The roots and aerial shoots are produced at the numerous nodes of the vegetative portion of a typically submerged rhizome (Penfound and Earle, 1948). The aerial buds, from which flowers and fruit clusters develop, are produced from the reproductive portion of the rhizome. Occasionally, the internodes of the rhizome expand and form new offsets.

4






























LA /
/SP
-PT


i





F

ST
RHS AR -RA








Figure 2. A generalized diagram of a water hyacinth plant.
The major morphological structures are
adventitious roots (AR); root hairs (RA); rhizome (RH); stolon (ST); detritus tissue (DT) attached to the plant; float (F); leaf isthmus (IS);. leaf
petiole (PT); peduncle (PD); spathe (SP); leaf
lamina (LA); inflorescence (IN).






6


The elongated internodes were designated as stolons (Penfound and Earle, 1948).

The relatively rapid rate of colonization by water hyacinth is due primarily to vegetative reproduction (stolon and offset production). The plants reproduce sexually during warmer months until freezing terminates anthesis. The developing fruits containing the seeds are cast off onto the mat or into the water. They sink in water and remain in a viable condition for several years. Manson and Manson (1958) reported that each plant could produce 5000 to 6000 seeds which remained viable for at least 5 years.

The geographical distribution of water hyacinth is regulated by

temperature and salt concentration in the water. When average minimum temperature reached 10 C, productivity of water hyacinth approached zero (Reddy and Bagnall, 1981). Optimum growth was found in a temperature range of 25 to 300C (Bock, 1969; Knipling et al., 1970). Water hyacinth is basically a freshwater plant and will die in waters with sustained salt concentrations in excess of 2500 mg L-1 (Haller et al., 1974).

Water hyacinth growth is regulated by the nutrient composition of the water medium, temperature, solar radiation, and plant density. Water hyacinth potentially could be grown in nutrient-enriched waters such as sewage effluents, agricultural runoff and drainage effluents, methane digester effluents, and runoff from animal waste operations. Nitrogen is present as NH -N, NO3-N, and organic N in water media avaiable for water hyacinth production. Organic N often predominates in most water media and is not readily available for plant assimilation. Water hyacinths are efficient users of inorganic N and plant assimilation is one of the major processes of N removal in hyacinth ponds.






7


Water hyacinth adapted to light intensity and full sunlight elicted the greatest photosynthetic rate (Patterson and Duke, 1979). Optimum plant density to obtain maximum biomass yield varied with season and available plant nutrients in the water (Reddy and Sutton, 1984). DeBusk et al. (1981) and Reddy et al. (1983) established that optimum plant density for achieving maximum growth cultured in wastewaters was in the
-2
range of 15 to 35 kg wet wt m-2

Water hyacinth productivity has been evaluated in natural and

nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m-2 day-1 were reported for plants growing in natural waters of central and south Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide-range of productivity (5 to 42 g dry wt m-2 day-1) was recorded for plants cultured in nutrient-enriched waters (Schwegler and Kim, 1981; Hanisak et al., 1980). Reddy and DeBusk (1984) obtained an average of 52 and a
-2
maximum of 64 g dry wt m-2 day-i for water hyacinths grown in nutrient-nonlimiting conditions.

The effectiveness of water hyacinth in removing inorganic N was reported for several nutrient-enriched wastewaters. Sheffield (1967) and Clock (1968) reported a 75 to 94% reduction of inorganic N from secondary sewage effluent in systems containing water hyacinths. Reddy et al. (1982) observed a 78 to 81% reduction of inorganic N from organic soil drainage water containing water hyacinths. Hanisak et al. (1980) concluded that 65% of N in digester effluents could be assimilated when water hyacinths were grown in diluted effluents. Boyd (1976) calculated average rates of N and P removal were 3.4 and 0.43 kg ha-1 day-I in fertilized fish ponds. Rogers and Davis (1972) concluded that water hyacinth removal capacities were less effective with increasing nutrient concentrations.









The potential productivity and nutrient removal capacities of water hyacinth has led to its selection as a biomass feedstock for methane generation while providing a means for treatment of nutrient-enriched waters. Extensive research, both in laboratory and field applications, was conducted on the use of water hyacinth in wastewater treatment during the past 20 years (Sheffield, 1967; Boyd, 1970a; Steward, 1970; Scarsbrook and Davis, 1970; Rogers and Davis, 1971; Dunigan et al., 1975; Cornwell et al., 1977; McDonald and Woverton, 1980; Reddy et al., 1982; DeBusk et al., 1983). Water hyacinth was shown to be effective in removing N, P and other nutrients, and reducing biological oxygen demand and total suspended solids. Water hyacinth was also shown to readily absorb and concentrate heavy metals (Wolverton and McDonald, 1975a,b; Cooley et al., 1978).

Nitrogen Cycling in the Water Hyacinth Production System

Nitrogen transformations occurring in a water hyacinth production system include 1) plant uptake; 2) mineralization/immobilization; 3) nitrification; 4) denitrification; and 5) NH3-N volatilization (Fig. 3).

Plant uptake is one of the major processes for N removal from water hyacinth-based wastewater systems. Plant uptake is directly related to the growth rate and the nutrient composition of the water. Water hyacinth was more efficient in utilizing NH -N than NO3-N when both forms were supplied in equal proportions (Reddy and Tucker, 1983).

A dense cover of floating water hyacinths will regulate dissolved 02' temperature and pH of water which influences several N transformations. Generally, diel fluctuations of these water parameters were reported to be lower in areas covered with water hyacinths compared to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980; Reddy, 1981).









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10


High 02 consumption within a water hyacinth mat during microbial decomposition of plant detritus (dead and decaying plant debris) could create anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980;). The bulk of detritus was trapped within the mat and decomposed primarily at the water surface (DeBusk et al., 1983). Rate of inorganic N release from decomposing detritus depended on dissolved 02 concentration of the water, C/N ratio, and temperature (Ogwada, 1983).

Low 02 concentrations create conditions less favorable for

nitrification and promote denitrification which may proceed within the water hyacinth mat, in the water column, or in the underlying sediment. Denitrification occurred primarily in the underlying sediment and the rate depended on diffusion of NO3 -N from the water column to the sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981).

Water temperatures were lower in areas covered with plants compared to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980; Schreiner, 1980). A dense mat over the water surface served as a blanket barrier for exchange of heat between the atmosphere and the water (Rai and Munshi, 1979). Water hyacinths growing in either acid or alkaline water had a tendency to alter the pH towards neutrality (Haller and Sutton, 1973). A pH of 7.0 in water occurred in areas covered with plants with little diel variation (McDonald and Wolverton, 1980; Reddy, 1981) which suggests that NH3-N volatilization is minimal in these systems

Decomposition of Plant Tissue in Freshwater

Decomposition of plant tissue in a freshwater habitat commonly occurs in two stages. The first stage was attributed to leaching of the more soluble plant constituents while the second stage was









microbial-controlled degradation (Boyd, 1970b; Hunter, 1976; Godshalk and Wetzel, 1978a; Howard-Williams et al., 1983).

Otsuki and Wetzel (1974) reported a rapid leaching loss of

dissolved organic matter regardless of conditions of aerobiosis or whether plants were fresh or freeze-dried. Hill (1979) concluded that rapid leaching of soluble material accounted for a 21 to 60% dry weight loss of aquatic macrophytes during the first 8 days of incubation. Leaching was established as the major process in the decomposition of eelgrass and total loss of organic matter by leaching accounted for 82% of dried leaves and 65% of fresh leaves (Harrison and Mann, 1975). Leaching rates appeared to be independent of temperature (Carpenter, 1980).

Potassium, Na, Mg, and Ca have all been reported as being rapidly lost during the early leaching phase of plant decomposition (Boyd, 1970b; Davis and van der Valk, 1978; Puriveth, 1980). Carpenter (1980) found that the higher the initial P concentration, the more rapid was P leaching.

The second stage of decomposition is attributed to biological processes. Microbial-controlled decomposition was influenced by temperature (Carpenter, 1980; Puriveth, 1980), pH (Sompongse, 1982), available 02 (Godshalk and Wetzel, 1978a), and available nutrients (Carpenter and Adams, 1979; Puriveth, 1980). Godshalk and Wetzel (1978a) found that the presence of 02, regardless of temperatures of 10 or 250C, permitted rapid degradation of dissolved and particulate organic matter. Decomposition of water hyacinth was found to be faster under aerobic than anaerobic conditions (Reddy and Sacco, 1981). However, Sompongse (1982) determined that aeration did not have a






12


measurable effect on rate of plant decomposition and Nichols and Keeney (1973) reported a more rapid decomposition rate under non-aerated conditions compared to aerated conditions. Ogwada et al. (1984) found that approximately the same amounts of N and P were released from decaying plant tissue under aerobic or completely anoxic conditions, but the extent of nutrient release was dependent on water temperature.

The changes in plant C and N composition or concentration during decomposition have received considerable attention. Build-up of microbal biomass on decaying plant tissue caused a loss in the C content while increasing the N content which resulted in a decrease in the C/N ratio (De La Cruz and Gabriel, 1974; Odum and Heywood, 1978; Hill, 1979). Godshalk and Wetzel (1978b) found that lignin was very resistant to decomposition while the other structural carbohydrates gradually decreased with time.

Nitrogen was a limiting factor in decomposition of several aquatic plants (Nichols and Keeney, 1973; Almazan and Boyd, 1978; Godshalk and Wetzel, 1978b; Carpenter, 1980). Decay rates were correlated both to initial N content and to C/N ratios (Godshalk and Wetzel, 1978b; Carpenter and Adams, 1979; Ogwada et al., 1984).

Particle size also influenced decomposition. Generally, the rate of decomposition increased as the particle size decreased (Fenchel, 1970; Hargrave, 1972; Gosselink and Kirby, 1974). Harrison and Mann (1975) reported that a reduction in size of leaf material from 2 to 4 cm to <1 mm doubled the rate of organic matter loss.

Boyd (1970b) and Odum and Heywood (1978) concluded that submerged leaves decomposed more rapidly than those placed upon the water surface or suspended in air. Nichols and Keeney (1973) found more rapid dry





13



weight loss under aerated conditions in sediment-water systems than in water only. They attributed this difference to an additional supply of N from the sediments.



Anaerobic Digestion

The second component of the integrated "biomass for energy" system was anaerobic digestion of plant biomass for methane production. Anaerobic digestion is a biological process in which organic matter, in the absence of oxygen, is converted to methane and carbon dioxide (Toerien and Hattingh, 1969).

During the process of anaerobic digestion, waste organic C was stabilized by the nearly complete microbial fermentation of carbohydrates resulting in a reduction of volatile solids (Miller, 1974). Anaerobically digested sewage sludges were considered more stable to microbial degradation than were aerobically digested sludges (Sommers, 1977).

Processes which regulated anaerobic digestion include hydrolysis of polymers, the dissimilation of starting subtrates to the level of acetic acid, and the conversion of acetic acid to CH4 and CO2 (Mah et al., 1977). Factors which influenced anaerobic digestion include pH and temperature changes. All methanogens were reported to be strict anaerobes with an optimum pH of 6.7 to 7.4 (Bryant, 1979). The optimum temperature range was 30 to 350C (House, 1981).

Water hyacinth biomass could be anaerobically digested to produce methane. Hanisak et al. (1980) found average methane yields of 0.24 L
-l
g volatile solids (VS) of shredded water hyacinth in 162 L digesters. Chyoweth et al. (1983) reported methane yields of 0.19 and 0.28 L g-1 VS






14


for water hyacinths and a 3:1 blend of water hyacinths:domestic sewage sludge, respectively in 5 L digesters. Shiralipour and Smith (1984)
-1
reported average methane yields of 0.32 and 0.17 L g-1 VS water hyacinth shoot and root samples, respectively, in a bioassay test of 100 ml culture volume. They concluded that the addition of N to growth media for water hyacinth production increased methane yields.

The nutrients in the biomass are recovered in the waste material after the digestion process. The recovered nutrients are distributed between the effluent and the digested biomass sludge. The total N recovery was usually 100% after digestion, but much of the organic N was converted to NH -N (Hashimoto et al., 1980; Field et al., 1984). Most of the K (Field et al., 1984) and Na (Atalay and Blanchar, 1984) were solubilized and remained in the digester effluent.

The digestion process increased the sorption of some nutrients (P, Ca, and Mg) by the sludge fraction such that fewer were available for dilute acid extraction and perhaps for crop recovery (Field et al., 1984). Field et al. (1984) hypothesized that sorption may have been increased by particle surface area increases due to size reduction of solids.

Nitrogen Cycling During Anaerobic Digestion

Nitrogen transformations occurring during anaerobic digestion

include 1) mineralization/immobilization; and 2) NH3-N volatilization (Fig. 4).

The mineralization or immobilization of N depends on the initial N content of the biomass feedstock. The C/N ratio of the biomass is often used as a guideline for prediction of net mineralization or immobilization. The optimal C/N ratio of the added biomass was 30:1






15




0




N .











L.. .- W..ozz
*O



-o J
a

Oc

o









0-4

C CJ SDo





I W0


z
*H -






16


(Hughes, 1981). Bioconversion of biomass with a higher C/N ratio was limited by N. A lower initial C/N ratio resulted in mineralization of organic N during digestion. The C/N ratio of the digester effluent was lower than the C/N ratio of the fresh slurry because of the release of C as CO2 and CH4 (House, 1981).

Anaerobic digestion of plant biomass resulted in high

concentrations of NH -N in the digester (Hashimoto et al., 1980; Field et al., 1984). Ammonium was toxic to methogens at concentrations > 3.0 g L-l, regardless of pH (Hashimoto et al., 1980). Losses of NH3-N through volatilization should be low in digesters operating at the optimum pH of 6.7 to 7.2 unless NH N concentrations are high.



Waste By-Product Recycling

The final component of the integrated "biomass for energy" system was recycling of the waste by-product generated from the anaerobic digestion of plant biomass. The waste by-product from the anaerobic digester was screened to separate the digested biomass sludge from the effluent. The effluent was recycled in the water hyacinth biomass production system discussed earlier. Methane digester effluent contains high levels of NH -N (> 200 mg L-1) which may inhibit plant growth. Dilution of the effluent is required before use in a water hyacinth production system. Optimum dilution of the effluent for maximum water hyacinth yields has not been established. Nitrogen cycling in a water hyacinth production system was presented earlier.

The sludge was added to soil as an organic amendment. A

consequence of anaerobic digestion was a reduction of the readily decomposable C of the plant tissue during production of CH4 and CO2'





17



Anaerobically digested sludge was considered to be stable and resist further decomposition (Sommers, 1977). Nitrogen Cycling in Soil Treated with Plant Residues

The land application of fresh or anaerobically digested plant biomass has significant implications on N cycling. Nitrogen transformations occurring after residue additions include 1) mineralization/immobilization; 2) microbial or plant assimilation; 3) nitrification; 4) denitrification; and 5) NH3-N volatilization (Fig. 5).

Since most of the N in fresh or digested plant biomass is in

organic forms, the rate of mineralization becomes the rate limiting step for all transformations that follow. Mineralization or immobilization depends on the initial N concentration of the plant biomass as well as the composition of the C constituents. A low N content or a wide C/N ratio was associated with slow decomposition and rates of decomposition were proportional to lignin content (Alexander, 1977). A wide C/N ratio (> 30:1) favored N immobilization whereas a narrower ratio (< 20:1) resulted in N mineralization (Alexander, 1977).

Mineralization of N from anaerobically digested sewage sludges was reported to be affected by the rate of application (Ryan et al., 1973; Stark and Clapp, 1980). However, Epstein et al. (1978) found that irrespective of the amount of material (sewage sludge and sludge compost) applied, the percentage of added N mineralized remained essentially constant.

The mineralization of NH+-N from organic N is accompanied by

microbial assimilation or plant uptake. In aerobic environments the NH -N was quickly converted to NO -N (nitrification) which could also be
4 3






18




C



So ) c Fr .- z I- "





4-




< *
I-- I I -I





a c









Ia U
o ud
U) 4.J Q
r_ cad








0 CM 4-0 Zz





19


assimilated by microbes or plants (Ryan et al., 1973). A high organic loading rate may result in 02 depletion during decomposition which promotes the denitrification process (Epstein et al., 1978; Hsieh et al., 1981b).

Decomposition of Plant Residues in Soil

Decomposition of plant residues in soil occurs in two stages. The first stage was attributed to loss of the easily decomposable labile fraction which was followed by the second stage of slow decomposition of a resistant residual fraction (Shields and Paul, 1973; Reddy et al., 1980). Both stages were thought to be controlled by two simultaneously occurring superimposed first-order kinetic reactions (Sinha et al., 1977).

Fresh and anaerobically digested plant biomass differ widely in their chemical composition. Anaerobic digestion converts most of the easily-decomposable plant C constituents into CH4 and CO2. The digested biomass sludge has a higher lignin content and is more resistant to decomposition. There is little information available on decomposition of anaerobically digested plant biomass added to soil. However, the rates and the factors which influence decomposition of fresh plant biomass added to soil have been well-established.

Tenny and Waksman (1929) concluded that water-soluble organic substances were first to be decomposed in the soil, followed by hemicellulose and at the same time, or immediately after, cellulose. Lignin was very resistant to decomposition and may even delay the disintegration of cellulose or hemicellulose because of the structural proximity of these C constituents in the cell wall (Tenny and Waksman, 1929; Peevey and Norman, 1948; Berg et al., 1982).





20


Application rates were shown to have insignificant effects on rate of fresh plant biomass decomposition (Jenkinson, 1965; Nyhan, 1975). However, several studies suggested that small amounts of fresh or anaerobically digested plant biomass decomposed more rapidly than large quantities (Broadbent and Bartholomew, 1948; Jenkinson, 1971; Atalay and Blanchar, 1984).

Miller and Johnson (1964) found an increasing rate of CO2

production with increasing moisture content up to a tension of 0.05 to 0.015 MPa and then a decreasing rate with further increases in tension. They concluded that maximum biological activity could be expected at the lowest tension when aeration was sufficient. Orchard and Cook (1983) found a log-linear relation between water potential and microbial activity in the range of 0.005 to 0.5 MPa.

Sain and Broadbent (1977) concluded that low temperatures

influenced decomposition rate more than excessive moisture. However, Nyhan (1976) found a pronounced decrease in rates of C loss with an increase in soil water tension even when temperature (100C) was limiting microbial activity. Miller (1974) determined that soil temperature was the major factor influencing the rate of decomposition of anaerobically digested sewage sludge.

Decomposition was generally considered to be initially slower in acid than neutral soil (Jenkinson, 1971). Addition of organic material altered the pH of a soil, particularly when the amount added was large relative to the amount of native organic matter present (Jenkinson, 1966). Atalay and Blanchar (1984) found that addition of anaerobically digested biomass sludge to soil increased the pH from 5.5 to 7.6 and they attributed this to a limestone buffer used during the digestion process.





21


Jenkinson (1965, 1971), using different plants and soils,

determined that the proportion of added plant C retained in the soil under different climatic conditions was remarkably similar over time. Generally, one-third of the added plant C remained after one year, falling to one-fifth after 5 years.

Atalay and Blanchar (1984) determined that anaerobically digested biomass sludge decomposed rapidly in soil as evidenced by nearly 40% of the C added evolved as CO2 during 100 days of decomposition. However, Miller (1974) concluded that anaerobic digested sewage sludge was resistant to further decomposition with a maximum of 20% of the added C evolved as CO2 during a 6-month incubation. Terry et al. (1979) found that 26 to 42% of anaerobically digested sewage sludge C was evolved as CO2 during incubation. Generally, the majority of the CO2 produced in incubation studies was evolved in the first 30 days (Miller, 1974; Terry et al., 1979; Ataway and Blanchar, 1984).

Other soil properties influenced by plant biomass additions included increasing water-holding capacity, CEC, and electrical conductivity (Stark and Clapp, 1980; Atalay and Blanchar, 1984). Epstein et al. (1976) found levels of salinity and chloride in sewage sludge applied to soils increased to a level which may affect salt-sensitive plants.



Conclusions

Although information is available on N cycling for each component of the system, no attempt has been made to follow N transformations in an integrated "biomass for energy" system. Evaluation of N cycling was





22


chosen because N is often identified as a limiting factor for plant growth and is used to establish loading rates in the disposal of solid and liquid waste.

Plant uptake was established as a major N removal process during water hyacinth biomass production (Reddy and Sutton, 1984). However, the role of water hyacinth detritus as a N source or sink has not been established. Methane yields during anaerobic digestion of water hyacinth were enhanced with increasing N content (Shiralipour and Smith, 1984). However, N mineralization rates were not investigated. Limited information was available on disposal or utilization of digester effluent or sludge from anaerobically digested plant biomass (Hanisak etal., 1980; Atalay and Blanchar, 1984). The overall objective of this research was to integrate the three components of biomass production, anaerobic digestion of biomass, and digester waste recycling with respect to N cycling.















WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION



Water hyacinth is one of the most productive aquatic macrophytes

found throughout the tropical and subtropical regions of the world. The plant has been used extensively for treatment of nutrient-enriched waters and currently there are a number of wastewater treatment systems in the U. S. utilizing water hyacinths for secondary and tertiary treatment (Cornwell et al.,1977; Dinges, 1978; Wolverton and McDonald, 1979; Reddy et al., 1985).

Water hyacinth productivity has been evaluated in natural and

nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m-2 day-i were reported for plants growing in natural waters of central and south Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide range of productivity (5 to 42 g dry wt m-2 day-1) was recorded for plants cultured in nutrient-enriched waters (Hanisak et al., 1980; Reddy and Bagnall, 1981; Reddy, 1984). Maximum growth rates provided an average of 52 and a maximum of 64 g dry wt m-2 day-i for plants cultured under nutrient-nonlimiting conditions (Reddy and DeBusk, 1984).

Plant detritus (dead and decaying plant debris) is an integral part of water hyacinth mats. Detritus is usually derived from natural aging of plants, biological or chemical control, and frost damage. Decomposition of detritus releases nutrients which can be subsequently utilized by water hyacinths. Information on water hyacinth productivity was extensive (Reddy et al., 1983), but research on detritus production and its role as a nutrient sink or source was limited.

23





24


DeBusk et al. (1983) measured detritus production in harvested and nonharvested water hyacinth based sewage treatment systems. Detritus

-2 -1
production in both systems averaged 2 g dry wt m day-. More than 80% of the detritus consisted of root material. The bulk of the standing crop detritus remained trapped in the floating plant mat. However, this study did not reveal the potential of detritus as a nutrient input to the water hyacinth ponds.

The objectives of this study were to 1) measure productivity and detritus (shoot and root) production of water hyacinths grown in eutrophic lake water with and without added nutrients and 2) determine the potential of detritus as a nutrient source or sink to the ponds.



Materials and Methods

The study was conducted in two reservoirs located at the Central

Florida Research and Education Center research farm near Lake Apopka in Zellwood, Florida. The reservoirs were constructed with 2.0 m high levees of a Lauderhill organic soil (Lithic medisaprists) and with bottoms composed of calcareous clay. The water depth was 60 cm and the dimensions of the reservoirs were 7.6 m by 61 m (total surface area of 465 m2). Both reservoirs were filled with water from nearby Lake Apopka, and were sectioned into four equal areas for replication and stocked with water hyacinths.

A total of eight 0.25 m2 cages (Vexar mesh screen connected to 5 cm diameter PVC pipe) were stocked with water hyacinth at an initial density of 16 kg (fresh wt) m-2. The cages were placed within the four density of 16 kg (fresh wt) m The cages were placed within the four





25


replicated areas of each reservoir. Each cage was lined with 1.00 mm fiberglass screen to retain any detritus dislodged from the plant mat. The fiberglass screen was positioned 15 cm below the PVC frame to allow normal waterflow within the root mat.

One reservoir was fertilized monthly by broadcasting a 10-4-10

fertilizer to add 100 kg N ha-1 from October 1981 to February 1982, and 50 kg N ha-1 from March 1982 to September 1982. The change in
-1
fertilizer rate was due to an excess of 10 mg N L-1 found in reservoir water several days after fertilization during winter. The second reservoir contained Lake Apopka water with no added nutrients and served as the control. Both reservoirs were drained and refilled with Lake Apopka water monthly (24 hr prior to plant sampling and fertilization).

Plant productivity and detritus production were monitored at monthly intervals for one year. The cages were removed from each section, drained for 5 min, and weighed. The plant material was divided into healthy plants and detritus. Detritus was defined as that collected from the fiberglass screen and dead shoot and root material remaining within the plant mat. Dead shoots were defined as material visably devoid of chlorophyll. Three plants were removed for analyses and the cages were restocked to the initial plant density and placed in the reservoirs.

The shoot and root lengths were recorded for each plant sample and separated for dry weight ratios. The plant tissue and detritus were oven-dried at 700C, weighed, and ground to pass a 0.84 mm Wiley Mill screen. All plant and detritus samples were analyzed for total Kjeldahl nitrogen (TKN) using a modified micro Kjeldahl procedure (Nelson and





26

Sommers, 1973). Solar radiation and high and low daily temperatures were recorded. The results were statistically analyzed for a randomized block design with the fertilized and control reservoirs as treatments.



Results and Discussion

The weekly averages of daily maximum and minimum air temperatures and solar radiation are-shown in Fig. 6. Maximum temperatures ranged from 21.90C during January to March and 36.50C during July to September. Minimum temperatures for these time periods were 8.20C and 20.30C, respectively. Maximum and minimum temperatures for the rest of the year were similar (high= 300C, low= 13.50C). Maximum and minimum solar radiation occurred from April to September and from November to March, respectively.

The monthly averages of daily primary productivity and detritus production of water hyacinth are presented in Fig. 7. Maximum daily water hyacinth productivity during this study was observed in August for
-2
the fertilized reservoir (28.3 g dry wt m-2 day- ) compared to June for
-2
the control reservoir (14.7 g dry wt m-2 day- ). Detritus production remained fairly consistent with time for both reservoirs. Detritus production in the fertilized reservoir increased noticeably in September when plant productivity began to decline. The average daily detritus production in the fertilized reservoir was 3.7 g dry wt m day compared to 3.5 g dry wt m-2 day-1 in the control reservoir. DeBusk et al. (1983) found that detritus production occurred at a relatively constant rate regardless of harvested or nonharvested conditions.

Monthly data for the plant parameters have been summarized by seasons: 1) autumn (October, November, and December); 2) winter





27


40

O" 35
C max. 3025
ILl




0-6 LLI
I I I ILi 5 SD



I>,

T( n 218 7)197

Z 176
O
155

I 134

I 13

< 92 0 71
U)



TIME (months)


Figure 6. Weekly averages of daily temperatures and solar
radiation.





28

Fertilized 43.2

30- Growth Rate Detritus 25


-- 20
I
0' 15



E 10
-A
5 -_ -.-0

20o- Control CI
C) 15



O o




0
s --I------ ------i--E--j





N D J F M A M J J A S 0
TIME (months)


Figure 7. Monthly averages of daily primary productivity and detritus production.





29


(January, February, and March; 3) spring (April, May, and June); and 4) summer (July, August, and September). The seasons were chosen to coincide with changes in water hyacinth productivity and temperature and solar radiation changes. The autumn season combines data for November and December of 1982 and October of 1983.

The effects of temperature and solar radiation were evident since the lowest net productivity occurred during winter and the highest during spring and summer (Table 1). The net primary productivity in winter for both reservoirs was lower than the production of detritus. Reddy and Bagnall (1981) reported that at average temperatures of 100C, productivity of water hyacinth approached zero. The majority of the detritus during winter came from the destruction of aerial shoots caused by freezing temperatures. Although a majority of the aerial shoots were destroyed, the plants survived, and noticeable gains in dry weight began in March.

There were significant differences in yields between seasons and reservoirs. Seasonal yields ranged from 1.9 to 23.1 Mg (dry wt) ha-1 for the fertilized reservoir and -0.2 to 10.2 Mg ha-1 for the control reservoir. Over 75% of the biomass production occurred during spring and summer for both reservoirs. Differences in detritus production were not significant between reservoirs or between seasons. Although the annual yield of water hyacinth in the fertilized reservoir was double that of the control reservoir, the detritus production in both reservoirs was similar (Table 1). There were significant differences in the shoot/root dry weight ratios between reservoirs, but not between seasons (Table 1). The average shoot/root dry weight ratio was 1.93 (1.64 to 2.46) for the fertilized reservoir and 1.14 (0.79 to 1.67) for the control reservoir.





30



Table 1. Seasonal water hyacinth yield and detritus production.



Fertilized reservoir Control reservoir Season Yield Detritus S/R Yield Detritus S/R

---Mg ha- --- Mg ha-1

Autumn (78) i 9.5 2.8 1.85 3.3 2.2 1.67 Winter (90) 1.9 2.7 1.75 -0.2 3.2 1.25 Spring (84) 14.7 2.7 1.64 9.7 3.3 0.86 Summer (88) 23.1 4.1 2.46 10.2 3.3 0.79



Test of Significance $ Yield Detritus S/R

Season ** NS NS Month (season) NS NS NS Reservoir ** NS **



Number of days in season. tSignificant at 0.01 level (**) or not significant (NS). S/R = Shoot/root dry wt ratio





31


Shoot and root lengths were similar for plants in both reservoirs during autumn and winter (Table 2). During spring, the water hyacinth root lengths were shorter and shoot lengths longer in the fertilized reservoir compared to the plants in the control reservoir. An interesting development in plant morphology was the dislodging of practically the entire root system from plants in the fertilized reservoir beginning in March after daily temperatures began to increase. The majority of plants were typified by a small root system compared to plants in the control reservoir. Some root dislodging was noticed in the control reservoir during spring and summer but was not as characteristically uniform as in the fertilized reservoir. Root lengths in the fertilized reservoir began to increase during summer, but the shoot lengths were double those in the control reservoir.

Under nutrient-limiting conditions, water hyacinths produce a large volume of root material presumably to increase their nutrient absorption capacity. With nutrient-enriched media, water hyacinth use more photosynthetic energy in shoot production. Cornwell et al. (1977) measured shoot lengths in excess of 1 m in wastewater media. Penfound and Earle (1948) recorded root lengths of 0.1 to 1 m. Maximum shoot length recorded during this study was 55 cm during summer for fertilized plants compared to 28 cm during summer for control plants.

The plant tissue N content is shown in Fig. 8. Fertilization

resulted in increases in shoot, root, and detritus N content compared to plants in the control reservoir. Maximum tissue N content for fertilized plants occurred during winter when plant productivity was low. The increase in plant productivity in spring and summer diluted the N content of the tissue although total N assimilation by the plants





32





Table 2. Seasonal water hyacinth shoot and root lengths.



Fertilized reservoir Control reservoir

Season Shoots Roots Shoots Roots

-------------------cm------------------Autumn (78)t 39.6 21.5 35.7 22.9 Winter (90) 25.2 14.2 23.3 20.2 Spring (84) 26.6 9.9 18.8 14.8 Summer (88) 54.9 21.3 27.8 27.1 Test of significance Shoot length Root length

Season ** ** Month (Season) NS ** Reservoir **



t Number of days in Season.
tSignificant at 0.05 (*) or 0.01 (**) level, or not significant
(NS).






33
























U)
1O 11 .


~ oLL





CL c

0
-C




z





ooA,- oz
eaa,


0 0 W- iN ) l
o 0 . -ve




(Io ) 0 ) 0 N 1N C) oI N- IC N t t ~s=~~ 0: :~ N- U) N
N - -





34


was much greater during this time period (Table 3). Plant tissue N content remained nearly consistent with time in the control reservoir and root tissue N content generally exceeded that of the shoot tissue (Fig. 8).

Seasonal N assimilation by water hyacinth ranged from 34 to 242 kg N ha-I for plants in the fertilizer reservoir and from <0 to 104 kg N h-1
ha-1 for plants in the control reservoir (Table 3). There were significant differences between seasons and reservoirs in water hyacinth N assimilation. The detritus N content was significantly greater for fertilized than control plants, but there were no significant differences in detritus N content between seasons.

Data on mass balance of N in both reservoirs are shown in Table 4. Nitrogen input from the lake was 238 kg N ha-I with 89% of the N in the organic fraction. Total amount of fertilizer applied during the study period was 781 kg N ha with NH -N, NO3-N, and organic N representing 55, 30, and 15% of total fertilizer applied, respectively.

The total N assimilated by water hyacinth (live plants and detritus) was 720 and 325 kg ha-1 yr- for fertilized and control reservoirs, respectively (Table 4). Annual net N loading by detritus was 148 and 92 kg ha-1 for fertilized and control reservoirs, respectively (Table 4). Maximum detritus N loading occurred during winter for the fertilized reservoir and during spring for the control reservoir. This corresponded to the time of root dislodging from plants in the two reservoirs.

The annual net N immobilized by detritus represented 21 and 28% of the total N removed by water hyacinth in the fertilized and control reservoirs, respectively. DeBusk et al. (1983) concluded that





35




Table 3. Seasonal nitrogen uptake by water hyacinth and detritus.



Fertilized reservoir Control reservoir Season Water hyacinth Detritus Water hyacinth Detritus

-------------------kg N ha----------------------1

Autumn (78) t 127.8 28.2 30.9 14.6 Winter (90) 33.9 42.4 -1.2 23.7 Spring (84) 167.7 37.5 104.3 29.1 Summer (88) 242.4 39.8 98.7 24.4



Test of significance $ Water hyacinth Detritus Season ** NS Month (season) NS Reservoir ** **



t Number of days in season.
$ Significant at 0.05 (*) or 0.01 (**) level or not significant (NS).






36


immobilization of N as plant detritus was 3 and 33% of standing crop assimilation for harvested and non-harvested water hyacinth plants, respectively. However, they did not include plant detritus trapped within the water hyacinth mat.

The annual N assimilation by water hyacinth is low compared to N removal rates reported for plants growing in nutrient-enriched waters. Reddy et al. (1985) found annual N removal rates of 1726 and 1193 kg N
-1 -1
ha yr for water hyacinths growing in primary and secondary sewage effluent, respectively. Rogers and Davis (1972) concluded that water
-1 -1
hyacinths could remove 2500 kg N ha yr if maximum growth could be sustained. Sato and Kondo (1981) measured a maximum removal rate of
-1 -1
4782 kg N ha yr for plants growing in a nutrient medium. The low annual N assimilation reported in the present study was due to low rates of fertilization.

Plant uptake played a major role in removing N in both the

reservoirs (Table 4). A large portion of lake water N was present as organic N, which was not readily available to plants. In both reservoirs, plants derived N from mineralization of lake water organic N, N release from underlying sediments, and mineralization of organic N in detritus. In the fertilized reservoirs, plants also derived N from the fertilizer N applied. Nitrogen assimiliation by water hyacinth from the added fertilizer was calculated as follows: (Total N assimilation by plants in the fertilized reservoir Total N assimilation by plants in the control reservoir / Total fertilizer N added) 100.

About 51% of the added fertilizer N was taken up by the plants in the fertilized reservoir, and the remaining 49% may have been lost through denitrification. Reddy et al. (1982) observed a reduction of 78





37



Table 4. Nitrogen balance for the two reservoirs.


Fertilized Control reservoir reservoir
-i
-----------kg ha-----------Nitrogen added
Fertilizer

NH -N 430 NO3-N 234 Organic N 117 -Total 781

Lake water

NH -N 13 13 NO3-N 12 12 Organic N 213 213

Total 238 238 Total added 1019 238 Nitrogen removed
Water hyacinth

Shoots 354 95 Roots 218 137 Detritus 148 92

Total 720 325

Reservoir water

NH+ -N 13 8

NO3-N 13 5 Organic N 167 156

Total 193 169 Total N accounted 913 494





38


to 81% of the agricultural drainage effluent NO3-N and NH -N in 3.6 days in a reservoir containing water hyacinths. DeBusk et al. (1983) calculated that 45% of the N removed from wastewater was immobilized in water hyacinth standing crop and detritus. About 30% of the fertilizer N was added as NO3-N, which could be potentially lost due to denitrification. Since water hyacinth plants prefer NH -N over NO3-N (Reddy and Tucker, 1983), the majority of the plant N uptake probably came from NH -N added through fertilizer. The role of underlying sediment in the immobilization/mineralization, and denitrification of N from these systems needs further investigation.

Total N recovery in the fertilized reservoir was about 90%, and plant uptake represented about 71% of total N inputs. In the control reservoir, total N recovery was higher than the N inputs. Plants removed 325 kg N ha as compared to 238 kg N ha added. Release of N from sediment or mineralization of N during decomposition of detritus may account for the higher N recovery compared to total N inputs. Ogwada (1983) found a yearly average of 150 + 34 kg KCl-extractable inorganic N ha-1 sediment using monthly sediment N concentrations of the same reservoirs.



Conclusions

Primary productivity of water hyacinths was influenced by ambient air temperature, solar radiation, and nutrient composition of the culture medium. Net detritus production (total detritus detritus lost through decomposition) was relatively constant throughout the year and represented 3.5 to 14.0% of the total standing crop. Detritus plant tissue of the fertilized reservoir contained higher tissue N, compared





39


to the detritus in the control reservoir. Fertilization and increases in ambient air temperature resulted in dislodging of root biomass.
-1 -1
Net N loading from detritus was 92 to 148 kg N ha yr which is potentially available upon decomposition. The N immobilized by detritus represented 21 and 28% of the total N removed by water hyacinths in the fertilized and control reservoirs, respectively.

Approximately 51% of the added fertilizer N was assimilated by

plants. The remaining 49% may have been lost through denitrification. Total N recovery was nearly 90% in the fertilized reservoir. More N was accounted for in the control reservoir than was added. Release of N from the sediment or mineralization of N during decomposition of detritus may account for the additional N recovery.















EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER HYACINTH SYSTEMS



Plant detritus (dead and decaying plant debris) is an integral part of water hyacinth mats and comprises 3 to 14% of the total biomass (see p. 39). It is usually derived from natural aging of plants, biological or chemical control, and frost damage. The addition of detritus to an aquatic system influenced several C and N transformations (Fenchel and Jorgenson, 1977).

Nitrogen is present as NH -N, NO3-N, and organic N in water media available for water hyacinth production. Organic N predominates in most water media and is not readily available for plant assimilation. Water hyacinths were efficient users of inorganic N and plant assimilation was a major process of N removal in aquatic systems containing water hyacinth (Reddy and Sutton, 1984). Other N transformations in aquatic systems resulting in removal of NO3-N or NH+-N include microbial assimilation, nitrification/denitrification, and NH3-N volatilization (Keeney, 1973; Bouldin et al., 1974). Addition of detritus significantly alters the rates of these processes.

Mineralization or immobilization of N occurs during decomposition of detritus in water and sediment. Decomposition of detritus and subsequent N release was found to be related to C/N ratio, initial N and fiber contents (De La Cruz and Gabriel, 1974; Godshalk and Wetzel, 1978b; Odum and Heywood, 1978; Ogwada et al., 1984).


40





41


A dense cover of floating water hyacinth depleted dissolved 02 of the underlying water, thus creating anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980; Reddy, 1981). Decomposition of plant detritus also consumed 02 (Nichols and Keeney, 1973; Rai and Munshi, 1979). Anaerobic conditions may restrict nitrification and favor denitrification, which may proceed within the water hyacinth mat, in the water column, or in the underlying sediment. Detritus also provides energy source for denitrification. Denitrification occurred primarily in the underlying sediment and the rate depended on NO3-N diffusion from the water column to the sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981).

Volatilization becomes increasingly important as the water pH

increases. The partial pressure of NH3-N in equilibrium with a solution of NH3-N increased rapidly in a pH range of 8.5 to 10.0 (Bouldin et al., 1974). A pH of 7.0 in water occurred in areas covered with water hyacinth with little diel variation (McDonald and Wolverton, 1980; Reddy, 1981) which suggests that NH3-N volatilization is minimal in areas covered with plants.

The relative role of N assimilation by water hyacinth on total N

removal from reservoirs was investigated by Reddy (1983). Approximately
15 + 15O40% of added NH -N or NO3-N was assimilated by plants. Less than 10% of the added 15N was found in the surface sediment layer. Over 40% of the added 15N was unaccounted for.

Information on the role of detritus in aquatic systems on

immobilization or mineralization of inorganic N is limited. The overall objective of this study was to determine the effect of detritus on selected N transformations in water columns with and without water





42


hyacinths. Specifically, the objectives were 1) to determine the regulatory function of detritus on dissolved 02 and pH of water and 2) 150
to determine the influence of detritus on the fate of 15N3 -N and 15NH-N in sediment- water-plant systems.



Materials and Methods

Two greenhouse studies were conducted to evaluate the effect of 15 +
detritus on the fate of N labeled NO3-N or NH -N in water with and without water hyacinth plants. Treatments evaluated were: 1) with and without underlying sediment, 2) with and without water hyacinth plant cover, and, 3) three rates of added water hyacinth detritus. There were 24 tanks in each study having dimensions of 50 cm 50 cm 25 cm depth. Twelve of the 24 tanks contained a 2.5 cm sediment layer (1.875 kg soil). The sediment was a Lauderhill organic soil (Lithic medisaprists) collected at the Central Florida REC research farm in Zellwood, Florida. The soil was air-dried and passed through a 2 mm sieve. Fifty liters of tap water were added to sediment tanks to obtain a 20 cm water depth.

The greenhouse studies were initiated after sediment/water

equilibration of 1 week. A nutrient medium (a modified 10% Hoagland's solution) was added to all tanks to obtain nutrient concentrations of: 15NH+N or 15NO3-N = 20.0 mg L- K = 23.5 mg L; PO4-P = 3.1 mg L-I Ca = 20.0 mg L-1; Mg = 4.8 mg L-I; SO4-S = 6.4 mg L1 ; Fe = 0.6 mg L1 and micronutrients. Micronutrients were applied through commercially available liquid fertilizer (Nutrispray-Sunniland, Chase and Co.,
-I
Sanford, Florida) to obtain concentrations of 0.2 mg Cu L -1; 1.5 mg Mn L- 0.04 mg B L; and 0.02 mg Mo L.





43


Water hyacinth detritus (shoot and root material) was added at the rates of 0, 100, and 400 mg C L-1. The detritus was chopped manually to 15
lengths of -2 cm. The detritus for treatments with added NO3-N was collected from a natural water hyacinth stand in Zellwood, Florida and
-1
had an initial N content of 5.6 mg g dry tissue. The detritus for 15 +
treatments with added NH -N was collected from a water hyacinth stand located in a wastewater stabilization pond at the University of Florida wastewater treatment plant in Gainesville, Florida and had an initial N
-1
content of 23.1 mg g-1 dry tissue.
-2
Water hyacinths, at an initial density of 10 kg (fresh wt) mwere placed in 12 of 24 tanks. The plants were collected from the University of Florida's Bivens Arm research reservoirs in Gainesville, Florida. The plants were clipped of dead tissue and rinsed with tap water prior to placement in tanks.

The disappearance of added inorganic N was determined by collecting water samples at 0, 1, 2, 3, 4, 8, 15, 28 days and measuring NH -N, NO3-N, and total Kjeldahl N (TKN). The changes in water hyacinth fresh weight were measured weekly. Plant samples and detritus were analyzed for TKN. The sediment was characterized for organic and inorganic N prior to and at the conclusion of each study. Fifty grams (dry wt) of moist sediment samples were extracted with 2 M KCl and analyzed for NH -N and NO3-N. Sediment samples were air-dried, ground by mortar and pestle, and analyzed for TKN. The inorganic N for all samples was determined by steam distillation (Keeney and Nelson, 1982). The TKN of water, plant, and sediment samples were determined by micro-Kjeldahl procedures (Nelson and Sommers, 1972; 1973; 1975). The 15N analyses on water, sediment, plant and detritus samples were conducted using a Micro Mass 602 spectrometer.





44



Water pH (Orion Model 404 Specific Ion Meter), dissolved 02 (Yellow Springs Instrument Model 54 02 meter) and temperature were measured every other day. Electrical conductivity (Hach Mini Conductivity Meter) was measured weekly.



Results and Discussion

Effect of Detritus on Water Dissolved 02

The dissolved 02 concentrations of water with added 15NO3-N and 15NH 4-N are shown in Figs. 9 and 10, respectively. Dissolved 02 concentrations remained < 5 mg LI in water having plant cover but lower dissolved 02 concentrations were recorded as the rate of detritus increased. This reflected increasing microbial 02 demand for respiratory functions with increasing C source (Fenchel and Jorgensen, 1977). For water without plant cover (open water), the dissolved 02 concentrations were scattered more with time. The dissolved 02 measurements were taken between 2:30 and 3:30 pm and should represent near maximum concentrations on a diurnal basis (Howeler, 1972). The increased dissolved 02 concentrations of open water were due to an increased rate of photosynthesis by algae during the day compared to respiration (Reddy, 1981).

Generally the effect of decreasing dissolved 02 concentrations with increasing detritus was seen for open water with or without sediment. Dissolved 02 concentrations were generally lower in open water with sediment compared to open water without sediment. Nichols and Keeney (1973) reported lower dissolved 02 concentrations for sediment-water systems than water only. Although detritus appeared to have a role in 02 dynamics in water, plant cover was the primary regulator.






45















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46


















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47


Effect of Detritus on Water pH
15 15 +
The pH of water with added 15NO3-N or NH4-N is shown in Figs. 11 and 12, respectively. A fairly constant pH of 7.0 was noted in water having plant cover and sediment regardless of detritus additions. The pH -decreased in water with plant cover but without sediment. The 15 +
decreasing pH was noted immediately for added 15NH-N and after 20 days for added 15NO3-N. The decrease in pH was less as the detritus rate increased.

The immediate pH decrease in water with plants and added NH -N was probably due to production of H+ during plant NH -N assimilation (Raven and Smith, 1976). The H+ generated is actively exuded, partly in exchange for cations (Franco and Munns, 1982). Plant NO3-N assimilation occurs by exchange with another anion or by simultaneous cation assimilation to maintain ion equilibrium (Kirkby and Mengel, 1967; Mengel, 1974). The decreasing pH in water with plant cover but without sediment suggested that the underlying sediment had a buffering role in pH regulation.

The pH of water without plant cover were generally higher and more variable than water with plant cover. Reddy (1981) reported high mid-day pH values in ponds where algal activity was high. Bouldin et al. (1974) found high pH values (> 8.5) for ponds containing submersed macrophytes during sunlight hours. The pH of open water was generally lower as the rate of detritus increased. Effect of Detritus on Nitrogen Loss
15 15 +
Nitrogen loss from water with added 15NO -N or NH -N is shown in Figs. 13 and 14, respectively. Sediment or detritus had no apparent






48















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52


effect on rate of N loss in water with plant cover. Nitrate and NH -N disappeared within 2 and 4 days, respectively, in water with water hyacinths.

Nitrogen loss in open water was influenced by the underlying

sediment and detritus additions. Nitrate disappeared more rapidly in open water with sediment than without sediment. An increase in detritus resulted in a more rapid NO3-N loss in water with or without sediment. A rapid decrease in NO3-N after 2 days was followed by an accumulation of NO3-N at 4 days for open water. Accumulation of NO3-N after 4 days in open water was probably due to rapid algal assimilation followed by turnover (death) of the algae and leaching of NO -N from the dead algal cells. Surface algal mats developed in open water within 2 days.

Ammonium disappeared more rapidly in open water with sediment than without sediment. Apparently detritus did not influence NH -N loss in open water with sediment. However, detritus additions resulted in a more rapid NH +-N loss in open water without sediment. Loss of NH -N
4 4
followed by accumulation of inorganic N during the first 4 days was not as striking as seen for NO3-N.

Plant Nitrogen Assimilation

Total plant N assimilation and the contribution of added 15NO -N
3
and NH4-N to total N assimilation is presented in Tables 5 and 6, respectively. Plant N assimilation was always greater for water with underlying sediment. Part of the increased plant N assimilation in water with sediment was due to release of N from the sediment.
15 15 +
Generally the contribution of added 15NO3-N or 15NH4-N to total plant N assimilation also decreased with increasing detritus. Mineralization of





53




Table 5. Total plant N and 15NO3-N assimilation.


Treatment Total uptake Labeled Other


--------mg (% of added 15N)-------Without sediment

0 mg C L-1 t 767 + 86 652 (85) 115 (15) 100 mg C L-1 1088 + 87 675 (62) 413 (38)
-1 2(1
400 mg C L-1 1027 + 268 606 (59) 421 (41) With sediment
-1 50(1
0 mg C L-1 1220 + 29 720 (59) 500 (41) 100 mg C L-1 1171 + 235 656 (56) 515 (44)
-1
400 mg C L-1 1112 + 163 567 (51) 545 (49)


tCarbon source was plant detritus. tOther N sources include sediment and detritus.





54



Table 6. Total plant N and 15NH-N assimilation.


Treatment Total uptake Labeled OtherS

--------mg (% of added 15N)-------Without sediment

0 mg C L-it 903 + 44 822 (91) 81 (9) 100 mg C L-1 768 + 103 697 (91) 71 (9) 400 mg C L-1 1133 + 81 807 (71) 326 (29) With sediment

0 mg C L-1 1390 + 194 891 (64) 499 (36) 100 mg C L-1 1255 + 89 845 (67) 410 (33)
-1 2 5 8 (9
400 mg C L-1 1629 + 65 828 (51) 801 (49) t Carbon source was plant detritus. SOther N sources include sediment and detritus.





55



detritus N was a potential N source for plant assimilation. Reddy (1983) found that 60 to 64% of total N assimilation by water hyacinths was derived from added 15N, while 36 to 40% was derived from sediment and from decomposition of detritus.

The 15N recovered by plant tissue (mg) was fairly consistent for 15 + 15
added labeled fertilizer but plant NH4-N uptake exceeded NO3-N uptake. Water hyacinth appeared to be more efficient in utilizing NH -N than NO3-N (Reddy and Tucker, 1983). When water hyacinth growth is not restricted by climate, rapid assimilation of added inorganic N would be expected.

Nitrogen-15 Balance for Water Columns

A 15N balance for water with added 15NO3-N and 15NH4-N is presented in Tables 7 and 8, respectively. Total 15N recovery by water hyacinths 15
ranged from 57 to 72% and 70 to 89% in water with added NO3-N and 15 +
NH -N, respectively. Reddy (1983) conluded that water hyacinth N 15 15 +
assimilation accounted for only 40% of added NO -N or NH -N in a
3 4
reservoir. Algal surface mats accounted for -8% of added 15NO3-N and up to 15% of added 15NH-N. The algal surface mats represented a minor portion of total microbial N assimilation. Algal activity was noted in open water, and the four sides and bottom of the microcosm tanks were colonized by algae.

The 15N associated with detritus was determined for water with added 15NH-N but not for added 15NO3-N. Less than 10% of the added
add3
15NH-N was immobilized by detritus in water with plant cover. Newly formed water hyacinth detritus from the plant cover was deposited during 28 days and accounted for 7% 15N recovery in water without added detritus or sediment.





56






Table 7. Mass balance of added 15NO3-N in sediment-water-plant systems.


Plant Sedimentt Unaccounted Treatment or algae org inorg Water Total For


----------------% Recovery of 15N------------------PLANTS
Without sediment

0 mg C L-it 65.2 --- --- ND 65.2 34.8 100 mg C L-1 67.5 --- --- ND 67.5 32.5
-1
400 mg C L-1 60.6 --- --- ND 60.6 39.4 With sediment

0 mg C L-1 72.0 3.2 0.4 ND 75.6 24.4
-1
100 mg C L 65.6 4.4 0.4 ND 70.4 29.6
-1
400 mg C L1 56.7 3.6 0.6 ND 59.9 40.1 NO PLANTS
Without sediment
-1
0 mg C L ND --- --- 6.2 6.2 93.8
-1
100 mg C L 6.8 --- --- ND 6.8 93.2
-1
400 mg C L 8.2 --- --- ND 8.2 91.8 With sediment
-1
0 mg C L-1 ND 11.4 0.4 5.1 16.9 83.1 100 mg C L-1 8.1 10.1 0.4 2.0 20.6 79.4

400 mg C L-1 8.4 4.9 0.5 0.5 14.3 85.7


t Carbon source was plant detritus. IOrg, Inorg = organic and inorganic N, respectively. ND = Not detectable.





57







15 +
Table 8. Mass balance of added NH -N in sediment-water-plant systems.


Plant Sediment-t Unaccounted Treatment or algae Detritus Org Inorg Water Total For


-----------------------% Recovery of 15N-------------------PLANTS
Without sediment

0 mg C L 82.2 7.3 --- --- 2.3 91.8 8.2
-1
100 mg C L 69.6 3.6 --- --- 5.8 79.0 21.0

400 mg C L-1 80.7 7.1 --- --- 3.8 91.6 8.4

With sediment
-1 F
0 mg C L-1 89.1 ND 4.3 2.6 1.0 97.0 3.0 100 mg C L-1 84.5 1.7 3.0 2.9 1.5 93.6 6.4
-I
400 mg C L-1 82.8 3.9 3.3 3.5 1.5 95.0 5.0 NO PLANTS
Without sediment
-1
0 mg C L-1 ND ND --- --- 9.6 9.6 90.4
-1
100 mg C L-1 ND 14.7 --- --- 17.5 32.2 67.8
-1
400 mg C L-1 6.1 34.7 --- --- 5.2 46.0 54.0

With sediment
-1
0 mg C L 14.9 ND 10.2 7.1 2.3 34.5 65.5
-i
100 mg C L 15.0 5.9 11.0 8.8 5.8 46.5 53.5 400 mg C L-1 11.1 26.2 3.9 6.9 3.8 51. 9 48.1


1Carbon source was plant detritus. Org, Inorg = organic and inorganic N, respectively. ND = Not detectable.





58



Detritus 15N recovery in water without plant cover increased with increasing rate of detritus. This suggests that during periods of low water hyacinth productivity, i.e. winter, detritus will be an important sink for inorganic N removal. The high 15N recovery in detritus was
-1
suprising since the original detritus had a high N content (23 mg g dry tissue). Therefore, the detritus used in water with added 15NO3-N 15
probably accounted for even more N immobilization due to a low initial N content (5 mg g-I dry tissue).

Generally 15N recovery in sediment was primarily organic N. Less than 1% of the added 15NO3-N was recovered as sediment inorganic N.
15 +
However, between 3 and 7% of the added NH -N was recovered as KC1 extractable inorganic N in the sediment. The lower recovery of sediment inorganic 15N in water with added 15NO3-N was probably due to reduction to gaseous N via denitrification (Engler and Patrick, 1973). Some of
15
the added 15N was recovered as organic N in the water.

Plant uptake was the primary mechanism of N removal in water having water hyacinths. The 15N unaccounted for was lost from the systems through a variety of possible transformations. A more thorough investigation would be required to establish the extent of algal N assimilation. Volatilization of NH3-N in water without plant cover and denitrification in water with sediment are two possible mechanisms for N removal.



Conclusions

Generally as the rate of detritus addition increased, dissolved 02 concentrations decreased in water with or without sediment and with or without plant cover. The decreasing dissolved 02 concentrations were





59



attributed to increasing heterotrophic respiration due to increasing amounts of C. Although this general relationship existed for all treatments, plant cover and sediment layer appeared to have more of a regulatory role in dissolved 02 dynamics than detritus.

Water pH was constant in water having plant cover and sediment. The decreasing pH of water with plant cover and no sediment was attributed to NH -N assimilation by plants in exchange for H +. The pH of open water was generally lower as the rate of detritus increased.

Detritus had no apparent effect on rate of N loss in water with water hyacinths. However, N loss was more rapid in open water as the rate of detritus increased.

Total plant 15NH -N uptake exceeded 15NO -N uptake. Both sediment
4 3
and detritus appeared to be a potential N source for water hyacinths. Total 15N recovered by water hyacinths ranged from 57 to 72% for added 15 15 +
NO-N and 70 to 89% for added NH -N.
15 +
Less than 10% of the added NH -N was immobilized by detritus in water with plant cover. However, in water without plant cover, up to 35% of the added 15NH -N was associated with detritus. This suggests that during periods of low water hyacinth productivity, typical in cold weather conditions, detritus is an important sink for added N.














ANAEROBIC DIGESTION OF WATER HYACINTH



The potential productivity of water hyacinth has led to its

selection as a biomass feedstock for methane generation while providing a means for treatment of nutrient-enriched waters. Methane yields during anaerobic digestion depended on characteristics of the feedstock (Stack et al., 1978; Wolverton and McDonald, 1981) as well as digester operating conditions (Hashimoto et al., 1980). Sievers and Brune (1978) reported higher methane yields for digesters operating on swine waste as the C/N ratio increased. They concluded that the optimum C/N range for maximum methane production was 15.5/1 to 19/1. The optimum pH and temperature range for anaerobic digestion was 6.7 to 7.4 (Bryant, 1979) and 30 to 350C (House, 1981), respectively.

Biogas and methane yields have been reported for water hyacinths using a variety of digesters. Wolverton and McDonald (1981) reported
-1
methane yields of 0.07 to 0.20 L g-1 total solids (TS) for blended water hyacinths. Hanisak et al. (1980) found average methane yields of 0.24 L
-1
g volatile solids (VS) from shredded water hyacinths in 162 L digesters at loading rates of 1.10 to 1.38 g VS L day and residence times of 30 to 38 days.

Chynoweth et al. (1983) reported methane yields of 0.19 and 0.28 L
-1
g-1 VS of water hyacinth and a 3:1 water hyacinth/primary sewage sludge blend, respectively, in 5 L daily-fed digesters with a loading rate of


60





61


1.6 g VS L-1 day-. Shiralipour and Smith (1984) reported average
-1
methane yields of 0.32 and 0.17 L g-1 VS water hyacinth shoot and root samples, respectively, in a bioassay test of 100 ml culture volume. They also concluded that the addition of N in growth media for water hyacinth production increased methane yields of both shoot and root samples.

Inoculum from operating anaerobic digesters is commonly added as a bacterial seed to initiate anaerobic digestion in new digesters (Sievers and Brune, 1978; Wolverton and McDonald, 1981; Field et al., 1984). Information on the effect of inoculum volume on gas production is limited.

The objectives of this study were 1) to determine C and N

mineralization during anaerobic digestion of water hyacinth; 2) to determine the effect of inoculum volume on gas production; and 3) to evaluate effluent (solids and liquid) composition based on inoculum volume.



Materials and Methods

Water hyacinths, with either high or low tissue N content, were

anaerobically digested at 350C in 55 L batch digesters containing 2.5, 5, or 10 L of inoculum. Water hyacinths with a high N content (-34 g kg-1 dry wt plant tissue) were obtained from the wastewater treatment plant of the Reedy Creek Utility Company, Inc., at Walt Disney World near Orlando, Florida. Water hyacinths with a low N content (-10 g kg-I dry wt plant tissue) were grown in nutrient-depleted water at Sanford, Florida. Both types of hyacinths were grown in 15N labeled (NH 4)2S4 for two weeks, frozen and chopped to 1.6 mm length using a Hobart T 215 food processor.





62


The digesters received 4.7 kg fresh weight of the 15N labeled water hyacinths and an inoculum volume of 2.5, 5 or 10 L. A control digester received 10 L of inoculum and no plant material. The inoculum used for plants with high N content was obtained from an operating continuous-fed upflow digester receiving a feedstock of water hyacinth and domestic sewage sludge in a blend ratio of 3:1 (Chynoweth et al., 1983). The inoculum used for the plants with low N content was obtained from a non-operating continuously-fed tank digester receiving water hyacinth as feedstock. Each digester was buffered with 210 g NaHCO3 and tap water was used to bring each batch digester to 54.7 kg.

Gas production was monitored for 60 days. At the end of the digestion period, each digester was thoroughly mixed and the total contents were emptied into a 60 L tub. The digested materials were passed through a 1.00 mm fiberglass screen into a second 60 L tub to separate the digested biomass sludge from the effluent. The sludge was drained for 7 minutes and transferred into a polyethylene bag and placed directly into a freezer. The effluent was transferred to a water hyacinth production system.

The liquid samples from the digester effluents and screened

effluents (sludge removed) were analyzed for pH, electrical conductivity

(EC), total solids (TS), fixed solids (FS), volatile solids (VS) (APHA, 1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH -N and NO3-N by steam distillation (Keeney and Nelson, 1982), and chemical oxygen demand (COD) (APHA, 1980). The screened effluent was also filtered through a 0.2 um membrane filter and analyzed for Ca, K, Na and Mg by atomic absorption and P by an autoanalyzer.





63


The fresh plant material and digested sludge were freeze-dried

(Thermovac-T) and analyzed for the following: TS, FS, VS, TKN (Nelson and Sommers, 1973), total carbon (TC) (LECO Induction Furnace 523-300), lignin, cellulose and hemicellulose (Goering and Van Soest, 1970), and ashed mineral constituents (Gaines and Mitchell, 1979).



Results and Discussion

Characteristics of Inocula

Characteristics of the two inocula varied considerably (Table 9). The inoculum used for plants with high N content (high N plants) contained higher levels of TS, VS, NH -N, TKN, and COD than the inoculum used for plants with low N content (low N plants). Characteristics of inoculum depended on the type of feedstock used for digestion (Stack et al., 1978) as well as digester operating conditions (Hashimoto et al., 1980). The inoculum used for high and low N plants came from digesters with feedstocks of a 3:1 water hyacinth/domestic sewage sludge blend and water hyacinths, respectively. Ammonium accounted for 68 and 92% of the total N of the inoculum from the water hyacinth/sewage sludge and water hyacinth feedstocks, respectively. Ammonium was the primary N source for methanogenic bacteria (Zeikus, 1977). Carbon and Nitrogen Mineralization During Digestion

Biogas (CH4 and CO2) production, corrected to standard conditions (00C and 0.1 MPa), is given in Table 10. Gas production essentially ceased after 60 days of digestion. Cumulative biogas production at 60 days for high N plants was approximately 21% less for 2.5 L of inoculum compared to 10 L. Furthermore, cumulative biogas production at 15 days





64







Table 9. Characteristics of the inocula used in the batch digesters.





Inocula characteristics t

NH -N NO3-N TKN COD pH TS FS VS


-i
-------- mg L1-----------------%-- ---% of TS--High N plant material

1072 48 1530 14200 6.3 1.75 33.5 66.5 Low N plant material

535 21 562 784 7.7 0.29 83.1 16.9



tCOD = Chemical oxygen demand, TS, FS, and VS = Total, fixed and volatile solids, respectively.











Table 10. Gas production during anaerobic digestion of high
and low N water hyacinth plants.


Inoculum Cumulative biogas production Total Gas Yields .volume 15 days 30 days 60 days biogas methane

--L-- ---------Liters-------------- ---L g-1 VS---High N plant material

2.5 16.4 40.8 60.3 0.21 0.14

5 28.1 53.5 73.0 0.23 0.15 10 34.5 59.1 75.4 0.20 0.13 Low N plant material

2.5 16.9 45.5 67.9 0.25 0.16 5 20.0 51.6 72.6 0.27 0.17 10 14.7 52.2 67.4 0.25 0.16




66



was over twice as great for the digester receiving the largest amount of inoculum. However, for low N plants, the amount of inoculum did not appreciably affect cumulative biogas production during digestion.

Cumulative biogas production at 60 days was similar for both high and low N plants. Biogas production at 15 days was generally greater for high N plants. It appeared that N was not a limiting factor for total gas production in either digestion test.

Converting 60 day biogas production to biogas or methane yields (L g-1 VS added) is also presented in Table 10. Volatile solids included inputs from water hyacinths and inoculum. The average methane content of the biogas was 63.7 + 5.2 % based on 18 samples. Surprisingly, biogas and methane yields were higher for the low N plants. This was caused by an increase of VS from inoculum used in digesters for high N plants. The inoculum used for high N plants contained 1.75% TS (66.5% VS of TS) (Table 9). The inoculum for low N plants contained 0.29% TS (16.9% VS of TS). Gas production expressed in these units suggested that inoculum volume did not appreciably affect total biogas or methane yields.
-l
The average methane yields were 0.14 and 0.16 L g-1 VS added for high and low N water hyacinth plants, respectively. The methane yields were lower than those reported for continuously-fed digesters (Hanisak et al., 1980; Chynoweth et al., 1983). Batch digestion (once fed and sealed) would not promote maximum gas yields as frequent addition of fresh substrate enhances gas production (Price and Cheremisinoff, 1981).

Shiralipour and Smith (1984) reported that methane production for water hyacinth roots was lower than for shoots and that increasing N in water hyacinth growth media increased methane yields. Water hyacinths typically produced longer roots as water fertility declined (see p. 32).





67


Shoot:root dry weight ratios of water hyacinth were higher when nutrients were not limiting and decreased significantly when plants grew in nutrient-poor waters (Reddy, 1984). It was assumed in the present study that gas production, both cumulative and yields, would be greater for the high N plants.

A mass balance of N is presented in Table 11. The organic N content decreased after anaerobic digestion for each treatment. Mineralization of organic N to NH -N was the primary N transformation occurring during digestion. The total N recovered was lower for low N water hyacinth plants. The majority of the N was recovered in the effluent as NH -N. Most of the N placed in digesters was recovered in
+
the effluent, although the proportion of NH -N of the total N tended to increase (Hashimoto et al., 1980; Field et al., 1984).

Approximately 30% of the organic N placed in the digesters was

recovered as organic N in the digested sludge for both high and low N plants. The organic N recovered in the screened effluent was 15 and 36% of the added organic N for high and low N plants, respectively. The total organic N recovered as effluent or sludge organic N was 45 and 66% of added organic N for high and low N plants, respectively. Therefore, a high N content of water hyacinth resulted in more mineralization of added organic N.
15 15 +
Total 15N recovered as NH -N in the screened effluent was 72 + 4% for high N plants compared to 35 + 9% for low N plants (Table 12). The organic 15N recovered in digested sludge accounted for 20 + 5% of the added 15N from fresh water hyacinth plants regardless of N content. Approximately 11 and 20% of the added 15N was recovered as organic N in the screened effluent for digested high and low N plants, respectively.





68




Table 11. Nitrogen balance for the batch digesters.

High N plant material Low N plant material

2.5 Li 5 L 10 L 2.5 L5 L 10 L
--------------------- g -------------------------Nitrogen added
Water hyacinth
Organic N 10.39 10.39 10.39 3.24 3.24 3.24 Inoculum
Organic N 1.15 2.31 4.61 0.07 0.14 0.27 Inorganic N 2.68 5.36 10.72 1.34 2.68 5.35 Total
Organic N 11.54 12.69 14.99 3.31 3.37 3.51 Inorganic N 2.68 5.36 10.72 1.34 2.68 5.35 14.22 18.05 25.72 4.64 6.05 8.86

Nitrogen recovered
Screened effluent
Organic N 1.48 2.41 2.02 1.26 1.31 1.09 Inorganic N 8.81 11.60 15.81 1.20 2.63 4.98 Digested sludge
Organic N 2.86 4.91 4.94 1.17 0.85 0.87 Total
Organic N 4.34 7.32 6.96 2.43 2.16 1.96 Inorganic N 8.81 11.60 15.81 1.20 2.63 4.98 13.15 18.91 22.77 3.63 4.79 6.94


% Recovered 92 105 89 78 79 78


tVolume (liters) of inoculum.





69




Table 12. Nitrogen-15 balance for the batch digesters.

High N plant material Low N plant material

2.5 Lt 5 L 10 L 2.5 L 5 L 10 L

--------------------- g --------------------15N Added
Water hyacinth
Organic N 10.39 10.39 10.39 3.24 3.24 3.24 N Recovered
Screened effluent
Organic N 1.18 1.48 0.93 0.84 0.65 0.34 Inorganic N 7.06 7.78 7.68 0.79 1.25 1.34
Digested Sludge
Organic N 1.67 2.53 2.21 0.87 0.57 0.52 Total, 9.91 11.79 10.82 2.50 2.46 2.20




% Recovered 95 113 104 77 76 68

Volume (liters) of inoculum.





70


The low total recovery of N and 15N for low N plants after

anaerobic digestion is difficult to explain. Nitrogen cycling during anaerobic digestion was primarily mineralization of organic N or immobilization of inorganic N. Volatilization of NH3-N may occur but the potential increases as NH -N concentrations increase or at higher pH values (Freney et al., 1983). Each digester received 210 g NaHCO3 as a buffer and the pH after digestion was similar for all digester effluents. The NH -N concentrations after digestion were much higher for the high N plants (Table 13).

Effluent composition

Characteristics of the digester effluents prior to sludge separation are presented in Table 13. Generally, as the rate of inoculum increased, there were increases in EC, NH -N, TKN, and TS. The COD increased with increasing inoculum volume for digesters with high N plants.

Characteristics of the screened effluent (sludge removed) are

reported in Table 14. Removing the digested biomass sludge from the digester effluents decreased the EC, NH -N and TKN. The screened effluent from the low N plants contained more Ca, Mg, K, and Na, and less P than the screened effluent from the high N plants.

Characteristics of the fresh plant biomass and digested biomass

sludge are given in Table 15. Anaerobic digestion resulted in increases in TC and TKN of sludge compared to fresh plant biomass. The increases in sludge TKN after digestion of low N plants caused a reduction of the C/N ratio from 35 to 16. The changes in TC or TKN of the digested high N plants did not appreciably alter the C/N ratio. The digested sludge




71





Table 13. Characteristics of digester effluents before sludge removal. Inoculum Digester Effluent Characteristicst volume pH EC COD NH -N TKN TS VS FS


--L-- dS ------ mg L- ------ % -% of TSHigh N plant material

2.5 7.4 4.5 3030 189 238 0.498 40.3 59.7

5 7.6 5.1 4290 210 315 0.570 38.7 61.3 10 7.4 5.6 5050 294 406 0.610 38.9 61.1 Control 7.7 NA$ 4170 205 259 0.535 36.6 63.4 Low N plant material

2.5 7.3 6.6 1690 50 70 0.445 28.8 71.1 5 7.2 6.9 2340 70 112 NA 37.3 62.7 10 7.3 7.8 1640 112 154 0.488 28.8 71.2 Control 7.8 NA 109 98 120 0.315 14.0 86.0



COD = Chemical oxygen demand, TS, FS and VS = Total, fixed and volatile solids, respectively.
tNA = Not available.







72













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74


had a higher lignin content compared to fresh-plant biomass. The increase in lignin was due to the loss of readily decomposable C during anaerobic digestion. Lignin appears to be practically inert to anaerobic digestion (Hashimoto et al., 1980) Generally, there was a decrease in cellulose after digestion. The hemicellulose remained similar for the fresh plant biomass and digested sludge. Anaerobic digestion resulted in losses of K and Mg from fresh plant biomass, but increased the concentration of sludge Ca, Na, Fe, and Zn.



Conclusions

Cumulative biogas production at 60 days was similar for high (-34 g N kg-1 dry wt tissue) and low (-10 g N kg- ) N plants suggesting that long term digestion of water hyacinth was not influenced by initial N content. Effects of inoculum volume on cumulative biogas production were seen at 15 days for high N plants but not low N plants. Conversion of cumulative biogas production into biogas and methane yields (L g-1 VS added) showed that low N plants produced more biogas and methane than high N plants. This was due to increase of TS (and consequently VS) in digesters of high N plants from the inoculum source, since cumulative gas production was similar for both types of plants.
15 15 +
Mineralization of organic N to NH -N accounted for 72 and 35% of
15
added N for high and low N plants, respectively. Approximately 20% of the added 15N was recovered as organic N in sludge for both types of plants. A low 15N recovery was observed for low N plants.

Increasing inoculum volume increased electrical conductivity, NH -N, TKN, and TS of the digester effluents. The digested biomass sludge had higher levels of TC, TKN, lignin, Ca, Na, Fe, and Zn, and lower levels





75


of K and Mg compared to the fresh plant biomass. The C/N ratio of the fresh plant biomass with a low tissue N content decreased from 35 to 16 after digestion. The C/N ratio of the fresh plant biomass with a high tissue N content was the same as the digested biomass sludge (C/N=12).














TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTHS



An integrated approach of wastewater renovation using aquatic macrophytes with utilization of biomass for energy production is economically appealing. The plant biomass produced in these systems, along with other wastes such as sewage sludge or animal waste could be anaerobically digested to produce methane (Stack et al., 1981; Shiralipour and Smith, 1984). This process generates a waste by-product which must be disposed of, or preferably utilized to reduce the cost of energy production, in an environmentally-safe manner. The waste by-product consists of digested sludge and a large volume of effluent. Integrating wastewater renovation through water hyacinth production provides an internal option for the disposal of effluent generated during conversion of biomass into methane.

The effluent composition of anaerobic digesters varied with type of feedstock used in digestion (Stack et al., 1981). Information on chemical composition of effluents from sewage or animal wastes was readily available (Sommers, 1977; Field et al., 1984). However, anaerobic digestion of plant biomass has only recently gained attention in the United States and information on composition or disposal of the effluent was limited (Hanisak et al., 1980; Atalay and Blanchar, 1984).

Digester effluents have high concentrations of BOD, NH -N, K and Na (Atalay and Blanchar, 1984; Field et al., 1984), while the divalent cations and metals were concentrated in the sludge (Sommers, 1977; Field et al., 1984).

76





77


Water hyacinth-based wastewater treatment systems have already been evaluated for use in treating primary and secondary sewage effluents (Wolverton and McDonald, 1979; Reddy et al., 1985) and anaerobic digester effluent (Hanisak et al., 1980). The potential productivity of water hyacinth in nutrient-enriched waters has led to its selection in alternative methods of wastewater renovation, particularly in areas where growth is not restricted by climatic limitations.

Use of water hyacinth for digester effluent treatment is

particularly attractive, because of its ability to grow in waters with high elemental concentrations. The biomass produced could be returned to the digester as a feedstock for methane production. Hanisak et al. (1980) determined that 64.5% of (liquid and sludge) N in diluted effluents from anaerobically digested water hyacinth could be reassimilated by water hyacinths. Diluting the effluent does not address the full potential of water hyacinth to grow under these nutrient and salt enriched conditions. Haller et al. (1974) concluded that water hyacinth will not live in waters containing sustained salt concentrations in excess of 2500 mg L-1. Optimal dilution of these concentrated effluents to obtain maximum water hyacinth yields and nutrient removal was not reported.

The objectives of this study were to 1) evaluate water hyacinth productivity in anaerobic digester effluents obtained from digesters receiving different types of water hyacinth as feedstock, and, 2) determine 15N recovery by water hyacinth growing in digester effluents from digested 15N labeled water hyacinth biomass.





78


Materials and Methods

Anaerobic digester effluents were obtained from six 55 L batch

digesters containing water hyacinth with a high or low tissue N content as feedstock. Water hyacinths with low (-10 g N kg-1 dry plant tissue) and high (-34 g N kg-1) tissue N content were grown in nutrient-depleted water and sewage effluent, respectively. After removal from their respective growth media, the hyacinths were grown in 15N labeled (NH4)2SO4 nutrient solution for two weeks, frozen, and chopped to 1.6 mm length using a Hobart T 215 food processor.

The water hyacinths were anaerobically digested for four months in 55 L batch digesters. Each digester received 4.7 kg (fresh weight) of the 15N labeled water hyacinth, 2.5, 5, or 10 L volume of inoculum from anaerobic digesters receiving water hyacinth as feedstock, and were buffered with 210 g NaHCO3. After digestion, the biomass sludge was separated from the effluent by passing the total contents of the digesters through a 1.00 mm fiberglass screen.

The screened effluents (sludge removed) were analyzed for total

solids (TS), volatile solids (VS), fixed solids (FS) (APHA, 1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH -N and NO3-N
4 by steam
distillation (Keeney and Nelson, 1982), electrical conductivity (EC) (Hach Mini Conductivity Meter) and pH (Orion Model 404 Specific Ion Meter). Samples passed through a 0.2 pm membrane filter were analyzed for Na, K, Mg and Ca by atomic absorption and ortho P colorimetrically after reacting with ammonium molybdate.

Six water hyacinth plants were placed in 10 L of undiluted or

diluted effluents in containers having a surface area of 0.051 m2. The water hyacinth plants were collected from the University of Florida's





79


Bivens Arm research reservoirs in Gainesville, Florida. The plants were clipped of dead tissue and acclimated to greenhouse conditions for two weeks prior to treatments. The studies to evaluate the potential of water hyacinth to treat digester effluents were conducted for a period of 22 days in March and September, 1984. The daily maximum greenhouse temperatures ranged from 19 to 37 C in March and 27 to 36 C in September.

There were a total of 10 treatments, replicated three times, as described below. Treatments 1 to 3 were diluted effluents from digesters containing high N water hyacinths and inoculum volumes of 2.5, '5, and 10 L, respectively. The dilutions were 1:8, 1:4, and 1:3 effluent : tap water for digester effluents with inoculum volumes of

2.5, 5, and 10 L, respectively. Treatments 4 to 6 were undiluted effluents from digesters containing high N plants and inoculum volumes of 2.5, 5, and 10 L, respectively. Treatments 7 to 9 were undiluted effluents from digesters containing low N plants and inoculum volumes of 2.5, 5, and 10 L, respectively. The final treatment was a modified 10% Hoagland's solution (p. 42) to serve as a control. Characteristics of the inoculum and effluents are given in the Anaerobic Digestion of Water Hyacinth chapter (pp. 60 to 75).

Plant samples collected initially and at the conclusion of the experiment were analyzed for dry weights, TKN (Nelson and Sommers, 1973), Na, K, Ca and Mg by atomic absorption and P by an autoanalyzer. Water samples were collected at 0, 1, 2, 3, 4, 8, 15, and 22 days and analyzed for NH -N and NO3-N by steam distillation (Keeney and Nelson, 1982), TKN (Nelson and Sommers, 1975), Ca, Mg, K, Na by atomic absorption and P by an autoanalyzer. Electrical conductivity (Hach Mini





80


Conductivity Meter), pH (Orion Model 404 Specific Ion Meter) and dissolved 02 (Yellow Springs Instrument Model 54 02 Meter) were measured 15
every other day. The N analyses of plant and water samples were conducted on a Micro Mass 602 spectrometer.



Results and Discussion

Chemical Composition of the Effluents

Characteristics of digester effluents used in the study are given

in Table 16. The initial pH of the effluent sources and nutrient medium were similar. The digester effluents had a wide range of EC, NH -N, TKN and other nutrients. This provided an opportunity to evaluate water hyacinth growth under diverse media conditions. The EC of the nutrient
-l
medium and diluted effluents ranged from 0.7 to 2.3 dS m-. The EC of
-i
the undiluted effluents ranged from 4.3 to 6.7 dS m-1

The NH -N and TKN concentrations of the undiluted effluents from

digested plants with a high N content (high N plants) were greater than those of the undiluted effluents of digested plants with a low N content (low N plants). The NH -N concentrations ranged from 23 to 104 mg L-1
-1
for diluted effluents and 24 to 289 mg L-1 for undiluted effluents, respectively.

High Na and K concentrations were noted for undiluted effluents. The highest levels of P were in undiluted effluents of digested high N plants and the highest levels of Ca and Mg were in undiluted effluents of digested low N plants. The critical levels of Na, K, Ca and Mg needed to achieve maximum water hyacinth yields are relatively unknown. The NH -N, K and Mg concentrations were higher than levels reported for water hyacinths cultured in primary or secondary sewage effluent (Reddy et al., 1985).






81






















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Productivity of Water Hyacinths

Total dry weight gains of water hyacinths were consistantly less in the undiluted effluents (Fig 15). Complete death of plants was observed in four undiluted effluents. The loss of dry weight for these treatments was probably due to leaching of soluble plant constituents after plant death.

The highest dry weight gains were associated with the lowest EC.

However, plants survived in undiluted effluents having EC levels of 5.6 and 5.9 dS m-1 (5600 and 5900 pmhos cm-1 ). These EC levels were
-1
equivalent to -2900 and 3200 mg NaCl L- and were higher salt concentrations reported for water hyacinth survival (Penfound and Earle, 1948; Haller et al., 1974). The undiluted effluents had Na levels in excess of 1100 mg L-1. Apparently water hyacinth has a wide range of adaptability to media composition and, therefore, total salt concentration is not a good criterium for determining plant survival.

The diluted effluents were excellent media for plant growth and the gains in dry weight were consistently higher compared to the nutrient medium (Fig. 15). The highest dry weight gain was in the diluted effluent having N and P concentrations of 65 and 5.5 mg L-1 respectively, and a N/P ratio of 11.8:1. Sato and Kondo (1981) reported maximum yields of water hyacinth at a N and P concentration of 50 and 13.8 mg L-1, respectively, and a N/P ratio of 3.7:1. Dry weight gains were noted for two of the undiluted effluents. However, tissue damage was noted in all undiluted effluents.

Tissue damage in undiluted effluents was observed within 24 hr

after study initiation. Two types of leaf tissue damage were observed. Damaged leaves on younger shoots had burnt (brown) tips which curled up





83















4@
CE)




-00
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o 0 0 0 N I (c. I) NIVO W IJ-IM AI





84


towards the center of the plant.- The other leaf tissue damage on young and older leaves was curling of the entire leaf towards the center of the leaf. Stem damage in undiluted effluents was noticeable after 2 days. Damaged stems collasped under slight manual pressure. The destruction of chlorophyll in stems and leaves was widely observed in the undiluted effluents.

The extent and spread of tissue damage increased in undiluted

effluents with increasing EC and NH -N concentrations. At the end of one week, all plants in four undiluted effluents were dead. The EC and NH -N concentrations of these effluents ranged from 4.3 to 6.7 dS m-1 and 87 to 289 mg N L-1, respectively. The shoots began to separate from the roots at the water surface and the submerged roots sank. Root separation also accounted for the negative dry weight gains of plants in undiluted effluents.

Three treatments showed occasional visual signs of tissue damage, i.e., the nutrient medium and 2 diluted effluents. Visual signs of plant damage but noticeable gains in plant dry weights were observed in the diluted effluent having an initial NH -N concentration of 104 mg L-1 and two undiluted effluents having NH -N concentrations of 24 and 49 mg
-l
L-1. All remaining treatments resulted in plant death, apparently due to high EC or high NH -N concentrations, or a combination of both.

Although water hyacinth plants struggled to survive in the

undiluted effluents, algal activity was noted in all undiluted effluent treatments. Upon emptying the containers, algae were found attached to the side and bottom surfaces of the plastic containers.





85


Nitrogen Removal

First-order kinetic equations were used to described NH -N loss from the effluents. An integrated rate equation for a first-order reaction is expressed as:

In C / Ct = kt
o t
where C = initial NH -N concentration in the effluent,
o 4
Ct = final concentration at time = t,

k = first order rate constant (days-l).

The rate constant is calculated by solving for k: k = In C / Ct 1/t
o
The rate constants for diluted effluents ranged from 0.228 to 0.593 day-i (Table 17). The rate constants for undiluted effluents ranged
-1
from 0.175 to 0.446 day-1

The time required for a 50% reduction in initial NH -N

concentration generally increased with decreasing dry weight gains and increasing NH -N concentrations. The shortest reduction times were associated with rapid plant growth (50% reduction in 1.12 to 3.04 days). Plant assimilation was probably the primary mechanism of NH -N loss in these treatments. Reddy (1983) reported a 50% reduction of inorganic N from agricultural drainage water in 18 days. The time required for a 50% reduction of NH -N in effluents resulting in plant death was 1.98 to
4
3.96 days. Mechanisms of NH -N loss in treatments resulting in plant death included microbial assimilation and NH3-N volatilization. The pH of the effluents ranged from 7.0 to 7.9 in treatments with actively growing plants but increased from 8.2 to 9.3 in treatments where plant death was observed and algal growth increased (APPENDIX A, Table 27). The potential of NH3-N volatilization increases as NH -N concentrations increase or at higher pH values (Freney et al., 1983).





86








Table 17. First-order kinetic descriptions of NH -N
loss with time.

Ini ial
Inoculum NH -N Reduction
volume conc. k time R2

--L-- mg L-1 day-1 days

Diluted effluents from high N plants

2.5 23 0.593 1.12 0.722 5 65 0.449 1.54 0.917 10 104 0.228 3.04 0.951

Undiluted effluents from high N plants

2.5 161 0.232 2.98 0.938 5 212 0.207 3.35 0.947 10 289 0.175 3.96 0.982

Undiluted effluents from low N plants

2.5 24 0.446 1.55 0.850 5 49 0.325 2.13 0.885 10 87 0.350 1.98 0.845 Nutrient medium

20 0.281 2.47 0.973


t Time required for 50% reduction in initial NH -N concentration.





87


Clock (1968) reported a 75% reduction of NO3-N in 5 days for water hyacinths growing in secondary sewage effluent. A 75% reduction of NH -N in digester effluents required 2.3 to 6 days for treatments where positive dry weight gains were observed. For treatments resulting in plant death, 4 to 8 days were required to remove 75% of the NH4-N.

Nitrogen-15 plant assimilation was observed for all treatments

although a low recovery was observed in treatments resulting in plant death (2 to 16%) (Table 18). The 15N recovery by plants for the other treatments ranged from 36 to 77%. The majority of the 15N was found in the shoot material for all treatments (54 to 73%). Approximately 75% of
15
the N was unaccounted for in treatments resulting in plant death. Microbial assimilation and NH3-N volatilization were probably important NH -N removal processes in undiluted effluents where plant death was observed.

Plant Tissue Chemical Composition

Plants survived in the diluted effluents but death was noted for plants in the undiluted effluents from digested high N plants. Mineral constituents from plants growing in these effluents were analyzed to isolate individual cation and P assimilation or loss from living and dead plant tissue.

The concentrations of plant tissue (root and shoot fractions) Na,

K, P, Ca and Mg are reported in Table 19. The original plant tissue had low concentrations of Na and P in both shoot and root material, but higher concentrations of K, Ca and Mg in the shoots compared to the roots. There were large increases in Na for both shoots and roots of the surviving and dead plants. The root K concentrations increased for surviving plants but decreased for dead plants. The P concentrations













Table 18. Nitrogen-15 balance for labeled effluents.


Recovered by
Inoculum Available plants 5N Recovery
volume 15N Roots Shoots Plants Water Unaccounted

15
--L-- -----------mg------------ ------% of applied 15N-----Diluted effluents from high N plants

2.5 225 44 103 66 7 27 5 649 80 199 43 9 48 10 1044 101 272 36 3 61

Undiluted effluents from high N plants

2.5 1610 20 45 4 19 77 5 2120 18 41 3 27 40 10 2890 19 46 2 25 74

Undiluted effluents from low N plants

2.5 236 62 119 77 6 17 5 487 71 124 40 4 56 10 869 63 73 16 6 78




89






Table 19. Distribution of nutrients in water hyacinth shoots
and roots in diluted or undiluted effluents of
digested high N plants.

.Inoculum
volume Na K P Ca Mg

-l
--L-- ---------------g kg

SHOOTS

Diluted effluents from high N plants

2.5 13.3 13.3 2.4 16.9 9.2 5 17.5 19.3 3.9 17.8 9.1 10 18.0 23.2 5.0 15.6 8.6

Undiluted effluents from high N plants

2.5 21.0 18.0 3.5 18.2 5.6 5 20.5 17.7 3.9 22.6 6.2 10 19.2 14.5 3.8 19.3 5.7 LSD (0.05) 4.4 7.1 1.3 3.7 1.9 Original shoot tissue

2.8 22.0 2.7 15.4 5.7 ROOTS

Diluted effluents from high N plants

2.5 14.5 7.1 2.1 6.1 4.6 5 17.8 8.9 2.6 6.0 4.7 10 16.0 12.4 3.4 9.0 4.2

Undiluted effluents from high N plants

2.5 16.2 3.1 11.2 26.5 4.6
5 16.8 3.3 10.3 23.6 4.3 10 17.2 3.3 10.2 24.6 4.1 LSD (0.05) 3.0 3.7 1.9 3.0 0.7 Original root tissue

4.6 5.5 2.8 6.6 2.6





90


increased for the dead plant root material but remained similar for the other materials. The Ca content increased for the shoots of all plants and the roots of the dead plant material. The Mg content increased for roots of all plants and shoots of the surviving plants.

The net assimilation or loss of plant nutrients from plants in

diluted or undiluted effluents from digested high N plants are reported in Table 20. The plants in diluted effluents assimilated large amounts of Na and K, apparently because these nutrients move rapidly with the transpiration stream. The net shoot assimilation of all nutrients was greater than the net root assimilation.

The dead plants from undiluted effluents showed a net loss of K but net gains of the other nutrients. Potassium, Na, Ca and Mg were reported as being rapidly lost during the early leaching phase of plant decomposition in fresh water (Boyd, 1970b; Davis and van der Valk, 1978). Most of the Na moved into the shoot region. Generally the net gains of P and Ca were found in dead plant roots compared to shoots of surviving plants in diluted effluents. Final Chemical Composition of the Effluents

Characteristics of the digester effluents and nutrient medium after water hyacinth treatment are given in Table 21. For treatments where plant dry weight gains were observed, generally there was large reductions of the elements analyzed in the effluents. For the treatments resulting in plant death, K and Mg increased and the reductions of other elements were of lesser magnitude.

The largest reductions of EC (49 + 7% reduction), K (93 + 3%) and P (92 + 3%) were observed in diluted effluents. The highest gains of




Full Text
Ill
Conclusions
After 90 days of incubation, approximately 20% of the added C of the
digested biomass sludges had evolved as CO^ compared to 39 and 50% of the
fresh plant biomass with a low and high N content, respectively.
Decomposition of fresh plant biomas followed a three stage first-order
kinetic description. Decomposition of digested sludge was adequately
described by two stage first-order kinetics.
Mineralization of organic N to ^NO^-N accounted for approximately
8% of applied N for both digested biomass sludges at the end of 90 days.
Nitrogen mineralization accounted for 3 and 33% of applied N for fresh
plant biomass with a low and high N content, respectively.
The soil pH increased after addition of digested biomass sludge,
but was not appreciately altered after addition of fresh plant biomass.
The Na content of digested sludges was attributed as the primary factor
for increasing soil pH.
Increases in Mehlich I extractable soil constituents were a direct
reflection of the mineral composition of fresh or digested plant
biomass. The high Na concentration of digested biomass sludge suggests
pretreating the sludge to remove some of the salts and selecting salt
tolerant plants if the sludge is used as a soil amendment.


Table 5. Total plant N and ^NO^-N assimilation.
Treatment
Total uptake Labeled
Other *
(% of added
15N)
mg
rij
Without sediment
0 mg C L_1t
767
+
86
652 (85)
115
(15)
100 mg C L_1
1088
+
87
675 (62)
413
(38)
400 mg C L_1
1027
+
268
606 (59)
421
(41)
With sediment
0 mg C L_1
1220
+
29
720 (59)
500
(41)
100 mg C L_1
1171
+
235
656 (56)
515
(44)
400 mg C L_1
1112
+
163
567 (51)
545
(49)
^Carbon source was plant detritus.
^Other N sources include sediment and detritus.


DECOMPOSITION OF FRESH AND ANAEROBICALLY DIGESTED PLANT
BIOMASS IN SOIL 95
Materials and Methods . 96
Results and Discussion 98
Conclusions Ill
MASS BALANCE OF NITROGEN IN AN INTEGRATED "BIOMASS FOR ENERGY"
SYSTEM 112
Nutrient-Enriched Systems 112
Nutrient-Limited Systems 116
CONCLUSIONS 119
Water Hyacinth Productivity and Detritus Production 119
Detritus and Nitrogen Transformations 120
Anaerobic Digestion of Water Hyacinth 120
Digester Effluent Recycling 121
Digester Sludge Recycling 122
APPENDICES
A DIGESTER EFFLUENT CHARACTERISTICS DURING WATER
HYACINTH TREATMENT 124
B SOIL CHARACTERISTICS FROM ADDED FRESH AND ANAEROBICALLY
DIGESTED PLANT BIOMASS 126
BIBLIOGRAPHY 131
BIOGRAPHICAL SKETCH 141
iv


79
Bivens Arm research reservoirs in Gainesville, Florida. The plants were
clipped of dead tissue and acclimated to greenhouse conditions for two
weeks prior to treatments. The studies to evaluate the potential of
water hyacinth to treat digester effluents were conducted for a period
of 22 days in March and September, 1984. The daily maximum greenhouse
temperatures ranged from 19 to 37C in March and 27 to 36C in
September.
There were a total of 10 treatments, replicated three times, as
described below. Treatments 1 to 3 were diluted effluents from
digesters containing high N water hyacinths and inoculum volumes of 2.5,
'5, and 10 L, respectively. The dilutions were 1:8, 1:4, and 1:3
effluent : tap water for digester effluents with inoculum volumes of
2.5, 5, and 10 L, respectively. Treatments 4 to 6 were undiluted
effluents from digesters containing high N plants and inoculum volumes
of 2.5, 5, and 10 L, respectively. Treatments 7 to 9 were undiluted
effluents from digesters containing low N plants and inoculum volumes of
2.5, 5, and 10 L, respectively. The final treatment was a modified 10%
Hoagland's solution (p. 42) to serve as a control. Characteristics of
the inoculum and effluents are given in the Anaerobic Digestion of Water
Hyacinth chapter (pp. 60 to 75).
Plant samples collected initially and at the conclusion of the
experiment were analyzed for dry weights, TKN (Nelson and Sommers,
1973), Na, K, Ca and Mg by atomic absorption and P by an autoanalyzer.
Water samples were collected at 0, 1, 2, 3, 4, 8, 15, and 22 days and
analyzed for NH^-N and NO^-N by steam distillation (Keeney and Nelson,
1982), TKN (Nelson and Sommers, 1975), Ca, Mg, K, Na by atomic
absorption and P by an autoanalyzer. Electrical conductivity (Hach Mini


85
Nitrogen Removal
First-order kinetic equations were used to described NH*-N loss
from the effluents. An integrated rate equation for a first-order
reaction is expressed as:
In C / C = kt
o t
where C = initial NhT-N concentration in the effluent,
o A
C = final concentration at time = t,
k = first order rate constant (days ^).
The rate constant is calculated by solving for k:
k = In C / C 1/t
o t
The rate constants for diluted effluents ranged from 0.228 to 0.593
day1 (Table 17). The rate constants for undiluted effluents ranged
from 0.175 to 0.AA6 day
The time required for a 50% reduction in initial NH^-N
concentration generally increased with decreasing dry weight gains and
increasing NH+-N concentrations. The shortest reduction times were
associated with rapid plant growth (50% reduction in 1.12 to 3.04 days).
Plant assimilation was probably the primary mechanism of NH+-N loss in
these treatments. Reddy (1983) reported a 50% reduction of inorganic N
from agricultural drainage water in 18 days. The time required for a
50% reduction of NH^-N in effluents resulting in plant death was 1.98 to
3.96 days. Mechanisms of NH+-N loss in treatments resulting in plant
death included microbial assimilation and NH^-N volatilization. The pH
of the effluents ranged from 7.0 to 7.9 in treatments with actively
growing plants but increased from 8.2 to 9.3 in treatments where plant
death was observed and algal growth increased (APPENDIX A, Table 27).
The potential of NH^-N volatilization increases as NH+-N concentrations
increase or at higher pH values (Freney et al., 1983).


Figure 19. Nitrogen cycling in an integrated system for water hyacinths growing
in nutrient-limited systems. Numbers in parentheses are percentages
of the initial 10 g N placed in the anaerobic digester.
117


105
Table 23.
Soil NO^-N concentration from added fresh and digested
plant biomass.
Low N plant biomass High N plant biomass
Day Control Fresh Digested Fresh Digested
. -1
. mg kg soil
0
9.1
a +
8.2
ab
7.9
abc
8.4
ab
6.6
c
30
11.0
c
0.9
d
20.0
b
51.4
a
17.2
b
60
13.2
c
1.0
d
24.4
b
70.3
a
24.8
b
90
14.5
c
4.8
d
28.4
b
68.8
a
30.3
b
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.


107
mineralized and 20% of the initial N was recovered in sorhgum-
sudangrass plants.
After 90 days of incubation, approximately 20% of the added C of
digested biomass sludges had evolved as CO^ compared to 39 and 50% of
fresh plant biomass with a low and high N content, respectively. More
than half of the C evolved from fresh plant biomass occurred within the
first 10 days.
The percentage of C evolved with time from anaerobically digested
biomass sludge is similar to results of Miller (1974) and Tester et al.
(1977) for anaerobically digested and composted sewage sludge. Miller
(1974) reported a maximum of 20% of the added C was evolved as CC^ during
a 6-month incubation period. Tester et al. (1977) reported that 16% of
the added C was evolved as CO^ from composted sewage sludge during 54
days of incubation. However, Terry et al. (1979) found a total of 26 to
42% of C was evolved as CO^ from anaerobically digested sewage sludge
during 130 days of incubation.
Carbon/nitrogen ratio is commonly used as a guideline for
predicting the relative decomposability or mineralization potential of
organic materials added to soil (Reddy et al., 1980). The high C/N ratio
of low N fresh plant biomass (C/N = 35) resulted in immobilization of
inorganic N. The low C/N ratio of high N fresh plant biomass (C/N = 12)
t
resulted in rapid mineralization of organic N. However, C/N ratios may
have limited applicability for predicting N transformations of digested
biomass sludges. Both high N fresh biomass and digested biomass sludge
had C/N ratios of 12, but only 8% of the applied N was recovered as
^NO^-N from digested sludge compared to 33% from fresh plant biomass.


NITROGEN CYCLING IN AN INTEGRATED
"BIOMASS FOR ENERGY" SYSTEM
By
KEVIN KEITH MOORHEAD
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS
FOR THE DEGREE OF DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA
1986

ACKNOWLEDGMENTS
This dissertation reports results from a project that contributes
to a cooperative program between the Institute of Food and Agricultural
Sciences (IFAS) of the University of Florida and the Gas Research
institute (GRI), entitled "Methane from Biomass and Waste." Financial
support from the IFAS/GRI cooperative program is gratefully
acknowledged.
Dr. D. A. Graetz was chairman and Dr. K. R. Reddy was cochairman Gf
my supervisory committee. Their friendship and guidance will always be
cherished. The other members of the committee were Dr. G. E. Bowes,
Dr. J. G. A. F.iskell, and Dr. R. A. Nordstedt.
Special thanks go to Bill F'othier who ran numerous L,N samples for
this research. Other people who provided assistance during the estudies
include Bill Christy, Stephen McCracken, Peter Krottje, Terry Siean,
Veronica Campbell, Rremila Rao. Ed Hopwood and Dave Cartiin. Steve
Linda designed the statistical analyses. I appreciate the use of Dr.
John Moore's facilities for fiber analyses. Carolyn Pickles and Brenda
Clutter typed the majority of this dissertation on an IBM computer.
Finally, this package is dedicated to my parents, sisters and brother.
A trip home always gave me a boost to carry on.
ir

TABLE OF CONTENTS
PaSe
ACKNOWLEDGEMENTS . ii
LIST OF TABLES v
LIST OF FIGURES vii
ABSTRACT ix
INTRODUCTION 1
LITERATURE REVIEW A
Water Hyacinth Biomass Production A
Anaerobic Digestion 13
Waste By-Product Recycling 16
Conclusions 21
WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION 23
Materials and Methods 2A
Results and Discussion 26
Conclusions 38
EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER
HYACINTH SYSTEMS AO
Materials and Methods A2
Results and Discussion AA
Conclusions 58
ANAEROBIC DIGESTION OF WATER HYACINTH 60
Materials and Methods 61
Results and Discussion 63
Conclusions 7A
TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTH 76
Materials and Methods 78
Results and Discussion 80
Conclusions 93
iii

DECOMPOSITION OF FRESH AND ANAEROBICALLY DIGESTED PLANT
BIOMASS IN SOIL 95
Materials and Methods . 96
Results and Discussion 98
Conclusions Ill
MASS BALANCE OF NITROGEN IN AN INTEGRATED "BIOMASS FOR ENERGY"
SYSTEM 112
Nutrient-Enriched Systems 112
Nutrient-Limited Systems 116
CONCLUSIONS 119
Water Hyacinth Productivity and Detritus Production 119
Detritus and Nitrogen Transformations 120
Anaerobic Digestion of Water Hyacinth 120
Digester Effluent Recycling 121
Digester Sludge Recycling 122
APPENDICES
A DIGESTER EFFLUENT CHARACTERISTICS DURING WATER
HYACINTH TREATMENT 124
B SOIL CHARACTERISTICS FROM ADDED FRESH AND ANAEROBICALLY
DIGESTED PLANT BIOMASS 126
BIBLIOGRAPHY 131
BIOGRAPHICAL SKETCH 141
iv

LIST OF TABLES
TABLE PAGE
1. Seasonal water hyacinth yield and detritus production ... 30
2. Seasonal water hyacinth shoot and root lengths 32
3. Seasonal nitrogen uptake by water hyacinth and detritus . 35
A. Nitrogen balance for the two reservoirs 37
5. Total plant N and ^NO^N assimilation 53
6. Total plant N and ^NH+-N assimilation 5A
7. Mass balance of added ^NO^-N in sediment-water-plant
systems 56
8. Mass balance of added ^NH*-N in sediment-water-plant
systems 57
9. Characteristics of the inoculum used in the batch
digesters 64
10. Gas production during anaerobic digestion of high and
low N water hyacinth plants 65
11. Nitrogen balance for the batch digesters 68
12. Nitrogen-15 balance for the batch digesters 69
13. Characteristics of digester effluents before sludge
removal 71
14. Characteristics of screened effluents (sludge removed)
after digestion 72
15. Characteristics of fresh and digested biomass residues. . 73
16. Initial characteristics of the digester effluents
and nutrient medium 81
17. First-order kinetic descriptions of NH+-N loss with time. 86
18. Nitrogen-15 balance for labeled effluents 88
v

TABLE
PAGE
19. Distribution of nutrients in water hyacinth shoots
and roots in diluted and undiluted effluents of
digested high N plants 89
20. Net assimilation or loss of plant nutrients in diluted
or undiluted effluents from digested high N plants 91
21. Characteristics of the digester effluents and nutrient
medium after water hyacinth treatment 92
22. Characteristics of the fresh and digested plant biomass . 99
23. Soil NO^-N concentration from added fresh and digested
plant biomass 105
24. Carbon and mineralization from added fresh and
digested plant biomass 106
25. Soil pH (1:2 w/v) from added fresh and digested plant
biomass 109
26. Mehlich I extractable constituents at Day 90 from
added fresh and digested plant biomass 110
27. Effluent pH during water hyacinth treatment 124
28. Effluent dissolved 0^ concentration during water
hyacinth treatment 125
29. Soil ammonium concentrations from added fresh and
digested plant biomass 127
30. Mehlich I extractable constituents at Day 0 from
added fresh and digested plant biomass 128
31. Mehlich I extractable constituents at Day 30 from
added fresh and digested plant biomass 129
32. Mehlich I extractable constituents at Day 60 from
added fresh and digested plant biomass 130

LIST OF FIGURES
FIGURE PAGE
1. Integrated water hyacinth aquaculture system
of biomass production, bioconversion to methane
and digester waste recycling 2
2. A generalized diagram of a water hyacinth plant 5
3. Nitrogen cycling in a water hyacinth production sytem ... 9
4. Nitrogen cycling during anaerobic digestion 15
5. Nitrogen cycling in soil treated with plant residues. ... 18
6. Weekly averages of daily temperatures and solar
radiation 27
7. Monthly averages of daily primary productivity and
detritus production 28
8. Seasonal plant tissue nitrogen content 33
9. Dissolved 0 in sediment-water-plant systems with
added nitrate 45
10. Dissolved 0^ in sediment-water-plant systems with
added ammonium 46
11. The pH of sediment-water-plant systems with
added nitrate 48
12. The pH of sediment-water-plant systems with
added ammonium 49
13. Nitrogen loss from sediment-water-plant systems
with added nitrate 50
14. Nitrogen loss from sediment-water-plant systems
with added ammonium 51
15. Dry weight gains of water hyacinths in digester
effluents and nutrient medium 83
vii

FIGURE
PAGE
16. Carbon evolution from soil applied fresh and
digested plant biomass . 101
17. Decomposition stages and rate constants of fresh
and digested plant biomass added to soil 103
18." Nitrogen cycling in an integrated system for water
hyacinths growing in nutrient-enriched systems 113
19. Nitrogen cycling in an integrated system for water
hyacinths growing in nutrient-limited systems 117
viii

Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosopy
NITROGEN CYCLING IN AN INTEGRATED
"BIOMASS FOR ENERGY" SYSTEM
By
Kevin Keith Moorhead
May, 1986
Chairman: Dr. D. A. Graetz
Cochairman: Dr. K. R. Reddy
Major Department: Soil Science
A series of experiments were conducted to evaluate N cycling in
three components of an integrated "biomass for energy" system, i.e.
water hyacinth production, anaerobic digestion of hyacinth biomass, and
recycling of digester effluent and sludge. Plants assimilated 50 to 90%
of added N in hyacinth production systems. Up to 28% of the total plant
N was contained in hyacinth detritus. Nitrogen loading as plant
detritus into hyacinth ponds was 92 to 148 kg N ha ^ yr ^.
Net mineralization of plant organic ^N during anaerobic digestion
was 35 and 70% for water hyacinth plants with low (10 g N kg ^ dry
tissue) and high (35 g N kg ) N content, respectively. Approximately
20% of the ^N was recovered in the digested sludge while the remaining
^~*N was recovered in the effluent.
Water hyacinth growth in digester effluents was affected by
electrical conductivity (0.7 to 6.7 dS m and ^NIij"-N concentration
ix

(23 to 289 mg N L ^). Biomass yields were maximum at electrical
conductivities of < 2.5 dS m 1 and ^NH^-N concentrations of < 100 mg N
L_1.
Addition of water hyacinth biomass to soil resulted in
decomposition of 39 to 50% of added C for fresh plant biomass and 19 to
15.
23% of added C for digested biomass sludge. Only 8% of added N in
V
digested sludges was mineralized to ^N0_-N despite differences in
initial N content (27 and 39 g N kg 1 dry sludge). In contrast, 3 and
33% of added ^ N in fresh biomass with low and high N content,
respectively, was recovered as ^NO^-N.
Total recovery after anaerobic digestion ranged from 70 to 100%
of the initial plant biomass Land application of digester sludge
resulted in the mineralization of 2% of initial biomass into plant
available form. Use of water hyacinth for digester effluent treatment
resulted in recycling of 21 to 38% of the initial biomass ^N. Total N
recovery by sludge and effluent recycling in the integrated "biomass for
energy" system was 48 to 60% of the initial plant biomass The
remaining was lost from the system during anaerobic digestion and
effluent recycling.
x

INTRODUCTION
Several types of aquatic plants are widely distributed in
freshwater lakes and streams. These plants assimilate nutrients and
produce biomass, which could potentially be used for beneficial
purposes. Water hyacinth (Eichhornia crassipes [Mart] Solms) is one of
the dominant aquatic plants distributed throughout the tropical and
subtropical regions of the world. This freshwater macrophyte has
already been evaluated for use in treating nutrient-enriched waters such
as sewage effluent (Cornwell et al., 1977; Wolverton and McDonald, 1979;
Reddy et al., 1985), agricultural drainage water (Reddy and Bagnall,
1981; Reddy et al., 1982), anaerobic digester effluent (Hanisak et al.,
1980), and fertilized fish ponds (Boyd, 1976). The characteristics that
make this plant grow rapidly in polluted waters make it an ideal
candidate for large-scale nutrient removal and water purification (Reddy
and Sutton, 1984).
An integrated aquaculture system has been developed using water
hyacinth for water treatment and for total resource recovery. The
components of an integrated aquaculture system are schematically
illustrated in Fig. 1. Water hyacinth plants have been used for
wastewater treatment while the biomass produced was harvested
periodically and processed through anaerobic digestion to produce
methane. The process produced a waste by-product which must be disposed
of, or preferably utilized, in an environmentally-safe manner.
1

Figure 1. Integrated water hyacinth aquaculture system of biomass
production, bioconversion to methane and digester waste
recycling.

3
The waste by-product contains digested biomass sludge (solid) and
effluent (liquid). The digested biomass sludge was applied to soil as a
nutrient source for plants. The effluent was recycled in water hyacinth
ponds for nutrient recovery by plants. This type of integrated system
will provide low cost water treatment and total resource recovery.
Efficient utilization of by-products could potentially reduce the cost
effectiveness of the system.
The overall objective of this study was to assess nitrogen cycling
in the three components of an integrated "biomass for energy" system.
Nitrogen is often identified as a limiting factor for plant growth and
is used to establish loading rates in the disposal of solid and liquid
waste. Information on N cycling is limited to studies on the individual
components of the integrated system, i.e. the water hyacinth production
system (Boyd, 1976; DeBusk et al., 1983; Reddy, 1983), anaerobic
digestion (Hashimoto et al., 1980; Field et al., 1984), and effluent and
sludge recycling (Ryan et al, 1973; Hanisak et al., 1980; Terry et al.,
1981; Atalay and Blanchar, 1984). No attempt has been made to establish
N cycling within the entire integrated system.
The specific objectives of this study were 1) to determine growth
rate and detritus production of water hyacinth grown in eutrophic lake
water; 2) to determine the effect of detritus on N transformations in
water hyacinth systems; 3) to evaluate N and C mineralization during
anaerobic digestion of water hyacinth biomass; 4) to evaluate the
potential of water hyacinth to grow in anaerobic digester effluents for
N recovery; and 5) to determine N and C mineralization during
decomposition of fresh and digested biomass added to soil.

LITERATURE REVIEW
The three components of the integrated "biomass for energy" system
were 1) the water hyacinth production system; 2) anaerobic digestion of
water hyacinth biomass; and 3) recycling of digested biomass sludge and
effluent. An integrated approach of wastewater renovation using aquatic
macrophytes with utilization of biomass for energy production is
economically appealing.
Water Hyacinth Biomass Production
The first component of the integrated "biomass for energy" system
was an aquatic system for the production of biomass as well as water
quality improvement. Although several aquatic plants naturally grow in
polluted waters, one the most productive plants appears to be water
hyacinth (Reddy et al., 1983).
Water hyacinth is a mat-forming, free-floating vascular aquatic
plant with wide distribution in sub-tropical and tropical regions. The
plant consists of a submerged rooting system and an aerial
photosynthetic petiole and leaf (shoot) system (Fig. 2). The roots and
aerial shoots are produced at the numerous nodes of the vegetative
portion of a typically submerged rhizome (Penfound and Earle, 1948).
The aerial buds, from which flowers and fruit clusters develop, are
produced from the reproductive portion of the rhizome. Occasionally,
the internodes of the rhizome expand and form new offsets.
4

5
Figure 2. A generalized diagram of a water hyacinth plant.
The major morphological structures are
adventitious roots (AR); root hairs (RA); rhizome
(RH); stolon (ST); detritus tissue (DT) attached
to the plant; float (F); leaf isthmus (IS);, leaf
petiole (PT); peduncle (PD); spathe (SP); leaf
lamina (LA); inflorescence (IN).

6
The elongated internodes were designated as stolons (Penfound and Earle,
1948).
The relatively rapid rate of colonization by water hyacinth is due
primarily to vegetative reproduction (stolon and offset production).
The plants reproduce sexually during warmer months until freezing
terminates anthesis. The developing fruits containing the seeds are
cast off onto the mat or into the water. They sink in water and remain
in a viable condition for several years. Manson and Manson (1958)
reported that each plant could produce 5000 to 6000 seeds which remained
viable for at least 5 years.
The geographical distribution of water hyacinth is'regulated by
temperature and salt concentration in the water. When average minimum
temperature reached 10C, productivity of water hyacinth approached zero
(Reddy and Bagnall, 1981). Optimum growth was found in a temperature
range of 25 to 30C (Bock, 1969; Knipling et al., 1970). Water hyacinth
is basically a freshwater plant and will die in waters with sustained
salt concentrations in excess of 2500 mg L (Haller et al., 1974).
Water hyacinth growth is regulated by the nutrient composition of
the water medium, temperature, solar radiation, and plant density.
Water hyacinth potentially could be grown in nutrient-enriched waters
such as sewage effluents, agricultural runoff and drainage effluents,
methane digester effluents, and runoff from animal waste operations.
Nitrogen is present as NH*-N, NO^-N, and organic N in water media
avaiable for water hyacinth production. Organic N often predominates in
most water media and is not readily available for plant assimilation.
Water hyacinths are efficient users of inorganic N and plant
assimilation is one of the major processes of N removal in hyacinth
ponds.

7
Water hyacinth adapted to light intensity and full sunlight elicted
the greatest photosynthetic rate (Patterson and Duke, 1979). Optimum
plant density to obtain maximum biomass yield varied with season and
available plant nutrients in the water (Reddy and Sutton, 1984). DeBusk
et al. (1981) and Reddy et al. (1983) established that optimum plant
density for achieving maximum growth cultured in wastewaters was in the
-2
range of 15 to 35 kg wet wt m
Water hyacinth productivity has been evaluated in natural and
-2 -1
nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day
were reported for plants growing in natural waters of central and south
Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide-range
-2 -1
of productivity (5 to 42 g dry wt m day ) was recorded for plants
cultured in nutrient-enriched waters (Schwegler and Kim, 1981; Hanisak
et al., 1980). Reddy and DeBusk (1984) obtained an average of 52 and a
-2 -1
maximum of 64 g dry wt m day for water hyacinths grown in
nutrient-nonlimiting conditions.
The effectiveness of water hyacinth in removing inorganic N was
reported for several nutrient-enriched wastewaters. Sheffield (1967)
and Clock (1968) reported a 75 to 94% reduction of inorganic N from
secondary sewage effluent in systems containing water hyacinths. Reddy
et al. (1982) observed a 78 to 81% reduction of inorganic N from organic
soil drainage water containing water hyacinths. Hanisak et al. (1980)
concluded that 65% of N in digester effluents could be assimilated when
water hyacinths were grown in diluted effluents. Boyd (1976) calculated
average rates of N and P removal were 3.4 and 0.43 kg ha1 day1 in
fertilized fish ponds. Rogers and Davis (1972) concluded that water
hyacinth removal capacities were less effective with increasing nutrient
concentrations.

8
The potential productivity and nutrient removal capacities of water
hyacinth has led to its selection as a biomass feedstock for methane
generation while providing a means for treatment of nutrient-enriched
waters. Extensive research, both in laboratory and field applications,
was conducted on the use of water hyacinth in wastewater treatment
during the past 20 years (Sheffield, 1967; Boyd, 1970a; Steward, 1970;
Scarsbrook and Davis, 1970; Rogers and Davis, 1971; Dunigan et al.,
1975; Cornwell et al., 1977; McDonald and Woverton, 1980; Reddy et al.,
1982; DeBusk et al., 1983). Water hyacinth was shown to be effective in
removing N, P and other nutrients, and reducing biological oxygen demand
and total suspended solids.' Water hyacinth was also shown to readily
absorb and concentrate heavy metals (Wolverton and McDonald, 1975a,b;
Cooley et al., 1978).
Nitrogen Cycling in the Water Hyacinth Production System
Nitrogen transformations occurring in a water hyacinth production
system include 1) plant uptake; 2) mineralization/immobilization; 3)
nitrification; 4) denitrification; and 5) NH^-N volatilization (Fig. 3).
Plant uptake is one of the major processes for N removal from water
hyacinth-based wastewater systems. Plant uptake is directly related to
the growth rate and the nutrient composition of the water. Water
hyacinth was more efficient in utilizing NH+-N than NO^-N when both
forms were supplied in equal proportions (Reddy and Tucker, 1983).
A dense cover of floating water hyacinths will regulate dissolved
0^, temperature and pH of water which influences several N
transformations. Generally, diel fluctuations of these water parameters
were reported to be lower in areas covered with water hyacinths compared
to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980;
Reddy, 1981).

Organic
Carbon
Organic
Nitrogen
/
t
/ /
tv////
\//
Organic
Carbon
/
/
Organic
Nitrogen
2. ,
/ y
/
t /
N TRANSFORMATIONS
1. Rant Uptake
2. Mineralization /
Immobilization
3. Nitrification
4. Denitrification
5. NH^ Volatilization
Figure 3. Nitrogen cycling in a water hyacinth production system.
Nitrogen transformations investigated during this study
are indicated with larger arrows.

10
High 0^ consumption within a water hyacinth mat during microbial
decomposition of plant detritus (dead and decaying plant debris) could
create anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980;).
The bulk of detritus was trapped within the mat and decomposed primarily
at the water surface (DeBusk et al., 1983). Rate of inorganic N release
from decomposing detritus depended on dissolved 0^ concentration of the
water, C/N ratio, and temperature (Ogwada, 1983).
Low 0^ concentrations create conditions less favorable for
nitrification and promote denitrification which may proceed within the
water hyacinth mat, in the water column, or in the underlying sediment.
Denitrification occurred primarily in the underlying sediment and the
rate depended on diffusion of NO^ -N from the water column to the
sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981).
Water temperatures were lower in areas covered with plants compared
to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980;
Schreiner, 1980). A dense mat over the water surface served as a
blanket barrier for exchange of heat between the atmosphere and the
water (Rai and Munshi, 1979). Water hyacinths growing in either acid or
alkaline water had a tendency to alter the pH towards neutrality (Haller
and Sutton, 1973). A pH of 7.0 in water occurred in areas covered with
plants with little diel variation (McDonald and Wolverton, 1980; Reddy,
1981) which suggests that NH^-N volatilization is minimal in these
systems .
Decomposition of Plant Tissue in Freshwater
Decomposition of plant tissue in a freshwater habitat commonly
occurs in two stages. The first stage was attributed to leaching
of the more soluble plant constituents while the second stage was

11
microbial-controlled degradation (Boyd, 1970b; Hunter, 1976; Godshalk
and Wetzel, 1978a; Howard-Williams et al., 1983).
Otsuki and Wetzel (1974) reported a rapid leaching loss of
dissolved organic matter regardless of conditions of aerobiosis or
whether plants were fresh or freeze-dried. Hill (1979) concluded that
rapid leaching of soluble material accounted for a 21 to 60% dry weight
loss of aquatic macrophytes during the first 8 days of incubation.
Leaching was established as the major process in the decomposition of
eelgrass and total loss of organic matter by leaching accounted for 82%
of dried leaves and 65% of fresh leaves (Harrison and Mann, 1975).
Leaching rates appeared to be independent of temperature (Carpenter,
1980).
Potassium, Na, Mg, and Ca have all been reported as being rapidly
lost during the early leaching phase of plant decomposition (Boyd,
1970b; Davis and van der Valk, 1978; Puriveth, 1980). Carpenter (1980)
found that the higher the initial P concentration, the more rapid was P
leaching.
The second stage of decomposition is attributed to biological
processes. Microbial-controlled decomposition was influenced by
temperature (Carpenter, 1980; Puriveth, 1980), pH (Sompongse, 1982),
available 0^ (Godshalk and Wetzel, 1978a), and available nutrients
(Carpenter and Adams, 1979; Puriveth, 1980). Godshalk and Wetzel
(1978a) found that the presence of O2, regardless of temperatures of 10
or 25 C, permitted rapid degradation of dissolved and particulate
organic matter. Decomposition of water hyacinth was found to be faster
under aerobic than anaerobic conditions (Reddy and Sacco, 1981).
However, Sompongse (1982) determined that aeration did not have a

12
measurable effect on rate of plant decomposition and Nichols and Keeney
(1973) reported a more rapid decomposition rate under non-aerated
conditions compared to aerated conditions. Ogwada et al. (1984) found
that approximately the same amounts of N and P were released from
decaying plant tissue under aerobic or completely anoxic conditions, but
the extent of nutrient release was dependent on water temperature.
The changes in plant C and N composition or concentration during
decomposition have received considerable attention. Build-up of
microbal biomass on decaying plant tissue caused a loss in the C content
while increasing the N content which resulted in a decrease in the C/N
ratio (De La Cruz and Gabriel, 1974; Odum and Heywood, 1978; Hill,
1979). Godshalk and Wetzel (1978b) found that lignin was very resistant
to decomposition while the other structural carbohydrates gradually
decreased with time.
Nitrogen was a limiting factor in decomposition of several aquatic
plants (Nichols and Keeney, 1973; Almazan and Boyd, 1978; Godshalk and
Wetzel, 1978b; Carpenter, 1980). Decay rates were correlated both to
initial N content and to C/N ratios (Godshalk and Wetzel, 1978b;
Carpenter and Adams, 1979; Ogwada et al., 1984).
Particle size also influenced decomposition. Generally, the rate
of decomposition increased as the particle size decreased (Fenchel,
1970; Hargrave, 1972; Gosselink and Kirby, 1974). Harrison and Mann
(1975) reported that a reduction in size of leaf material from 2 to 4 cm
to <1 mm doubled the rate of organic matter loss.
Boyd (1970b) and Odum and Heywood (1978) concluded that submerged
leaves decomposed more rapidly than those placed upon the water surface
or suspended in air. Nichols and Keeney (1973) found more rapid dry

13
weight loss under aerated conditions in sediment-water systems than in
water only. They attributed this difference to an additional supply of
N from the sediments.
Anaerobic Digestion
The second component of the integrated "biomass for energy" system
was anaerobic digestion of plant biomass for methane production.
Anaerobic digestion is a biological process in which organic matter, in
the absence of oxygen, is converted to methane and carbon dioxide
(Toerien and Hattingh, 1969).
During the process of anaerobic digestion, waste organic C was
stabilized by the nearly complete microbial fermentation of
carbohydrates resulting in a reduction of volatile solids (Miller,
1974). Anaerobically digested sewage sludges were considered more
stable to microbial degradation than were aerobically digested sludges
(Sommers, 1977).
Processes which regulated anaerobic digestion include hydrolysis of
polymers, the dissimilation of starting subtrates to the level of acetic
acid, and the conversion of acetic acid to CH^ and CO^ (Mah et al.,
1977). Factors which influenced anaerobic digestion include pH and
temperature changes. All methanogens were reported to be strict
i
anaerobes with an optimum pH of 6.7 to 7.4 (Bryant, 1979). The optimum
temperature range was 30 to 35C (House, 1981).
Water hyacinth biomass could be anaerobically digested to produce
methane. Hanisak et al. (1980) found average methane yields of 0.24 L
g volatile solids (VS) of shredded water hyacinth in 162 L digesters.
Chyoweth et al. (1983) reported methane yields of 0.19 and 0.28 L g ^ VS

14
for water hyacinths and a 3:1 blend of water hyacinths:domestic sewage
sludge, respectively in 5 L digesters. Shiralipour and Smith (1984)
reported average methane yields of 0.32 and 0.17 L g ^ VS water hyacinth
shoot and root samples, respectively, in a bioassay test of 100 ml
culture volume. They concluded that the addition of N to growth media
for water hyacinth production increased methane yields.
The nutrients in the biomass are recovered in the waste material
after the digestion process. The recovered nutrients are
distributed between the effluent and the digested biomass sludge. The
total N recovery was usually 100% after digestion, but much of the
organic N was converted to NH*-N (Hashimoto et al., 1980; Field et al.,
1984). Most of the K (Field et al., 1984) and Na (Atalay and Blanchar,
1984) were solubilized and remained in the digester effluent.
The digestion process increased the sorption of some nutrients (P,
Ca, and Mg) by the sludge fraction such that fewer were available for
dilute acid extraction and perhaps for crop recovery (Field et al.,
1984). Field et al. (1984) hypothesized that sorption may have been
increased by particle surface area increases due to size reduction of
solids.
Nitrogen Cycling During Anaerobic Digestion
Nitrogen transformations occurring during anaerobic digestion
include 1) mineralization/immobilization; and 2) NH^-N volatilization
(Fig. 4).
The mineralization or immobilization of N depends on the initial N
content of the biomass feedstock. The C/N ratio of the biomass is often
used as a guideline for prediction of net mineralization or
immobilization. The optimal C/N ratio of the added biomass was 30:1

N TRANSFORMATIONS
ANAEROBIC
DIGESTION
Organic
Acids
NH
1. Mineralization /
Immobilization
2. NH^ Volatilization
Figure 4. Nitrogen cycling during anaerobic digestion.
Nitrogen and carbon transformations
investigated during this study are indicated
with larger arrows.

16
(Hughes, 1981). Bioconversion of biomass with a higher C/N ratio was
limited by N. A lower initial C/N ratio resulted in mineralization of
organic N during digestion. The C/N ratio of the digester effluent was
lower than the C/N ratio of the fresh slurry because of the release of C
as CO^ and CH^ (House, 1981).
Anaerobic digestion of plant biomass resulted in high
concentrations of NH*-N in the digester (Hashimoto et al., 1980; Field
et al., 1984). Ammonium was toxic to methogens at concentrations > 3.0
g L-l, regardless of pH (Hashimoto et al., 1980). Losses of NH^-N
through volatilization should be low in digesters operating at the
optimum pH of 6.7 to 7.2 unless NH^-N concentrations are high.
Waste By-Product Recycling
The final component of the integrated "biomass for energy" system
was recycling of the waste by-product generated from the anaerobic
digestion of plant biomass. The waste by-product from the anaerobic
digester was screened to separate the digested biomass sludge from the
effluent. The effluent was recycled in the water hyacinth biomass
production system discussed earlier. Methane digester effluent contains
high levels of NH+-N (> 200 mg L ^) which may inhibit plant growth.
Dilution of the effluent is required before use in a water hyacinth
production system. Optimum dilution of the effluent for maximum water
hyacinth yields has not been established. Nitrogen cycling in a water
hyacinth production system was presented earlier.
The sludge was added to soil as an organic amendment. A
consequence of anaerobic digestion was a reduction of the readily
decomposable C of the plant tissue during production of CH^ and CO^-

17
Anaerobically digested sludge was considered to be stable and resist
further decomposition (Sommers, 1977).
Nitrogen Cycling in Soil Treated with Plant Residues
The land application of fresh or anaerobically digested plant
biomass has significant implications on N cycling. Nitrogen
transformations occurring after residue additions include 1)
mineralization/immobilization; 2) microbial or plant assimilation; 3)
nitrification; 4) denitrification; and 5) NH^-N volatilization (Fig. 5).
Since most of the N in fresh or digested plant biomass is in
organic forms, the rate of mineralization becomes the rate limiting step
for all transformations that follow. Mineralization or immobilization
depends on the initial N concentration of the plant biomass as well as
the composition of the C constituents. A low N content or a wide C/N
ratio was associated with slow decomposition and rates of decomposition
were proportional to lignin content (Alexander, 1977). A wide C/N ratio
(> 30:1) favored N immobilization whereas a narrower ratio (< 20:1)
resulted in N mineralization (Alexander, 1977).
Mineralization of N from anaerobically digested sewage sludges was
reported to be affected by the rate of application (Ryan et al., 1973;
Stark and Clapp, 1980). However, Epstein et al. (1978) found that
irrespective of the amount of material (sewage sludge and sludge
|
compost) applied, the percentage of added N mineralized remained
essentially constant.
The mineralization of NH+-N from organic N is accompanied by
microbial assimilation or plant uptake. In aerobic environments the
NH^-N was quickly converted to NC^-N (nitrification) which could also be

N TRANSFORMATIONS
1. Mineralization /
Immobilization
2. Plant Uptake
3. Nitrification
4. Denitrification
5. NH Volatilization
Figure 5. Nitrogen cycling in soil treated with plant residues.
Nitrogen and carbon transformations investigated
during this study are indicated with larger arrows.

19
assimilated by microbes or plants (Ryan et al., 1973). A high organic
loading rate may result in 0^ depletion during decomposition which
promotes the denitrification process (Epstein et al., 1978; Hsieh et
al., 1981b).
Decomposition of Plant Residues in Soil
Decomposition of plant residues in soil occurs in two stages. The
first stage was attributed to loss of the easily decomposable labile
fraction which was followed by the second stage of slow decomposition of
a resistant residual fraction (Shields and Paul, 1973; Reddy et al.,
1980). Both stages were thought to be controlled by two simultaneously
occurring superimposed first-order kinetic reactions (Sinha et al.,
1977).
Fresh and anaerobically digested plant biomass differ widely in
their chemical composition. Anaerobic digestion converts most of the
easily-decomposable plant C constituents into CH^ and CC^. The digested
biomass sludge has a higher lignin content and is more resistant to
decomposition. There is little information available on decomposition
of anaerobically digested plant biomass added to soil. However, the
rates and the factors which influence decomposition of fresh plant
biomass added to soil have been well-established.
Tenny and Waksman (1929) concluded that water-soluble organic
substances were first to be decomposed in the soil, followed by
hemicellulose and at the same time, or immediately after, cellulose.
Lignin was very resistant to decomposition and may even delay the
disintegration of cellulose or hemicellulose because of the structural
proximity of these C constituents in the cell wall (Tenny and Waksman,
1929; Peevey and Norman, 1948; Berg et al., 1982).

20
Application rates were shown to have insignificant effects on rate
of fresh plant biomass decomposition (Jenkinson, 1965; Nyhan, 1975).
However, several studies suggested that small amounts of fresh or
anaerobically digested plant biomass decomposed more rapidly than large
quantities (Broadbent and Bartholomew, 1948; Jenkinson, 1971; Atalay and
Blanchar, 1984).
Miller and Johnson (1964) found an increasing rate of CO^
production with increasing moisture content up to a tension of 0.05 to
0.015 MPa and then a decreasing rate with further increases in tension.
They concluded that maximum biological activity could be expected at the
lowest tension when aeration was sufficient. Orchard and Cook (1983)
found a log-linear relation between water potential and microbial
activity in the range of 0.005 to 0.5 MPa.
Sain and Broadbent (1977) concluded that low temperatures
influenced decomposition rate more than excessive moisture. However,
Nyhan (1976) found a pronounced decrease in rates of C loss with an
increase in soil water tension even when temperature (10C) was limiting
microbial activity. Miller (1974) determined that soil temperature was
the major factor influencing the rate of decomposition of anaerobically
digested sewage sludge.
Decomposition was generally considered to be initially slower in
acid than neutral soil (Jenkinson, 1971). Addition of organic material
altered the pH of a soil, particularly when the amount added was large
relative to the amount of native organic matter present (Jenkinson,
1966). Atalay and Blanchar (1984) found that addition of anaerobically
digested biomass sludge to soil increased the pH from 5.5 to 7.6 and
they attributed this to a limestone buffer used during the digestion
process.

21
Jenkinson (1965, 1971), using different plants and soils,
determined that the proportion of added plant C retained in the soil
under different climatic conditions was remarkably similar over time.
Generally, one-third of the added plant C remained after one year,
falling to one-fifth after 5 years.
Atalay and Blanchar (1984) determined that anaerobically digested
biomass sludge decomposed rapidly in soil as evidenced by nearly 40% of
the C added evolved as CO^ during 100 days of decomposition. However,
Miller (1974) concluded that anaerobic digested sewage sludge was
resistant to further decomposition with a maximum of 20% of the added C
evolved as C0 during a 6-month incubation. Terry et al. (1979) found
that 26 to 42% of anaerobically digested sewage sludge C was evolved as
CO^ during incubation. Generally, the majority of the CO2 produced in
incubation studies was evolved in the first 30 days (Miller, 1974; Terry
et al., 1979; Ataway and Blanchar, 1984).
Other soil properties influenced by plant biomass additions
included increasing water-holding capacity, CEC, and electrical
conductivity (Stark and Clapp, 1980; Atalay and Blanchar, 1984).
Epstein et al. (1976) found levels of salinity and chloride in sewage
sludge applied to soils increased to a level which may affect
salt-sensitive plants.
Conclusions
Although information is available on N cycling for each component
of the system, no attempt has been made to follow N transformations in
an integrated "biomass for energy" system. Evaluation of N cycling was

22
chosen because N is often identified as a limiting factor for plant
growth and is used to establish loading rates in the disposal of solid
and liquid waste.
Plant uptake was established as a major N removal process during
water hyacinth biomass production (Reddy and Sutton, 198A). However,
the role of water hyacinth detritus as a N source or sink has not been
established. Methane yields during anaerobic digestion of water
hyacinth were enhanced with increasing N content (Shiralipour and Smith,
1984). However, N mineralization rates were not investigated. Limited
information was available on disposal or utilization of digester
effluent or sludge from anaerobically digested plant biomass (Hanisak et
al., 1980; Atalay and Blanchar, 1984). The overall objective of this
research was to integrate the three components of biomass production,
anaerobic digestion of biomass, and digester waste recycling with
respect to N cycling.

WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION
Water hyacinth is one of the most productive aquatic macrophytes
found throughout the tropical and subtropical regions of the world. The
plant has been used extensively for treatment of nutrient-enriched
waters and currently there are a number of wastewater treatment systems
in the U. S. utilizing water hyacinths for secondary and tertiary
treatment (Cornwell et al.,1977; Dinges, 1978; Wolverton and McDonald,
1979; Reddy et al., 1985).
Water hyacinth productivity has been evaluated in natural and
-2 -1
nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day
were reported for plants growing in natural waters of central and south
Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide range
-2 -1
of productivity (5 to 42 g dry wt m day ) was recorded for plants
cultured in nutrient-enriched waters (Hanisak et al., 1980; Reddy and
Bagnall, 1981; Reddy, 1984). Maximum growth rates provided an average
-2 -1
of 52 and a maximum of 64 g dry wt m day for plants cultured under
nutrient-nonlimiting conditions (Reddy and DeBusk, 1984).
Plant detritus (dead and decaying plant debris) is an integral part
of water hyacinth mats. Detritus is usually derived from natural aging
of plants, biological or chemical control, and frost damage.
Decomposition of detritus releases nutrients which can be subsequently
utilized by water hyacinths. Information on water hyacinth productivity
was extensive (Reddy et al., 1983), but research on detritus production
and its role as a nutrient sink or source was limited.
23

24
DeBusk et al. (1983) measured detritus production in harvested and
nonharvested water hyacinth based sewage treatment systems. Detritus
-2 -1
production in both systems averaged 2 g dry wt m day More than 80%
of the detritus consisted of root material. The bulk of the standing
crop detritus remained trapped in the floating plant mat. However, this
study did not reveal the potential of detritus as a nutrient input to
the water hyacinth ponds.
The objectives of this study were to 1) measure productivity and
detritus (shoot and root) production of water hyacinths grown in
eutrophic lake water with and without added nutrients and 2) determine
the potential of detritus as a nutrient source or sink to the ponds.
Materials and Methods
The study was conducted in two reservoirs located at the Central
Florida Research and Education Center research farm near Lake Apopka in
Zellwood, Florida. The reservoirs were constructed with 2.0 m high
levees of a Lauderhill organic soil (Lithic medisaprists) and with
bottoms composed of calcareous clay. The water depth was 60 cm and the
dimensions of the reservoirs were 7.6 m by 61 m (total surface area of
2
465 m ). Both reservoirs were filled with water from nearby Lake
Apopka, and were sectioned into four equal areas for replication and
stocked with water hyacinths.
2
A total of eight 0.25 m cages (Vexar mesh screen connected to 5 cm
diameter PVC pipe) were stocked with water hyacinth at an initial
_2
density of 16 kg (fresh wt) m The cages were placed within the four

25
replicated areas of each reservoir. Each cage was lined with 1.00 mm
fiberglass screen to retain any detritus dislodged from the plant mat.
The fiberglass screen was positioned 15 cm below the PVC frame to allow
normal waterflow within the root mat.
One reservoir was fertilized monthly by broadcasting a 10-4-10
fertilizer to add 100 kg N ha ^ from October 1981 to February 1982, and
50 kg N ha from March 1982 to September 1982. The change in
fertilizer rate was due to an excess of 10 mg N L ^ found in reservoir
water several days after fertilization during winter. The second
reservoir contained Lake Apopka water with no added nutrients and served
as the control. Both reservoirs were drained and refilled with Lake
Apopka water monthly (24 hr prior to plant sampling and fertilization).
Plant productivity and detritus production were monitored at
monthly intervals for one year. The cages were removed from each
section, drained for 5 min, and weighed. The plant material was divided
into healthy plants and detritus. Detritus was defined as that
collected from the fiberglass screen and dead shoot and root material
remaining within the plant mat. Dead shoots were defined as material
visably devoid of chlorophyll. Three plants were removed for analyses
and the cages were restocked to the initial plant density and placed in
the reservoirs.
*
The shoot and root lengths were recorded for each plant sample and
separated for dry weight ratios. The plant tissue and detritus were
oven-dried at 70 C, weighed, and ground to pass a 0.84 mm Wiley Mill
screen. All plant and detritus samples were analyzed for total Kjeldahl
nitrogen (TKN) using a modified micro Kjeldahl procedure (Nelson and

26
Sommers, 1973). Solar radiation and high and low daily temperatures
were recorded. The results were statistically analyzed for a randomized
block design with the fertilized and control reservoirs as treatments.
Results and Discussion
The weekly averages of daily maximum and minimum air temperatures
and solar radiation are shown in Fig. 6. Maximum temperatures ranged
from 21.9C during January to March and 36.5C during July to September.
Minimum temperatures for these time periods were 8.2C and 20.3C,
respectively. Maximum and minimum temperatures for the rest of the year
were similar (high= 30C, low= 13.5C). Maximum and minimum solar
radiation occurred from April to September and from November to March,
respectively.
The monthly averages of daily primary productivity and detritus
production of water hyacinth are presented in Fig. 7. Maximum daily
water hyacinth productivity during this study was observed in August for
_2
the fertilized reservoir (28.3 g dry wt m day ) compared to June for
the control reservoir (14.7 g dry wt m day ). Detritus production
remained fairly consistent with time for both reservoirs. Detritus
production in the fertilized reservoir increased noticeably in September
when plant productivity began to decline. The average daily detritus
-2 -1
production in the fertilized reservoir was 3.7 g dry wt m day
-2 -1
compared to 3.5 g dry wt m day in the control reservoir. DeBusk et
al. (1983) found that detritus production occurred at a relatively
constant rate regardless of harvested or nonharvested conditions.
Monthly data for the plant parameters have been summarized by
seasons: 1) autumn (October, November, and December); 2) winter

27
Figure 6. Weekly averages of daily temperatures and solar
radiation.

25
20
I 5
I 0
5
0
20
I 5
10
5
0
Fertilized
TIME (months)
. Monthly averages of daily primary productivity and
detritus production.

29
(January, February, and March; 3) spring (April, May, and June); and 4)
summer (July, August, and September). The seasons were chosen to
coincide with changes in water hyacinth productivity and temperature and
solar radiation changes. The autumn season combines data for November
and December of 1982 and October of 1983.
The effects of temperature and solar radiation were evident since
the lowest net productivity occurred during winter and the highest
during spring and summer (Table 1). The net primary productivity in
winter for both reservoirs was lower than the production of detritus.
Reddy and Bagnall (1981) reported that at average temperatures of 10C,
productivity of water hyacinth approached zero. The majority of the
detritus during winter came from the destruction of aerial shoots caused
by freezing temperatures. Although a majority of the aerial shoots were
destroyed, the plants survived, and noticeable gains in dry weight began
in March.
There were significant differences in yields between seasons and
reservoirs. Seasonal yields ranged from 1.9 to 23.1 Mg (dry wt) ha ^
for the fertilized reservoir and -0.2 to 10.2 Mg ha ^ for the control
reservoir. Over 75% of the biomass production occurred during spring
and summer for both reservoirs. Differences in detritus production were
not significant between reservoirs or between seasons. Although the
annual yield of water hyacinth in the fertilized reservoir was double
that of the control reservoir, the detritus production in both
reservoirs was similar (Table 1). There were significant differences in
the shoot/root dry weight ratios between reservoirs, but not between
seasons (Table 1). The average shoot/root dry weight ratio was 1.93
(1.6A to 2.A6) for the fertilized reservoir and 1.14 (0.79 to 1.67) for
the control reservoir.

30
Table 1. Seasonal water hyacinth yield and detritus production.
Fertilized reservoir
Control
reservoir
Season
Yield
Detritus
S/R §
Yield Detritus
S/R

Mg ha 1
Mg ha
-1
Autumn (78) *
9.5
2.8
1.85
3.3
2.2
1.67
Winter (90)
1.9
2.7
1.75
-0.2
3.2
1.25
Spring (84)
14.7
2.7
1.64
9.7
3.3
0.86
Summer (88)
23.1
4.1
2.46
10.2
3.3
0.79
Test of Significance ^
Yield
Detritus
S/R
Season
**
NS
NS
Month (season
)
NS
NS
NS
Reservoir
**
NS
**
TNumber of days in season.
Significant at 0.01 level (**) or not significant (NS).
§S/R = Shoot/root dry wt ratio

31
Shoot and root lengths were similar for plants in both reservoirs
during autumn and winter (Table 2). During spring, the water hyacinth
root lengths were shorter and shoot lengths longer in the fertilized
reservoir compared to the plants in the control reservoir. An
interesting development in plant morphology was the dislodging of
practically the entire root system from plants in the fertilized
reservoir beginning in March after daily temperatures began to increase.
The majority of plants were typified by a small root system compared to
plants in the control reservoir. Some root dislodging was noticed in
the control reservoir during spring and summer but was not as
characteristically uniform as in the fertilized reservoir. Root lengths
in the fertilized reservoir began to increase during summer, but the
shoot lengths were double those in the control reservoir.
Under nutrient-limiting conditions, water hyacinths produce a large
volume of root material presumably to increase their nutrient absorption
capacity. With nutrient-enriched media, water hyacinth use more
photosynthetic energy in shoot production. Cornwell et al. (1977)
measured shoot lengths in excess of 1 m in wastewater media. Penfound
and Earle (1948) recorded root lengths of 0.1 to 1 m. Maximum shoot
length recorded during this study was 55 cm during summer for fertilized
plants compared to 28 cm during summer for control plants.
i
The plant tissue N content is shown in Fig. 8. Fertilization
resulted in increases in shoot, root, and detritus N content compared to
plants in the control reservoir. Maximum tissue N content for
fertilized plants occurred during winter when plant productivity was
low. The increase in plant productivity in spring and summer diluted
the N content of the tissue although total N assimilation by the plants

Table 2. Seasonal water hyacinth shoot and root lengths.
Fertilized reservoir Control reservoir
Season Shoots Roots Shoots Roots
cm
Autumn (78)
39.6
21.5
35.7
22.9
Winter (90)
25.2
14.2
23.3
20.2
Spring (84)
26.6
9.9
18.8
14.8
Summer (88)
54.9
21.3
27.8
27.1
Test of significance *
Shoot
length
Root length
Season
**
**
Month (Season)
NS
**
Reservoir
*
**
t Number of days in Season.
Significant at 0.05 (*) or 0.01 (**) level, or not significant
(NS).

~ 20.0
CD
7.5
CD I 5.0
12.5
10.0
h-
7.5
f
¡Z
<
_J
CL
5.0
2.5
Fertilized Control Fertilized Control
AUTUMN WINTER
S R D S R D
Fertilized Control
SPRING
Figure 8. Seasonal plant tissue nitrogen content.
Fertilized Control
SUMMER
u>
OJ

34
was much greater during this time period (Table 3). Plant tissue N
content remained nearly consistent with time in the control reservoir
and root tissue N content generally exceeded that of the shoot tissue
(Fig. 8).
Seasonal N assimilation by water hyacinth ranged from 34 to 242 kg
N ha ^ for plants in the fertilizer reservoir and from <0 to 104 kg N
ha ^ for plants in the control reservoir (Table 3). There were
significant differences between seasons and reservoirs in water hyacinth
N assimilation. The detritus N content was significantly greater for
fertilized than control plants, but there were no significant
differences in detritus N content between seasons.
Data on mass balance of N in both reservoirs are shown in Table 4.
Nitrogen input from the lake was 238 kg N ha ^ with 89% of the N in the
organic fraction. Total amount of fertilizer applied during the study
period was 781 kg N ha ^, with NH^-N, NO^-N, and organic N representing
55, 30, and 15% of total fertilizer applied, respectively.
The total N assimilated by water hyacinth (live plants and
detritus) was 720 and 325 kg ha ^ yr ^ for fertilized and control
reservoirs, respectively (Table 4). Annual net N loading by detritus
was 148 and 92 kg ha ^ for fertilized and control reservoirs,
respectively (Table 4). Maximum detritus N loading occurred during
i
winter for the fertilized reservoir and during spring for the control
reservoir. This corresponded to the time of root dislodging from plants
in the two reservoirs.
The annual net N immobilized by detritus represented 21 and 28% of
the total N removed by water hyacinth in the fertilized and control
reservoirs, respectively. DeBusk et al. (1983) concluded that

Table 3. Seasonal nitrogen uptake by water hyacinth and detritus.
Fertilized
reservoir
Control reservoir
Season Water hyacinth Detritus
Water hyacinth
Detritus
1. ~ XT T ~ ^
Autumn (78) ^ 127.8
28.2
30.9
14.6
Winter (90) 33.9
42.4
-1.2
23.7
Spring (84) 167.7
37.5
104.3
29.1
Summer (88) 242.4
39.8
98.7
24.4
Test of significance t
Water hyacinth
Detritus
Season
**
NS
Month (season)
NS
*
Reservoir
**
**
^ Number of days in season.
Significant at 0.05 (*) or 0.01 (**) level or not significant (NS).

36
immobilization of N as plant detritus was 3 and 33% of standing crop
assimilation for harvested and non-harvested water hyacinth plants,
respectively. However, they did not include plant detritus trapped
within the water hyacinth mat.
The annual N assimilation by water hyacinth is low compared to N
removal rates reported for plants growing in nutrient-enriched waters.
Reddy et al. (1985) found annual N removal rates of 1726 and 1193 kg N
ha ^ yr ^ for water hyacinths growing in primary and secondary sewage
effluent, respectively. Rogers and Davis (1972) concluded that water
hyacinths could remove 2500 kg N ha ^ yr ^ if maximum growth could be
sustained. Sato and Kondo (1981) measured a maximum removal rate of
4782 kg N ha ^ yr ^ for plants growing in a nutrient medium. The low
annual N assimilation reported in the present study was due to low rates
of fertilization.
Plant uptake played a major role in removing N in both the
reservoirs (Table 4). A large portion of lake water N was present as
organic N, which was not readily available to plants. In both
reservoirs, plants derived N from mineralization of lake water organic
N, N release from underlying sediments, and mineralization of organic N
in detritus. In the fertilized reservoirs, plants also derived N from
the fertilizer N applied. Nitrogen assimiliation by water hyacinth from
the added fertilizer was calculated as follows: (Total N assimilation
by plants in the fertilized reservoir Total N assimilation by plants
in the control reservoir / Total fertilizer N added) 100.
About 51% of the added fertilizer N was taken up by the plants in
the fertilized reservoir, and the remaining 49% may have been lost
through denitrification. Reddy et al. (1982) observed a reduction of 78

Table 4. Nitrogen balance for the two reservoirs.
Fertilized Control
reservoir reservoir
Fertilizer
kg ha
Nitrogen added
NH+-N
4
430

NO~-N
234
--
Organic N
117
--
Total
781
Lake water
NHt-N
4
13
13
NO~-N
12
12
Organic N
213
213
Total
238
238
Total added
1019
238
Nitrogen removed
Water hyacinth
Shoots
354
95
Roots
218
137
Detritus
148
92
Total
720
325
Reservoir water
NH+-N
4
13
8
NO~-N
13
5
Organic N
167
156
Total
193
169
Total N accounted
913
494

38
to 81% of the agricultural drainage effluent NO^-N and NH^-N in 3.6 days
in a reservoir containing water hyacinths. DeBusk et al. (1983)
calculated that 45% of the N removed from wastewater was immobilized in
water hyacinth standing crop and detritus. About 30% of the fertilizer
N was added as NO^-N, which could be potentially lost due to
denitrification. Since water hyacinth plants prefer NH+-N over NO^-N
(Reddy and Tucker, 1983), the majority of the plant N uptake probably
came from NH*-N added through fertilizer. The role of underlying
sediment in the immobilization/mineralization, and denitrification of N
from these systems needs further investigation.
Total N recovery in the fertilized reservoir was about 90%, and
plant uptake represented about 71% of total N inputs. In the control
reservoir, total N recovery was higher than the N inputs. Plants
removed 325 kg N ha ^, as compared to 238 kg N ha ^ added. Release of N
from sediment or mineralization of N during decomposition of detritus
may account for the higher N recovery compared to total N inputs.
Ogwada (1983) found a yearly average of 150 + 34 kg KCl-extractable
inorganic N ha ^ sediment using monthly sediment N concentrations of the
same reservoirs.
Conclusions
Primary productivity of water hyacinths was influenced by ambient
air temperature, solar radiation, and nutrient composition of the
culture medium. Net detritus production (total detritus detritus lost
through decomposition) was relatively constant throughout the year and
represented 3.5 to 14.0% of the total standing crop. Detritus plant
tissue of the fertilized reservoir contained higher tissue N, compared

39
to the detritus in the control reservoir. Fertilization and increases
in ambient air temperature resulted in dislodging of root biomass.
Net N loading from detritus was 92 to 148 kg N ha ^ yr \ which is
potentially available upon decomposition. The N immobilized by detritus
represented 21 and 28% of the total N removed by water hyacinths in the
fertilized and control reservoirs, respectively.
Approximately 51% of the added fertilizer N was assimilated by
plants. The remaining 49% may have been lost through denitrification.
Total N recovery was nearly 90% in the fertilized reservoir. More N was
accounted for in the control reservoir than was added. Release of N
from the sediment or mineralization of N during decomposition of
detritus may account for the additional N recovery.

EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER HYACINTH SYSTEMS
Plant detritus (dead and decaying plant debris) is an integral part
of water hyacinth mats and comprises 3 to 14% of the total biomass (see
p. 39). It is usually derived from natural aging of plants, biological
or chemical control, and frost damage. The addition of detritus to an
aquatic system influenced several C and N transformations (Fenchel and
Jorgenson, 1977).
Nitrogen is present as NH*-N, NO^-N, and organic N in water media
available for water hyacinth production. Organic N predominates in most
water media and is not readily available for plant assimilation. Water
hyacinths were efficient users of inorganic N and plant assimilation was
a major process of N removal in aquatic systems containing water
hyacinth (Reddy and Sutton, 1984). Other N transformations in aquatic
systems resulting in removal of NO^-N or NH+-N include microbial
assimilation, nitrification/denitrification, and NH^-N volatilization
(Keeney, 1973; Bouldin et al., 1974). Addition of detritus
significantly alters the rates of these processes.
Mineralization or immobilization of N occurs during decomposition
of detritus in water and sediment. Decomposition of detritus and
subsequent N release was found to be related to C/N ratio, initial N and
fiber contents (De La Cruz and Gabriel, 1974; Godshalk and Wetzel,
1978b; Odum and Heywood, 1978; Ogwada et al., 1984).
40

41
A dense cover of floating water hyacinth depleted dissolved 0^ of
the underlying water, thus creating anaerobic conditions (Boyd, 1970;
McDonald and Wolverton, 1980; Reddy, 1981). Decomposition of plant
detritus also consumed 0^ (Nichols and Keeney, 1973; Rai and Munshi,
1979). Anaerobic conditions may restrict nitrification and favor
denitrification, which may proceed within the water hyacinth mat, in the
water column, or in the underlying sediment. Detritus also provides
energy source for denitrification. Denitrification occurred primarily
in the underlying sediment and the rate depended on NO^-N diffusion from
the water column to the sediment (Engler and Patrick, 1974; Reddy and
Graetz, 1981).
Volatilization becomes increasingly important as the water pH
increases. The partial pressure of NH^-N in equilibrium with a solution
of NH^-N increased rapidly in a pH range of 8.5 to 10.0 (Bouldin et al.,
1974). A pH of 7.0 in water occurred in areas covered with water
hyacinth with little diel variation (McDonald and Wolverton, 1980;
Reddy, 1981) which suggests that NH^-N volatilization is minimal in
areas covered with plants.
The relative role of N assimilation by water hyacinth on total N
removal from reservoirs was investigated by Reddy (1983). Approximately
40% of added ^NH^-N or ^no^-N was assimilated by plants. Less than
15 I
10% of the added N was found in the surface sediment layer. Over 40%
of the added was unaccounted for.
Information on the role of detritus in aquatic systems on
immobilization or mineralization of inorganic N is limited. The overall
objective of this study was to determine the effect of detritus on
selected N transformations in water columns with and without water

42
hyacinths. Specifically, the objectives were 1) to determine the
regulatory function of detritus on dissolved 0^ and pH of water and 2)
15 -
to determine the influence of detritus on the fate of N0^ -N and
^NH+-N in sediment- water-plant systems.
Materials and Methods
Two greenhouse studies were conducted to evaluate the effect of
detritus on the fate of labeled N0--N or NH+-N in water with and
3 4
without water hyacinth plants. Treatments evaluated were: 1) with and
without underlying sediment, 2) with and without water hyacinth plant
cover, and, 3) three rates of added water hyacinth detritus. There were
\
24 tanks in each study having dimensions of 50 cm 50 cm 25 cm depth.
Twelve of the 24 tanks contained a 2.5 cm sediment layer (1.875 kg
soil). The sediment was a Lauderhill organic soil (Lithic medisaprists)
collected at the Central Florida REC research farm in Zellwood, Florida.
The soil was air-dried and passed through a 2 mm sieve. Fifty liters of
tap water were added to sediment tanks to obtain a 20 cm water depth.
The greenhouse studies were initiated after sediment/water
equilibration of 1 week. A nutrient medium (a modified 10% Hoagland's
solution) was added to all tanks to obtain nutrient concentrations of:
15NH+-N or 15N0-N = 20.0 mg L_1; K = 23.5 mg L_1; PC^-P = 3.1 mg L_1j
Ca = 20.0 mg L ; Mg = 4.8 mg L SO^-S = 6.4 mg L ^; Fe = 0.6 mg L ^
and micronutrients. Micronutrients were applied through commercially
available liquid fertilizer (Nutrispray-Sunniland, Chase and Co.,
Sanford, Florida) to obtain concentrations of 0.2 mg Cu L ^; 1.5 mg Mn
L ^; 0.04 mg B L ^; and 0.02 mg Mo L ^.

43
Water hyacinth detritus (shoot and root material) was added at the
rates of 0, 100, and 400 mg C L *. The detritus was chopped manually to
lengths of -2 cm. The detritus for treatments with added ^NO^-N was
collected from a natural water hyacinth stand in Zellwood, Florida and
had an initial N content of 5.6 mg g 1 dry tissue. The detritus for
15 +
treatments with added NH^-N was collected from a water hyacinth stand
located in a wastewater stabilization pond at the University of Florida
wastewater treatment plant in Gainesville, Florida and had an initial N
content of 23.1 mg g dry tissue.
_2
Water hyacinths, at an initial density of 10 kg (fresh wt) m ,
were placed in 12 of 24 tanks. The plants were collected from the
University of Florida's Bivens Arm research reservoirs in Gainesville,
Florida. The plants were clipped of dead tissue and rinsed with tap
water prior to placement in tanks.
The disappearance of added inorganic N was determined by collecting
water samples at 0, 1, 2, 3, 4, 8, 15, 28 days and measuring NH+-N,
NO^-N, and total Kjeldahl N (TKN). The changes in water hyacinth fresh
weight were measured weekly. Plant samples and detritus were analyzed
for TKN. The sediment was characterized for organic and inorganic N
prior to and at the conclusion of each study. Fifty grams (dry wt) of
moist sediment samples were extracted with 2 M KC1 and analyzed for
NH^-N and NO^-N. Sediment samples were air-dried, ground by mortar and
pestle, and analyzed for TKN. The inorganic N for all samples was
determined by steam distillation (Keeney and Nelson, 1982). The TKN of
water, plant, and sediment samples were determined by micro-Kjeldahl
procedures (Nelson and Sommers, 1972; 1973; 1975). The analyses on
water, sediment, plant and detritus samples were conducted using a Micro
Mass 602 spectrometer.

44
Water pH (Orion Model 404 Specific Ion Meter), dissolved 02 (Yellow
Springs Instrument Model 54 0^ meter) and temperature were measured
every other day. Electrical conductivity (Hach Mini Conductivity Meter)
was measured weekly.
Results and Discussion
Effect of Detritus on Water Dissolved 02
The dissolved 0^ concentrations of water with added ^NO^-N and
15 +
NH^-N are shown in Figs. 9 and 10, respectively. Dissolved
concentrations remained < 5 mg L ^ in water having plant cover but lower
dissolved O2 concentrations were recorded as the rate of detritus
increased. This reflected increasing microbial 0^ demand for
respiratory functions with increasing C source (Fenchel and Jorgensen,
1977). For water without plant cover (open water), the dissolved 0^
concentrations were scattered more with time. The dissolved 0^
measurements were taken between 2:30 and 3:30 pm and should represent
near maximum concentrations on a diurnal basis (Howeler, 1972). The
increased dissolved 0^ concentrations of open water were due to an
increased rate of photosynthesis by algae during the day compared to
respiration (Reddy, 1981).
Generally the effect of decreasing dissolved 0^ concentrations with
increasing detritus was seen for open water with or without sediment.
Dissolved 0^ concentrations were generally lower in open water with
sediment compared to open water without sediment. Nichols and Keeney
(1973) reported lower dissolved concentrations for sediment-water
systems than water only. Although detritus appeared to have a role in
C>2 dynamics in water, plant cover was the primary regulator.

TIME (days)
Figure 9. Dissolved 0^ in sediment-water-plant systems with added nitrate.
Ln

DISSOLVED
20
CO
E
V-/
CVJ
O
Plants
0 mg C
L_l T T
No Sediment
1 00 mg C
L~1 *

400 mg C
L~ 1
TIME (days)
Figure 10. Dissolved in sedimentwaterplant systems with added ammonium.

47
Effect of Detritus on Water pH
The pH of water with added ^NO^-N or ^NH+-N is shown in Figs. 11
and 12, respectively. A fairly constant pH of 7.0 was noted in water
having plant cover and sediment regardless of detritus additions. The
pH decreased in water with plant cover but without sediment. The
decreasing pH was noted immediately for added ^NH^-N and after 20 days
for added ^NO^-N. The decrease in pH was less as the detritus rate
increased.
The immediate pH decrease in water with plants and added NH^-N was
probably due to production of H+ during plant NH^-N assimilation (Raven
and Smith, 1976). The H+'generated is actively exuded, partly in
exchange for cations (Franco and Munns, 1982). Plant NO^-N assimilation
occurs by exchange with another anion or by simultaneous cation
assimilation to maintain ion equilibrium (Kirkby and Mengel, 1967;
Mengel, 1974). The decreasing pH in water with plant cover but without
sediment suggested that the underlying sediment had a buffering role in
pH regulation.
The pH of water without plant cover were generally higher and more
variable than water with plant cover. Reddy (1981) reported high
mid-day pH values in ponds where algal activity was high. Bouldin et
al. (1974) found high pH values (> 8.5) for ponds containing submersed
macrophytes during sunlight hours. The pH of open water was generally
lower as the rate of detritus increased.
Effect of Detritus on Nitrogen Loss
Nitrogen loss from water with added ^N0_-N or ^NhI'-N is shown in
3 4
Figs. 13 and 14, respectively. Sediment or detritus had no apparent

TIME (days)
Figure 11. The pH of sediment-water-plant systems with added nitrate.

10
8
6
4
2
10
8
6
4
2
v-
Planta
S#d¡mnt
1 .1 1.1 i
) 4 8
-Tt+-~r v '
i i I -i, l I ,I.J
12 16 20 24 28
TIME (days)
12. The pH of sediment-water-plant systems with added ammonium.

NITRATE NITROGEN (mg
TIME (days)
Figure 13. Nitrogen loss from sediment-water-plant systems with added nitrate.

AMMONIUM NITROGEN Cmg LT
TIME (days)
Figure 14. Nitrogen loss from sediment-water-plant systems with added ammonium.

52
effect on rate of N loss in water with plant cover. Nitrate and NH^-N
disappeared within 2 and 4 days, respectively, in water with water
hyacinths.
Nitrogen loss in open water was influenced by the underlying
sediment and detritus additions. Nitrate disappeared more rapidly in
open water with sediment than without sediment. An increase in detritus
resulted in a more rapid NO^-N loss in water with or without sediment.
A rapid decrease in NO^-N after 2 days was followed by an accumulation
of NO^'N at 4 days for open water. Accumulation of NO^-N after 4 days
in open water was probably due to rapid algal assimilation followed by
turnover (death) of the algae and leaching of NO^-N from the dead algal
cells. Surface algal mats developed in open water within 2 days.
Ammonium disappeared more rapidly in open water with sediment than
without sediment. Apparently detritus did not influence NH+-N loss in
open water with sediment. However, detritus additions resulted in a
more rapid NH+-N loss in open water without sediment. Loss of NH+-N
followed by accumulation of inorganic N during the first 4 days was not
as striking as seen for NO^-N.
Plant Nitrogen Assimilation
Total plant N assimilation and the contribution of added ^NO^-N
and ^NH^-N to total N assimilation is presented in Tables 5 and 6,
respectively. Plant N assimilation was always greater for water with
underlying sediment. Part of the increased plant N assimilation in
water with sediment was due to release of N from the sediment.
Generally the contribution of added ^NO^-N or ^NH^-N to total plant N
assimilation also decreased with increasing detritus. Mineralization of

Table 5. Total plant N and ^NO^-N assimilation.
Treatment
Total uptake Labeled
Other *
(% of added
15N)
mg
rij
Without sediment
0 mg C L_1t
767
+
86
652 (85)
115
(15)
100 mg C L_1
1088
+
87
675 (62)
413
(38)
400 mg C L_1
1027
+
268
606 (59)
421
(41)
With sediment
0 mg C L_1
1220
+
29
720 (59)
500
(41)
100 mg C L_1
1171
+
235
656 (56)
515
(44)
400 mg C L_1
1112
+
163
567 (51)
545
(49)
^Carbon source was plant detritus.
^Other N sources include sediment and detritus.

Table 6. Total plant N and ^NH^-N assimilation.
Treatment
Total uptake
Labeled
Other +
15n)
mg (% of added
Vi)
Without sediment
0 mg C L ^
903
+
44
822 (91)
81
(9)
100 mg C L-1
768
+
103
697 (91)
71
(9)
400 mg C L~1
1133
+
81
807 (71)
326
(29)
With sediment
0 mg C L ^
1390
+
194
891 (64)
499
(36)
100 mg C L'1
1255
+
89
845 (67)
410
(33)
400 mg C L_1
1629
+
65
828 (51)
801
(49)
Carbon source was plant detritus.
^Other N sources include sediment and detritus.

55
detritus N was a potential N source for plant assimilation. Reddy
(1983) found that 60 to 64% of total N assimilation by water hyacinths
was derived from added ^N, while 36 to 40% was derived from sediment
and from decomposition of detritus.
The recovered by plant tissue (mg) was fairly consistent for
added labeled fertilizer but plant ^NH^-N uptake exceeded ^NO^-N
uptake. Water hyacinth appeared to be more efficient in utilizing NH+-N
than N0^-N (Reddy and Tucker, 1983). When water hyacinth growth is not
restricted by climate, rapid assimilation of added inorganic N would be
expected.
Nitrogen-15 Balance for Water Columns
A N balance for water with added ^no^-N and ^^NH^-N is presented
in Tables 7 and 8, respectively. Total recovery by water hyacinths
ranged from 57 to 72% and 70 to 89% in water with added ^no^-N and
respectively. Reddy (1983) conluded that water hyacinth N
assimilation accounted for only 40% of added ^no^-N or ^^NH^-N in a
reservoir. Algal surface mats accounted for ~8% of added ^no^-N and up
to 15% of added The algal surface mats represented a minor
portion of total microbial N assimilation. Algal activity was noted
in open water, and the four sides and bottom of the microcosm tanks were
colonized by algae.
The 15$ associated with detritus was determined for water with
added but not for added ^no -N. Less than 10% of the added
4 3
NH^-N was immobilized by detritus in water with plant cover. Newly
formed water hyacinth detritus from the plant cover was deposited during
28 days and accounted for 7% recovery in water without added
detritus or sediment.

56
15
Table 7. Mass balance of added NO^-N in sediment-waterwplant systems.
Plant
Sediment $
Unaccounted
Treatment or
algae
org
inorg
Water
Total
For
15,
PLANTS
Without
sediment
0
mg
C L_lt
65.2


ND §
65.2
34.8
100
mg
c l'1
67.5


ND
67.5
32.5
400
mg
C L_1
60.6


ND
60.6
39.4
With
sediment
0
mg
C L-1
72.0
3.2
0.4
ND
75.6
24.4
100
mg
C L-1
65.6
4.4
0.4
ND
70.4
29.6
400
mg
CL_1
56.7
3.6
0.6
ND
59.9
40.1
NO PLANTS
Without
sediment
0
mg
C L_1
ND


6.2
6.2
93.8
100
mg
c l"1
6.8


ND
6.8
93.2
400
mg
C L_1
8.2


ND
8.2
91.8
With
sediment
0
mg
c l1
ND
11.4
0.4
5.1
16.9
83.1
100
mg
cl"1
8.1
10.1
0.4
2.0
20.6
79.4
400
mg
C L-1
8.4
4.9
0.5
0.5
14.3
85.7
^Carbon source was plant detritus.
^Org, Inorg = organic and inorganic N, respectively.
§ND = Not detectable.

57
Table 8. Mass
balance
of added
^NH^-N in sediment-water
4
-plant systems.
-
Plant
Sediment:^
Unaccounted
Treatment
or algae
Detritus
Org Inorg
Water
Total
For
/o IVU v w 1. y ui
PLANTS
Without sediment
0 mg C L_1
82.2
7.3
2.3
91.8
8.2
100 mg C L_1
69.6
3.6
5.8
79.0
21.0
400 mg C L_1
80.7
7.1
3.8
91.6
8.4
With sediment
0 mg C L-1
89.1
ND §
4.3 2.6
1.0
97.0
3.0
100 mg C L"1
84.5
1.7
3.0 2.9
1.5
93.6
6.4
400 mg C L_1
82.8
3.9
3.3 3.5
1.5
95.0
5.0
NO PLANTS
Without sediment
0 mg C L_1
ND
ND
9.6
9.6
90.4
100 mg C L_1
ND
14.7
17.5
32.2
67.8
400 mg C L"1
6.1
34.7
5.2
46.0
54.0
With sediment
0 mg C L_1
14.9
ND
10.2 7.1
2.3
34.5
65.5
100 mg C L'1
15.0
5.9
11.0 8.8
5.8
46.5
53.5
400 mg C L_1
11.1
26.2
3.9 6.9
3.8
51. 9
48.1
t

§
Carbon source was plant detritus.
Org, Inorg = organic and inorganic N,
ND = Not detectable.
respectively.

58
Detritus recovery in water without plant cover increased with
increasing rate of detritus. This suggests that during periods of low
water hyacinth productivity, i.e. winter, detritus will be an important
sink for inorganic N removal. The high recovery in detritus was
suprising since the original detritus had a high N content (23 mg g
dry tissue). Therefore, the detritus used in water with added ^NO^-N
probably accounted for even more immobilization due to a low initial
N content (5 mg g ^ dry tissue).
Generally recovery in sediment was primarily organic N. Less
than 1% of the added ^NO^-N was recovered as sediment inorganic N.
However, between 3 and 7% of the added ^NH"!"-N was recovered as KC1
4
extractable inorganic N in the sediment. The lower recovery of sediment
15 15 -
inorganic N in water with added NO^-N was probably due to reduction
to gaseous N via denitrification (Engler and Patrick, 1973). Some of
the added was recovered as organic N in the water.
Plant uptake was the primary mechanism of N removal in water
having water hyacinths. The unaccounted for was lost from the
systems through a variety of possible transformations. A more thorough
investigation would be required to establish the extent of algal N
assimilation. Volatilization of NH^-N in water without plant cover and
denitrification in water with sediment are two possible mechanisms for N
removal.
Conclusions
Generally as the rate of detritus addition increased, dissolved
concentrations decreased in water with or without sediment and with or
without plant cover. The decreasing dissolved 0^ concentrations were

59
attributed to increasing heterotrophic respiration due to increasing
amounts of C. Although this general relationship existed for all
treatments, plant cover and sediment layer appeared to have more of a
regulatory role in dissolved 0^ dynamics than detritus.
Water pH was constant in water having plant cover and sediment.
The decreasing pH of water with plant cover and no sediment was
attributed to NH^-N assimilation by plants in exchange for H+. The pH
of open water was generally lower as the rate of detritus increased.
Detritus had no apparent effect on rate of N loss in water with
water hyacinths. However, N loss was more rapid in open water as the
rate of detritus increased.
Total plant ^NH^-N uptake exceeded ^N0^-N uptake. Both sediment
and detritus appeared to be a potential N source for water hyacinths.
Total recovered by water hyacinths ranged from 57 to 72% for added
15N0"-N and 70 to 89% for added 15NH+-N.
3 4
Less than 10% of the added ^NH^-N was immobilized by detritus in
water with plant cover. However, in water without plant cover, up to
35% of the added ^NH*-N was associated with detritus. This suggests
that during periods of low water hyacinth productivity, typical in cold
weather conditions, detritus is an important sink for added N.

ANAEROBIC DIGESTION OF WATER HYACINTH
The potential productivity of water hyacinth has led to its
selection as a biomass feedstock for methane generation while providing
a means for treatment of nutrient-enriched waters. Methane yields
during anaerobic digestion depended on characteristics of the feedstock
(Stack et al., 1978; Wolverton and McDonald, 1981) as well as digester
operating conditions (Hashimoto et al., 1980). Sievers and Brue (1978)
reported higher methane yields for digesters operating on swine waste as
the C/N ratio increased. They concluded that the optimum C/N range for
maximum methane production was 15.5/1 to 19/1. The optimum pH and
temperature range for anaerobic digestion was 6.7 to 7.4 (Bryant, 1979)
and 30 to 35C (House, 1981), respectively.
Biogas and methane yields have been reported for water hyacinths
using a variety of digesters. Wolverton and McDonald (1981) reported
methane yields of 0.07 to 0.20 L g 1 total solids (TS) for blended water
hyacinths. Hanisak et al. (1980) found average methane yields of 0.24 L
g volatile solids (VS) from shredded water hyacinths in 162 L digesters
at loading rates of 1.10 to 1.38 g VS L ^ day ^ and residence times of 30
to 38 days.
Chynoweth et al. (1983) reported methane yields of 0.19 and 0.28 L
g VS of water hyacinth and a 3:1 water hyacinth/primary sewage sludge
blend, respectively, in 5 L daily-fed digesters with a loading rate of
60

61
1.6 g VS L ^ day Shiralipour and Smith (1984) reported average
methane yields of 0.32 and 0.17 L g 1 VS water hyacinth shoot and root
samples, respectively, in a bioassay test of 100 ml culture volume. They
also concluded that the addition of N in growth media for water hyacinth
production increased methane yields of both shoot and root samples.
Inoculum from operating anaerobic digesters is commonly added as a
bacterial seed to initiate anaerobic digestion in new digesters (Sievers
and Brue, 1978; Wolverton and McDonald, 1981; Field et al., 1984).
Information on the effect of inoculum volume on gas production is
limited.
The objectives of this study were 1) to determine C and N
mineralization during anaerobic digestion of water hyacinth; 2) to
determine the effect of inoculum volume on gas production; and 3) to
evaluate effluent (solids and liquid) composition based on inoculum
volume.
Materials and Methods
Water hyacinths, with either high or low tissue N content, were
anaerobically digested at 35C in 55 L batch digesters containing 2.5, 5,
or 10 L of inoculum. Water hyacinths with a high N content (-34 g kg ^
dry wt plant tissue) were obtained from the wastewater treatment plant of
the Reedy Creek Utility Company, Inc., at Walt Disney World near Orlando,
Florida. Water hyacinths with a low N content (-10 g kg ^ dry wt plant
tissue) were grown in nutrient-depleted water at Sanford, Florida. Both
types of hyacinths were grown in labeled (NH^^SO^ for two weeks,
frozen and chopped to 1.6 mm length using a Hobart T 215 food processor.

62
The digesters received A.7 kg- fresh weight of the N labeled water
hyacinths and an inoculum volume of 2.5, 5 or 10 L. A control digester
received 10 L of inoculum and no plant material. The inoculum used for
plants with high N content was obtained from an operating continuous-fed
upflow digester receiving a feedstock of water hyacinth and domestic
sewage sludge in a blend ratio of 3:1 (Chynoweth et al., 1983). The
inoculum used for the plants with low N content was obtained from a
non-operating continuously-fed tank digester receiving water hyacinth as
feedstock. Each digester was buffered with 210 g NaHCO^ and tap water
was used to bring each batch digester to 54.7 kg.
Gas production was monitored for 60 days. At the end of the
digestion period, each digester was thoroughly mixed and the total
contents were emptied into a 60 L tub. The digested materials were
passed through a 1.00 mm fiberglass screen into a second 60 L tub to
separate the digested biomass sludge from the effluent. The sludge was
drained for 7 minutes and transferred into a polyethylene bag and placed
directly into a freezer. The effluent was transferred to a water
hyacinth production system.
The liquid samples from the digester effluents and screened
effluents (sludge removed) were analyzed for pH, electrical conductivity
(EC), total solids (TS), fixed solids (FS), volatile solids (VS) (APHA,
1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH^-N and NO^-N
by steam distillation (Keeney and Nelson, 1982), and chemical oxygen
demand (COD) (APHA, 1980). The screened effluent was also filtered
through a 0.2 jum membrane filter and analyzed for Ca, K, Na and Mg by
atomic absorption and P by an autoanalyzer.

63
The fresh plant material and digested sludge were freeze-dried
(Thermovac-T) and analyzed for the following: TS, FS, VS, TKN (Nelson
and Sommers, 1973), total carbon (TC) (LECO Induction Furnace 523-300),
lignin, cellulose and hemicellulose (Goering and Van Soest, 1970), and
ashed mineral constituents (Gaines and Mitchell, 1979).
Results and Discussion
Characteristics of Inocula
Characteristics of the two inocula varied considerably (Table 9).
The inoculum used for plants with high N content (high N plants)
contained higher levels of TS, VS, NH+-N, TKN, and COD than the inoculum
used for plants with low N content (low N plants). Characteristics of
inoculum depended on the type of feedstock used for digestion (Stack et
al., 1978) as well as digester operating conditions (Hashimoto et al.,
1980). The inoculum used for high and low N plants came from digesters
with feedstocks of a 3:1 water hyacinth/domestic sewage sludge blend and
water hyacinths, respectively. Ammonium accounted for 68 and 92% of the
total N of the inoculum from the water hyacinth/sewage sludge and water
hyacinth feedstocks, respectively. Ammonium was the primary N source for
methanogenic bacteria (Zeikus, 1977).
Carbon and Nitrogen Mineralization During Digestion
Biogas (CH^ and CC^) production, corrected to standard conditions
(0C and 0.1 MPa), is given in Table 10. Gas production essentially
ceased after 60 days of digestion. Cumulative biogas production at 60
days for high N plants was approximately 21% less for 2.5 L of inoculum
compared to 10 L. Furthermore, cumulative biogas production at 15 days

64
Table 9. Characteristics of the inocula used in the batch digesters.
Inocula characteristics t
NH+-N
4
no3-n
TKN COD
pH TS
FS
VS
T-1
-% of
TS---
mg L
High N plant
material
1072
48
1530 1-4200
6.3 1.75
33.5
66.5
Low N plant
material
535
21
562 784
7.7 0.29
83.1
16.9
^ COD = Chemical
oxygen demand,
TS, FS, and VS
= Total,
fixed and
volatile solids, respectively.

Table 10. Gas production during anaerobic digestion of high
and low N water hyacinth plants.
Inoculum Cumulative biogas production Total Gas Yields
.volume 15 days 30 days 60 days biogas methane
--L-- Liters L g-1 VS
High N plant material
2.5 16.4 40.8 60.3 0.21 0.14
5 28.1 53.5 73.0 0.23 0.15
10 34.5 59.1 75.4 0.20 0.13
Low N plant material
2.5 16.9 45.5 67.9 0.25 0.16
5 20.0 51.6 72.6 0.27 0.17
14.7 52.2 67.4 0.25 0.16
10

66
was over twice as great for the digester receiving the largest amount of
inoculum. However, for low N plants, the amount of inoculum did not
appreciably affect cumulative biogas production during digestion.
Cumulative biogas production at 60 days was similar for both high
and low N plants. Biogas production at 15 days was generally greater
for high N plants. It appeared that N was not a limiting factor for
total gas production in either digestion test.
Converting 60 day biogas production to biogas or methane yields (L
g ^ VS added) is also presented in Table 10. Volatile solids included
inputs from water hyacinths and inoculum. The average methane content
of the biogas was 63.7 + 5.2 % based on 18 samples. Surprisingly, biogas
and methane yields were higher for the low N plants. This was caused by
an increase of VS from inoculum used in digesters for high N plants. The
inoculum used for high N plants contained 1.75% TS (66.5% VS of TS)
(Table 9). The inoculum for low N plants contained 0.29% TS (16.9% VS
of TS). Gas production expressed in these units suggested that inoculum
volume did not appreciably affect total biogas or methane yields.
The average methane yields were 0.14 and 0.16 L g ^ VS added for
high and low N water hyacinth plants, respectively. The methane yields
were lower than those reported for continuously-fed digesters (Hanisak et
al., 1980; Chynoweth et al., 1983). Batch digestion (once fed and
t
sealed) would not promote maximum gas yields as frequent addition of
fresh substrate enhances gas production (Price and Cheremisinoff, 1981).
Shiralipour and Smith (1984) reported that methane production for
water hyacinth roots was lower than for shoots and that increasing N in
water hyacinth growth media increased methane yields. Water hyacinths
typically produced longer roots as water fertility declined (see p. 32).

67
Shoot:root dry weight ratios of water hyacinth were higher when nutrients
were not limiting and decreased significantly when plants grew in
nutrient-poor waters (Reddy, 1984). It was assumed in the present study
that gas production, both cumulative and yields, would be greater for the
high N plants.
A mass balance of N is presented in Table 11. The organic N
content decreased after anaerobic digestion for each treatment.
Mineralization of organic N to NH+-N was the primary N transformation
occurring during digestion. The total N recovered was lower for low N
water hyacinth plants. The majority of the N was recovered in the
effluent as NH+-N. Most of the N placed in digesters was recovered in
the effluent, although the proportion of NH^-N of the total N tended to
increase (Hashimoto et al., 1980; Field et al., 1984).
Approximately 30% of the organic N placed in the digesters was
recovered as organic N in the digested sludge for both high and low N
plants. The organic N recovered in the screened effluent was 15 and 36%
of the added organic N for high and low N plants, respectively. The
total organic N recovered as effluent or sludge organic N was 45 and 66%
of added organic N for high and low N plants, respectively. Therefore,
a high N content of water hyacinth resulted in more mineralization of
added organic N.
Total recovered as ^Nh1"-N in the screened effluent was 72 + 4%
4
for high N plants compared to 35 + 9% for low N plants (Table 12). The
organic recovered in digested sludge accounted for 20 + 5% of the
added from fresh water hyacinth plants regardless of N content.
Approximately 11 and 20% of the added was recovered as organic N in
the screened effluent for digested high and low N plants, respectively.

68
Table 11. Nitrogen balance for the batch digesters.
High N plant material Low N plant material
2.5 L+ 5 L 10 L 2.5 L 5 L 10 L
Nitrogen added
Water hyacinth
Organic N
Inoculum
Organic N
Inorganic N
Total
Organic N
Inorganic N
Nitrogen recovered
Screened effluent
Organic N
Inorganic N
Digested sludge
Organic N
Total
Organic N
Inorganic N
g
10.39
10.39
10.39
1.15
2.31
4.61
2.68
5.36
10.72
11.54
12.69
14.99
2.68
5.36
10.72
14.22
18.05
25.72
1.48
2.41
2.02
8.81
11.60
15.81
2.86
4.91
4.94
4.34
7.32
6.96
8.81
11.60
15.81
13.15
18.91
22.77
3.24
3.24
3.24
0.07
0.14
0.27
1.34
2.68
5.35
3.31
3.37
3.51
1.34
2.68
5.35
4.64
6.05
8.86
1.26
1.31
1.09
1.20
2.63
4.98
1.17
0.85
0.87
2.43
2.16
1.96
1.20
2.63
4.98
3.63
4.79
6.94
% Recovered 92 105 89 78 79 78
t
Volume (liters) of inoculum.

69
Table 12. Nitrogen-15 balance for the batch digesters.
High N plant material Low N plant material
2.5 L+ 5 L 10 L 2.5 L 5 L 10 L
g
15N Added
Water hyacinth
Organic N
10.39
10.39
10.39
3.24
3.24
3.24
Recovered
Screened effluent
Organic N
1.18
1.48
0.93
0.84
0.65
0.34
Inorganic N
7.06
7.78
7.68
0.79
1.25
1.34
Digested Sludge
Organic N
1.67
2.53
2.21
0.87
0.57
0.52
Total
9.91
11.79
10.82
2.50
2.46
2.20
% Recovered
95
113
104
77
76
68
t
Volume (liters) of inoculum.

70
The low total recovery of N and N for low N plants after
anaerobic digestion is difficult to explain. Nitrogen cycling during
anaerobic digestion was primarily mineralization of organic N or
immobilization of inorganic N. Volatilization of NH3~N may occur but the
* 4.
potential increases as NH^-N concentrations increase or at higher pH
values (Freney et al., 1983). Each digester received 210 g NaHCO^ as a
buffer and the pH after digestion was similar for all digester
effluents. The NH+-N concentrations after digestion were much higher for
the high N plants (Table 13).
Effluent composition
Characteristics of the digester effluents prior to sludge
separation are presented in Table 13. Generally, as the rate of
inoculum increased, there were increases in EC, NH+-N, TKN, and TS. The
COD increased with increasing inoculum volume for digesters with high N
plants.
Characteristics of the screened effluent (sludge removed) are
reported in Table 14. Removing the digested biomass sludge from the
digester effluents decreased the EC, NH^-N and TKN. The screened
effluent from the low N plants contained more Ca, Mg, K, and Na, and less
P than the screened effluent from the high N plants.
Characteristics of the fresh plant biomass and digested biomass
sludge are given in Table 15. Anaerobic digestion resulted in increases
in TC and TKN of sludge compared to fresh plant biomass. The increases
in sludge TKN after digestion of low N plants caused a reduction of the
C/N ratio from 35 to 16. The changes in TC or TKN of the digested high
N plants did not appreciably alter the C/N ratio. The digested sludge

71
Table 13. Characteristics of digester effluents before sludge removal.
Inoculum
volume
Digester
Effluent
Characteristics^
pH
EC
COD
NH*-N
4
TKN
TS
VS
FS
-i
--L--
dS
mg L
%
-% of
TS-
High N plant material
2.5
7.4
4.5
3030
189
238
0.498
40.3
59.7
5
7.6
5.1
4290
210
315
0.570
38.7
61.3
10
7.4
5.6
5050
294
406
0.610
38.9
61.1
Control
7.7
NA
4170
205
259
0.535
36.6
63.4
Low N plant material
2.5
7.3
6.6
1690
50
70
0.445
28.8
71.1
5
7.2
6.9
2340
70
112
NA
37.3
62.7
10
7.3
7.8
1640
112
154
0.488
28.8
71.2
Control
7.8
NA
109
98
120
0.315
14.0
86.0
^ COD = Chemical
oxygen
demand,
TS, FS
and VS
- Total
, fixed
and
volatile solids, respectively.
*NA = Not available.

Table 14. Characteristics of screened effluents (sludge removed) after digestion.
Inoculum Screened Effluent Characteristics^
volume pH EC NH^-N TKN Ca Mg K Na P TS VS FS
-L- dS mg L_1 % --% of TS
High N plant material
2.5 7.6 4.3 161 188 32 13 82 1140 12.8 0.357 32.4 67.6
5 7.6 4.7 212 256 30 12 80 1160 12.8 0.342 41.5 58.5
10 7.6 5.3 289 326 23 14 123 1160 11.5 0.369 41.0 59.0
Low N plant material
2.5 7.5 5.6 22 45 147 50 195 1220 3.1 0.367 30.3 69.7
5 7.4 5.9 48 72 53 53 245 1240 4.6 0.408 30.5 69.5
10 7.5 6.7 91 111 61 63 325 1200 4.5 0.425 28.1 71.9
-j-TS, VS, FS = Total, volatile and fixed solids, respectively.

Table 15. Characteristics of fresh and digested biomass residues.
Biomass
Inoculum
_
residue
volume
VS
FS
TC
TKN
C/N
Lig t Cell
Heme
Ca
K
Mg
Na
Fe
Zn
--L--
r> 1
ke"1-
g kg
8 1
High N
plant material
Fresh
834
166
385
34.0
12
43
167
182
17.6
23.5
3.2
8.0
1.60
0.52
Digested
2.5
843
157
449
37.2
12
136
159
NA t
17.6
2.2
1.8
15.3
4.04
1.20
Digested
5
827
173
446
39.3
12
130
207
180
18.4
3.8
2.2
26.0
4.76
0.92
Digested
10
846
154
441
30.8
14
111
168
NA
17.8
3.6
2.0
20.8
3.88
1.00
Low N plant material
Fresh
839
161
373
10.6
35
83
266
247
21.0
22.0
6.7
10.9
1.88
0.64
Digested
2.5
879
121
441
27.4
16
149
177
NA
22.1
2.8
2.8
14.8
5.72
1.92
Digested
5
866
134
425
26.6
16
145
181
242
24.2
2.8
2.2
17.4
7.16
2.04
Digested
10
847
153
433
25.2
17
163
163
NA
26.8
4.6
2.7
19.5
8.40
2.16
tLig, Cell, Heme = Lignin, cellulose and hemicellulose
' NA = Not available

74
had a higher lignin content compared to fresh plant biomass. The
increase in lignin was due to the loss of readily decomposable C during
anaerobic digestion. Lignin appears to be practically inert to
anaerobic digestion (Hashimoto et al., 1980) Generally, there was a
decrease in cellulose after digestion. The hemicellulose remained
similar for the fresh plant biomass and digested sludge. Anaerobic
digestion resulted in losses of K and Mg from fresh plant biomass, but
increased the concentration of sludge Ca, Na, Fe, and Zn.
Conclusions
Cumulative biogas production at 60 days was similar for high (-34 g
N kg ^ dry wt tissue) and low (-10 g N kg ^) N plants suggesting that
long term digestion of water hyacinth was not influenced by initial N
content. Effects of inoculum volume on cumulative biogas production
were seen at 15 days for high N plants but not low N plants. Conversion
of cumulative biogas production into biogas and methane yields (L g ^ VS
added) showed that low N plants produced more biogas and methane than
high N plants. This was due to increase of TS (and consequently VS) in
digesters of high N plants from the inoculum source, since cumulative gas
production was similar for both types of plants.
Mineralization of organic to ^NH^-N accounted for 72 and 35% of
15 I
added N for high and low N plants, respectively. Approximately 20% of
15
the added N was recovered as organic N in sludge for both types of
plants. A low recovery was observed for low N plants.
Increasing inoculum volume increased electrical conductivity, NH+-N,
TKN, and TS of the digester effluents. The digested biomass sludge had
higher levels of TC, TKN, lignin, Ca, Na, Fe, and Zn, and lower levels

75
of K and Mg compared to the fresh plant biomass. The C/N ratio of the
fresh plant biomass with a low tissue N content decreased from 35 to 16
after digestion. The C/N ratio of the fresh plant biomass with a high
tissue N content was the same as the digested biomass sludge (C/N=12).

TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTHS
An integrated approach of wastewater renovation using aquatic
macrophytes with utilization of biomass for energy production is
economically appealing. The plant biomass produced in these systems,
along with other wastes such as sewage sludge or animal waste could be
anaerobically digested to produce methane (Stack et al., 1981;
Shiralipour and Smith, 1984). This process generates a waste by-product
which must be disposed of, or preferably utilized to reduce the cost of
energy production, in an environmentally-safe manner. The waste
by-product consists of digested sludge and a large volume of effluent.
Integrating wastewater renovation through water hyacinth production
provides an internal option for the disposal of effluent generated
during conversion of biomass into methane.
The effluent composition of anaerobic digesters varied with type of
feedstock used in digestion (Stack et al., 1981). Information on
chemical composition of effluents from sewage or animal wastes was
readily available (Sommers, 1977; Field et al., 1984). However,
anaerobic digestion of plant biomass has only recently gained attention
in the United States and information on composition or disposal of the
effluent was limited (Hanisak et al., 1980; Atalay and Blanchar, 1984).
Digester effluents have high concentrations of BOD, NH+-N, K and Na
(Atalay and Blanchar, 1984; Field et al., 1984), while the divalent
cations and metals were concentrated in the sludge (Sommers, 1977; Field
et al., 1984).
76

77
Water hyacinth-based wastewater treatment systems have already been
evaluated for use in treating primary and secondary sewage effluents
(Wolverton and McDonald, 1979; Reddy et al., 1985) and anaerobic
digester effluent (Hanisak et al., 1980). The potential productivity of
water hyacinth in nutrient-enriched waters has led to its selection in
alternative methods of wastewater renovation, particularly in areas
where growth is not restricted by climatic limitations.
Use of water hyacinth for digester effluent treatment is
particularly attractive, because of its ability to grow in waters with
high elemental concentrations. The biomass produced could be returned
to the digester as a feedstock for methane production. Hanisak et al.
(1980) determined that 64.5% of (liquid and sludge) N in diluted
effluents from anaerobically digested water hyacinth could be
reassimilated by water hyacinths. Diluting the effluent does not
address the full potential of water hyacinth to grow under these
nutrient and salt enriched conditions. Haller et al. (1974) concluded
that water hyacinth will not live in waters containing sustained salt
concentrations in excess of 2500 mg L ^. Optimal dilution of these
concentrated effluents to obtain maximum water hyacinth yields and
nutrient removal was not reported.
The objectives of this study were to 1) evaluate water hyacinth
productivity in anaerobic digester effluents obtained from digesters
receiving different types of water hyacinth as feedstock, and, 2)
determine recovery by water hyacinth growing in digester effluents
from digested labeled water hyacinth biomass.

78
Materials and Methods
Anaerobic digester effluents were obtained from six 55 L batch
digesters containing water hyacinth with a high or low tissue N content
as feedstock. Water hyacinths with low (-10 g N kg ^ dry plant tissue)
and high (-34 g N kg ) tissue N content were grown in nutrient-depleted
water and sewage effluent, respectively. After removal from their
respective growth media, the hyacinths were grown in ^N labeled
(NH^)^SO^ nutrient solution for two weeks, frozen, and chopped to 1.6 mm
length using a Hobart T 215 food processor.
The water hyacinths were anaerobically digested for four months in
J v
55 L batch digesters. Each digester received 4.7 kg (fresh weight) of
the labeled water hyacinth, 2.5, 5, or 10 L volume of inoculum from
anaerobic digesters receiving water hyacinth as feedstock, and were
buffered with 210 g NaHCO^. After digestion, the biomass sludge was
separated from the effluent by passing the total contents of the
digesters through a 1.00 mm fiberglass screen.
The screened effluents (sludge removed) were analyzed for total
solids (TS), volatile solids (VS), fixed solids (FS) (APHA, 1980), total
Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH+-N and NO^-N by steam
distillation (Keeney and Nelson, 1982), electrical conductivity (EC)
(Hach Mini Conductivity Meter) and pH (Orion Model 404 Specific Ion
Meter). Samples passed through a 0.2 pm membrane filter were analyzed
for Na, K, Mg and Ca by atomic absorption and ortho P colorimetrically
after reacting with ammonium molybdate.
Six water hyacinth plants were placed in 10 L of undiluted or
2
diluted effluents in containers having a surface area of 0.051 m The
water hyacinth plants were collected from the University of Florida's

79
Bivens Arm research reservoirs in Gainesville, Florida. The plants were
clipped of dead tissue and acclimated to greenhouse conditions for two
weeks prior to treatments. The studies to evaluate the potential of
water hyacinth to treat digester effluents were conducted for a period
of 22 days in March and September, 1984. The daily maximum greenhouse
temperatures ranged from 19 to 37C in March and 27 to 36C in
September.
There were a total of 10 treatments, replicated three times, as
described below. Treatments 1 to 3 were diluted effluents from
digesters containing high N water hyacinths and inoculum volumes of 2.5,
'5, and 10 L, respectively. The dilutions were 1:8, 1:4, and 1:3
effluent : tap water for digester effluents with inoculum volumes of
2.5, 5, and 10 L, respectively. Treatments 4 to 6 were undiluted
effluents from digesters containing high N plants and inoculum volumes
of 2.5, 5, and 10 L, respectively. Treatments 7 to 9 were undiluted
effluents from digesters containing low N plants and inoculum volumes of
2.5, 5, and 10 L, respectively. The final treatment was a modified 10%
Hoagland's solution (p. 42) to serve as a control. Characteristics of
the inoculum and effluents are given in the Anaerobic Digestion of Water
Hyacinth chapter (pp. 60 to 75).
Plant samples collected initially and at the conclusion of the
experiment were analyzed for dry weights, TKN (Nelson and Sommers,
1973), Na, K, Ca and Mg by atomic absorption and P by an autoanalyzer.
Water samples were collected at 0, 1, 2, 3, 4, 8, 15, and 22 days and
analyzed for NH^-N and NO^-N by steam distillation (Keeney and Nelson,
1982), TKN (Nelson and Sommers, 1975), Ca, Mg, K, Na by atomic
absorption and P by an autoanalyzer. Electrical conductivity (Hach Mini

80
Conductivity Meter), pH (Orion Model 404 Specific Ion Meter) and
dissolved 0^ (Yellow Springs Instrument Model 54 O2 Meter) were measured
every other day. The analyses of plant and water samples were
conducted on a Micro Mass 602 spectrometer.
Results and Discussion
Chemical Composition of the Effluents
Characteristics of digester effluents used in the study are given
in Table 16. The initial pH of the effluent sources and nutrient medium
were similar. The digester effluents had a wide range of EC, NH^-N, TKN
and other nutrients. This provided an opportunity to evaluate water
hyacinth growth under diverse media conditions. The EC of the nutrient
medium and diluted effluents ranged from 0.7 to 2.3 dS m ^. The EC of
the undiluted effluents ranged from 4.3 to 6.7 dS m ^.
The NH+-N and TKN concentrations of the undiluted effluents from
4
digested plants with a high N content (high N plants) were greater than
those of the undiluted effluents of digested plants with a low N content
(low N plants). The NH^-N concentrations ranged from 23 to 104 rag L ^
for diluted effluents and 24 to 289 mg L ^ for undiluted effluents,
respectively.
High Na and K concentrations were noted for undiluted effluents.
The highest levels of P were in undiluted effluents of digested high N
plants and the highest levels of Ca and Mg were in undiluted effluents
of digested low N plants. The critical levels of Na, K, Ca and Mg
needed to achieve maximum water hyacinth yields are relatively unknown.
The NH*-N, K and Mg concentrations were higher than levels reported for
water hyacinths cultured in primary or secondary sewage effluent (Reddy
et al., 1985).

Table 16. Initial characteristics of the digester effluents and nutrient medium.
Inoculum
volume
pH
EC
NH+-N
4
TKN
Na
K
P
Ca
Mg
T-1
i_j
QO
mg u
Diluted effluents from high N plants
2.5
7.8
0.7
23
26
166
20
2.8
24
17
5
7.7
1.6
65
72
360
44
5.5
18
18
10
7.7
2.3
104
124
450
62
4.7
13
13
Undiluted effluents from high N plants
2.5
7.6
4.3
161
188
1140
82
12.8
32
13
5
7.6
4.7
212
256
1160
80
12.8
30
12
10
7.6
5.3
289
326
1160
123
11.5
23
14
Undiluted effluents from low N plants
2.5
7.5
5.6
24
45
1220
195
3.1
147
50
5
7.4
5.9
49
72
1240
245
4.6
53
53
10
7.5
6.7
87
111
1200
325
4.5
61
63
Nutrient medium
7.5
0.7
20
21
11
24
3.6
20
5

82
Productivity of Water Hyacinths
Total dry weight gains of water hyacinths were consistently less in
the undiluted effluents (Fig 15). Complete death of plants was observed
in four undiluted effluents. The loss of dry weight for these
treatments was probably due to leaching of soluble plant constituents
after plant death.
The highest dry weight gains were associated with the lowest EC.
However, plants survived in undiluted effluents having EC levels of 5.6
and 5.9 dS m ^ (5600 and 5900 pmhos cm ^). These EC levels were
equivalent to -2900 and 3200 mg NaCl L ^ and were higher salt
concentrations reported for water hyacinth survival (Penfound and Earle,
1948; Haller et al., 1974). The undiluted effluents had Na levels in
excess of 1100 mg L ^. Apparently water hyacinth has a wide range of
adaptability to media composition and, therefore, total salt
concentration is not a good criterium for determining plant survival.
The diluted effluents were excellent media for plant growth and the
gains in dry weight were consistently higher compared to the nutrient
medium (Fig. 15). The highest dry weight gain was in the diluted
effluent having N and P concentrations of 65 and 5.5 mg L
respectively, and a N/P ratio of 11.8:1. Sato and Kondo (1981) reported
maximum yields of water hyacinth at a N and P concentration of 50 and
13.8 mg L 1, respectively, and a N/P ratio of 3.7:1. Dry weight gains
were noted for two of the undiluted effluents. However, tissue damage
was noted in all undiluted effluents.
Tissue damage in undiluted effluents was observed within 24 hr
after study initiation. Two types of leaf tissue damage were observed.
Damaged leaves on younger shoots had burnt (brown) tips which curled up

DRY WEIGHT GAIN (g
/~\
Diluted Effluent of Undiluted Effluent of Undiluted Effluent of
High N Planta High N Plants Low N Planta Nutrient
400
300
200
100
0
-50
0^7 L6 2^3 4 4V7 3 5^6 5 6^7 07
2.5 5 10 2.5 5 10 2.5 5 10 Medium
i 4 i i 1 1 * 1,. l X - ..
-
L
JL
_L,
-
u
p
4
:T :
23 65 104 161 212 289 24 49 87 20
NH* CONCENTRATION (mg LT1)
ELECTRICAL CONDUCTIVITY CdS rrT1)
Figure 15. Dry weight gains of water hyacinths in digester effluents
and nutrient medium.

84
towards the center of the plant.- The other leaf tissue damage on young
and older leaves was curling of the entire leaf towards the center of
the leaf. Stem damage in undiluted effluents was noticeable after 2
days. Damaged stems collasped under slight manual pressure. The
destruction of chlorophyll in stems and leaves was widely observed in
the undiluted effluents.
The extent and spread of tissue damage increased in undiluted
effluents with increasing EC and NH+-N concentrations. At the end of
one week, all plants in four undiluted effluents were dead. The EC and
NH+-N concentrations of these effluents ranged from 4.3 to 6.7 dS m ^
and 87 to 289 mg N L \ respectively. The shoots began to separate from
the roots at the water surface and the submerged roots sank. Root
separation also accounted for the negative dry weight gains of plants in
undiluted effluents.
Three treatments showed occasional visual signs of tissue damage,
i.e., the nutrient medium and 2 diluted effluents. Visual signs of
plant damage but noticeable gains in plant dry weights were observed in
the diluted effluent having an initial NH+-N concentration of 104 mg L ^
and two undiluted effluents having NH^-N concentrations of 24 and 49 mg
L ^. All remaining treatments resulted in plant death, apparently due to
high EC or high NH^-N concentrations, or a combination of both.
Although water hyacinth plants struggled to survive in the
undiluted effluents, algal activity was noted in all undiluted effluent
treatments. Upon emptying the containers, algae were found attached to
the side and bottom surfaces of the plastic containers.

85
Nitrogen Removal
First-order kinetic equations were used to described NH*-N loss
from the effluents. An integrated rate equation for a first-order
reaction is expressed as:
In C / C = kt
o t
where C = initial NhT-N concentration in the effluent,
o A
C = final concentration at time = t,
k = first order rate constant (days ^).
The rate constant is calculated by solving for k:
k = In C / C 1/t
o t
The rate constants for diluted effluents ranged from 0.228 to 0.593
day1 (Table 17). The rate constants for undiluted effluents ranged
from 0.175 to 0.AA6 day
The time required for a 50% reduction in initial NH^-N
concentration generally increased with decreasing dry weight gains and
increasing NH+-N concentrations. The shortest reduction times were
associated with rapid plant growth (50% reduction in 1.12 to 3.04 days).
Plant assimilation was probably the primary mechanism of NH+-N loss in
these treatments. Reddy (1983) reported a 50% reduction of inorganic N
from agricultural drainage water in 18 days. The time required for a
50% reduction of NH^-N in effluents resulting in plant death was 1.98 to
3.96 days. Mechanisms of NH+-N loss in treatments resulting in plant
death included microbial assimilation and NH^-N volatilization. The pH
of the effluents ranged from 7.0 to 7.9 in treatments with actively
growing plants but increased from 8.2 to 9.3 in treatments where plant
death was observed and algal growth increased (APPENDIX A, Table 27).
The potential of NH^-N volatilization increases as NH+-N concentrations
increase or at higher pH values (Freney et al., 1983).

Table 17. First-order kinetic descriptions of NH^-N
loss with time.
Inoculum
Initial
nh4-n
Reduction
volume
cone.
k
time^
R2
--L--
T-1
mg L
day ^
days
Diluted effluents from high N plants
2.5
23
0.593
1.12
0.722
5
65
0.449
1.54
0.917
10
104
0.228
3.04
0.951
Undiluted effluents from high N plants
2.5
161
0.232
2.98
0.938
5
212
0.207
3.35
0.947
10
289
0.175
3.96
0.982
Undiluted effluents from
low N plants
2.5
24
0.446
1.55
0.850
5
49
0.325
2.13
0.885
10
87
0.350
1.98
0.845
Nutrient medium
20
0.281
2.47
0.973
^ Time required for 50% reduction in initial NH^-N
concentration.

67
Clock (1968) reported a 75% reduction of NO^_N in 5 days for water
hyacinths growing in secondary sewage effluent. A 75% reduction of
NH^-N in digester effluents required 2.3 to 6 days for treatments where
positive dry weight gains were observed. For treatments resulting in
+
plant death, 4 to 8 days were required to remove 75% of the NH^-N.
Nitrogen-15 plant assimilation was observed for all treatments
although a low recovery was observed in treatments resulting in plant
death (2 to 16%) (Table 18). The recovery by plants for the other
treatments ranged from 36 to 77%. The majority of the was found in
the shoot material for all treatments (54 to 73%). Approximately 75% of
the was unaccounted for in treatments resulting in plant death.
Microbial assimilation and NH^-N volatilization were probably important
NH*-N removal processes in undiluted effluents where plant death was
observed.
Plant Tissue Chemical Composition
Plants survived in the diluted effluents but death was noted for
plants in the undiluted effluents from digested high N plants. Mineral
constituents from plants growing in these effluents were analyzed to
isolate individual cation and P assimilation or loss from living and
dead plant tissue.
The concentrations of plant tissue (root and shoot fractions) Na,
t
K, P, Ca and Mg are reported in Table 19. The original plant tissue had
low concentrations of Na and P in both shoot and root material, but
higher concentrations of K, Ca and Mg in the shoots compared to the
roots. There were large increases in Na for both shoots and roots of the
surviving and dead plants. The root K concentrations increased for
surviving plants but decreased for dead plants. The P concentrations

88
Table 18. Nitrogen-15 balance for labeled effluents.
Recovered by
Inoculum Available plants N Recovery
volume 15N Roots Shoots Plants Water Unaccounted
L
mg
% of applied
Diluted effluents from high N plants
2.5
225
44
103
66
7
27
5
649
80
199
43
9
48
10
1044
101
272
36
3
61
Undiluted effluents from high N plants
2.5
1610
20
45
4
19
77
5
2120
18
41
3
27
40
10
2890
19
46
2
25
74
Undiluted <
affluents from
low N plants
2.5
236
62
119
77
6
17
5
487
71
124
40
4
56
10
869
63
73
16
6
78

89
Table 19. Distribution of nutrients in water hyacinth shoots
and roots in diluted or undiluted effluents of
digested high N plants.
.Inoculum
volume Na K P Ca Mg
--L-- g kg
SHOOTS
Diluted effluents from high N plants
2.5
13.3
13.3
2.4
16.9
9.2
5
17.5
19.3
3.9
17.8
9.1
10
18.0
23.2
5.0
15.6
8.6
Undiluted effluents
from high N plants
2.5
21.0
18.0
3.5
18.2
5.6
5
20.5
17.7
3.9
22.6
6.2
10
19.2
14.5
3.8
19.3
5.7
LSD (0.05)
4.4
7.1
1.3
3.7
1.9
Original shoot tissue
2.8
22.0
2.7
15.4
5.7
ROOTS
Diluted effluents
from high N
plants
2.5
14.5
7.1
2.1
6.1
4.6
5
17.8
8.9
2.6
6.0
4.7
10
16.0
12.4
3.4
9.0
4.2
Undiluted effluents
from high N plants
2.5
16.2
3.1
11.2
26.5
4.6
5
16.8
3.3
10.3
23.6
4.3
10
17.2
3.3
10.2
24.6
4.1
LSD (0.05)
3.0
3.7
1.9
3.0
0.7
Original root tissue
4.6
5.5
2.8
6.6
2.6

90
increased for the dead plant root material but remained similar for the
other materials. The Ca content increased for the shoots of all plants
and the roots of the dead plant material. The Mg content increased for
roots of all plants and shoots of the surviving plants.
The net assimilation or loss of plant nutrients from plants in
diluted or undiluted effluents from digested high N plants are reported
in Table 20. The plants in diluted effluents assimilated large amounts
of Na and K, apparently because these nutrients move rapidly with the
transpiration stream. The net shoot assimilation of all nutrients was
greater than the net root assimilation.
The dead plants from undiluted effluents showed a net loss of K but
net gains of the other nutrients. Potassium, Na, Ca and Mg were
reported as being rapidly lost during the early leaching phase of plant
decomposition in fresh water (Boyd, 1970b; Davis and van der Valk,
1978). Most of the Na moved into the shoot region. Generally the net
gains of P and Ca were found in dead plant roots compared to shoots of
surviving plants in diluted effluents.
Final Chemical Composition of the Effluents
Characteristics of the digester effluents and nutrient medium after
water hyacinth treatment are given in Table 21. For treatments where
plant dry weight gains were observed, generally there was large
reductions of the elements analyzed in the effluents. For the
treatments resulting in plant death, K and Mg increased and the
reductions of other elements were of lesser magnitude.
The largest reductions of EC (49 + 7% reduction), K (93 + 3%) and P
(92 + 3%) were observed in diluted effluents. The highest gains of

91
Table 20. Net assimilation or loss of plant nutrients in
diluted or undiluted effluents from digested '
high N plants.
Inoculum
.volume
Na
K
P
Ca
Mg
--L--
-2
A ^
-mg m
aay
SHOOTS
Diluted effluents
from high N plants
2.5
240
35
19
179
126
5
353
187
57
223
138
10
333
220
69
148
110
Undiluted
effluents
from high N plants
\
2.5
190
-23
10
42
4
5
201
-36
15
89
9
10
181
-92
11
35
-4
LSD (0.05)
64
101
17
62
27
ROOTS
Diluted effluents
from high N plants
2.5
217
80
14
53
59
5
273
111
22
50
61
10
227
155
32
94
48
Undiluted
effluents
from high N plants
2.5
40
-26
31
73
2
5
50
-29
32
72
1
10
70
-29
42
103
3
LSD (0.05)
57
50
27
60
14

Table 21. Characteristics of the digester effluents and nutrient medium after
water hyacinth treatment.
Inoculum
volume pH EC NH*-N TKN Na K P Ca Mg
--L-- dS mg L
Diluted effluents from high N plants
2.5
7.2
0.4
< 1
4.3
80
2
0.3
14
11
5
7.5
0.7
< 1
6.0
173
2
0.4
16
12
10
7.5
1.2
< 1
3.8
297
4
0.3
19
18
Undiluted effluents
from hiRh
N plants
2.5
8.7
4.2
22.2
39.4
873
103
10.5
11
28
5
8.6
4.4
31.9
80.2
903
134
9.9
9
24
10
8.4
4.9
49.9
87.9
950
163
7.1
11
22
Undiluted effluents
from low
N plants
2.5
8.8
4.2
< 1
4.3
450
72
1.2
23
24
5
9.2
4.6
< 1
5.1
523
98
1.2
10
33
10
9.3
4.8
< 1
8.6
547
159
1.3
10
38
Nutrient
Medium
3.5
0.7
< 1
1.8
10
1
0.5
42
17

93
plant dry weight were also noted for these effluents. Ammonium was >1.0
mg L ^ and TKN was reduced 83 to 97% in these effluents after 22 days.
Sodium reduction was in the range of 34 to 52%. Calcium and Mg
decreased for the 2.5 and 5 L inoculum diluted effluents but increased
for the 10 L inoculum diluted effluent.
Intermediate reductions of EC (25 + 3%), K (58 + 6%) and P (69 +
7%) were observed in undiluted effluents from digested low N plants.
Ammonium was < 1.0 mg L ^ and TKN was reduced 90 to 93% in these
effluents after 22 days. Sodium decreased 54 to 63% after 22 days. The
largest reductions of Ca (82 + 4%) and Mg (43 + 8%) were found in these
effluents.
Plant death was noted in all undiluted effluents from digested high
N plants. These treatments had the lowest reductions of EC (5 + 3%) and
P (26 + 10%). Plants were removed after 10 days following complete
death of aerial tissue and separation of root masses. The death of
plant tissue resulted in an increase of K and Mg in the effluents.
Sodium was reduced 18 to 23%. Calcium reduction was in the range of 52
to 70%. The loss of NH^-N (83 to 86% reduction) was probably due to NH^
volatilization and microbial assimilation.
Conclusions
The highest dry weight gains were for plants growing in diluted
effluents with EC levels ranging from 0.7 to 2.3 dS m ^ and NH")"-N
4
concentrations of 23 to 104 mg L Plants survived and grew in two
undiluted effluents with EC levels of 5.6 and 5.9 dS m'1 and Nh|-N
A
concentrations of 24 and 49 mg L_1. All other EC and NH+-N combinations
A
of undiluted effluents resulted in plant death.

94
A first-order kinetic equation was used to describe NH^-N loss with
time. Rate constants for diluted effluents ranged from 0.228 to 0.593
day 1. Rate constants for undiluted effluents ranged from 0.175 to
0.446 day ^. The time required for a 50% reduction of NH+-N was 1.12 to
3.04 days for treatments with positive water hyacinth dry weight gains.
A 50% reduction of NH*-N in treatments resulting in plant death required
1.98 to 3.96 days. Plant assimilation was one of the primary mechanisms
of NH^-N loss in the systems with actively growing plants. Microbial
assimilation and NH^-N volatilization were probably important mechanisms
of NH+-N removal for treatments resulting in plant death.
Plant assimilation accounted for a 36 to 77% recovery of effluent
for surviving plants. Only 2 to 16% of the was recovered in
dead plant tissue. Approximately 75% of the was unaccounted for in
effluents resulting in plant death.
Surviving plants assimilated large amounts of Na, K, Ca and Mg
while dead plants lost K and had small gains of Ca and P. Sodium
accumulated in dead plant tissue. Death was attributed to an
indiscriminate salt injury and/or NH^-N toxicity.
The largest reductions of EC, K and P were observed in diluted
effluents. The highest plant dry weight gains were also found in these
effluents. Potassium and Mg increased in effluents where plant death
was noted.

DECOMPOSITION OF FRESH AND ANAEROBICALLY
DIGESTED PLANT BIOMASS IN SOIL
Anaerobic digestion of plant biomass, sewage sludge, or animal
wastes generates a waste by-product which must be disposed of, or
preferably utilized, in an environmentally-safe manner. Disposal of the
digested sludge by land application is one option often considered
(Miller, 1974; Terry et al., 1979; Atalay and Blanchar, 1984). The
digested biomass sludge differs chemically from the fresh plant biomass.
A consequence of anaerobic digestion is a reduction of the more readily
decomposable C of the plant tissue during production of CH^ and CC^.
Many sludges contained relatively large amounts of Ca, Mg, P and Zn and
lower contents of soluble elements such as K (Sommers, 1977)
Miller (1974) concluded that anaerobically digested sewage sludge
was rather resistant to further decomposition with a maximum of 20% of
the added C was evolved as CO^ during a 6-month incubation. Tester et
al. (1977) reported 16% of added C from composted sewage sludge was
evolved as CO^ during 54 days of incubation. Hsieh et al. (1981a) showed
that activated sludge had a much higher C mineralization rate compared to
digested sludge due to a larger portion of active organic C.
Epstein et al. (1978) determined that the percentage of added N
mineralized from digested sludge remained essentially constant
irrespective of application rate. However, Ryan et al. (1973) and Stark
95

96
and Clapp (1980) observed an enhanced rate of N mineralization with
increasing rate of sewage sludge application. Nitrogen mineralization
potential was found to be 30 and 38% of organic N in activated and
digested sludge, respectively, during 60 days of incubation (Hsieh et al.
1981b).
Much of the available information dealt with land application of
anaerobically digested sewage sludge (Miller, 1974; Terry et al., 1979),
and limited data was reported on the decomposition of sludge obtained
from the anaerobic digestion of plant biomass (Atalay and Blanchar,
1984). The objective of this study was to evaluate the decomposition and
N mineralization rates of anaerobically digested plant biomass added to
soil. Four materials were evaluated: fresh plant biomass with a low or
high tissue N content, and their respective anerobically digested
residues.
Materials and Methods
Surface (0-15 cm depth) soil samples of a Kendrick fine sand
(Arenic paleudult) were collected at the Agronomy Farm, University of
Florida in Gainesville, Florida. The soil was air-dried and passed
through a 2 mm sieve. The soil had a particle size distribution of
92.9% sand, 4.6% silt, and 2.5% clay. The CEC was 3.44 cmol(+) kg *
soil with a base saturation of 47%.
Water hyacinths with low (-10 g N kg 1 dry plant tissue) and high
(-34 g N kg ^) tissue N content were grown in nutrient-depleted water and
sewage effluent, respectively. After removal from their respective
growth media, the hyacinths were grown in 15N labeled (NH,)S0. nutrient
4 2 4
solution for two weeks, frozen, and chopped to 1.6 mm length using a
Hobart T 215 food processor.

97
The water hyacinths were anaerobically digested for four months in
55 L batch digesters. Each digester received 4.7 kg (fresh weight) of
the labeled water hyacinth, 5 L of an inoculum from anaerobic
digesters receiving water hyacinth as feedstock, and were buffered with
210 g NaHCO^. After digestion, the biomass sludge was separated from
the effluent by passing the total contents of the digester through a
1.00 mm fiberglass screen.
Samples of the fresh water hyacinth and anaerobically digested
water hyacinth sludge were freeze-dried (Thermovac T) and ground through
a 0.84 mm screen of a Wiley Mill. The freeze-dried materials were
characterized for lignin, cellulose and hemicellulose (Goering and Van
Soest, 1970), ashed mineral constituents (Gaines and Mitchell, 1979),
total solids (TS), volatile solids (VS), total C (TC) (LECO Induction
Furnace 523-300), and total Kjeldahl N (TKN) (Nelson and Sommers, 1973).
Carbon/nitrogen (C/N) ratios of the four residues were calculated from
percentage TC and TKN.
Fifty gram soil samples were preincubated for 5 days at a water
content adjusted to 0.01 MPa before addition of the residues. The
freeze-dried materials were added to the soil at a rate of 5 g (dry wt)
kg 1 soil (10 Mg ha ^) and incubated for 90 days at 27C. Water content
was adjusted to 0.01 MPa every 15 days. Ambient laboratory air, with CO^
and NH^-N removed by 3 M NaOH and 4 M H^SO^ traps, respectively, was
pumped through the incubation flask at a rate of 50 ml min ^. The CO^
evolved from soil samples was collected in 0.1 M NaOH traps and
determined by titration with acid after reacting with saturated BaC^.
The percentage C evolved with time was calculated by subtracting C
evolved as CO^ of the control soil (no organic C amendment) from the

98
various treatments and dividing by the amount of C added for each
residue.
Soil samples were analyzed at 0, 30, 60, and 90 days for 2 M
KCl-extractable NH+-N and NO^-N by steam distillation (Keeney and Nelson,
1982), TKN (Nelson and Sommers, 1972), Mehlich I extractable Ca, K, Mg,
Na, Fe, Zn, and P (Melich, 1953), organic C by the Walkley-Black method
(Nelson and Sommers, 1982), and pH. The ^N analyses were conducted on
a Micro Mass 602 spectrometer.
Results and Discussion
Plant Residue Characterization
Characteristics of the fresh plant biomass and anaerobically
digested biomass sludge are presented in Table 22. The TC and TKN
concentrations of the digested sludges were higher than their respective
fresh plant materials. Total C was not significantly different for low
and high N fresh plant biomass or digested sludge. The C/N ratio of the
fresh plant biomass with a low N content (low N plant biomass) decreased
from 35 to 16 after digestion. The C/N ratio of fresh plant biomass
with a high N content (high N plant biomass) did not change during
digestion.
Lignin content was significantly higher in digested sludges due to
loss of readily-decomposable C during anaerobic digestion. The low N
fresh plant biomass contained approximately twice as much lignin as the
high N fresh plant biomass. Moore and Bjorndal (1984, Unpublished
results, Univ. of Florida, Gainesville) concluded that water hyacinth
roots, in general, have higher lignin content than shoots. The lignin
content of the low N fresh plant biomass was approximately double that of

99
Table 22. Characteristics of the fresh and digested plant biomass
Chemical Low N plant biomass High N plant biomass
constituent Fresh Digested Fresh Digested
g kg of biomass
Volatile solids
839 bf
866 a
837 b
825 c
Ash
161 b
134 c
166 b
173 a
Total carbon
373 b
425 a
385 b
446 a
Lignin
83 b
145 a
43 c
130 a
Cellulose
266 a
180 b
167 b
206 b
Hemicellulose
247 a
243 a
183 b
180 b
Total nitrogen
10.6 c
26.6 b
34.0 a
39.3 a
Calcium
21.0 b
24.2 a
17.6 c
18.4 c
Potassium
22.2 a
2.8 b
23.5 a
3.8 b
Magnesium
6.7 a
2.2 a
3.2 b
2.2 c
Sodium
10.9 c
17.4 b
8.0 d
26.0 a
Iron
1.9 c
7.2 a
1.6 c
4.7 b
Zinc
0.7 c
2.0 a
0.5 d
0.9 b
C/N ratio
t
35 a
16 b
12 c
12 c
'Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.

100
the high N fresh biomass due to a larger rooting mass associated with
plants grown in nutrient-depleted water. The low N fresh and digested
plant biomass contained more hemicellulose than high N materials.
Cellulose content decreased for low N fresh plant biomass after
digestion, but increased for high N fresh plant biomass.
Anaerobic digestion resulted in a large loss of K from fresh plant
biomass. Potassium is relatively soluble and is used for translocation
of anions via the zylem and phloem, enzyme activation, and stomatal
movements. Additional K may be required to enhance the microbial
degradation of digested biomass sludges when added to soil. The increase
in Na concentration in the digested sludges was due to the addition of
NaHCO^ buffer to stabilize digester pH. The digestion process resulted
in increases in the relative amounts of Ca, Na, Fe, and Zn in the sludge
compared to fresh plant biomass.
Carbon and Nitrogen Mineralization
Carbon evolution from the four materials, reported as mg C evolved
as CO^ per g residue C added, is shown in Fig. 16. The soils with fresh
plant biomass additions always released more CO^ compared to that from
digested biomass sludges. The two sludges showed similar evolution rates
during the first 40 days of incubation despite their difference in TKN.
After 40 days, the CO^ evolution of high N digested sludge increased and
i
continued to increase for the remaining incubation period. This suggests
a possible lag period during which the microbial population is adjusting
to those species which tolerate high levels of Na. However, there were
no significant differences in CO^ evolution from the two sludges
throughout the incubation period (Duncan's multiple range test).
Overall C decomposition cannot be described by simple kinetic
equations. However, first-order kinetics describe plant residue or

Figure 16. Carbon evolution from soil applied fresh
and digested plant biomass.

102
animal waste decomposition if the overall decomposition sequence is
presented as occurring in stages (Gilmour et al., 1977; Hunt, 1977;
Reddy et al., 1980). Each stage is thought to represent the sequential
ease of C constituent decomposition, i.e. soluble sugars and starch,
cellulose and hemicellulose, and lignin.
The rate equation for a first-order reaction is expressed as
-dC./dt = k.C.
i li
where subscript i refers to a particular stage of C decomposition. The
integrated first-order rate equation is
Ct. = C.exp(-k.t)
i i r i
where C. = C at beginning of a decomposition stage,
Ct^ = C remaining at end of a decomposition stage at time = t,
k. = first-order rate constant.
i
Therefore, a rate constant can be calculated for each decomposition
stage of an organic C material.
A graphical representation of the stages and their respective rate
constants of the materials used in this study are shown in Fig. 17.
Decomposition of fresh plant biomass required a three stage first-order
kinetic description. Rate constants for the first stage (soluble sugars
and starch) were 0.0441 and 0.0222 day ^ for high and low N plant
biomass, respectively. This stage of decomposition was essentially
completed in 4 days. A longer time was required during the second stage
(cellulose and hemicellulose) of decomposition for low N plant material
due to a higher content of these C constituents (Table 22). The rate
constants for the final stage (lignin) of decomposition were low for
both materials.

PERCENT C REMAINING
TIME (days)
Figure 17. Decomposition stages and rate constants
of fresh and digested plant biomass added
to soil

104
Decomposition of digested biomass sludges was adequately described
in two stages. The conversion of soluble sugars and starch to CH^ and
CO^ during anaerobic digestion eliminated the first stage of soluble C
degradation for the digested sludges. Therefore, the first stage of
sludge decomposition corresponded to the second stage (cellulose and
hemicellulose) of fresh plant biomass. The longer second stage
corresponded to the final stage (lignin) of fresh plant biomass.
Soil NO^-N concentrations during incubation are presented in Table
23. Soil NH*-N concentrations (APPENDIX B, Table 29) were < 2 mg kg 1
during the incubation period. Initial NO^-N concentrations were similar
for all treatments. No NO^-N accumulated in the soil amended with low N
fresh plant biomass until 60 days had elapsed. The concentrations of
soil NO^-N were similar for both digested biomass sludges throughout the
incubation despite the higher TKN concentration of the high N sludge.
Although the C/N ratios of high N fresh biomass or digested biomass
sludge were the same, the amount of NO^-N that accumulated in the soil
was more than double for fresh biomass compared to digested biomass
sludge.
A summary of C and ^N mineralization during the incubation period
is presented in Table 24. Approximately 8% of the applied N was
recovered as 15N03-N at 90 days for both sludges. Tester et al. (1977)
reported 6% of composted sewage sludge N mineralized to NO^-N in 54
days. In contrast, 3 and 33% of applied N was recovered as ^NO^-N for
fresh plant biomass with a low and high N content, respectively. Ogwada
(1983) incorporated shredded 15N labeled water hyacinth with a C/N ratio
of 35 into a Myakka fine sand at a rate of 20 Mg dry wt ha'1. At 60
days, 33% of N from the soil-incorporated water hyacinth was

105
Table 23.
Soil NO^-N concentration from added fresh and digested
plant biomass.
Low N plant biomass High N plant biomass
Day Control Fresh Digested Fresh Digested
. -1
. mg kg soil
0
9.1
a +
8.2
ab
7.9
abc
8.4
ab
6.6
c
30
11.0
c
0.9
d
20.0
b
51.4
a
17.2
b
60
13.2
c
1.0
d
24.4
b
70.3
a
24.8
b
90
14.5
c
4.8
d
28.4
b
68.8
a
30.3
b
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.

Table 24. Carbon and N mineralization from added
fresh and digested plant biomass.
Low N plant
biomass
High N
plant
biomass
Fresh
Digested
Fresh
Digested
/o OX aflQGQ
l. o tr
N
Day
30
c
22.4 b t
11.2 c
34.2
a
9.8c
15n
ND *
4.7 b
24.8
a
3.8 b
Day
60
c
32.5 b
14.9 c
41.6
a
16.1 c
15n
ND
6.2 b
34.3
a
6.0 b
Day
90
c
39.0 b
19.1 c
49.9
a
23.1 c
15n
3.3 c
7.7 b
33.3
a
7.7 b
t

Values with same letter within rows are not
significantly different at 0.05 level by Duncan
Multiple Range Test.
ND = not detectable.

107
mineralized and 20% of the initial N was recovered in sorhgum-
sudangrass plants.
After 90 days of incubation, approximately 20% of the added C of
digested biomass sludges had evolved as CO^ compared to 39 and 50% of
fresh plant biomass with a low and high N content, respectively. More
than half of the C evolved from fresh plant biomass occurred within the
first 10 days.
The percentage of C evolved with time from anaerobically digested
biomass sludge is similar to results of Miller (1974) and Tester et al.
(1977) for anaerobically digested and composted sewage sludge. Miller
(1974) reported a maximum of 20% of the added C was evolved as CC^ during
a 6-month incubation period. Tester et al. (1977) reported that 16% of
the added C was evolved as CO^ from composted sewage sludge during 54
days of incubation. However, Terry et al. (1979) found a total of 26 to
42% of C was evolved as CO^ from anaerobically digested sewage sludge
during 130 days of incubation.
Carbon/nitrogen ratio is commonly used as a guideline for
predicting the relative decomposability or mineralization potential of
organic materials added to soil (Reddy et al., 1980). The high C/N ratio
of low N fresh plant biomass (C/N = 35) resulted in immobilization of
inorganic N. The low C/N ratio of high N fresh plant biomass (C/N = 12)
t
resulted in rapid mineralization of organic N. However, C/N ratios may
have limited applicability for predicting N transformations of digested
biomass sludges. Both high N fresh biomass and digested biomass sludge
had C/N ratios of 12, but only 8% of the applied N was recovered as
^NO^-N from digested sludge compared to 33% from fresh plant biomass.

108
Other Soil Parameters
Soil pH increased by one and two pH units after addition of
anaerobically digested biomass sludge with a low and high N content,
respectively (Table 25). The addition of fresh plant biomass did not
appreciately alter the initial soil pH. During the 90 day incubation,
soil pH generally decreased with all residue additions, except for low N
fresh plant biomass.
The Na content of the digested biomass sludges explains, in part,
the increase in the initial soil pH. Atalay and Blanchar (1984) found
that addition of anaerobically digested plant residue to soil increased
pH from 5.5 to 7.6. They attributed the pH increase to a limestone
*,
buffer used during anaerobic digestion.
The addition of fresh or anaerobically-digested water hyacinth
increased Mehlich I extractable soil constituents after 90 days of
incubation (Table 26). Mehlich I extractable soil constituents at 0, 30
and 60 days of incubation are presented in APPENDIX B, Tables 30 to 32.
The increases were a direct reflection of the mineral composition of the
respective residue (Table 22). There was a large increase in soil K with
fresh plant biomass additions compared to digested biomass sludges. All
treatments resulted in large increases of soil Na and Ca. Parra and
Hortenstine (1976) concluded that fresh water hyacinths contained
appreciable amounts of soluble salts and that crops susceptible to salt
injury should not be planted immediately after soil additions.

109
Table 25. Soil pH (1:2 w/v) from added fresh and digested
plant biomass.
Low N plant biomass
High N plant biomass
Day
Control
Fresh
Digested
Fresh
Digested
0
5.44
d +
5.38 d
6.49 b
5.73 c
7.34 a
30
5.48
c
5.99 b
6.27 a
4.90 d
6.21 a
60
4.99
c
5.96 ab
6.03 a
4.30 d
5.77 b
90
5.42
b
5.94 a
6.03 a
4.72 c
6.02 a
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.

110
Table 26. Mehlich I extractable constituents at Day 90 from added
fresh and digested plant biomass.
Chemical
constituent
Control
Low N plant biomass
High N plant
biomass
Fresh
Digested
Fresh
Digested
, -1
mg Kg soil
Calcium
252 e f
361 ab
373 a
327 d
348 be
Potassium
25 e
115 b
40 d
119 a
47 c
Magnesium
24 d
59 a
35 c
39 b
35 c
Sodium
3 e
53 c
72 b
41 d
117 a
Iron
14 d
14 d
20 a
15 c
18 b
Zinc
3.3 e
5.7 d
11.3 a
6.0 c
7.2 1
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.

Ill
Conclusions
After 90 days of incubation, approximately 20% of the added C of the
digested biomass sludges had evolved as CO^ compared to 39 and 50% of the
fresh plant biomass with a low and high N content, respectively.
Decomposition of fresh plant biomas followed a three stage first-order
kinetic description. Decomposition of digested sludge was adequately
described by two stage first-order kinetics.
Mineralization of organic N to ^NO^-N accounted for approximately
8% of applied N for both digested biomass sludges at the end of 90 days.
Nitrogen mineralization accounted for 3 and 33% of applied N for fresh
plant biomass with a low and high N content, respectively.
The soil pH increased after addition of digested biomass sludge,
but was not appreciately altered after addition of fresh plant biomass.
The Na content of digested sludges was attributed as the primary factor
for increasing soil pH.
Increases in Mehlich I extractable soil constituents were a direct
reflection of the mineral composition of fresh or digested plant
biomass. The high Na concentration of digested biomass sludge suggests
pretreating the sludge to remove some of the salts and selecting salt
tolerant plants if the sludge is used as a soil amendment.

MASS BALANCE OF NITROGEN IN AN INTEGRATED
"BIOMASS FOR ENERGY SYSTEM
A series of experiments were conducted to evaluate N cycling in
three components of an integrated "biomass for energy" system, i.e.
water hyacinth production, anaerobic digestion of hyacinth biomass, and
recycling of digester effluent and sludge. Two types of water hyacinth
biomass production systems were evaluated; 1) plants growing in
t
nutrient-enriched waters representing wastewater treatment systems
(sewage and digester effluents, animal wastes or agricultural drainage
water), and plants growing in nutrient-limited systems representing
natural waters receiving low external N inputs.
Nutrient-Enriched Systems
Nitrogen cycling in an integrated system for water hyacinths
growing in nutrient-enriched systems is summarized in Fig. 18.
Water Hyacinth Production
-2 -1
A wide range of productivity (5 to 64 g dry wt m day ) has been
recorded for plants growing in nutripnt-enriched waters (Boyd, 1976;
Hanisak et al., 1980; Reddy and DeBusk, 1983; Reddy et al., 1985).
Plants growing in nutrient-enriched media generally have an N content of
30 to 40 g N kg 1 of dry tissue (Boyd, 1976; Wolverton and McDonald,
1979; Reddy et al., 1985). In this study the tissue N content of plants
growing in sewage effluent was 35 g N kg ^ of dry biomass.
112

Figure 18. Nitrogen cycling in an integrated system for water hyacinths growing
in nutrient-enriched systems. Numbers in parentheses are percentages
of the initial 35 g N placed in the anaerobic digester.
113

114
Although plant assimilation is a major process for N removal in
ponds of water hyacinths, other N transformations may occur in the root
zone, water column and underlying sediment. These processes account for
much of the loss of N from the system. For example, a dense mat of
water hyacinth could potentially enhance denitrification of NO^-N
present in the water column (Boyd, 1970; Reddy, 1981). Total N
reduction in water hyacinth systems was reported in a range from 65 to
94% of various wastewaters (Sheffield, 1967; Clock, 1968; Hanisak et
al., 1980; Reddy et al., 1982).
Total fertilizer N recovered by plants in this study ranged from
50% in a field study to 90% in controlled-greenhouse studies. Results
indicated that detritus accounted from 3 to 14% of the total biomass
produced in water hyacinth ponds. Up to 28% of the total plant N was
found in detritus, which could potentially be deposited at the sediment-
water interface.
Anaerobic Digestion of Hyacinth Biomass
Anaerobic digestion of biomass with high tissue N and low C/N ratio
resulted in mineralization of 70% of plant organic N to NH+-N. Total N
recovered for digested high N plants was -100% of the added N which
agreed with results by Hashimoto et al. (1980) and Field et al. (1984).
After digestion, 80% of the plant organic N was recovered in the
t
effluent and 20% was recovered in the sludge.
Waste By-Product Recycling
Undiluted and diluted effluents were used as nutrient sources to
produce additional water hyacinth biomass and for potential N recovery.
Plants growing in undiluted effluents did not survive the combination of
high salt and NH^-N concentrations. However, diluting the digester
effluents provided a media which stimulated water hyacinth growth.

115
Plant assimilation of effluent N recycled 38% of the initial N
placed in the digester. Nitrogen loss from this component of the
integrated system accounted for 42% of the N placed in the digester.
Possible N loss mechanisms during recycling of digester effluent include
algal assimilation and NH^-N volatilization.
The digested sludge contained 20% of the initial N placed in the
digester. The sludge N could potentially be recovered during crop
production after land application. The sludge was fairly resistant to
decomposition in soil and only 20% of the sludge C was evolved as CO^ in
90 days of incubation. A low decomposition rate of sludge applied to
soil was attributed to loss of the more readily-decomposable C
constituents during anaerobic digestion. The decomposition rates
observed in this study for digested biomass sludge applied to soil were
found to be similar to those reported for digested sewage sludge
(Miller, 1974; Tester et al., 1977). Nitrogen mineralization of sludge
organic N during 90 days of decomposition in soil accounted for only 2%
of the initial N placed in the digester.
Fresh water hyacinth biomass was also added to soil to compare its
decomposition to that of digested sludge. Approximately 50% of the
biomass C was evolved as CO^ during 90 days of incubation. Nitrogen
mineralization of biomass organic N accounted for 34% of the initial
plant N placed in the soil. Mixing the fresh water hyacinth biomass
with anaerobically digested sludge may enhance the decomposition of the
sludge.
Total N recovery by sludge and effluent recycling in the integrated
"biomass for energy" system was 60% of the initial N placed in the
digester for high N plants. The remaining 40% was lost from the system
during effluent recycling in a water hyacinth production system.

116
Nutrient-Limited Systems
Nitrogen cycling in an integrated system for water hyacinths
growing in nutrient-limited systems is summarized in Fig. 19.
Water Hyacinth Production
-2 -1
Growth rates of 2 to 29 g dry wt m day have been reported for
plants in natural waters of central and south Florida (Yount and
Crossman, 1970; DeBusk et al., 1981). Plants growing in nutrient-
limited systems generally have a low N content. In this study, tissue N
content of plants grown in tap water without nutrients was 10 g N kg ^
of dry biomass. In addition to a low N content, the shoot:root dry
weight ratio decreased as nutrient availability decreased in the water
media (Reddy, 1984). The shoot:root dry weight ratio of plants with a
low N content was 1.6 compared to 4.2 of plants with a high N content.
Anaerobic Digestion of Plant Biomass
Shiralipour and Smith (1984) concluded that increasing N in the
growth medium used for water hyacinth production increased methane
yields during anaerobic digestion. Surprisingly, gas yields were higher
for plants with a low N content compared to plants with a high N
content. However, the gas yield differences were attributed to
characteristics of inoculum used during digestion. Only 75% of the
initial N placed in the digester was recovered in the effluent and
sludge for low N plants. The low total N recovery is difficult to
explain. Only 35% of the plant organic N placed in the digester was
mineralized to NH+-N compared to 70% mineralization for high N plants.
Approximately 40% of the initial N placed in the digester remained as
organic N after digestion. After digestion, 54% of the initial plant
organic N was recovered in the effluent while 20% was recovered in the
digested sludge.

Figure 19. Nitrogen cycling in an integrated system for water hyacinths growing
in nutrient-limited systems. Numbers in parentheses are percentages
of the initial 10 g N placed in the anaerobic digester.
117

118
Waste By-Product Recycling
The undiluted effluent from digested plants with a low N content
was recycled directly to a water hyacinth production system. Plants
survived in undiluted effluents due to lower NH+-N concentrations.
4
Approximately 21% of the N placed in the digester was reassimilated by
water hyacinths during effluent recycling. Nitrogen loss during
digestion and effluent recycling accounted for 52% of the initial N
placed in the digester.
The digested sludge contained 20% of the initial N placed in the
digester. Similar decomposition rates were noted for both digested
sludges applied to soil despite their differences of initial N content
(27 and 39 g N kg ^ dry sludge). The mineralization of sludge organic N
accounted for 2% of the initial N placed in the digester.
The fresh water hyacinth biomass decomposed rapidly in soil but
only 3% of the applied N was mineralized to NO^'N. Total N recovery by
sludge and effluent recycling was 48% of the initial N placed in the
digester for low N plants. The remaining N was lost from the system
during anaerobic digestion and effluent recycling.

CONCLUSIONS
The three components of the integrated "biomass for energy" system
were 1) the water hyacinth biomass production system; 2) anaerobic
digestion of water hyacinth biomass, and 3) waste recycling of digested
biomass sludge and effluent. Nitrogen cycling was investigated for each
component of the integrated system.
Water Hyacinth Productivity and Detritus Production
Productivity of water hyacinth was influenced by ambient air
temperature, solar radiation, and nutrient compostion of the medium.
The highest net productivity occurred during spring and summer and over
75% of the biomass produced was recorded during this time period. The
detritus production exceeded net biomass production during winter
regardless of water fertility.
Seasonal yields of water hyacinth ranged from 1.9 to 23.1 Mg (dry
wt) ha ^ and -0.2 to 10.2 Mg ha ^ for plants growing in eutrophic lake
water with and without added nutrient, respectively. Detritus comprised
3 to 14% of the total biomass and detritus production was not
significant between reservoirs or seasons. Although detritus production
was similar for both reservoirs, fertilization resulted in significant
increases in detritus N content. Nitrogen loading to the reservoirs
from detritus was 148 and 92 kg N ha 1 yr 1 for plants grown in
eutrophic lake water with and without added nutrients, respectively.
119

120
Approximately 50% of the fertilizer N was recovered by plants.
Total N assimilated by water hyacinth (live plants and detritus) was 720
and 325 kg N ha ^ yr ^ for plants grown in eutrophic lake water with and
without added nutrients, respectively.
Detritus and Nitrogen Transfromations
Detritus had no apparent effect on rate of N loss in water with
water hyacinth plant cover due to rapid plant assimilation. However, N
loss in water without plant cover was more rapid with detritus
additions. Both sediment and detritus appeared to be potential N
sources for plant assimilation.
Total N recovered by water hyacinth ranged from 57 to 72% for added
15N0l-N and 70 to 89% for added 15NHt-N. Less than 10% of added 15Nh1'-N
3 4 4
was immobilized by detritus in water with plant cover. However, up to
35% of the added ^NH"!"-N was associated with detritus in water without
4
plant cover. This suggests that during periods of low water hyacinth
productivity, i.e. winter, detritus is an important sink for added N.
Increasing amounts of detritus generally decreased dissolved 0^
concentrations of water. The pH of water without plant cover generally
was lower when detritus was added.
t
Anaerobic Digestion of Water Hyacinth
Initial water hyacinth N content and volume of inoculum did not
affect long term (60 days) biogas production. At 15 days, biogas
production was generally greater for plants with a high N content.
Inoculum volume showed little effect of biogas production for low N
plants throughout incubation. However, a larger volume of inoculum

121
increased biogas production for plants with a high N content after 15
days of digestion.
Removing the biomass sludge from digester effluents decreased the
electrical conductivity, NH^-N and TKN of the effluent. Anaerobic
digestion resulted in a loss of K and Mg from fresh plant biomass but
increased lignin, total C, TKN, Ca, Na, Fe and Zn of the digested
sludge. Generally cellulose decreased during digestion and
hemicellulose remained unchanged.
Mineralization of organic N to NH+-N was the primary N
transformation occurring during anaerobic digestion. Approximately 20%
of the organic N placed in the digesters was recovered as sludge organic
N. Net mineralization of organic N to NH^-N was 70 and 35% of the added
organic N of plants with a high and low N content, respectively.
Digester Effluent Recyling
The initial electrical conductivities and NH+-N concentrations of
A
the digester effluents ranged from 0.7 to 6.7 dS m ^ and 23 to 289 mg N
L The highest water hyacinth dry weight gains were associated with
the lowest electrical conductivities. However, plants survived in
effluents having electrical conductivities of 5.6 and 5.9 dS m Plant
death was observed in four undiluted effluents. Additional information
t
would be needed to establish optimum dilution of anaerobic digester
effluents to promote maximum biomass yields.
First-order kinetic equations were used to describe NH+-N loss from
effluent with k values ranging from 0.175 to 0.593 day ^. Generally the
highest k values were associated with rapid plant growth. Plant ^N
assimilation was observed for all effluents although plant death

122
resulted in low recoveries (2 to 16%). The N recovery for other
15
effluents ranged from 36 to 77%. The majority of the was recovered
in shoot material.
Plants grown in diluted effluents assimilated large amounts of Na
and K and net shoot assimilation of all nutrients was greater than net
root assimilation. Dead plants showed a net loss of K but net gains of
other nutrients.
Digested Sludge Recycling
Decomposition of fresh and anaerobically digested water hyacinth
biomass added to soil was evaluated by CO^ evolution and
mineralization. Approximately 39 and 50% of the added C was evolved as
CO^ in 90 days for fresh plant biomass with a low and high tissue N
content, respectively. Approximately 20% of the added C was evolved as
CO^ for both digested sludges in 90 days. Decomposition of fresh plant
biomass required a three stage first-order kinetic description of C loss
compared to a two stage first-order description for digested sludge.
Apparently the loss of readily-decomposable C during anaerobic digestion
eliminated the first stage of decomposition in soil.
Only 8% of the applied was mineralized to NO^-N for digested
sludges despite their differences of initial N content. In contrast, 3
and 33% of the applied was recovered as NO^-N for fresh biomass with
a low and high N content, respectively. The C/N ratio of digested
sludge was not a predictive guide for N mineralization potential.
Mixing the fresh biomass with digested sludge may enhance the
decomposition of the sludge in soil.

APPENDIX A
DIGESTER EFFLUENT CHARACTERISTICS DURING WATER HYACINTH TREATMENT

124
Table 27. Effluent pH during water hyacinth treatment.
Inoculum
volume
Day
0
2
4
6
8
10
12
14
16
18
20
'22
T
--pH
Diluted effluents from high N plants
2.5
7.8
7.7
7.7
7.6
7.5
7.3
7.5
7.2
7.4
7.3
7.5
7.2
5
7.7
7.7
7.9
7.8
7.8
7.8
7.6
7.4
7.6
7.4
7.5
7.5
10
7.7
8.0
8.1
7.9
7.9
7.6
7.7
7.4
7.8
7.6
7.7
7.5
Undiluted
effluents from high N plants
2.5
7.6
8.1
8.3
8.4
8.7
8.4
8.0
8.0
8.3
8.2
8.3
8.3
5
7.6
8.0
8.2
8.2
8.3
8.6
8.7
8.3
8.8
8.8
9.0
8.8
10
7.6
8.0
8.2
8.2
8.2
8.4
8.3
8.0
8.3
8.5
8.8
8.8
Undiluted
effluents from
low N
plants
2.5
7.5
8.1
8.3
8.4
8.5
NAt
NA
8.8
8.8
8.7
8.8
8.8
5
7.4
8.2
8.3
8.5
8.6
NA
NA
9.2
9.1
9.2
9.2
9.1
10
7.5
8.1
8.3
8.5
8.6
NA
NA
9.3
9.3
NA
NA
NA
Nutrient medium
7.5
6.4
4.8
3.9
3.6
NA
NA
3.3
3.5
3.4
3.5
3.5
t NA = Not available.

Table 28. Effluent dissolved 0^ concentration during water hyacinth treatment.
Inoculum
volume
0
2
4
6
8
10
12
14
16
18
20
22
T --
r-1
-mg 02
Li
Diluted i
effluents
from high N
plants
2.5
6.2
4.6
5.4
5.2
4.9
4.7
5.3
5.6
3.9
3.8
3.7
3.3
5
5.4
4.4
3.2
4.0
4.0
4.1
4.5
4.6
3.2
3.5
3.4
2.8
10
4.4
3.9
3.6
2.8
3.5
1.8
4.6
4.3
3.0
3.2
3.7
3.3
Undiluted
effluents
from
ilgh.
N plants
2.5
1.8
2.6
2.9
0.6
0.5
1.0
2.6
2.3
2.9
2.9
3.9
3.1
5
1.7
2.9
2.1
1.0
0.4
0.8
2.9
2.5
2.8
2.5
3.8
2.1
10
0.4
2.1
1.4
0.7
0.2
0.3
2.2
1.3
1.1
1.3
1.5
1.0
Undiluted
effluents
from
low N plants
2.5
4.1
4.5
3.1
2.7
5.1
NA1"
NA
8.0
7.0
8.0
5.9
7.2
5
4.1
4.7
3.7
2.1
3.8
NA
NA
11.9
12.1
11.4
6.9
9.1
10
3.9
4.8
3.5
1.1
1.4
NA
NA
9.1
11.0
NA
NA
NA
Nutrient
medium
7.1
6.9
6.3
5.7
7.0
NA
NA
6.7
7.0
7.2
6.9
6.4
NA = Not available.
125

APPENDIX B
SOIL CHARACTERISTICS FROM ADDED FRESH AND
ANAEROBICALLY DIGESTED PLANT BIOMASS

Table 29. Soil ammonium concentrations from added fresh and digested
plant biomass.
Day
Control
Low N plant biomass
High N plant biomass
Fresh
Digested
Fresh
Digested
-1
0
1.2 b t
2.2 b
1.7 b
25.1 a
1.9 b
30
1.2 a
1.4 a
1.8 a
1.8 a
1.3 a
60
1.1 a
1.2 a
1.7 a
1.0 a
1.0 a
90
1.0 a
1.0 a
1.0 a
1.0 a
1.0 d
^ Values
with same letter
within rows
are not significantly different at
0.05 level by Duncan's Multiple Range Test.
127

128
Table 30. Mehlich I extractable constituents at day 0 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant biomass
constituent
Control
Fresh
Digested
Fresh
Digested
- -mg kg
Calcium
257 cf
377 a
344 ab
332 b
351 ab
Potassium
27 d
128 a
44 c
127 a
53 b
Magnesium
25 d
55 a
34 c
37 b
33 c
Sodium
7 e
60 c
79 b
56 d
133 a
Iron
18 d
25 b
35 a
21 c
26 b
Zinc
4.1 d
7.3 b
13.3 a
6.1 c
7.2 b
^Values with same letter within rows are not significantly different at
0.05 level by Duncan's Multiple Range Test.

129
Table 31. Mehlich I extractable constituents at day 30 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant
biomass
constituent
Control
Fresh
Digested
Fresh Digested
-l
- mg kg
Calcium
245 ct
396 a
383 a
340 b
379 a
Potassium
39 c
139 a
55 c
124 b
49 c
Magnesium
26 c
68 a
39 b
39 b
34 be
Sodium
3 e
57 c
75 b
41 d
114 a
Iron
17 c
18 b
25 a
15 d
18 b
Zinc
4.1 d
6.1 c
11.9 a
6.1 c
7.7 b
^Values with same letter within rows are not significantly different at
0.05 level by Duncan's Multiple Range Test.

130
Table 32. Mehlich I extractable constituents at day 60 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant biomass
constituent
Control
Fresh
Digested
Fresh
Digested
, -1
--mg Kg
Calcium
276 c f
359 b
393 a
343 b
357 b
Potassium
27 d
113 b
44 c
123 a
48 c
Magnesium
27 d
56 a
39 b
40 b
34 c
Sodium
4 e
55 c
74 b
38 d
119 a
Iron
16 c
16 c
23 a
16 c
18 b
Zinc
4.5 d
6.5 be
12.8 a
6.0
c 7.3b
^Values with
same letter
within rows
are not significantly
different at
0.05 level
by Duncan's ]
Multiple Range Test.

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BIOGRAPHICAL SKETCH
Kevin Moorhead was born in Findlay, Ohio, on May 22, 1956, and spent
the first 18 years of his life there. After high school graduation and a
summer in California, Kevin moved to Swannanoa, North Carolina, and began
his college career at Warren Wilson College. He graduated in December,
1978, with a Bachelor of Arts degree in biology. Kevin moved to Pintlala,
Alabama, and worked for Alaga-Whitfield food company in Montgomery,
Alabama, for 7 months. In August, 1979, he moved to Columbus, Ohio, to
begin a graduate program in agronomy at the Ohio State University. Kevin
received his Master of Science degree in December, 1981, with an emphasis
in soil fertility, under the direction of the late Dr. E. 0. McLean.
Since January, 1982, he has pursued a Ph.D. degree at the University of
Florida in the Department of Soil Science. He is presently a candidate
for the degree of Doctor of Philosophy with an emphasis in soil
biochemistry. Kevin has 7 sisters, a brother looking for Sasquatch, and
(at this point) 3 nieces and 1 nephew.
141

I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
A. ^Graet'z ,'^Chadm^^^
Associate Professor of Soil Science
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
K. R. Reddy, CocKairman
Professor of Soil Science
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
J. G. A. Fiskell
Professor of Soil Science

I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
R. A. dtfordstedt
Associate Professor of Agricultural
Engineering
This dissertation was submitted to the Graduate Faculty of the College
of Agriculture and to the Graduate School and was accepted as partial
fulfillment of the requirements for the degree of Doctor of Philosophy.
May 1986
[c vZ/ty
Dean, College of Agrictfftc
ture
Dean, Graduate School



61
1.6 g VS L ^ day Shiralipour and Smith (1984) reported average
methane yields of 0.32 and 0.17 L g 1 VS water hyacinth shoot and root
samples, respectively, in a bioassay test of 100 ml culture volume. They
also concluded that the addition of N in growth media for water hyacinth
production increased methane yields of both shoot and root samples.
Inoculum from operating anaerobic digesters is commonly added as a
bacterial seed to initiate anaerobic digestion in new digesters (Sievers
and Brue, 1978; Wolverton and McDonald, 1981; Field et al., 1984).
Information on the effect of inoculum volume on gas production is
limited.
The objectives of this study were 1) to determine C and N
mineralization during anaerobic digestion of water hyacinth; 2) to
determine the effect of inoculum volume on gas production; and 3) to
evaluate effluent (solids and liquid) composition based on inoculum
volume.
Materials and Methods
Water hyacinths, with either high or low tissue N content, were
anaerobically digested at 35C in 55 L batch digesters containing 2.5, 5,
or 10 L of inoculum. Water hyacinths with a high N content (-34 g kg ^
dry wt plant tissue) were obtained from the wastewater treatment plant of
the Reedy Creek Utility Company, Inc., at Walt Disney World near Orlando,
Florida. Water hyacinths with a low N content (-10 g kg ^ dry wt plant
tissue) were grown in nutrient-depleted water at Sanford, Florida. Both
types of hyacinths were grown in labeled (NH^^SO^ for two weeks,
frozen and chopped to 1.6 mm length using a Hobart T 215 food processor.


109
Table 25. Soil pH (1:2 w/v) from added fresh and digested
plant biomass.
Low N plant biomass
High N plant biomass
Day
Control
Fresh
Digested
Fresh
Digested
0
5.44
d +
5.38 d
6.49 b
5.73 c
7.34 a
30
5.48
c
5.99 b
6.27 a
4.90 d
6.21 a
60
4.99
c
5.96 ab
6.03 a
4.30 d
5.77 b
90
5.42
b
5.94 a
6.03 a
4.72 c
6.02 a
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.


104
Decomposition of digested biomass sludges was adequately described
in two stages. The conversion of soluble sugars and starch to CH^ and
CO^ during anaerobic digestion eliminated the first stage of soluble C
degradation for the digested sludges. Therefore, the first stage of
sludge decomposition corresponded to the second stage (cellulose and
hemicellulose) of fresh plant biomass. The longer second stage
corresponded to the final stage (lignin) of fresh plant biomass.
Soil NO^-N concentrations during incubation are presented in Table
23. Soil NH*-N concentrations (APPENDIX B, Table 29) were < 2 mg kg 1
during the incubation period. Initial NO^-N concentrations were similar
for all treatments. No NO^-N accumulated in the soil amended with low N
fresh plant biomass until 60 days had elapsed. The concentrations of
soil NO^-N were similar for both digested biomass sludges throughout the
incubation despite the higher TKN concentration of the high N sludge.
Although the C/N ratios of high N fresh biomass or digested biomass
sludge were the same, the amount of NO^-N that accumulated in the soil
was more than double for fresh biomass compared to digested biomass
sludge.
A summary of C and ^N mineralization during the incubation period
is presented in Table 24. Approximately 8% of the applied N was
recovered as 15N03-N at 90 days for both sludges. Tester et al. (1977)
reported 6% of composted sewage sludge N mineralized to NO^-N in 54
days. In contrast, 3 and 33% of applied N was recovered as ^NO^-N for
fresh plant biomass with a low and high N content, respectively. Ogwada
(1983) incorporated shredded 15N labeled water hyacinth with a C/N ratio
of 35 into a Myakka fine sand at a rate of 20 Mg dry wt ha'1. At 60
days, 33% of N from the soil-incorporated water hyacinth was


137
Peevey, W. J., and A. G. Norman. 1948. Influence of composition of
plant materials on properties of the decomposed residues. Soil
Sci. 65:209-226.
Penfound, W. T. and T. T. Earle. 1948. The biology of the water
hyacinth. Ecol. Monogr. 18:447-472.
Ponnamperuma, F. N. 1972. The chemistry of submerged soils. Adv.
Agron. 24:29-96.
Price, E. C., and P. N. Cheremisinoff. 1981. Biogas production and
utilization. Ann Arbor Science Publishers, Inc., Ann Arbor, MI.
Puriveth, P. 1980. Decomposition of emergent macrophytes in a
Wisconsin marsh. Hydrobiol. 72:231-242.
Rai, D. N., and J. D. Munshi. 1979. The influence of thick floating
vegetation (water hyacinth: Eichhornia crassipes) on the
physico-chemical environment of a freshwater wetland.
Hydrobiol. 62:65-69.
\
Raven, J. A., and F. A. Smith. 1976. Nitrogen assimilation and
transport in vascular land plants in relation to intracellular pH
regulation. New Phytol. 76:415-431.
Reddy, K. R. 1981. Diel variations in physico-chemical parameters
of water in selected aquatic systems. Hydrobiol. 85:201-207.
Reddy, K. R. 1983. Fate of nitrogen and phosphorus in a waste-water
retention reservoir containing aquatic macrophytes. J. Environ.
Qual. 12:137-141.
Reddy, K. R. 1984. Water hyacinth (Eichhornia crassipes) biomass
production in Florida. Biomass 6:167-181.
Reddy, K. R., and L. 0. Bagnall. 1981. Biomass production of aquatic
plants used in agricultural drainage water treatment. In 1981
International Gas Res. Conf. Proc. Govt. Inst. Inc., Rockville, MD.
Reddy, K. R., and W. F. DeBusk. 1984. Growth characteristics of
aquatic macrophytes cultured in nutrient-enriched water: I. Water
hyacinth, water lettuce, and pennywort. Econ. Bot. 38:229-239.
Reddy, K. R., and D. A. Graetz. 1981. Use of shallow reservoir and
flooded soil systems for wastewater treatment: Nitrogen and
phosphorus transformations. J. Environ. Qual. 10:113-119.
Reddy, K. R., F. M. Hueston, and T. McKim. 1985. Biomass production
and nutrient removal potential of water hyacinth cultured in sewage
effluent. J. Solar Energy Eng. 107:128-135.


58
Detritus recovery in water without plant cover increased with
increasing rate of detritus. This suggests that during periods of low
water hyacinth productivity, i.e. winter, detritus will be an important
sink for inorganic N removal. The high recovery in detritus was
suprising since the original detritus had a high N content (23 mg g
dry tissue). Therefore, the detritus used in water with added ^NO^-N
probably accounted for even more immobilization due to a low initial
N content (5 mg g ^ dry tissue).
Generally recovery in sediment was primarily organic N. Less
than 1% of the added ^NO^-N was recovered as sediment inorganic N.
However, between 3 and 7% of the added ^NH"!"-N was recovered as KC1
4
extractable inorganic N in the sediment. The lower recovery of sediment
15 15 -
inorganic N in water with added NO^-N was probably due to reduction
to gaseous N via denitrification (Engler and Patrick, 1973). Some of
the added was recovered as organic N in the water.
Plant uptake was the primary mechanism of N removal in water
having water hyacinths. The unaccounted for was lost from the
systems through a variety of possible transformations. A more thorough
investigation would be required to establish the extent of algal N
assimilation. Volatilization of NH^-N in water without plant cover and
denitrification in water with sediment are two possible mechanisms for N
removal.
Conclusions
Generally as the rate of detritus addition increased, dissolved
concentrations decreased in water with or without sediment and with or
without plant cover. The decreasing dissolved 0^ concentrations were


121
increased biogas production for plants with a high N content after 15
days of digestion.
Removing the biomass sludge from digester effluents decreased the
electrical conductivity, NH^-N and TKN of the effluent. Anaerobic
digestion resulted in a loss of K and Mg from fresh plant biomass but
increased lignin, total C, TKN, Ca, Na, Fe and Zn of the digested
sludge. Generally cellulose decreased during digestion and
hemicellulose remained unchanged.
Mineralization of organic N to NH+-N was the primary N
transformation occurring during anaerobic digestion. Approximately 20%
of the organic N placed in the digesters was recovered as sludge organic
N. Net mineralization of organic N to NH^-N was 70 and 35% of the added
organic N of plants with a high and low N content, respectively.
Digester Effluent Recyling
The initial electrical conductivities and NH+-N concentrations of
A
the digester effluents ranged from 0.7 to 6.7 dS m ^ and 23 to 289 mg N
L The highest water hyacinth dry weight gains were associated with
the lowest electrical conductivities. However, plants survived in
effluents having electrical conductivities of 5.6 and 5.9 dS m Plant
death was observed in four undiluted effluents. Additional information
t
would be needed to establish optimum dilution of anaerobic digester
effluents to promote maximum biomass yields.
First-order kinetic equations were used to describe NH+-N loss from
effluent with k values ranging from 0.175 to 0.593 day ^. Generally the
highest k values were associated with rapid plant growth. Plant ^N
assimilation was observed for all effluents although plant death


20
Application rates were shown to have insignificant effects on rate
of fresh plant biomass decomposition (Jenkinson, 1965; Nyhan, 1975).
However, several studies suggested that small amounts of fresh or
anaerobically digested plant biomass decomposed more rapidly than large
quantities (Broadbent and Bartholomew, 1948; Jenkinson, 1971; Atalay and
Blanchar, 1984).
Miller and Johnson (1964) found an increasing rate of CO^
production with increasing moisture content up to a tension of 0.05 to
0.015 MPa and then a decreasing rate with further increases in tension.
They concluded that maximum biological activity could be expected at the
lowest tension when aeration was sufficient. Orchard and Cook (1983)
found a log-linear relation between water potential and microbial
activity in the range of 0.005 to 0.5 MPa.
Sain and Broadbent (1977) concluded that low temperatures
influenced decomposition rate more than excessive moisture. However,
Nyhan (1976) found a pronounced decrease in rates of C loss with an
increase in soil water tension even when temperature (10C) was limiting
microbial activity. Miller (1974) determined that soil temperature was
the major factor influencing the rate of decomposition of anaerobically
digested sewage sludge.
Decomposition was generally considered to be initially slower in
acid than neutral soil (Jenkinson, 1971). Addition of organic material
altered the pH of a soil, particularly when the amount added was large
relative to the amount of native organic matter present (Jenkinson,
1966). Atalay and Blanchar (1984) found that addition of anaerobically
digested biomass sludge to soil increased the pH from 5.5 to 7.6 and
they attributed this to a limestone buffer used during the digestion
process.


3
The waste by-product contains digested biomass sludge (solid) and
effluent (liquid). The digested biomass sludge was applied to soil as a
nutrient source for plants. The effluent was recycled in water hyacinth
ponds for nutrient recovery by plants. This type of integrated system
will provide low cost water treatment and total resource recovery.
Efficient utilization of by-products could potentially reduce the cost
effectiveness of the system.
The overall objective of this study was to assess nitrogen cycling
in the three components of an integrated "biomass for energy" system.
Nitrogen is often identified as a limiting factor for plant growth and
is used to establish loading rates in the disposal of solid and liquid
waste. Information on N cycling is limited to studies on the individual
components of the integrated system, i.e. the water hyacinth production
system (Boyd, 1976; DeBusk et al., 1983; Reddy, 1983), anaerobic
digestion (Hashimoto et al., 1980; Field et al., 1984), and effluent and
sludge recycling (Ryan et al, 1973; Hanisak et al., 1980; Terry et al.,
1981; Atalay and Blanchar, 1984). No attempt has been made to establish
N cycling within the entire integrated system.
The specific objectives of this study were 1) to determine growth
rate and detritus production of water hyacinth grown in eutrophic lake
water; 2) to determine the effect of detritus on N transformations in
water hyacinth systems; 3) to evaluate N and C mineralization during
anaerobic digestion of water hyacinth biomass; 4) to evaluate the
potential of water hyacinth to grow in anaerobic digester effluents for
N recovery; and 5) to determine N and C mineralization during
decomposition of fresh and digested biomass added to soil.


ANAEROBIC DIGESTION OF WATER HYACINTH
The potential productivity of water hyacinth has led to its
selection as a biomass feedstock for methane generation while providing
a means for treatment of nutrient-enriched waters. Methane yields
during anaerobic digestion depended on characteristics of the feedstock
(Stack et al., 1978; Wolverton and McDonald, 1981) as well as digester
operating conditions (Hashimoto et al., 1980). Sievers and Brue (1978)
reported higher methane yields for digesters operating on swine waste as
the C/N ratio increased. They concluded that the optimum C/N range for
maximum methane production was 15.5/1 to 19/1. The optimum pH and
temperature range for anaerobic digestion was 6.7 to 7.4 (Bryant, 1979)
and 30 to 35C (House, 1981), respectively.
Biogas and methane yields have been reported for water hyacinths
using a variety of digesters. Wolverton and McDonald (1981) reported
methane yields of 0.07 to 0.20 L g 1 total solids (TS) for blended water
hyacinths. Hanisak et al. (1980) found average methane yields of 0.24 L
g volatile solids (VS) from shredded water hyacinths in 162 L digesters
at loading rates of 1.10 to 1.38 g VS L ^ day ^ and residence times of 30
to 38 days.
Chynoweth et al. (1983) reported methane yields of 0.19 and 0.28 L
g VS of water hyacinth and a 3:1 water hyacinth/primary sewage sludge
blend, respectively, in 5 L daily-fed digesters with a loading rate of
60


LITERATURE REVIEW
The three components of the integrated "biomass for energy" system
were 1) the water hyacinth production system; 2) anaerobic digestion of
water hyacinth biomass; and 3) recycling of digested biomass sludge and
effluent. An integrated approach of wastewater renovation using aquatic
macrophytes with utilization of biomass for energy production is
economically appealing.
Water Hyacinth Biomass Production
The first component of the integrated "biomass for energy" system
was an aquatic system for the production of biomass as well as water
quality improvement. Although several aquatic plants naturally grow in
polluted waters, one the most productive plants appears to be water
hyacinth (Reddy et al., 1983).
Water hyacinth is a mat-forming, free-floating vascular aquatic
plant with wide distribution in sub-tropical and tropical regions. The
plant consists of a submerged rooting system and an aerial
photosynthetic petiole and leaf (shoot) system (Fig. 2). The roots and
aerial shoots are produced at the numerous nodes of the vegetative
portion of a typically submerged rhizome (Penfound and Earle, 1948).
The aerial buds, from which flowers and fruit clusters develop, are
produced from the reproductive portion of the rhizome. Occasionally,
the internodes of the rhizome expand and form new offsets.
4


N TRANSFORMATIONS
1. Mineralization /
Immobilization
2. Plant Uptake
3. Nitrification
4. Denitrification
5. NH Volatilization
Figure 5. Nitrogen cycling in soil treated with plant residues.
Nitrogen and carbon transformations investigated
during this study are indicated with larger arrows.


91
Table 20. Net assimilation or loss of plant nutrients in
diluted or undiluted effluents from digested '
high N plants.
Inoculum
.volume
Na
K
P
Ca
Mg
--L--
-2
A ^
-mg m
aay
SHOOTS
Diluted effluents
from high N plants
2.5
240
35
19
179
126
5
353
187
57
223
138
10
333
220
69
148
110
Undiluted
effluents
from high N plants
\
2.5
190
-23
10
42
4
5
201
-36
15
89
9
10
181
-92
11
35
-4
LSD (0.05)
64
101
17
62
27
ROOTS
Diluted effluents
from high N plants
2.5
217
80
14
53
59
5
273
111
22
50
61
10
227
155
32
94
48
Undiluted
effluents
from high N plants
2.5
40
-26
31
73
2
5
50
-29
32
72
1
10
70
-29
42
103
3
LSD (0.05)
57
50
27
60
14


MASS BALANCE OF NITROGEN IN AN INTEGRATED
"BIOMASS FOR ENERGY SYSTEM
A series of experiments were conducted to evaluate N cycling in
three components of an integrated "biomass for energy" system, i.e.
water hyacinth production, anaerobic digestion of hyacinth biomass, and
recycling of digester effluent and sludge. Two types of water hyacinth
biomass production systems were evaluated; 1) plants growing in
t
nutrient-enriched waters representing wastewater treatment systems
(sewage and digester effluents, animal wastes or agricultural drainage
water), and plants growing in nutrient-limited systems representing
natural waters receiving low external N inputs.
Nutrient-Enriched Systems
Nitrogen cycling in an integrated system for water hyacinths
growing in nutrient-enriched systems is summarized in Fig. 18.
Water Hyacinth Production
-2 -1
A wide range of productivity (5 to 64 g dry wt m day ) has been
recorded for plants growing in nutripnt-enriched waters (Boyd, 1976;
Hanisak et al., 1980; Reddy and DeBusk, 1983; Reddy et al., 1985).
Plants growing in nutrient-enriched media generally have an N content of
30 to 40 g N kg 1 of dry tissue (Boyd, 1976; Wolverton and McDonald,
1979; Reddy et al., 1985). In this study the tissue N content of plants
growing in sewage effluent was 35 g N kg ^ of dry biomass.
112


114
Although plant assimilation is a major process for N removal in
ponds of water hyacinths, other N transformations may occur in the root
zone, water column and underlying sediment. These processes account for
much of the loss of N from the system. For example, a dense mat of
water hyacinth could potentially enhance denitrification of NO^-N
present in the water column (Boyd, 1970; Reddy, 1981). Total N
reduction in water hyacinth systems was reported in a range from 65 to
94% of various wastewaters (Sheffield, 1967; Clock, 1968; Hanisak et
al., 1980; Reddy et al., 1982).
Total fertilizer N recovered by plants in this study ranged from
50% in a field study to 90% in controlled-greenhouse studies. Results
indicated that detritus accounted from 3 to 14% of the total biomass
produced in water hyacinth ponds. Up to 28% of the total plant N was
found in detritus, which could potentially be deposited at the sediment-
water interface.
Anaerobic Digestion of Hyacinth Biomass
Anaerobic digestion of biomass with high tissue N and low C/N ratio
resulted in mineralization of 70% of plant organic N to NH+-N. Total N
recovered for digested high N plants was -100% of the added N which
agreed with results by Hashimoto et al. (1980) and Field et al. (1984).
After digestion, 80% of the plant organic N was recovered in the
t
effluent and 20% was recovered in the sludge.
Waste By-Product Recycling
Undiluted and diluted effluents were used as nutrient sources to
produce additional water hyacinth biomass and for potential N recovery.
Plants growing in undiluted effluents did not survive the combination of
high salt and NH^-N concentrations. However, diluting the digester
effluents provided a media which stimulated water hyacinth growth.


75
of K and Mg compared to the fresh plant biomass. The C/N ratio of the
fresh plant biomass with a low tissue N content decreased from 35 to 16
after digestion. The C/N ratio of the fresh plant biomass with a high
tissue N content was the same as the digested biomass sludge (C/N=12).


N TRANSFORMATIONS
ANAEROBIC
DIGESTION
Organic
Acids
NH
1. Mineralization /
Immobilization
2. NH^ Volatilization
Figure 4. Nitrogen cycling during anaerobic digestion.
Nitrogen and carbon transformations
investigated during this study are indicated
with larger arrows.


39
to the detritus in the control reservoir. Fertilization and increases
in ambient air temperature resulted in dislodging of root biomass.
Net N loading from detritus was 92 to 148 kg N ha ^ yr \ which is
potentially available upon decomposition. The N immobilized by detritus
represented 21 and 28% of the total N removed by water hyacinths in the
fertilized and control reservoirs, respectively.
Approximately 51% of the added fertilizer N was assimilated by
plants. The remaining 49% may have been lost through denitrification.
Total N recovery was nearly 90% in the fertilized reservoir. More N was
accounted for in the control reservoir than was added. Release of N
from the sediment or mineralization of N during decomposition of
detritus may account for the additional N recovery.


13 A
Hanisak, M. D., L. D. Williams, and J. H. Ryther. 1980. Recycling the
nutrients in residues from methane digesters of aquatic macrophytes
for new biomass production. Resource Recovery Conser. 4:313-323.
Hargrave, B. T. 1972. Aerobic decomposition of sediment and detritus
as a function of particle surface area and organic content.
Limnol. Oceanogr. 17:583-596.
Harrison, P. G., and K. H. Mann. 1975. Detritus formation from
eelgrass (Zostera marina L.): The relative effects of
fragmentation, leaching, and decay. Limnol. Oceanogr. 20:924-934.
Hashimoto, A. G., Y. R. Chen, V. H. Varel, and R. L. Prior. 1980.
Anaerobic fermentation of agricultural residues. In Shuler, M.
Utilization and Recycle of Agricultural Wastes and Residues. CRC
Press, Inc., Boca Raton, FL.
Hill, B. H. 1979. Uptake and release of nutrients by aquatic
macrophytes. Aquat. Bot. 7:87-93.
House, D. 1981. The Biogas Handbook. Peace Press, Inc., San Francisco,
CA.
Howard-Williams, C., S. Pickmere, and J. Davies. 1983. Decay rates and
nitrogen dynamics of decomposing watercress (Nasturtium officinale
R.Br.). Hydrobiol. 99:207-214.
Howeler, R. H. 1972.
Qual. 1:366-371.
The oxygen status of lake sediments. J. Environ.
Hsieh, Y. P., L. A. Douglas, and H. L. Motto. 1981a. Modeling sewage
sludge decomposition in soil: I. Organic carbon transformations.
J. Environ. Qual. 10:54-59.
Hsieh, Y. P., L. A. Douglas, and H. L. Motto. 1981b. Modeling sewage
sludge decomposition in soil: II. Nitrogen transformations. J.
Environ. Qual. 10:59-64.
Hughes, W. L. (ed.). 1981. Supplement Energy for Rural Development.
Renewable Resources and Alternative Technologies for Developing
Countries. Natl. Acad. Press, Washington, DC.
Hunt, H. W. 1977. A simulation model for decomposition in grasslands.
Ecol. 58:469-484.
Hunter, R. D. 1976. Changes in carbon and nitrogen content during
decomposition of three macrophytes in freshwater and marine
environments. Hydrobiol. 51:119-128.
Jenkinson, D. S. 1965. Studies on the decomposition of plant material
in soils. I. Losses of carbon from C labelled ryegrass
incubated with soil in the field. J. Soil Sci. 16:104-115.


Table 6. Total plant N and ^NH^-N assimilation.
Treatment
Total uptake
Labeled
Other +
15n)
mg (% of added
Vi)
Without sediment
0 mg C L ^
903
+
44
822 (91)
81
(9)
100 mg C L-1
768
+
103
697 (91)
71
(9)
400 mg C L~1
1133
+
81
807 (71)
326
(29)
With sediment
0 mg C L ^
1390
+
194
891 (64)
499
(36)
100 mg C L'1
1255
+
89
845 (67)
410
(33)
400 mg C L_1
1629
+
65
828 (51)
801
(49)
Carbon source was plant detritus.
^Other N sources include sediment and detritus.


10
High 0^ consumption within a water hyacinth mat during microbial
decomposition of plant detritus (dead and decaying plant debris) could
create anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980;).
The bulk of detritus was trapped within the mat and decomposed primarily
at the water surface (DeBusk et al., 1983). Rate of inorganic N release
from decomposing detritus depended on dissolved 0^ concentration of the
water, C/N ratio, and temperature (Ogwada, 1983).
Low 0^ concentrations create conditions less favorable for
nitrification and promote denitrification which may proceed within the
water hyacinth mat, in the water column, or in the underlying sediment.
Denitrification occurred primarily in the underlying sediment and the
rate depended on diffusion of NO^ -N from the water column to the
sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981).
Water temperatures were lower in areas covered with plants compared
to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980;
Schreiner, 1980). A dense mat over the water surface served as a
blanket barrier for exchange of heat between the atmosphere and the
water (Rai and Munshi, 1979). Water hyacinths growing in either acid or
alkaline water had a tendency to alter the pH towards neutrality (Haller
and Sutton, 1973). A pH of 7.0 in water occurred in areas covered with
plants with little diel variation (McDonald and Wolverton, 1980; Reddy,
1981) which suggests that NH^-N volatilization is minimal in these
systems .
Decomposition of Plant Tissue in Freshwater
Decomposition of plant tissue in a freshwater habitat commonly
occurs in two stages. The first stage was attributed to leaching
of the more soluble plant constituents while the second stage was


67
Clock (1968) reported a 75% reduction of NO^_N in 5 days for water
hyacinths growing in secondary sewage effluent. A 75% reduction of
NH^-N in digester effluents required 2.3 to 6 days for treatments where
positive dry weight gains were observed. For treatments resulting in
+
plant death, 4 to 8 days were required to remove 75% of the NH^-N.
Nitrogen-15 plant assimilation was observed for all treatments
although a low recovery was observed in treatments resulting in plant
death (2 to 16%) (Table 18). The recovery by plants for the other
treatments ranged from 36 to 77%. The majority of the was found in
the shoot material for all treatments (54 to 73%). Approximately 75% of
the was unaccounted for in treatments resulting in plant death.
Microbial assimilation and NH^-N volatilization were probably important
NH*-N removal processes in undiluted effluents where plant death was
observed.
Plant Tissue Chemical Composition
Plants survived in the diluted effluents but death was noted for
plants in the undiluted effluents from digested high N plants. Mineral
constituents from plants growing in these effluents were analyzed to
isolate individual cation and P assimilation or loss from living and
dead plant tissue.
The concentrations of plant tissue (root and shoot fractions) Na,
t
K, P, Ca and Mg are reported in Table 19. The original plant tissue had
low concentrations of Na and P in both shoot and root material, but
higher concentrations of K, Ca and Mg in the shoots compared to the
roots. There were large increases in Na for both shoots and roots of the
surviving and dead plants. The root K concentrations increased for
surviving plants but decreased for dead plants. The P concentrations


ACKNOWLEDGMENTS
This dissertation reports results from a project that contributes
to a cooperative program between the Institute of Food and Agricultural
Sciences (IFAS) of the University of Florida and the Gas Research
institute (GRI), entitled "Methane from Biomass and Waste." Financial
support from the IFAS/GRI cooperative program is gratefully
acknowledged.
Dr. D. A. Graetz was chairman and Dr. K. R. Reddy was cochairman Gf
my supervisory committee. Their friendship and guidance will always be
cherished. The other members of the committee were Dr. G. E. Bowes,
Dr. J. G. A. F.iskell, and Dr. R. A. Nordstedt.
Special thanks go to Bill F'othier who ran numerous L,N samples for
this research. Other people who provided assistance during the estudies
include Bill Christy, Stephen McCracken, Peter Krottje, Terry Siean,
Veronica Campbell, Rremila Rao. Ed Hopwood and Dave Cartiin. Steve
Linda designed the statistical analyses. I appreciate the use of Dr.
John Moore's facilities for fiber analyses. Carolyn Pickles and Brenda
Clutter typed the majority of this dissertation on an IBM computer.
Finally, this package is dedicated to my parents, sisters and brother.
A trip home always gave me a boost to carry on.
ir


138
Reddy, K. R., R. Khaleel, and M. R. Overcash. 1980. Carbon
transformations in the land areas receiving organic wastes in
relation to nonpoint source pollution: A conceptual model. J.
Environ. Qual. 9:434-442.
Reddy, K. R., and P. D. Sacco. 1981. Decomposition of water hyacinth
in agricultural drainage water. J. Environ. Qual. 10:228-233.
Reddy, K. R., P. D. Sacco, D. A. Graetz, K. L. Campbell, and L. R.
Sinclair. 1982. Water treatment by aquatic ecosystems: Nutrient
removal by reservoirs and flooded fields. Environ. Manage.
6:261-271.
Reddy, K. R., and D. L. Sutton. 1984. Water hyacinths for water
quality improvement and biomass production. J. Environ. Qual.
13:1-8.
Reddy, K. R., D. L. Sutton, and G. Bowes. 1983. Freshwater aquatic
plant biomass production in Florida. Soil Crop Sci. Soc. Fla.
42:28-40.
Reddy, K. R., and J. C. Tucker. 1983. Productivity and nutrient uptake
of water hyacinth, Eichhornia crassipes. I. Effect of nitrogen
source. Econ. Bot. 37:237-247.
Rogers, H. H. and D. E. Davis. 1972. Nutrient removal by water
hyacinth. Weed Sci. 20:423-427.
Ryan, J. A., D. R. Keeney, and L. M. Walsh. 1973. Nitrogen
transformations and availability of an anaerobically digested
sewage sludge in soil. J. Environ. Qual. 2:489-492.
Sain, P., and F. E. Broadbent. 1977. Decomposition of rice straw in
soils as affected by some management factors. J. Environ. Qual.
6:96-100.
Sato, H., and T. Kondo. 1981. Biomass production of waterhyacinth and
its ability to remove inorganic minerals from water. I. Effect of
the concentration of culture solution on the rates of plant growth
and nutrient uptake. Japan. J. Ecol. 31:257-267.
Scarsbrook, E., and D. E. Davis. 1971. Effect of sewage effluent on
growth of five vascular aquatic species. Hyacinth Contr. J.
9:26-30.
Schreiner, S. D. 1980. Effects of water hyacinth on the
physio-chemistry of a south Georgia pond. J. Aquat. Plant Manage.
18:9-12.
Schwegler, B. R. and T. W. McKim. 1981. Reedy Creek Improvement
District Water Hyacinths Program. Report to EPA. Grant S805655.
Nov. 1981.


97
The water hyacinths were anaerobically digested for four months in
55 L batch digesters. Each digester received 4.7 kg (fresh weight) of
the labeled water hyacinth, 5 L of an inoculum from anaerobic
digesters receiving water hyacinth as feedstock, and were buffered with
210 g NaHCO^. After digestion, the biomass sludge was separated from
the effluent by passing the total contents of the digester through a
1.00 mm fiberglass screen.
Samples of the fresh water hyacinth and anaerobically digested
water hyacinth sludge were freeze-dried (Thermovac T) and ground through
a 0.84 mm screen of a Wiley Mill. The freeze-dried materials were
characterized for lignin, cellulose and hemicellulose (Goering and Van
Soest, 1970), ashed mineral constituents (Gaines and Mitchell, 1979),
total solids (TS), volatile solids (VS), total C (TC) (LECO Induction
Furnace 523-300), and total Kjeldahl N (TKN) (Nelson and Sommers, 1973).
Carbon/nitrogen (C/N) ratios of the four residues were calculated from
percentage TC and TKN.
Fifty gram soil samples were preincubated for 5 days at a water
content adjusted to 0.01 MPa before addition of the residues. The
freeze-dried materials were added to the soil at a rate of 5 g (dry wt)
kg 1 soil (10 Mg ha ^) and incubated for 90 days at 27C. Water content
was adjusted to 0.01 MPa every 15 days. Ambient laboratory air, with CO^
and NH^-N removed by 3 M NaOH and 4 M H^SO^ traps, respectively, was
pumped through the incubation flask at a rate of 50 ml min ^. The CO^
evolved from soil samples was collected in 0.1 M NaOH traps and
determined by titration with acid after reacting with saturated BaC^.
The percentage C evolved with time was calculated by subtracting C
evolved as CO^ of the control soil (no organic C amendment) from the


FIGURE
PAGE
16. Carbon evolution from soil applied fresh and
digested plant biomass . 101
17. Decomposition stages and rate constants of fresh
and digested plant biomass added to soil 103
18." Nitrogen cycling in an integrated system for water
hyacinths growing in nutrient-enriched systems 113
19. Nitrogen cycling in an integrated system for water
hyacinths growing in nutrient-limited systems 117
viii


70
The low total recovery of N and N for low N plants after
anaerobic digestion is difficult to explain. Nitrogen cycling during
anaerobic digestion was primarily mineralization of organic N or
immobilization of inorganic N. Volatilization of NH3~N may occur but the
* 4.
potential increases as NH^-N concentrations increase or at higher pH
values (Freney et al., 1983). Each digester received 210 g NaHCO^ as a
buffer and the pH after digestion was similar for all digester
effluents. The NH+-N concentrations after digestion were much higher for
the high N plants (Table 13).
Effluent composition
Characteristics of the digester effluents prior to sludge
separation are presented in Table 13. Generally, as the rate of
inoculum increased, there were increases in EC, NH+-N, TKN, and TS. The
COD increased with increasing inoculum volume for digesters with high N
plants.
Characteristics of the screened effluent (sludge removed) are
reported in Table 14. Removing the digested biomass sludge from the
digester effluents decreased the EC, NH^-N and TKN. The screened
effluent from the low N plants contained more Ca, Mg, K, and Na, and less
P than the screened effluent from the high N plants.
Characteristics of the fresh plant biomass and digested biomass
sludge are given in Table 15. Anaerobic digestion resulted in increases
in TC and TKN of sludge compared to fresh plant biomass. The increases
in sludge TKN after digestion of low N plants caused a reduction of the
C/N ratio from 35 to 16. The changes in TC or TKN of the digested high
N plants did not appreciably alter the C/N ratio. The digested sludge


25
replicated areas of each reservoir. Each cage was lined with 1.00 mm
fiberglass screen to retain any detritus dislodged from the plant mat.
The fiberglass screen was positioned 15 cm below the PVC frame to allow
normal waterflow within the root mat.
One reservoir was fertilized monthly by broadcasting a 10-4-10
fertilizer to add 100 kg N ha ^ from October 1981 to February 1982, and
50 kg N ha from March 1982 to September 1982. The change in
fertilizer rate was due to an excess of 10 mg N L ^ found in reservoir
water several days after fertilization during winter. The second
reservoir contained Lake Apopka water with no added nutrients and served
as the control. Both reservoirs were drained and refilled with Lake
Apopka water monthly (24 hr prior to plant sampling and fertilization).
Plant productivity and detritus production were monitored at
monthly intervals for one year. The cages were removed from each
section, drained for 5 min, and weighed. The plant material was divided
into healthy plants and detritus. Detritus was defined as that
collected from the fiberglass screen and dead shoot and root material
remaining within the plant mat. Dead shoots were defined as material
visably devoid of chlorophyll. Three plants were removed for analyses
and the cages were restocked to the initial plant density and placed in
the reservoirs.
*
The shoot and root lengths were recorded for each plant sample and
separated for dry weight ratios. The plant tissue and detritus were
oven-dried at 70 C, weighed, and ground to pass a 0.84 mm Wiley Mill
screen. All plant and detritus samples were analyzed for total Kjeldahl
nitrogen (TKN) using a modified micro Kjeldahl procedure (Nelson and


30
Table 1. Seasonal water hyacinth yield and detritus production.
Fertilized reservoir
Control
reservoir
Season
Yield
Detritus
S/R §
Yield Detritus
S/R

Mg ha 1
Mg ha
-1
Autumn (78) *
9.5
2.8
1.85
3.3
2.2
1.67
Winter (90)
1.9
2.7
1.75
-0.2
3.2
1.25
Spring (84)
14.7
2.7
1.64
9.7
3.3
0.86
Summer (88)
23.1
4.1
2.46
10.2
3.3
0.79
Test of Significance ^
Yield
Detritus
S/R
Season
**
NS
NS
Month (season
)
NS
NS
NS
Reservoir
**
NS
**
TNumber of days in season.
Significant at 0.01 level (**) or not significant (NS).
§S/R = Shoot/root dry wt ratio


Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosopy
NITROGEN CYCLING IN AN INTEGRATED
"BIOMASS FOR ENERGY" SYSTEM
By
Kevin Keith Moorhead
May, 1986
Chairman: Dr. D. A. Graetz
Cochairman: Dr. K. R. Reddy
Major Department: Soil Science
A series of experiments were conducted to evaluate N cycling in
three components of an integrated "biomass for energy" system, i.e.
water hyacinth production, anaerobic digestion of hyacinth biomass, and
recycling of digester effluent and sludge. Plants assimilated 50 to 90%
of added N in hyacinth production systems. Up to 28% of the total plant
N was contained in hyacinth detritus. Nitrogen loading as plant
detritus into hyacinth ponds was 92 to 148 kg N ha ^ yr ^.
Net mineralization of plant organic ^N during anaerobic digestion
was 35 and 70% for water hyacinth plants with low (10 g N kg ^ dry
tissue) and high (35 g N kg ) N content, respectively. Approximately
20% of the ^N was recovered in the digested sludge while the remaining
^~*N was recovered in the effluent.
Water hyacinth growth in digester effluents was affected by
electrical conductivity (0.7 to 6.7 dS m and ^NIij"-N concentration
ix


12
measurable effect on rate of plant decomposition and Nichols and Keeney
(1973) reported a more rapid decomposition rate under non-aerated
conditions compared to aerated conditions. Ogwada et al. (1984) found
that approximately the same amounts of N and P were released from
decaying plant tissue under aerobic or completely anoxic conditions, but
the extent of nutrient release was dependent on water temperature.
The changes in plant C and N composition or concentration during
decomposition have received considerable attention. Build-up of
microbal biomass on decaying plant tissue caused a loss in the C content
while increasing the N content which resulted in a decrease in the C/N
ratio (De La Cruz and Gabriel, 1974; Odum and Heywood, 1978; Hill,
1979). Godshalk and Wetzel (1978b) found that lignin was very resistant
to decomposition while the other structural carbohydrates gradually
decreased with time.
Nitrogen was a limiting factor in decomposition of several aquatic
plants (Nichols and Keeney, 1973; Almazan and Boyd, 1978; Godshalk and
Wetzel, 1978b; Carpenter, 1980). Decay rates were correlated both to
initial N content and to C/N ratios (Godshalk and Wetzel, 1978b;
Carpenter and Adams, 1979; Ogwada et al., 1984).
Particle size also influenced decomposition. Generally, the rate
of decomposition increased as the particle size decreased (Fenchel,
1970; Hargrave, 1972; Gosselink and Kirby, 1974). Harrison and Mann
(1975) reported that a reduction in size of leaf material from 2 to 4 cm
to <1 mm doubled the rate of organic matter loss.
Boyd (1970b) and Odum and Heywood (1978) concluded that submerged
leaves decomposed more rapidly than those placed upon the water surface
or suspended in air. Nichols and Keeney (1973) found more rapid dry


29
(January, February, and March; 3) spring (April, May, and June); and 4)
summer (July, August, and September). The seasons were chosen to
coincide with changes in water hyacinth productivity and temperature and
solar radiation changes. The autumn season combines data for November
and December of 1982 and October of 1983.
The effects of temperature and solar radiation were evident since
the lowest net productivity occurred during winter and the highest
during spring and summer (Table 1). The net primary productivity in
winter for both reservoirs was lower than the production of detritus.
Reddy and Bagnall (1981) reported that at average temperatures of 10C,
productivity of water hyacinth approached zero. The majority of the
detritus during winter came from the destruction of aerial shoots caused
by freezing temperatures. Although a majority of the aerial shoots were
destroyed, the plants survived, and noticeable gains in dry weight began
in March.
There were significant differences in yields between seasons and
reservoirs. Seasonal yields ranged from 1.9 to 23.1 Mg (dry wt) ha ^
for the fertilized reservoir and -0.2 to 10.2 Mg ha ^ for the control
reservoir. Over 75% of the biomass production occurred during spring
and summer for both reservoirs. Differences in detritus production were
not significant between reservoirs or between seasons. Although the
annual yield of water hyacinth in the fertilized reservoir was double
that of the control reservoir, the detritus production in both
reservoirs was similar (Table 1). There were significant differences in
the shoot/root dry weight ratios between reservoirs, but not between
seasons (Table 1). The average shoot/root dry weight ratio was 1.93
(1.6A to 2.A6) for the fertilized reservoir and 1.14 (0.79 to 1.67) for
the control reservoir.


55
detritus N was a potential N source for plant assimilation. Reddy
(1983) found that 60 to 64% of total N assimilation by water hyacinths
was derived from added ^N, while 36 to 40% was derived from sediment
and from decomposition of detritus.
The recovered by plant tissue (mg) was fairly consistent for
added labeled fertilizer but plant ^NH^-N uptake exceeded ^NO^-N
uptake. Water hyacinth appeared to be more efficient in utilizing NH+-N
than N0^-N (Reddy and Tucker, 1983). When water hyacinth growth is not
restricted by climate, rapid assimilation of added inorganic N would be
expected.
Nitrogen-15 Balance for Water Columns
A N balance for water with added ^no^-N and ^^NH^-N is presented
in Tables 7 and 8, respectively. Total recovery by water hyacinths
ranged from 57 to 72% and 70 to 89% in water with added ^no^-N and
respectively. Reddy (1983) conluded that water hyacinth N
assimilation accounted for only 40% of added ^no^-N or ^^NH^-N in a
reservoir. Algal surface mats accounted for ~8% of added ^no^-N and up
to 15% of added The algal surface mats represented a minor
portion of total microbial N assimilation. Algal activity was noted
in open water, and the four sides and bottom of the microcosm tanks were
colonized by algae.
The 15$ associated with detritus was determined for water with
added but not for added ^no -N. Less than 10% of the added
4 3
NH^-N was immobilized by detritus in water with plant cover. Newly
formed water hyacinth detritus from the plant cover was deposited during
28 days and accounted for 7% recovery in water without added
detritus or sediment.


52
effect on rate of N loss in water with plant cover. Nitrate and NH^-N
disappeared within 2 and 4 days, respectively, in water with water
hyacinths.
Nitrogen loss in open water was influenced by the underlying
sediment and detritus additions. Nitrate disappeared more rapidly in
open water with sediment than without sediment. An increase in detritus
resulted in a more rapid NO^-N loss in water with or without sediment.
A rapid decrease in NO^-N after 2 days was followed by an accumulation
of NO^'N at 4 days for open water. Accumulation of NO^-N after 4 days
in open water was probably due to rapid algal assimilation followed by
turnover (death) of the algae and leaching of NO^-N from the dead algal
cells. Surface algal mats developed in open water within 2 days.
Ammonium disappeared more rapidly in open water with sediment than
without sediment. Apparently detritus did not influence NH+-N loss in
open water with sediment. However, detritus additions resulted in a
more rapid NH+-N loss in open water without sediment. Loss of NH+-N
followed by accumulation of inorganic N during the first 4 days was not
as striking as seen for NO^-N.
Plant Nitrogen Assimilation
Total plant N assimilation and the contribution of added ^NO^-N
and ^NH^-N to total N assimilation is presented in Tables 5 and 6,
respectively. Plant N assimilation was always greater for water with
underlying sediment. Part of the increased plant N assimilation in
water with sediment was due to release of N from the sediment.
Generally the contribution of added ^NO^-N or ^NH^-N to total plant N
assimilation also decreased with increasing detritus. Mineralization of


120
Approximately 50% of the fertilizer N was recovered by plants.
Total N assimilated by water hyacinth (live plants and detritus) was 720
and 325 kg N ha ^ yr ^ for plants grown in eutrophic lake water with and
without added nutrients, respectively.
Detritus and Nitrogen Transfromations
Detritus had no apparent effect on rate of N loss in water with
water hyacinth plant cover due to rapid plant assimilation. However, N
loss in water without plant cover was more rapid with detritus
additions. Both sediment and detritus appeared to be potential N
sources for plant assimilation.
Total N recovered by water hyacinth ranged from 57 to 72% for added
15N0l-N and 70 to 89% for added 15NHt-N. Less than 10% of added 15Nh1'-N
3 4 4
was immobilized by detritus in water with plant cover. However, up to
35% of the added ^NH"!"-N was associated with detritus in water without
4
plant cover. This suggests that during periods of low water hyacinth
productivity, i.e. winter, detritus is an important sink for added N.
Increasing amounts of detritus generally decreased dissolved 0^
concentrations of water. The pH of water without plant cover generally
was lower when detritus was added.
t
Anaerobic Digestion of Water Hyacinth
Initial water hyacinth N content and volume of inoculum did not
affect long term (60 days) biogas production. At 15 days, biogas
production was generally greater for plants with a high N content.
Inoculum volume showed little effect of biogas production for low N
plants throughout incubation. However, a larger volume of inoculum


DISSOLVED
20
CO
E
V-/
CVJ
O
Plants
0 mg C
L_l T T
No Sediment
1 00 mg C
L~1 *

400 mg C
L~ 1
TIME (days)
Figure 10. Dissolved in sedimentwaterplant systems with added ammonium.


DRY WEIGHT GAIN (g
/~\
Diluted Effluent of Undiluted Effluent of Undiluted Effluent of
High N Planta High N Plants Low N Planta Nutrient
400
300
200
100
0
-50
0^7 L6 2^3 4 4V7 3 5^6 5 6^7 07
2.5 5 10 2.5 5 10 2.5 5 10 Medium
i 4 i i 1 1 * 1,. l X - ..
-
L
JL
_L,
-
u
p
4
:T :
23 65 104 161 212 289 24 49 87 20
NH* CONCENTRATION (mg LT1)
ELECTRICAL CONDUCTIVITY CdS rrT1)
Figure 15. Dry weight gains of water hyacinths in digester effluents
and nutrient medium.


Table 4. Nitrogen balance for the two reservoirs.
Fertilized Control
reservoir reservoir
Fertilizer
kg ha
Nitrogen added
NH+-N
4
430

NO~-N
234
--
Organic N
117
--
Total
781
Lake water
NHt-N
4
13
13
NO~-N
12
12
Organic N
213
213
Total
238
238
Total added
1019
238
Nitrogen removed
Water hyacinth
Shoots
354
95
Roots
218
137
Detritus
148
92
Total
720
325
Reservoir water
NH+-N
4
13
8
NO~-N
13
5
Organic N
167
156
Total
193
169
Total N accounted
913
494


78
Materials and Methods
Anaerobic digester effluents were obtained from six 55 L batch
digesters containing water hyacinth with a high or low tissue N content
as feedstock. Water hyacinths with low (-10 g N kg ^ dry plant tissue)
and high (-34 g N kg ) tissue N content were grown in nutrient-depleted
water and sewage effluent, respectively. After removal from their
respective growth media, the hyacinths were grown in ^N labeled
(NH^)^SO^ nutrient solution for two weeks, frozen, and chopped to 1.6 mm
length using a Hobart T 215 food processor.
The water hyacinths were anaerobically digested for four months in
J v
55 L batch digesters. Each digester received 4.7 kg (fresh weight) of
the labeled water hyacinth, 2.5, 5, or 10 L volume of inoculum from
anaerobic digesters receiving water hyacinth as feedstock, and were
buffered with 210 g NaHCO^. After digestion, the biomass sludge was
separated from the effluent by passing the total contents of the
digesters through a 1.00 mm fiberglass screen.
The screened effluents (sludge removed) were analyzed for total
solids (TS), volatile solids (VS), fixed solids (FS) (APHA, 1980), total
Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH+-N and NO^-N by steam
distillation (Keeney and Nelson, 1982), electrical conductivity (EC)
(Hach Mini Conductivity Meter) and pH (Orion Model 404 Specific Ion
Meter). Samples passed through a 0.2 pm membrane filter were analyzed
for Na, K, Mg and Ca by atomic absorption and ortho P colorimetrically
after reacting with ammonium molybdate.
Six water hyacinth plants were placed in 10 L of undiluted or
2
diluted effluents in containers having a surface area of 0.051 m The
water hyacinth plants were collected from the University of Florida's


Table 15. Characteristics of fresh and digested biomass residues.
Biomass
Inoculum
_
residue
volume
VS
FS
TC
TKN
C/N
Lig t Cell
Heme
Ca
K
Mg
Na
Fe
Zn
--L--
r> 1
ke"1-
g kg
8 1
High N
plant material
Fresh
834
166
385
34.0
12
43
167
182
17.6
23.5
3.2
8.0
1.60
0.52
Digested
2.5
843
157
449
37.2
12
136
159
NA t
17.6
2.2
1.8
15.3
4.04
1.20
Digested
5
827
173
446
39.3
12
130
207
180
18.4
3.8
2.2
26.0
4.76
0.92
Digested
10
846
154
441
30.8
14
111
168
NA
17.8
3.6
2.0
20.8
3.88
1.00
Low N plant material
Fresh
839
161
373
10.6
35
83
266
247
21.0
22.0
6.7
10.9
1.88
0.64
Digested
2.5
879
121
441
27.4
16
149
177
NA
22.1
2.8
2.8
14.8
5.72
1.92
Digested
5
866
134
425
26.6
16
145
181
242
24.2
2.8
2.2
17.4
7.16
2.04
Digested
10
847
153
433
25.2
17
163
163
NA
26.8
4.6
2.7
19.5
8.40
2.16
tLig, Cell, Heme = Lignin, cellulose and hemicellulose
' NA = Not available


69
Table 12. Nitrogen-15 balance for the batch digesters.
High N plant material Low N plant material
2.5 L+ 5 L 10 L 2.5 L 5 L 10 L
g
15N Added
Water hyacinth
Organic N
10.39
10.39
10.39
3.24
3.24
3.24
Recovered
Screened effluent
Organic N
1.18
1.48
0.93
0.84
0.65
0.34
Inorganic N
7.06
7.78
7.68
0.79
1.25
1.34
Digested Sludge
Organic N
1.67
2.53
2.21
0.87
0.57
0.52
Total
9.91
11.79
10.82
2.50
2.46
2.20
% Recovered
95
113
104
77
76
68
t
Volume (liters) of inoculum.


84
towards the center of the plant.- The other leaf tissue damage on young
and older leaves was curling of the entire leaf towards the center of
the leaf. Stem damage in undiluted effluents was noticeable after 2
days. Damaged stems collasped under slight manual pressure. The
destruction of chlorophyll in stems and leaves was widely observed in
the undiluted effluents.
The extent and spread of tissue damage increased in undiluted
effluents with increasing EC and NH+-N concentrations. At the end of
one week, all plants in four undiluted effluents were dead. The EC and
NH+-N concentrations of these effluents ranged from 4.3 to 6.7 dS m ^
and 87 to 289 mg N L \ respectively. The shoots began to separate from
the roots at the water surface and the submerged roots sank. Root
separation also accounted for the negative dry weight gains of plants in
undiluted effluents.
Three treatments showed occasional visual signs of tissue damage,
i.e., the nutrient medium and 2 diluted effluents. Visual signs of
plant damage but noticeable gains in plant dry weights were observed in
the diluted effluent having an initial NH+-N concentration of 104 mg L ^
and two undiluted effluents having NH^-N concentrations of 24 and 49 mg
L ^. All remaining treatments resulted in plant death, apparently due to
high EC or high NH^-N concentrations, or a combination of both.
Although water hyacinth plants struggled to survive in the
undiluted effluents, algal activity was noted in all undiluted effluent
treatments. Upon emptying the containers, algae were found attached to
the side and bottom surfaces of the plastic containers.


130
Table 32. Mehlich I extractable constituents at day 60 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant biomass
constituent
Control
Fresh
Digested
Fresh
Digested
, -1
--mg Kg
Calcium
276 c f
359 b
393 a
343 b
357 b
Potassium
27 d
113 b
44 c
123 a
48 c
Magnesium
27 d
56 a
39 b
40 b
34 c
Sodium
4 e
55 c
74 b
38 d
119 a
Iron
16 c
16 c
23 a
16 c
18 b
Zinc
4.5 d
6.5 be
12.8 a
6.0
c 7.3b
^Values with
same letter
within rows
are not significantly
different at
0.05 level
by Duncan's ]
Multiple Range Test.


136
Nelson, D. W. and L. E. Sommers. 1972. A simple digestion procedure
for estimation of total nitrogen in soils and sediments. J.
Environ. Qual. 1:423-425.
Nelson, D. W., and L. E. Sommers,
nitrogen in plant materials.
Nelson, D. W., and L. E. Sommers,
nitrogen in natural waters.
Nelson, D. W., and L. E. Sommers,
and organic matter.' In Page
Analysis. Part. 2. Second
1973. Determination of total
Agron. J. 65:109-112.
1975. Determination of total
J. Environ. Qual. 4:465-468.
1982. Total carbon, organic carbon,
A. L. (ed.). Methods of Soil
dition. Agronomy 9. ASA, Madison, WI.
Nichols, D. S., and D. R. Keeney. 1973. Nitrogen and phosphorus
release from decaying water milfoil. Hydrobiol. 42:509-525.
Nyhan, J. W. 1975. Decomposition of carbon-14 labeled plant materials
in a grassland soil under field conditions. Soil Sci. Soc. Am.
Proc. 39:643-648.
Nyhan, J. W. 1976. Influence of soil temperature and water tension on
the decomposition rate of carbon-14 labeled herbage. Soil Sci.
121:288-293.
Odum, W. E., and M. A. Heywood. 1978. Decomposition of intertidal
freshwater marsh plants. In Good, R. E., D. F. Whigham, and R. J.
Simpson (eds.). Freshwater Wetlands. Ecololgical Processes and
Management Potential. Academic Press, New York.
Ogwada, R. A. 1983. Growth, nutrient uptake, and nutrient regeneration
by selected aquatic macrophytes. M.S. Thesis. Univ. of Florida.
Ogwada, R. A., K. R. Reddy, and D. A. Graetz. 1984. Effects of
aeration and temperature on nutrient regeneration from selected
aquatic macrophytes. J. Environ. Qual. 13:239-243.
Orchard, V. A., and F. J. Cook. 1983. Relationship between soil
respiration and soil moisture. Soil Biol. Biochem. 15:447-453.
Otsuki, A., and R. G. Wetzel. 1974. Release of dissolved organic
matter by autoanalysis of a submerged macrophyte, Scriptus
subterminalis. Limnol. Oceanogr. 19:842-845.
Parra, J. V., and C. C. Hortenstine. 1976. Response by pearl millet to
soil incorporation of water hyacinths. J. Aquat. Plant Manage.
14:75-79.
Patterson, D. T., and S. 0. Duke. 1979. Effect of growth irradiance on
the maximum photosynthetic capacity of water hyacinth (Eichhornia
crassipes). Plant Cell Physiol. 20:117-184.


14
for water hyacinths and a 3:1 blend of water hyacinths:domestic sewage
sludge, respectively in 5 L digesters. Shiralipour and Smith (1984)
reported average methane yields of 0.32 and 0.17 L g ^ VS water hyacinth
shoot and root samples, respectively, in a bioassay test of 100 ml
culture volume. They concluded that the addition of N to growth media
for water hyacinth production increased methane yields.
The nutrients in the biomass are recovered in the waste material
after the digestion process. The recovered nutrients are
distributed between the effluent and the digested biomass sludge. The
total N recovery was usually 100% after digestion, but much of the
organic N was converted to NH*-N (Hashimoto et al., 1980; Field et al.,
1984). Most of the K (Field et al., 1984) and Na (Atalay and Blanchar,
1984) were solubilized and remained in the digester effluent.
The digestion process increased the sorption of some nutrients (P,
Ca, and Mg) by the sludge fraction such that fewer were available for
dilute acid extraction and perhaps for crop recovery (Field et al.,
1984). Field et al. (1984) hypothesized that sorption may have been
increased by particle surface area increases due to size reduction of
solids.
Nitrogen Cycling During Anaerobic Digestion
Nitrogen transformations occurring during anaerobic digestion
include 1) mineralization/immobilization; and 2) NH^-N volatilization
(Fig. 4).
The mineralization or immobilization of N depends on the initial N
content of the biomass feedstock. The C/N ratio of the biomass is often
used as a guideline for prediction of net mineralization or
immobilization. The optimal C/N ratio of the added biomass was 30:1



PAGE 1

NITROGEN CYCLING IN AN INTEGRATED "BIOMASS FOR ENERGY" SYSTEM By KEVIN KEITH MOORHEAD A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 1986

PAGE 2

AC;KNOWLEDGiJ>MENTS This di sserl'.ation reports results from a project that contributes to a cooperative program between the Institute of Food and Agricultural Sciences ^IFAS) of the University of Florida and the Gas Research Institute (GRI), entitled "Methane from Biomass and Waste." Financial support from the IFAS/GRI cooperative program is gratefully acknowledged Dr. D. A. Graetz was chairman and Dr. K. R. Reddy was cochairrnan of my supervisory committee. Their friendship and guidance will always be cherislied. The other members of the co.mmiltee were Dr. G. £. Bcwes, Dr. .J. G. A. Fiskell, and Dr. R. A. Ncrastedt. Special tlianks go to Bill Pctliier who ran nvjiiercus 'N samples for this research. Other people who provided assistance during the r.r.udies include Bill Christy, Snephen McCracki-jr:, Pecer Krottje, Terry Slean, Veronica C^iTlpbell, Premila Rao. Ed Hopwood and Dave Cartlin, Steve Linda designed the statistical a.".alyses. J appreciate the use of Dr. John Moore's facilities for fiber anaJyses. Carolyn Pickles a;id Brenda Clutter typed the majority of this dissertation on aa IBM computer. Finally, this package is dedicated to my parents, sisters and brother. A trip home always gave me a boost to carry on.

PAGE 3

TABLE OF CONTENTS Page ACKNOWLEDGEMENTS . • ii LIST OF TABLES v LIST OF FIGURES vii ABSTRACT ix INTRODUCTION 1 LITERATURE REVIEW 4 Water Hyacinth Biomass Production 4 Anaerobic Digestion 13 Waste By-Product Recycling 16 Conclusions 21 WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION 23 Materials and Methods 24 Results and Discussion 26 Conclusions 38 EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER HYACINTH SYSTEMS 40 Materials and Methods 42 Results and Discussion 44 Conclusions 58 ANAEROBIC DIGESTION OF WATER HYACINTH 60 Materials and Methods 61 Results and Discussion 63 Conclusions 74 TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTH 76 Materials and Methods 78 Results and Discussion 80 Conclusions 93 iii

PAGE 4

DECOMPOSITION OF FRESH AND ANAEROBICALLY DIGESTED PLANT BIOMASS IN SOIL 95 Materials and Methods ^ 96 Results and Discussion 98 Conclusions Ill MASS BALANCE OF NITROGEN IN AN INTEGRATED "BIOMASS FOR ENERGY" SYSTEM 112 Nutrient-Enriched Systems 112 Nutrient-Limited Systems 116 CONCLUSIONS 119 Water Hyacinth Productivity and Detritus Production 119 Detritus and Nitrogen Transformations 120 Anaerobic Digestion of Water Hyacinth 120 Digester Effluent Recycling 121 Digester Sludge Recycling 122 APPENDICES A DIGESTER EFFLUENT CHARACTERISTICS DURING WATER HYACINTH TREATMENT 124 B SOIL CHARACTERISTICS FROM ADDED FRESH AND ANAEROBICALLY DIGESTED PLANT BIOMASS 126 BIBLIOGRAPHY 131 BIOGRAPHICAL SKETCH 141 iv

PAGE 5

LIST OF TABLES TABLE PAGE 1. Seasonal water hyacinth yield and detritus production ... 30 2. Seasonal water hyacinth shoot and root lengths 32 3. Seasonal nitrogen uptake by water hyacinth and detritus 35 A. Nitrogen balance for the two reservoirs 37 5. Total plant N and ^^NO^-N assimilation 53 6. Total plant N and ''^NH^-N assimilation 54 7. Mass balance of added '^^NO^-N in sediment-water-plant systems 56 8. Mass balance of added NH^-N in sediment-water-plant systems 57 9. Characteristics of the inoculum used in the batch digesters 64 10. Gas production during anaerobic digestion of high and low N water hyacinth plants 65 11. Nitrogen balance for the batch digesters 68 12. Nitrogen-15 balance for the batch digesters 69 13. Characteristics of digester effluents before sludge removal 71 14. Characteristics of screened effluents (sludge removed) after digestion 72 15. Characteristics of fresh and digested biomass residues. 73 16. Initial characteristics of the digester effluents and nutrient medium 81 17. First-order kinetic descriptions of NH^-N loss with time. 86 18. Nitrogen-15 balance for labeled effluents 88 V

PAGE 6

TABLE PAGE 19. Distribution of nutrients in water hyacinth shoots and roots in diluted and undiluted effluents of digested high N plants 89 20. Net assimilation or loss of plant nutrients in diluted or undiluted effluents from digested high N plants 91 21. Characteristics of the digester effluents and nutrient medium after water hyacinth treatment 92 22. Characteristics of the fresh and digested plant biomass 99 23. Soil NO^-N concentration from added fresh and digested plant biomass 105 24. Carbon and ^^N mineralization from added fresh and digested plant biomass 106 25. Soil pH (1:2 w/v) from added fresh and digested plant biomass 109 26. Mehlich I extractable constituents at Day 90 from added fresh and digested plant biomass 110 27. Effluent pH during water hyacinth treatment 12A 28. Effluent di ssolved O2 concentration during water hyacinth treatment 125 29. Soil ammonium concentrations from added fresh and digested plant biomass 127 30. Mehlich I extractable constituents at Day 0 from added fresh and digested plant biomass 128 31. Mehlich I extractable constituents at Day 30 from added fresh and digested plant biomass 129 32. Mehlich I extractable constituents at Day 60 from added fresh and digested plant biomass 130 vi

PAGE 7

LIST OF FIGURES FIGURE PAGE 1. Integrated water hyacinth aquaculture system of biomass production, bioconversion to methane and digester waste recycling 2 2. A generalized diagram of a water hyacinth plant 5 3. Nitrogen cycling in a water hyacinth production sytem ... 9 4. Nitrogen cycling during anaerobic digestion 15 5. Nitrogen cycling in soil treated with plant residues. ... 18 6. Weekly averages of daily temperatures and solar radiation 27 7. Monthly averages of daily primary productivity and detritus production 28 8. Seasonal plant tissue nitrogen content 33 9. Dissolved 0in sediment-water-plant systems with added nitrate 45 10. Dissolved 0^ in sediment-water-plant systems with added ammonium 45 11. The pH of sediment-water-plant systems with added nitrate 43 12. The pH of sediment-water-plant systems with added ammonium 49 13. Nitrogen loss from sediment-water-plant systems with added nitrate 50 14. Nitrogen loss from sediment-water-plant systems with added ammonium 5j 15. Dry weight gains of water hyacinths in digester effluents and nutrient medium 83 vli

PAGE 8

FIGURE PAGE 16. Carbon evolution from soil applied fresh and digested plant biomass 101 17. Decomposition stages and rate constants of fresh and digested plant biomass added to soil 103 18. Nitrogen cycling in an integrated system for water hyacinths growing in nutrient-enriched systems 113 19. Nitrogen cycling in an integrated system for water hyacinths growing in nutrient -limited systems 117 viil

PAGE 9

Abstract of Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosopy NITROGEN CYCLING IN AN INTEGRATED "BIOMASS FOR ENERGY" SYSTEM By Kevin Keith Moorhead May, 1986 Chairman: Dr. D. A. Graetz Cochairman: Dr. K. R. Reddy Major Department: Soil Science A series of experiments were conducted to evaluate N cycling in three components of an integrated "biomass for energy" system, i.e. water hyacinth production, anaerobic digestion of hyacinth biomass, and recycling of digester effluent and sludge. Plants assimilated 50 to 90% of added N in hyacinth production systems. Up to 28% of the total plant N was contained in hyacinth detritus. Nitrogen loading as plant detritus into hyacinth ponds was 92 to 148 kg N ha ^ yr ^. Net mineralization of plant organic ^^N during anaerobic digestion was 35 and 70% for water hyacinth plants with low (10 g N kg ^ dry tissue) and high (35 g N kg ^) N content, respectively. Approximately 20% of the '^^N was recovered in the digested sludge while the remaining ^^N was recovered in the effluent. Water hyacinth growth in digester effluents was affected by electrical conductivity (0.7 to 6.7 dS m~^) and '^^NH^-N concentration Ix

PAGE 10

(23 to 289 mg N L Biomass yields were maximum at electrical conductivities of < 2.5 dS m ^ and ^^NH^-N concentrations of < 100 mg N Addition of water hyacinth biomass to soil resulted in decomposition of 39 to 50% of added C for fresh plant biomass and 19 to 23% of added C for digested biomass sludge. Only 8% of added ''^^N in digested sludges was mineralized to ^^NO^'N despite differences in initial N content (27 and 39 g N kg ^ dry sludge). In contrast, 3 and 33% of added ^^N in fresh biomass with low and high N content, respectively, was recovered as ^^NO^-N. Total ''^^N recovery after anaerobic digestion ranged from 70 to 100% of the initial plant biomass ''^^N. Land application of digester sludge resulted in the mineralization of 2% of initial biomass ^^N into plant available form. Use of water hyacinth for digester effluent treatment resulted in recycling of 21 to 38% of the initial biomass ^^N. Total N recovery by sludge and effluent recycling in the integrated "biomass for energy" system was 48 to 60% of the initial plant biomass ^^N. The remaining ^^N was lost from the system during anaerobic digestion and effluent recycling.

PAGE 11

INTRODUCTION Several types of aquatic plants are widely distributed in freshwater lakes and streams. These plants assimilate nutrients and produce biomass, which could potentially be used for beneficial purposes. Water hyacinth ( Eichhornia crassipes [Mart] Solms) is one of the dominant aquatic plants distributed throughout the tropical and subtropical regions of the world. This freshwater macrophyte has already been evaluated for use in treating nutrient-enriched waters such as sewage effluent (Cornwell et al., 1977; Wolverton and McDonald, 1979; Reddy et al., 1985), agricultural drainage water (Reddy and Bagnall, 1981; Reddy et al., 1982), anaerobic digester effluent (Hanisak et al., 1980), and fertilized fish ponds (Boyd, 1976). The characteristics that make this plant grow rapidly in polluted waters make it an ideal candidate for large-scale nutrient removal and water purification (Reddy and Sutton, 1984). An integrated aquaculture system has been developed using water hyacinth for water treatment and for total resource recovery. The components of an integrated aquaculture system are schematically illustrated in Fig. 1. Water hyacinth plants have been used for wastewater treatment while the biomass produced was harvested periodically and processed through anaerobic digestion to produce methane. The process produced a waste by-product which must be disposed of, or preferably utilized, in an environmentally-safe manner.

PAGE 12

2 o o o o CD LU O cr co LU LU < O AN Q < EW [< CO (U CO 4J CO CO B CO o > •H (U J-) o CD (U g OC (U •H 4J CO CO c CO (U 3 c 4J CO iH J= 3 4-1 O 0) CO B 3 XT o CO ^ c o c •H CO O i-i CO 0) o o (U o J-l •H CO c (U o 00 4J •H c CO i-l •H u o rH 00 3 (U •a •u o o c u M o. u 0) 3 oo .1

PAGE 13

3 The waste by-product contains digested biomass sludge (solid) and effluent (liquid). The digested biomass sludge was applied to soil as a nutrient source for plants. The effluent was recycled in water hyacinth ponds for nutrient recovery by plants. This type of integrated system will provide low cost water treatment and total resource recovery. Efficient utilization of by-products could potentially reduce the cost effectiveness of the system. The overall objective of this study was to assess nitrogen cycling in the three components of an integrated "biomass for energy" system. Nitrogen is often identified as a limiting factor for plant growth and is used to establish loading rates in the disposal of solid and liquid waste. Information on N cycling is limited to studies on the individual components of the integrated system, i.e. the water hyacinth production system (Boyd, 1976; DeBusk et al., 1983; Reddy, 1983), anaerobic digestion (Hashimoto et al., 1980; Field et al., 1984), and effluent and sludge recycling (Ryan et al, 1973; Hanisak et al., 1980; Terry et al., 1981; Atalay and Blanchar, 1984). No attempt has been made to establish N cycling within the entire integrated system. The specific objectives of this study were 1) to determine growth rate and detritus production of water hyacinth grown in eutrophic lake water; 2) to determine the effect of detritus on N transformations in water hyacinth systems; 3) to evaluate N and C mineralization during anaerobic digestion of water hyacinth biomass; 4) to evaluate the potential of water hyacinth to grow in anaerobic digester effluents for N recovery; and 5) to determine N and C mineralization during decomposition of fresh and digested biomass added to soil.

PAGE 14

LITERATURE REVIEW The three components of the integrated "biomass for energy" system were 1) the water hyacinth production system; 2) anaerobic digestion of water hyacinth biomass; and 3) recycling of digested biomass sludge and effluent. An integrated approach of wastewater renovation using aquatic macrophytes with utilization of biomass for energy production is economically appealing. Water Hyacinth Biomass Production The first component of the integrated "biomass for energy" system was an aquatic system for the production of biomass as well as water quality improvement. Although several aquatic plants naturally grow in polluted waters, one the most productive plants appears to be water hyacinth (Reddy et al., 1983). Water hyacinth is a mat-forming, free-floating vascular aquatic plant with wide distribution in sub-tropical and tropical regions. The plant consists of a submerged rooting system and an aerial photosynthetic petiole and leaf (shoot) system (Fig. 2). The roots and aerial shoots are produced at the numerous nodes of the vegetative portion of a typically submerged rhizome (Penfound and Earle, 19A8). The aerial buds, from which flowers and fruit clusters develop, are produced from the reproductive portion of the rhizome. Occasionally, the internodes of the rhizome expand and form new offsets. 4

PAGE 15

5 Figure 2. A generalized diagram of a water hyacinth plant. The major morphological structures are adventitious roots (AR) ; root hairs (RA) ; rhizome (RH); stolon (ST); detritus tissue (DT) attached to the plant; float (F) ; leaf isthmus (IS);, leaf petiole (PT) ; peduncle (PD) ; spathe (SP) ; leaf lamina (LA) ; inflorescence (IN)

PAGE 16

6 The elongated internodes were designated as stolons (Penfound and Earle, 19A8). The relatively rapid rate of colonization by water hyacinth is due primarily to vegetative reproduction (stolon and offset production). The plants reproduce sexually during warmer months until freezing terminates anthesis. The developing fruits containing the seeds are cast off onto the mat or into the water. They sink in water and remain in a viable condition for several years. Manson and Manson (1958) reported that each plant could produce 5000 to 6000 seeds which remained viable for at least 5 years. The geographical distribution of water hyacinth is regulated by temperature and salt concentration in the water. When average minimum temperature reached 10C, productivity of water hyacinth approached zero (Reddy and Bagnall, 1981). Optimiam growth was found in a temperature range of 25 to 30C (Bock, 1969; Knipling et al., 1970). Water hyacinth is basically a freshwater plant and will die in waters with sustained salt concentrations in excess of 2500 mg L ^ (Haller et al., 197A). Water hyacinth growth is regulated by the nutrient composition of the water medium, temperature, solar radiation, and plant density. Water hyacinth potentially could be grown in nutrient -enriched waters such as sewage effluents, agricultural runoff and drainage effluents, methane digester effluents, and runoff from animal waste operations. Nitrogen is present as NH^-N, NO^-N, and organic N in water media avaiable for water hyacinth production. Organic N often predominates in most water media and is not readily available for plant assimilation. Water hyacinths are efficient users of inorganic N and plant assimilation is one of the major processes of N removal in hyacinth ponds.

PAGE 17

7 Water hyacinth adapted to light intensity and full sunlight elicted the greatest photosynthetic rate (Patterson and Duke, 1979). Optimum plant density to obtain maximum biomass yield varied with season and available plant nutrients in the water (Reddy and Sutton, 198A). DeBusk et al. (1981) and Reddy et al. (1983) established that optimum plant density for achieving maximum growth cultured in wastewaters was in the -2 range of 15 to 35 kg wet wt m Water hyacinth productivity has been evaluated in natural and -2 -1 nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day were reported for plants growing in natural waters of central and south Florida (Yount and Grossman, 1970; DeBusk et al., 1981). A wide-range of productivity (5 to 42 g dry wt m day ) was recorded for plants cultured in nutrient-enriched waters (Schwegler and Kim, 1981; Hanisak et al., 1980). Reddy and DeBusk (198A) obtained an average of 52 and a -2 -1 maximum of 6A g dry wt m day for water hyacinths grown in nutrient-nonlimiting conditions. The effectiveness of water hyacinth in removing inorganic N was reported for several nutrient-enriched wastewaters. Sheffield (1967) and Clock (1968) reported a 75 to 9A% reduction of inorganic N from secondary sewage effluent in systems containing water hyacinths. Reddy et al. (1982) observed a 78 to 81Z reduction of inorganic N from organic soil drainage water containing water hyacinths. Hanisak et al. (1980) concluded that 65% of N in digester effluents could be assimilated when water hyacinths were grown in diluted effluents. Boyd (1976) calculated average rates of N and P removal were 3. A and 0.A3 kg ha"^ day"^ in fertilized fish ponds. Rogers and Davis (1972) concluded that water hyacinth removal capacities were less effective with increasing nutrient concentrations

PAGE 18

8 The potential productivity and nutrient removal capacities of water hyacinth has led to its selection as a biomass feedstock for methane generation while providing a means for treatment of nutrient-enriched waters. Extensive research, both in laboratory and field applications, was conducted on the use of water hyacinth in wastewater treatment during the past 20 years (Sheffield, 1967; Boyd, 1970a; Steward, 1970; Scarsbrook and Davis, 1970; Rogers and Davis, 1971; Dunigan et al., 1975; Cornwell et al., 1977; McDonald and Woverton, 1980; Reddy et al., 1982; DeBusk et al., 1983). Water hyacinth was shown to be effective in removing N, P and other nutrients, and reducing biological oxygen demand and total suspended solids.' Water hyacinth was also shown to readily absorb and concentrate heavy metals (Wolverton and McDonald, 1975a, b; Cooley et al. 1978) Nitrogen Cycling in the Water Hyacinth Production System Nitrogen transformations occurring in a water hyacinth production system include 1) plant uptake; 2) mineralization/ immobilization; 3) nitrification; 4) denitrif ication; and 5) NH^-N volatilization (Fig. 3). Plant uptake is one of the major processes for N removal from water hyacinthbased wastewater systems. Plant uptake is directly related to the growth rate and the nutrient composition of the water. Water hyacinth was more efficient in utilizing NH^-N than NO^-N when both forms were supplied in equal proportions (Reddy and Tucker, 1983). A dense cover of floating water hyacinths will regulate dissolved O^, temperature and pH of water which influences several N transformations. Generally, diel fluctuations of these water parameters were reported to be lower in areas covered with water hyacinths compared to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980; Reddy, 1981).

PAGE 19

9 2 43 .N IS o E E c o t E 2 S ro ro CO O ^ u ^ n •H O to (1) c •H a >. o C 0) 60 o •H z : (U 3 to 00 CO o >-l to CO
PAGE 20

10 High 2 consumption within a water hyacinth mat during microbial decomposition of plant detritus (dead and decaying plant debris) could create anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980;). The bulk of detritus was trapped within the mat and decomposed primarily at the water surface (DeBusk et al., 1983). Rate of inorganic N release from decomposing detritus depended on dissolved 0^ concentration of the water, C/N ratio, and temperature (Ogwada, 1983). Low 2 concentrations create conditions less favorable for nitrification and promote denitrif ication which may proceed within the water hyacinth mat, in the water column, or in the underlying sediment. Denitrif ication occurred primarily in the underlying sediment and the rate depended on diffusion of NO^ -N from the water column to the sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981). Water temperatures were lower in areas covered with plants compared to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980; Schreiner, 1980). A dense mat over the water surface served as a blanket barrier for exchange of heat between the atmosphere and the water (Rai and Munshi, 1979). Water hyacinths growing in either acid or alkaline water had a tendency to alter the pH towards neutrality (Haller and Sutton, 1973). A pH of 7.0 in water occurred in areas covered with plants with little diel variation (McDonald and Wolverton, 1980; Reddy, 1981) which suggests that NH^-N volatilization is minimal in these systems Decomposition of Plant Tissue in Freshwater Decomposition of plant tissue in a freshwater habitat commonly occurs in two stages. The first stage was attributed to leaching of the more soluble plant constituents while the second stage was

PAGE 21

11 microbial-controlled degradation (Boyd, 1970b; Hunter, 1976; Godshalk and Wetzel, 1978a; Howard-Williams et al., 1983). Otsuki and Wetzel (1974) reported a rapid leaching loss of dissolved organic matter regardless of conditions of aerobiosis or whether plants were fresh or f reeze-dried. Hill (1979) concluded that rapid leaching of soluble material accounted for a 21 to 60% dry weight loss of aquatic macrophytes during the first 8 days of incubation. Leaching was established as the major process in the decomposition of eelgrass and total loss of organic matter by leaching accounted for 82% of dried leaves and 65% of fresh leaves (Harrison and Mann, 1975). Leaching rates appeared to be independent of temperature (Carpenter, 1980). Potassium, Na, Mg, and Ca have all been reported as being rapidly lost during the early leaching phase of plant decomposition (Boyd, 1970b; Davis and van der Valk, 1978; Puriveth, 1980), Carpenter (1980) found that the higher the initial P concentration, the more rapid was P leaching. The second stage of decomposition is attributed to biological processes. Microbial-controlled decomposition was influenced by temperature (Carpenter, 1980; Puriveth, 1980), pH (Sompongse, 1982), available O2 (Godshalk and Wetzel, 1978a), and available nutrients (Carpenter and Adams, 1979; Puriveth, 1980). Godshalk and Wetzel (1978a) found that the presence of regardless of temperatures of 10 or 25C, permitted rapid degradation of dissolved and particulate organic matter. Decomposition of water hyacinth was found to be faster under aerobic than anaerobic conditions (Reddy and Sacco, 1981). However, Sompongse (1982) determined that aeration did not have a

PAGE 22

12 measurable effect on rate of plant decomposition and Nichols and Keeney (1973) reported a more rapid decomposition rate under non-aerated conditions compared to aerated conditions. Ogwada et al. (198A) found that approximately the ssune amounts of N and P were released from decaying plant tissue under aerobic or completely anoxic conditions, but the extent of nutrient release was dependent on water temperature. The changes in plant C and N composition or concentration during decomposition have received considerable attention. Build-up of microbal biomass on decaying plant tissue caused a loss in the C content while increasing the N content which resulted in a decrease in the C/N ratio (De La Cruz and Gabriel, 197A; Odum and Heywood, 1978; Hill, 1979). Godshalk and Wetzel (1978b) found that lignin was very resistant to decomposition while the other structural carbohydrates gradually decreased with time. Nitrogen was a limiting factor in decomposition of several aquatic plants (Nichols and Keeney, 1973; Almazan and Boyd, 1978; Godshalk and Wetzel, 1978b; Carpenter, 1980). Decay rates were correlated both to initial N content and to C/N ratios (Godshalk and Wetzel, 1978b; Carpenter and Adams, 1979; Ogwada et al., 1984). Particle size also influenced decomposition. Generally, the rate of decomposition increased as the particle size decreased (Fenchel, 1970; Hargrave, 1972; Gosselink and Kirby, 197A). Harrison and Mann (1975) reported that a reduction in size of leaf material from 2 to A cm to <1 mm doubled the rate of organic matter loss. Boyd (1970b) and Odum and Heywood (1978) concluded that submerged leaves decomposed more rapidly than those placed upon the water surface or suspended in air. Nichols and Keeney (1973) found more rapid dry

PAGE 23

13 weight loss under aerated conditions in sediment -water systems than in water only. They attributed this difference to an additional supply of N from the sediments. Anaerobic Digestion The second component of the integrated "biomass for energy" system was anaerobic digestion of plant biomass for methane production. Anaerobic digestion is a biological process in which organic matter, in the absence of oxygen, is converted to methane and carbon dioxide (Toerien and Hattingh, 1969). During the process of anaerobic digestion, waste organic C was stabilized by the nearly complete microbial fermentation of carbohydrates resulting in a reduction of volatile solids (Miller, 197A). Anaerobically digested sewage sludges were considered more stable to microbial degradation than were aerobically digested sludges (Sommers, 1977). Processes which regulated anaerobic digestion include hydrolysis of polymers, the dissimilation of starting subtrates to the level of acetic acid, and the conversion of acetic acid to CH^ and (Mah et al., 1977). Factors which influenced anaerobic digestion include pH and temperature changes. All methanogens were reported to be strict anaerobes with an optimum pH of 6.7 to 7. A (Bryant, 1979). The optimum temperature range was 30 to 35C (House, 1981). Water hyacinth biomass could be anaerobically digested to produce methane. Hanisak et al. (1980) found average methane yields of 0.24 L g ^ volatile solids (VS) of shredded water hyacinth in 162 L digesters. Chyoweth et al. (1983) reported methane yields of 0.19 and 0.28 L VS

PAGE 24

14 for water hyacinths and a 3:1 blend of water hyacinths: domestic sewage sludge, respectively in 5 L digesters. Shiralipour and Smith (1984) reported average methane yields of 0.32 and 0.17 L g ^ VS water hyacinth shoot and root samples, respectively, in a bioassay test of 100 ml culture volume. They concluded that the addition of N to growth media for water hyacinth production increased methane yields. The nutrients in the bioraass are recovered in the waste material after the digestion process. The recovered nutrients are distributed between the effluent and the digested biomass sludge. The total N recovery was usually 100% after digestion, but much of the organic N was converted to NH^-N (Hashimoto et al., 1980; Field et al., 1984). Most of the K (Field et al., 1984) and Na (Atalay and Blanchar, 1984) were solubilized and remained in the digester effluent. The digestion process increased the sorption of some nutrients (P, Ca, and Mg) by the sludge fraction such that fewer were available for dilute acid extraction and perhaps for crop recovery (Field et al., 1984). Field et al. (1984) hypothesized that sorption may have been increased by particle surface area increases due to size reduction of solids. Nitrogen Cycling During Anaerobic Digestion Nitrogen transformations occurring during anaerobic digestion include 1) mineralization/ immobilization; and 2) NH^-N volatilization (Fig. 4). The mineralization or immobilization of N depends on the initial N content of the biomass feedstock. The C/N ratio of the biomass is often used as a guideline for prediction of net mineralization or immobilization. The optimal C/N ratio of the added biomass was 30:1

PAGE 25

15 \ c o (d o +3 o E E o (d N o > ro X c o CO 00 -a 4-1 u •H c to n) o C M CO •H 4J •O o GO'S C to O io 60 (JO ? o 4J 4J 1 T3 4J CO CO •H 60 •H t-l 3 ID a) CO 60 60 to Q) >

PAGE 26

16 (Hughes, 1981). Bioconversion of biomass with a higher C/N ratio was limited by N. A lower initial C/N ratio resulted in mineralization of organic N during digestion. The C/N ratio of the digester effluent was lower than the C/N ratio of the fresh slurry because of the release of C as CO^ and CH^ (House, 1981). Anaerobic digestion of plant biomass resulted in high concentrations of NH^-N in the digester (Hashimoto et al., 1980; Field et al., 1984). Ammonium was toxic to methogens at concentrations > 3.0 g L-1, regardless of pH (Hashimoto et al., 1980). Losses of NH^-N through volatilization should be low in digesters operating at the optimum pH.of 6.7 to 7.2 unless NH^-N concentrations are high. Waste By-Product Recycling The final component of the integrated "biomass for energy" system was recycling of the waste by-product generated from the anaerobic digestion of plant biomass. The waste by-product from the anaerobic digester was screened to separate the digested biomass sludge from the effluent. The effluent was recycled in the water hyacinth biomass production system discussed earlier. Methane digester effluent contains high levels of NH^-N (> 200 mg L ^) which may inhibit plant growth. Dilution of the effluent is required before use in a water hyacinth production system. Optimum dilution of the effluent for maximum water hyacinth yields has not been established. Nitrogen cycling in a water hyacinth production system was presented earlier. The sludge was added to soil as an organic amendment. A consequence of anaerobic digestion was a reduction of the readily decomposable C of the plant tissue during production of CH, and C0„

PAGE 27

17 Anaerobically digested sludge was considered to be stable and resist further decomposition (Sommers, 1977). Nitrogen Cycling in Soil Treated with Plant Residues The land application of fresh or anaerobically digested plant biomass has significant implications on N cycling. Nitrogen transformations occurring after residue additions include 1) mineralization/ immobilization; 2) microbial or plant assimilation; 3) nitrification; 4) denitrif ication; and 5) NH^-N volatilization (Fig. 5). Since most of the N in fresh or digested plant biomass is in organic forms, the rate of mineralization becomes the rate limiting step for all transformations that follow. Mineralization or immobilization depends on the initial N concentration of the plant biomass as well as the composition of the C constituents. A low N content or a wide C/N ratio was associated with slow decomposition and rates of decomposition were proportional to lignin content (Alexander, 1977). A wide C/N ratio (> 30:1) favored N immobilization whereas a narrower ratio (< 20:1) resulted in N mineralization (Alexander, 1977). Mineralization of N from anaerobically digested sewage sludges was reported to be affected by the rate of application (Ryan et al., 1973; Stark and Clapp, 1980). However, Epstein et al. (1978) found that irrespective of the amount of material (sewage sludge and sludge compost) applied, the percentage of added N mineralized remained essentially constant. The mineralization of NH^-N from organic N is accompanied by microbial assimilation or plant uptake. In aerobic environments the NH^-N was quickly converted to NO"-N (nitrification) which could also be

PAGE 28

18 CO p CO < \ c o •43 a> c o o E ^ -2 o cvj ro o •43 cd o O i3 o > ro tf5 CO o u i-l 0) bO u cd CO (U 3 •U m (0 >^ to Cfl to •H ai •H CO -U
PAGE 29

19 assimilated by microbes or plants (Ryan et al., 1973). A high organic loading rate may result in 0^ depletion during decomposition which promotes the denitrif ication process (Epstein et al., 1978; Hsieh et al., 1981b). Decomposition of Plant Residues in Soil Decomposition of plant residues in soil occurs in two stages. The first stage was attributed to loss of the easily decomposable labile fraction which was followed by the second stage of slow decomposition of a resistant residual fraction (Shields and Paul, 1973; Reddy et al., 1980). Both stages were thought to be controlled by two simultaneously occurring superimposed first-order kinetic reactions (Sinha et al., 1977). Fresh and anaerobically digested plant biomass differ widely in their chemical composition. Anaerobic digestion converts most of the easily-decomposable plant C constituents into CH^ and CO^. The digested biomass sludge has a higher lignin content and is more resistant to decomposition. There is little information available on decomposition of anaerobically digested plant biomass added to soil. However, the rates and the factors which influence decomposition of fresh plant biomass added to soil have been well-established. Tenny and Waksman (1929) concluded that water-soluble organic substances were first to be decomposed in the soil, followed by hemicellulose and at the same time, or immediately after, cellulose. Lignin was very resistant to decomposition and may even delay the disintegration of cellulose or hemicellulose because of the structural proximity of these C constituents in the cell wall (Tenny and Waksman, 1929; Peevey and Norman, 1948; Berg et al., 1982).

PAGE 30

20 Application rates were shown to have insignificant effects on rate of fresh plant biomass decomposition (Jenkinson, 1965; Nyhan, 1975). However, several studies suggested that small amounts of fresh or anaerobically digested plant biomass decomposed more rapidly than large quantities (Broadbent and Bartholomew, 1948; Jenkinson, 1971; Atalay and Blanchar, 1984). Miller and Johnson (1964) found an increasing rate of production with increasing moisture content up to a tension of 0.05 to 0.015 MPa and then a decreasing rate with further increases in tension. They concluded that maximum biological activity could be expected at the lowest tension when aeration was sufficient. Orchard and Cook (1983) found a log-linear relation between water potential and microbial activity in the range of 0.005 to 0.5 MPa. Sain and Broadbent (1977) concluded that low temperatures influenced decomposition rate more than excessive moisture. However, Nyhan (1976) found a pronounced decrease in rates of C loss with an increase in soil water tension even when temperature (10C) was limiting microbial activity. Miller (1974) determined that soil temperature was the major factor influencing the rate of decomposition of anaerobically digested sewage sludge. Decomposition was generally considered to be initially slower in acid than neutral soil (Jenkinson, 1971). Addition of organic material altered the pH of a soil, particularly when the amount added was large relative to the amount of native organic matter present (Jenkinson, 1966). Atalay and Blanchar (1984) found that addition of anaerobically digested biomass sludge to soil increased the pH from 5.5 to 7.6 and they attributed this to a limestone buffer used during the digestion process.

PAGE 31

21 < Jenkinson (1965, 1971), using different plants and soils, determined that the proportion of added plant C retained in the soil under different climatic conditions was remarkably similar over time. Generally, onethird of the added plant C remained after one year, falling to one-fifth after 5 years. Atalay and Blanchar (1984) determined that anaerobically digested biomass sludge decomposed rapidly in soil as evidenced by nearly 40% of the C added evolved as CO^ during 100 days of decomposition. However, Miller (1974) concluded that anaerobic digested sewage sludge was resistant to further decomposition with a maximum of 20% of the added C evolved as CO^ during a 6-month incubation. Terry et al. (1979) found that 26 to 42% of anaerobically digested sewage sludge C was evolved as CO^ during incubation. Generally, the majority of the produced in incubation studies was evolved in the first 30 days (Miller, 1974; Terry et al., 1979; Ataway and Blanchar, 1984). Other soil properties influenced by plant biomass additions included increasing water-holding capacity, CEC, and electrical conductivity (Stark and Clapp, 1980; Atalay and Blanchar, 1984). Epstein et al. (1976) found levels of salinity and chloride in sewage sludge applied to soils increased to a level which may affect salt-sensitive plants. Conclusions Although information is available on N cycling for each component of the system, no attempt has been made to follow N transformations in an integrated "biomass for energy" system. Evaluation of N cycling was

PAGE 32

22 chosen because N is often identified as a limiting factor for plant growth and is used to establish loading rates in the disposal of solid and liquid waste. Plant uptake was established as a major N removal process during water hyacinth biomass production (Reddy and Sutton, 198A). However, the role of water hyacinth detritus as a N source or sink has not been established. Methane yields during anaerobic digestion of water hyacinth were enhanced with increasing N content (Shiralipour and Smith, 1984). However, N mineralization rates were not investigated. Limited information was available on disposal or utilization of digester effluent or sludge from anaerobically digested plant biomass (Hanisak et al., 1980; Atalay and Blanchar, 1984). The overall objective of this research was to integrate the three components of biomass production, anaerobic digestion of biomass, and digester waste recycling with respect to N cycling.

PAGE 33

WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION Water hyacinth is one of the most productive aquatic macrophytes found throughout the tropical and subtropical regions of the world. The plant has been used extensively for treatment of nutrient -enriched waters and currently there are a number of wastewater treatment systems in the U. S. utilizing water hyacinths for secondary and tertiary treatment (Cornwell et al.,1977; Dinges, 1978; Wolverton and McDonald, 1979; Reddy et al., 1985). Water hyacinth productivity has been evaluated in natural and -2 -1 nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day were reported for plants growing in natural waters of central and south Florida (Yount and Grossman, 1970; DeBusk et al., 1981). A wide range of productivity (5 to A2 g dry wt m day ) was recorded for plants cultured in nutrient -enriched waters (Hanisak et al., 1980; Reddy and Bagnall, 1981; Reddy, 198A). Maximum growth rates provided an average of 52 and a maximiam of 64 g dry wt m day for plants cultured under nutrient-nonlimiting conditions (Reddy and DeBusk, 1984). Plant detritus (dead and decaying plant debris) is an integral part of water hyacinth mats. Detritus is usually derived from natural aging of plants, biological or chemical control, and frost damage. Decomposition of detritus releases nutrients which can be subsequently utilized by water hyacinths. Information on water hyacinth productivity was extensive (Reddy et al., 1983), but research on detritus production and its role as a nutrient sink or source was limited, 23

PAGE 34

24 DeBusk et al. (1983) measured detritus production in harvested and nonharvested water hyacinth based sewage treatment systems. Detritus -2 -1 production in both systems averaged 2 g dry wt m day More than 80% of the detritus consisted of root material. The bulk of the standing crop detritus remained trapped in the floating plant mat. However, this study did not reveal the potential of detritus as a nutrient input to the water hyacinth ponds. The objectives of this study were to 1) measure productivity and detritus (shoot and root) production of water hyacinths grown in eutrophic lake water with and without added nutrients and 2) determine the potential of detritus as a nutrient source or sink to the ponds. Materials and Methods The study was conducted in two reservoirs located at the Central Florida Research and Education Center research farm near Lake Apopka in Zellwood, Florida. The reservoirs were constructed with 2.0 m high levees of a Lauderhill organic soil (Lithic medisaprists) and with bottoms composed of calcareous clay. The water depth was 60 cm and the dimensions of the reservoirs were 7.6 m by 51 m (total surface area of 2 465 m ). Both reservoirs were filled with water from nearby Lake Apopka, and were sectioned into four equal areas for replication and stocked with water hyacinths. 2 A total of eight 0.25 m cages (Vexar mesh screen connected to 5 cm diameter PVC pipe) were stocked with water hyacinth at an initial _2 density of 16 kg (fresh wt) m The cages were placed within the four

PAGE 35

replicated areas of each reservoir. Each cage was lined with 1.00 nm fiberglass screen to retain any detritus dislodged from the plant mat. The fiberglass screen was positioned 15 cm below ^:he PVC frame to allow normal waterflow within the root mat. One reservoir was fertilized monthly by broadcasting a 10-4-10 fertilizer to add 100 kg N ha ^ from October 1981 to February 1982, and 50 kg N ha ^ from March 1982 to September 1982. The change in fertilizer rate was due to an excess of 10 mg N L ''^ found in reservoir water several days after fertilization during winter. The second reservoir contained Lake Apopka water with no added nutrients and served as the control. Both reservoirs were drained and refilled with Lake Apopka water monthly (24 hr prior to plant sampling and fertilization). Plant productivity and detritus production were monitored at monthly intervals for one year. The cages were removed from each section, drained for 5 min, and weighed. The plant material was divided into healthy plants and detritus. Detritus was defined as that collected from the fiberglass screen and dead shoot and root material remaining within the plant mat. Dead shoots were defined as material visably devoid of chlorophyll. Three plants were removed for analyses and the cages were restocked to the initial plant density and placed in the reservoirs. The shoot and root lengths were recorded for each plant sample and separated for dry weight ratios. The plant tissue and detritus were oven-dried at 70C, weighed, and ground to pass a 0.84 mm Wiley Mill screen. All plant and detritus samples were analyzed for total Kjeldahl nitrogen (TKN) using a modified micro Kjeldahl procedure (Nelson and

PAGE 36

26 Sommers, 1973), Solar radiation and high and low daily temperatures were recorded. The results were statistically analyzed for a randomized block design with the fertilized and control reservoirs as treatments. Results and Discussion The weekly averages of daily maximum and minimum air temperatures and solar radiation are shown in Fig. 6. Maximvim temperatures ranged from 21.9C during January to March and 36.5C during July to September. Minimum temperatures for these time periods were 8.2C and 20.3C, respectively. Maximtim and minimum temperatures for the rest of the year were similar (high= 30C, low= 13.5C). Maximum and minimum solar radiation occurred from April to September and from November to March, respectively. The monthly averages of daily primary productivity and detritus production of water hyacinth are presented in Fig. 7. Maximum daily water hyacinth productivity during this study was observed in August for the fertilized reservoir (28.3 g dry wt m day ) compared to June for 2 1 the control reservoir (14.7 g dry wt m day ). Detritus production remained fairly consistent with time for both reservoirs. Detritus production in the fertilized reservoir increased noticeably in September when plant productivity began to decline. The average daily detritus production in the fertilized reservoir was 3.7 g dry wt m~f day"'^ compared to 3.5 g dry wt m day in the control reservoir. DeBusk et al. (1983) found that detritus production occurred at a relatively constant rate regardless of harvested or nonharvested conditions. Monthly data for the plant parameters have been summarized by seasons: 1) autumn (October, November, and December); 2) winter

PAGE 37

27 TIME (months) Figure 6. Weekly averages of daily temperatures and solar radiation. 1

PAGE 38

J t I I I I I I 1 1 1 L. NDJ FMAMJ JASO TIME (months) Figure 7. Monthly averages of daily primary productivity and detritus production.

PAGE 39

(January, February, and March; 3) spring (April, May, and June); and 4) summer (July, August, and September). The seasons were chosen to coincide with changes in water hyacinth productivity and temperature and solar radiation changes. The autumn season combines data for November and December of 1982 and October of 1983. The effects of temperature and solar radiation were evident since the lowest net productivity occurred during winter and the highest during spring and summer (Table 1). The net primary productivity in winter for both reservoirs was lower than the production of detritus. Reddy and Bagnall (1981) reported that at average temperatures of 10C, productivity of water hyacinth approached zero. The majority of the detritus during winter came from the destruction of aerial shoots caused by freezing temperatures. Although a majority of the aerial shoots were destroyed, the plants survived, and noticeable gains in dry weight began in March. There were significant differences in yields between seasons and reservoirs. Seasonal yields ranged from 1.9 to 23.1 Mg (dry wt) ha'''" for the fertilized reservoir and -0.2 to 10.2 Mg ha"^ for the control reservoir. Over 75% of the biomass production occurred during spring and sijmmer for both reservoirs. Differences in detritus production were not significant between reservoirs or between seasons. Although the annual yield of water hyacinth in the fertilized reservoir was double that of the control reservoir, the detritus production in both reservoirs was similar (Table 1). There were significant differences in the shoot/root dry weight ratios between reservoirs, but not between seasons (Table 1). The average shoot/root dry weight ratio was 1.93 (1.64 to 2.46) for the fertilized reservoir and 1.14 (0.79 to 1.67) for the control reservoir.

PAGE 40

Table 1. Seasonal water hyacinth yield and detritus production. Fertilized reservoir Control reservoir Season Yield Detritus S/R § Yield Detritus S/R --Mg ha ^ Mg ha -1_ Autumn (78) 9 .5 2.8 1.85 3.3 2. 2 1. 67 Winter (90) 1 .9 2.7 1.75 -0.2 3. 2 1. 25 Spring (8A) lA .7 2.7 1.6A 9.7 3. 3 0. 86 Summer (88) 23 .1 A.l 2.A6 10.2 3. 3 0. 79 Test of Significance Yield Detritus S/R Season NS NS Month (season) NS NS NS Reservoir NS ** Number of days in season. tSignificant at 0.01 level (**) or not significant (NS). §S/R = Shoot/root dry wt ratio

PAGE 41

31 Shoot and root lengths were similar for plants in both reservoirs during autumn and winter (Table 2). During spring, the water hyacinth root lengths were shorter and shoot lengths longer in the fertilized reservoir compared to the plants in the control reservoir. An interesting development in plant morphology was the dislodging of practically the entire root system from plants in the fertilized reservoir beginning in March after daily temperatures began to increase. The majority of plants were typified by a small root system compared to plants in the control reservoir. Some root dislodging was noticed in the control reservoir during spring and summer but was not as characteristically uniform as in the fertilized reservoir. Root lengths in the fertilized reservoir began to increase during summer, but the shoot lengths were double those in the control reservoir. Under nutrient-limiting conditions, water hyacinths produce a large volume of root material presumably to increase their nutrient absorption capacity. With nutrient-enriched media, water hyacinth use more photosynthetic energy in shoot production. Cornwell et al. (1977) measured shoot lengths in excess of 1 m in wastewater media. Penfound and Earle (19A8) recorded root lengths of 0.1 to 1 m. Maximum shoot length recorded during this study was 55 cm during summer for fertilized plants compared to 28 cm during summer for control plants. The plant tissue N content is shown in Fig. 8. Fertilization resulted in increases in shoot, root, and detritus N content compared to plants in the control reservoir. Maximum tissue N content for fertilized plants occurred during winter when plant productivity was low. The increase in plant productivity in spring and summer diluted the N content of the tissue although total N assimilation by the plants

PAGE 42

Table 2. Seasonal water hyacinth shoot and root lengths. Season Fertilized reservoir Control reservoir Shoots Roots Shoots Roots Autumn (78)"'' 39.6 21.5 Winter (90) 25.2 14.2 Spring (84) 26.6 9.9 Summer (88) 54.9 21.3 Test of significance Shoot length Season ** Month (Season) NS Reservoir -cm35.7 23.3 18.8 27.8 22.9 20.2 14.8 27.1 Root length ft* t Number of days in Season. t Significant at 0.05 (*) or 0.01 (**) level, or not significant (NS).

PAGE 43

-1 1 rCO 3 m o o o oj (O cr Q II :' II u) DC a o 1 CO CO CD *5 01 "S CL CO IS hCO CD o CO o ^ Z) 1< 6) N>il INVld

PAGE 44

34 was much greater during this time period (Table 3). Plant tissue N content remained nearly consistent with time in the control reservoir and root tissue N content generally exceeded that of the shoot tissue (Fig. 8). Seasonal N assimilation by water hyacinth ranged from 34 to 242 kg N ha ^ for plants in the fertilizer reservoir and from <0 to 104 kg N ha for plants in the control reservoir (Table 3). There were significant differences between seasons and reservoirs in water hyacinth N assimilation. The detritus N content was significantly greater for fertilized than control plants, but there were no significant differences in detritus N content between seasons. Data on mass balance of N in both reservoirs are shown in Table 4. Nitrogen input from the lake was 238 kg N ha"'^ with 89% of the N in the organic fraction. Total amount of fertilizer applied during the study period was 781 kg N ha ^, with NH^-N, NO^-N, and organic N representing 55, 30, and 15% of total fertilizer applied, respectively. The total N assimilated by water hyacinth (live plants and detritus) was 720 and 325 kg ha"-"^ yr'-^ for fertilized and control reservoirs, respectively (Table 4). Annual net N loading by detritus was 148 and 92 kg ha ^ for fertilized and control reservoirs, respectively (Table 4). Maximum detritus N loading occurred during winter for the fertilized reservoir and during spring for the control reservoir. This corresponded to the time of root dislodging from plants in the two reservoirs. The annual net N immobilized by detritus represented 21 and 28% of the total N removed by water hyacinth in the fertilized and control reservoirs, respectively. DeBusk et al. (1983) concluded that

PAGE 45

Table 3. Seasonal nitrogen uptake by water hyacinth and detritus. Fertilized reservoir Control reservoir Season Autumn (78) t Winter (90) Spring (8A) Summer (88) Water hyacinth Detritus Water hyacinth Detritus 127.8 33.9 167.7 2A2.4 — kg N ha 28.2 A2.4 37.5 39.8 1 30.9 -1.2 104.3 98.7 14.6 23.7 29.1 24.4 Test of significance i Season Month (season) Reservoir Water hyacinth ** NS Detritus NS ^ Number of days in season. t Significant at 0.05 (*) or 0.01 (**) level or not significant (NS),

PAGE 46

36 immobilization of N as plant detritus was 3 and 33% of standing crop assimilation for harvested and non-harvested water hyacinth plants, respectively. However, they did not include plant detritus trapped within the water hyacinth mat. The annual N assimilation by water hyacinth is low compared to N removal rates reported for plants growing in nutrient-enriched waters. Reddy et al. (1985) found annual N removal rates of 1726 and 1193 kg N ha ^ yr ^ for water hyacinths growing in primary and secondary sewage effluent, respectively. Rogers and Davis (1972) concluded that water hyacinths could remove 2500 kg N ha ^ yr ^ if maximvim growth could be sustained. Sato and Kondo (1981) measured a maximum removal rate of 4782 kg N ha ^ yr ^ for plants growing in a nutrient medium. The low annual N assimilation reported in the present study was due to low rates of fertilization. Plant uptake played a major role in removing N in both the reservoirs (Table 4). A large portion of lake water N was present as organic N, which was not readily available to plants. In both reservoirs, plants derived N from mineralization of lake water organic N, N release from underlying sediments, and mineralization of organic N in detritus. In the fertilized reservoirs, plants also derived N from the fertilizer N applied. Nitrogen assimiliation by water hyacinth from the added fertilizer was calculated as follows: (Total N assimilation by plants in the fertilized reservoir Total N assimilation by plants in the control reservoir / Total fertilizer N added) 100. About 51% of the added fertilizer N was taken up by the plants in the fertilized reservoir, and the remaining 49% may have been lost through denitrif ication. Reddy et al. (1982) observed a reduction of 78

PAGE 47

Table 4. Nitrogen balance for the two reservoirs. Fertilized reservoir Control reservoir Fertilizer nh'I'-n 4 NO^-N Organic N Total Lake water NH^-N 4 NO^-N Organic N Total Total added Water hyacinth Shoots Roots Detritus Total Reservoir water NH^-N 4 NO^-N Organic N Total Total N accounted kg ha Nitrogen added 430 234 117 781 13 12 213 238 1019 Nitrogen removed 354 218 148 720 13 13 167 193 913 13 12 213 238 238 95 137 92 325 8 5 156 169 494

PAGE 48

38 to 81% of the agricultural drainage effluent NO^-N and NH^-N in 3.6 days in a reservoir containing water hyacinths. DeBusk et al. (1983) calculated that A5% of the N removed from wastewater was immobilized in water hyacinth standing crop and detritus. About 30% of the fertilizer N was added as NO^-N, which could be potentially lost due to denitrif ication. Since water hyacinth plants prefer NH^-N over NO^-N (Reddy and Tucker, 1983), the majority of the plant N uptake probably came from NH^-N added through fertilizer. The role of underlying sediment in the immobilization/mineralization, and denitrif ication of N from these systems needs further investigation. Total N recovery in the fertilized reservoir was about 90%, and plant uptake represented about 71% of total N inputs. In the control reservoir, total N recovery was higher than the N inputs. Plants removed 325 kg N ha \ as compared to 238 kg N ha added. Release of N from sediment or mineralization of N during decomposition of detritus may account for the higher N recovery compared to total N inputs. Ogwada (1983) found a yearly average of 150 + 34 kg KCl-extractable inorganic N ha ^ sediment using monthly sediment N concentrations of the same reservoirs. Conclusions Primary productivity of water hyacinths was influenced by ambient air temperature, solar radiation, and nutrient composition of the culture medium. Net detritus production (total detritus detritus lost through decomposition) was relatively constant throughout the year and represented 3.5 to 14. 0% of the total standing crop. Detritus plant tissue of the fertilized reservoir contained higher tissue N, compared

PAGE 49

39 to the detritus in the control reservoir. Fertilization and increases in ambient air temperature resulted in dislodging of root biomass. Net N loading from detritus was 92 to 1A8 kg N ha ^ yr ^, which is potentially available upon decomposition. The N immobilized by detritus represented 21 and 28% of the total N removed by water hyacinths in the fertilized and control reservoirs, respectively. Approximately 51% of the added fertilizer N was assimilated by plants. The remaining 49% may have been lost through denitrif ication. Total N recovery was nearly 90% in the fertilized reservoir. More N was accounted for in the control reservoir than was added. Release of N from the sediment or mineralization of N during decomposition of detritus may account for the additional N recovery.

PAGE 50

EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER HYACINTH SYSTEMS Plant detritus (dead and decaying plant debris) is an integral part of water hyacinth mats and comprises 3 to 14% of the total biomass (see p. 39). It is usually derived from natural aging of plants, biological or chemical control, and frost damage. The addition of detritus to an aquatic system influenced several C and N transformations (Fenchel and Jorgenson, 1977). Nitrogen is present as NH^-N, NO^-N, and organic N in water media available for water hyacinth production. Organic N predominates in most water media and is not readily available for plant assimilation. Water hyacinths were efficient users of inorganic N and plant assimilation was a major process of N removal in aquatic systems containing water hyacinth (Reddy and Sutton, 1984). Other N transformations in aquatic systems resulting in removal of NO^-N or NH^-N include microbial assimilation, nitrif ication/denitrif ication, and NH^-N volatilization (Keeney, 1973; Bouldin et al., 1974). Addition of detritus significantly alters the rates of these processes. Mineralization or immobilization of N occurs during decomposition of detritus in water and sediment. Decomposition of detritus and subsequent N release was found to be related to C/N ratio, initial N and fiber contents (De La Cruz and Gabriel, 1974; Godshalk and Wetzel, 1978b; Odum and Heywood, 1978; Ogwada et al., 1984). 40

PAGE 51

41 A dense cover of floating water hyacinth depleted dissolved O2 of the underlying water, thus creating anaerobic conditions (Boyd, 1970; McDonald and Wolverton, 1980; Reddy, 1981). Decomposition of plant detritus also consumed O2 (Nichols and Keeney, 1973; Rai and Munshi, 1979). Anaerobic conditions may restrict nitrification and favor denitrif ication, which may proceed within the water hyacinth mat, in the water column, or in the underlying sediment. Detritus also provides energy source for denitrif ication. Denitrif ication occurred primarily in the underlying sediment and the rate depended on NO^-N diffusion from the water column to the sediment (Engler and Patrick, 1974; Reddy and Graetz, 1981). Volatilization becomes increasingly important as the water pH increases. The partial pressure of NH^-N in equilibrium with a solution of NH^-N increased rapidly in a pH range of 8.5 to 10.0 (Bouldin et al., 1974). A pH of 7.0 in water occurred in areas covered with water hyacinth with little diel variation (McDonald and Wolverton, 1980; Reddy, 1981) which suggests that NH^'N volatilization is minimal in areas covered with plants. The relative role of N assimilation by water hyacinth on total N removal from reservoirs was investigated by Reddy (1983). Approximately 40% of added ^^NH^-N or ''^^NO^-N was assimilated by plants. Less than 10% of the added ^^N was found in the surface sediment layer. Over 40% of the added ^^N was unaccounted for. Information on the role of detritus in aquatic systems on immobilization or mineralization of inorganic N is limited. The overall objective of this study was to determine the effect of detritus on selected N transformations in water columns with and without water

PAGE 52

A2 hyacinths. Specifically, the objectives were 1) to determine the regulatory function of detritus on dissolved 0^ and pH of water and 2) 15 to determine the influence of detritus on the fate of NO^ -N and ^^NhT-N in sedimentwater-plant systems. A Materials and Methods Two greenhouse studies were conducted to evaluate the effect of detritus on the fate of ^^N labeled NO^-N or NH^-N in water with and without water hyacinth plants. Treatments evaluated were: 1) with and without underlying sediment, 2) with and without water hyacinth plant cover, and, 3) three rates of added water hyacinth detritus. There were 2A tanks in each study having dimensions of 50 cm 50 cm 25 cm depth. Twelve of the 24 tanks contained a 2.5 cm sediment layer (1.875 kg soil). The sediment was a Lauderhill organic soil (Lithic raedisaprists) collected at the Central Florida REC research farm in Zellwood, Florida. The soil was air-dried and passed through a 2 mm sieve. Fifty liters of tap water were added to sediment tanks to obtain a 20 cm water depth. The greenhouse studies were initiated after sediment /water equilibration of 1 week. A nutrient medi\im (a modified 10% Hoagland's solution) was added to all tanks to obtain nutrient concentrations of: ^^NH^-N or ^^NO^-N = 20.0 mg l"-^; K = 23.5 mg l"-^; PO^-P = 3.1 mg l"S Ca = 20.0 mg L"S Mg = A. 8 mg l"-^; SO^-S = 6. A mg L~^; Fe = 0.6 mg L"-^ and micronutrients. Micronutrients were applied through commercially available liquid fertilizer (Nutrispray-Sunniland, Chase and Co., Sanford, Florida) to obtain concentrations of 0.2 mg Cu L ^ ; 1.5 mg Mn l"-^; O.OA mg B L~S and 0.02 mg Mo l"'^.

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43 Water hyacinth detritus (shoot and root material) was added at the rates of 0, 100, and 400 mg C l"''^. The detritus was chopped manually to lengths of -2 cm. The detritus for treatments with added ^^NO^-N was collected from a natural water hyacinth stand in Zellwood, Florida and had an initial N content of 5.6 mg g ^ dry tissue. The detritus for treatments with added ^^NH^-N was collected from a water hyacinth stand located in a wastewater stabilization pond at the University of Florida wastewater treatment plant in Gainesville, Florida and had an initial N content of 23.1 mg g ^ dry tissue. Water hyacinths, at an initial density of 10 kg (fresh wt) m were placed in 12 of 24 tanks. The plants were collected from the University of Florida's Bivens Arm research reservoirs in Gainesville, Florida. The plants were clipped of dead tissue and rinsed with tap water prior to placement in tanks. The disappearance of added inorganic N was determined by collecting water samples at 0, 1, 2, 3, 4, 8, 15, 28 days and measuring NH^-N, NO^-N, and total Kjeldahl N (TKN). The changes in water hyacinth fresh weight were measured weekly. Plant samples and detritus were analyzed for TKN. The sediment was characterized for organic and inorganic N prior to and at the conclusion of each study. Fifty grams (dry wt) of moist sediment samples were extracted with 2 M KCl and analyzed for NH^-N and NO^-N. Sediment samples were air-dried, ground by mortar and pestle, and analyzed for TKN. The inorganic N for all samples was detemined by steam distillation (Keeney and Nelson, 1982). The TKN of water, plant, and sediment samples were determined by micro-Kjeldahl procedures (Nelson and Sommers, 1972; 1973; 1975). The N analyses on water, sediment, plant and detritus samples were conducted using a Micro Mass 602 spectrometer.

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A4 Water pH (Orion Model A04 Specific Ion Meter), dissolved 0^ (Yellow Springs Instrtiment Model 54 O2 meter) and temperature were measured every other day. Electrical conductivity (Hach Mini Conductivity Meter) was measured weekly. Results and Discussion Effect of Detritus on Water Dissolved 02 The dissolved 0^ concentrations of water with added ^^NO^-N and NH^-N are shown in Figs. 9 and 10, respectively. Dissolved 0^ concentrations remained < 5 mg L ^ in water having plant cover but lower dissolved 0^ concentrations were recorded as the rate of detritus increased. This reflected increasing microbial 0^ demand for respiratory functions with increasing C source (Fenchel and Jorgensen, 1977). For water without plant cover (open water), the dissolved 0^ concentrations were scattered more with time. The dissolved 0^ measurements were taken between 2:30 and 3:30 pm and should represent near maximum concentrations on a diurnal basis (Howeler, 1972). The increased dissolved 0^ concentrations of open water were due to an increased rate of photosynthesis by algae during the day compared to respiration (Reddy, 1981). Generally the effect of decreasing dissolved 0^ concentrations with increasing detritus was seen for open water with or without sediment. Dissolved 2 concentrations were generally lower in open water with sediment compared to open water without sediment. Nichols and Keeney (1973) reported lower dissolved 0^ concentrations for sediment-water systems than water only. Although detritus appeared to have a role in O2 dynamics in water, plant cover was the primary regulator.

PAGE 56

I 46 ^uj) ^0 a3Aiossia

PAGE 57

Effect of Detritus on Water pH The pH of water with added ^\o^-N or ^^NH^-N is shown in Figs. 11 and 12, respectively. A fairly constant pH of 7.0 was" noted in water having plant cover and sediment regardless of detritus additions. The pH -decreased in water with plant cover but without sediment. The decreasing pH was noted immediately for added ^^NH^-N and after 20 days for added ^\o^-N. The decrease in pH was less as the detritus rate increased. The immediate pH decrease in water with plants and added NH"*'-N was A probably due to production of h"^ during plant NH^-N assimilation (Raven and Smith, 1976). The H"*" "generated is actively exuded, partly in exchange for cations (Franco and Munns. 1982). Plant NO^-N assimilation occurs by exchange with another anion or by simultaneous cation assimilation to maintain ion equilibrium (Kirkby and Mengel, 1967; Mengel, 1974). The decreasing pH in water with plant cover but without sediment suggested that the underlying sediment had a buffering role in pH regulation. The pH of water without plant cover were generally higher and more variable than water with plant cover. Reddy (1981) reported high mid-day pH values in ponds where algal activity was high. Bouldin et al. (1974) found high pH values (> 8.5) for ponds containing submersed macrophytes during sunlight hours. The pH of open water was generallylower as the rate of detritus increased. Effect of Detritus on Nitrogen Loss Nitrogen loss from water with added ^^O^-N or ^^NH^-N is shown in Figs. 13 and 14, respectively. Sediment or detritus had no apparent

PAGE 60

50

PAGE 61

51 INfllNOyNl^V

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52 effect on rate of N loss in water with plant cover. Nitrate and NH^-N disappeared within 2 and A days, respectively, in water with water hyacinths Nitrogen loss in open water was influenced by the underlying sediment and detritus additions. Nitrate disappeared more rapidly in open water with sediment than without sediment. An increase in detritus resulted in a more rapid NO^-N loss in water with or without sediment. A rapid decrease in NO^-N after 2 days was followed by an accumulation of NO^-N at A days for open water. Accumulation of NO^-N after 4 days in open water was probably due to rapid algal assimilation followed by turnover (death) of the algae and leaching of NO^-N from the dead algal cells. Surface algal mats developed in open water within 2 days. Ammonium disappeared more rapidly in open water with sediment than without sediment. Apparently detritus did not influence NH^-N loss in open water with sediment. However, detritus additions resulted in a more rapid NH^-N loss in open water without sediment. Loss of NH^-N followed by accumulation of inorganic N during the first A days was not as striking as seen for NO^-N. Plant Nitrogen Assimilation Total plant N assimilation and the contribution of added ''^^NO^-N and '^^NH^-N to total N assimilation is presented in Tables 5 and 6, respectively. Plant N assimilation was always greater for water with underlying sediment. Part of the increased plant N assimilation in water with sediment was due to release of N from the sediment. Generally the contribution of added ''"^O^-N or ^^NH^-N to total plant N assimilation also decreased with increasing detritus. Mineralization of

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Table 5. Total plant N and ^^NO^-N assimilation. Treatment Total uptake Labeled Other -mg (% of added ^^N)Without sediment 0 mg C L"-^"^ 767 + 86 652 (85) 115 (15) 100 mg C l"-^ 1088 + 87 675 (62) A13 (38) AOO mg C l"^ 1027 + 268 606 (59) 421 (41) With sediment 0 mg C l"-^ 1220 + 29 720 (59) 500 (41) 100 mg C l"-^ 1171 + 235 656 (56) 515 (44) 400 mg C L"^ 1112 + 163 567 (51) 545 (49) 'Carbon source was plant detritus. ^ Other N sources include sediment and detritus.

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Table 6. Total plant N and ^^NH^-N assimilation. Treatment Total uptake Labeled Other ^ -mg (% of added ^^N)Without sediment OmgCL'^^ 903+ AA 822(91) 81 (9) 100 mg C l"-^ 768 + 103 697 (91) 71 (9) AOO mg C l"-^ 1133 + 81 807 (71) 326 (29) With sediment 0 mg C L"-^ 1390 + 19A 891 (6A) A99 (36) 100 mg C l"^ 1255 + 89 8A5 (67) AlO (33) AOO mg C L"-^ 1629 + 65 828 (51) 801 (A9) "'^ Carbon source was plant detritus. Mother N sources include sediment and detritus.

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55 detritus N was a potential N source for plant assimilation. Reddy (1983) found that 60 to 64% of total N assimilation by water hyacinths was derived from added ''^^N, while 36 to 40% was derived from sediment and from decomposition of detritus. The ^''n recovered by plant tissue (mg) was fairly consistent for added labeled fertilizer but plant ^^NH^-N uptake exceeded '''^NO^-N uptake. Water hyacinth appeared to be more efficient in utilizing NH^-N than NO^-N (Reddy and Tucker, 1983). When water hyacinth growth is not restricted by climate, rapid assimilation of added inorganic N would be expected. Nitrogen15 Balance for Water Columns A ^^N balance for water with added ^^NO^-N and ^^NH^-N is presented in Tables 7 and 8, respectively. Total ^^N recovery by water hyacinths ranged from 57 to 72% and 70 to 89% in water with added ''^^NO^-N and ^^NH^-N, respectively. Reddy (1983) conluded that water hyacinth N assimilation accounted for only 40% of added ^^NO_-N or ^^NH^-N in a 3 4 reservoir. Algal surface mats accounted for -8% of added ^^NO^-N and up to 15% of added ^^NH^-N. The algal surface mats represented a minor portion of total microbial N assimilation. Algal activity was noted in open water, and the four sides and bottom of the microcosm tanks were colonized by algae. The ^^N associated with detritus was determined for water with added ''^NH'I'-N but not for added ''^^NO'-N. Less than 10% of the added 4 3 ^ NH^-N was immobilized by detritus in water with plant cover. Newly formed water hyacinth detritus from the plant cover was deposited during 28 days and accounted for 7% ^^N recovery in water without added detritus or sediment.

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56 Table 7. Mass balance of added NO_-N in sediment-water'-plant systems. Plant Sediment t Unaccounted Treatment or algae org inorg Water Total For % Recovery of N PLANTS Without sediment 0 mg 65.2 ND§ 65.2 34.8 100 mg 67.5 _._ ND 67.5 32.5 AOO mg 60.6 ND 60.6 39.4 With sediment 0 mg 72.0 3.2 0.4 ND 75.6 24.4 100 mg D J O / U H 400 mg 56.7 3.6 NO 0.6 PLANTS ND 59.9 40.1 Without sediment 0 mg ND 6.2 6.2 93.8 100 mg 6.8 ND 6.8 93.2 400 mg 8.2 ND 8.2 91.8 With sediment 0 mg ND 11.4 0.4 5.1 16.9 83.1 100 mg 8.1 10.1 0.4 2.0 20.6 79.4 400 mg C 8.4 4.9 0.5 0.5 14.3 85.7 T Carbon source was plant detritus. *Org, Inorg = organic and inorganic N, respectively. §ND = Not detectable.

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57 Table 8. Mass balance of added ^^NH^-N in sediment-water-plant systems. Plant Sediment t Treatment Unaccounted or algae Detritus Org Inorg Water Total For -% Recovery of ^^NPLANTS Without sediment 0 mg C l"-^ ^ 82.2 7.3 2.3 91.8 8.2 100 mg C L"-^ 69.6 3.6 5.8 79.0 21.0 AOO mg C L"-^ 80.7 7.1 3.8 91.6 8. A With sediment 0 mg C l"-^ 89.1 ND ^ A. 3 2.6 1.0 97.0 3.0 100 mg C l"-^ 8A.5 1.7 3.0 2.9 1.5 93.6 6. A AOO mg C L"-^ 82.8 3.9 3.3 3.5 1.5 95.0 5.0 Without sediment NO PLANTS 0 mg C L'-^ ND ND 9.6 9.6 90. A 100 mg C l"-^ ND 1A.7 17.5 32.2 67.8 AOO mg C l''^ 6.1 3A.7 5.2 A6.0 5A.0 With sediment 0 mg C L"-^ 1A.9 ND 10.2 7.1 2.3 3A.5 65.5 100 mg C L'-"" 15.0 5.9 11.0 8.8 5.8 A6.5 53.5 AOO mg C l"-^ 11.1 26.2 3.9 6.9 3.8 51. 9 A8.1 Carbon source was plant detritus. |Org, Inorg = organic and inorganic N, respectively. ^ND = Not detectable.

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58 Detritus ^^N recovery in water without plant cover increased with increasing rate of detritus. This suggests that during periods of low water hyacinth productivity, i.e. winter, detritus will be an important sink for inorganic N removal. The high '^^N recovery in detritus was -1 suprising since the original detritus had a high N content (23 mg g dry tissue). Therefore, the detritus used in water with added ^^NO^-N probably accounted for even more '^^N immobilization due to a low initial N content (5 mg g ^ dry tissue). Generally ^^N recovery in sediment was primarily organic N. Less than 1% of the added ^^NO^-N was recovered as sediment inorganic N. However, between 3 and 7% of the added ^^Nh1^-N was recovered as KCl 4 extractable inorganic N in the sediment. The lower recovery of sediment 15 15 inorganic N in water with added NO^-N was probably due to reduction to gaseous N via denitrif ication (Engler and Patrick, 1973). Some of the added ^^N was recovered as organic N in the water. Plant uptake was the primary mechanism of N removal in water having water hyacinths. The ^^N unaccounted for was lost from the systems through a variety of possible transformations. A more thorough investigation would be required to establish the extent of algal N assimilation. Volatilization of NH^-N in water without plant cover and denitrif ication in water with sediment are two possible mechanisms for N removal Conclusions Generally as the rate of detritus addition increased, dissolved 0. concentrations decreased in water with or without sediment and with or without plant cover. The decreasing dissolved 0_ concentrations were

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59 attributed to increasing heterotrophic respiration due to increasing amounts of C, Although this general relationship existed for all treatments, plant cover and sediment layer appeared to have more of a regulatory role in dissolved O2 dynamics than detritus. Water pH was constant in water having plant cover and sediment. The decreasing pH of water with plant cover and no sediment was attributed to NH^-N assimilation by plants in exchange for h"*". The pH of open water was generally lower as the rate of detritus increased. Detritus had no apparent effect on rate of N loss in water with water hyacinths. However, N loss was more rapid in open water as the rate of detritus increased. Total plant ^^NH^-N uptake exceeded ^^NO^-N uptake. Both sediment and detritus appeared to be a potential N source for water hyacinths. Total '^N recovered by water hyacinths ranged from 57 to 72% for added ^^NO^-N and 70 to 89% for added ^^NH^-N. Less than 10% of the added ^^NH^-N was immobilized by detritus in water with plant cover. However, in water without plant cover, up to 35% of the added ^^NH^-N was associated with detritus. This suggests that during periods of low water hyacinth productivity, typical in cold weather conditions, detritus is an important sink for added N.

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ANAEROBIC DIGESTION OF WATER HYACINTH The potential productivity of water hyacinth has led to its selection as a biomass feedstock for methane generation while providing a means for treatment of nutrient -enriched waters. Methane yields during anaerobic digestion depended on characteristics of the feedstock (Stack et al., 1978; Wolverton and McDonald, 1981) as well as digester operating conditions (Hashimoto et al., 1980). Sievers and Brune (1978) reported higher methane yields for digesters operating on swine waste as the C/N ratio increased. They concluded that the optimiim C/N range for maximum methane production was 15.5/1 to 19/1. The optimum pH and temperature range for anaerobic digestion was 6.7 to 7.4 (Bryant, 1979) and 30 to 35C (House, 1981), respectively. Biogas and methane yields have been reported for water hyacinths using a variety of digesters. Wolverton and McDonald (1981) reported methane yields of 0.07 to 0.20 L g"''' total solids (TS) for blended water hyacinths. Hanisak et al. (1980) found average methane yields of 0.24 L g ^ volatile solids (VS) from shredded water hyacinths in 162 L digesters at loading rates of 1.10 to 1.38 g VS L ''' day~^ and residence times of 30 to 38 days. Chynoweth et al. (1983) reported methane yields of 0.19 and 0.28 L g Ws of water hyacinth and a 3:1 water hyacinth/primary sewage sludge blend, respectively, in 5 L daily-fed digesters with a loading rate of 60

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61 1.6 g VS l''^ day'''^. Shiralipour and Smith (1984) reported average methane yields of 0.32 and 0.17 L g"-^ VS water hyacinth shoot and root samples, respectively, in a bioassay test of 100 ml culture volume. They also concluded that the addition of N in growth media for water hyacinth production increased methane yields of both shoot and root samples. Inoculum from operating anaerobic digesters is commonly added as a bacterial seed to initiate anaerobic digestion in new digesters (Sievers and Brune, 1978; Wolverton and McDonald, 1981; Field et al., 198A). Information on the effect of inoculum volume on gas production is limited. The objectives of this study were 1) to determine C and N mineralization during anaerobic digestion of water hyacinth; 2) to determine the effect of inoculum volume on gas production; and 3) to evaluate effluent (solids and liquid) composition based on inoculum volume. Materials and Methods Water hyacinths, with either high or low tissue N content, were anaerobically digested at 35C in 55 L batch digesters containing 2.5, 5, or 10 L of inoculum. Water hyacinths with a high N content (-34 g kg ^ dry wt plant tissue) were obtained from the wastewater treatment plant of the Reedy Creek Utility Company, Inc., at Walt Disney World near Orlando, Florida. Water hyacinths with a low N content (-10 g kg ^ dry wt plant tissue) were grown in nutrient-depleted water at Sanford, Florida. Both types of hyacinths were grown in ^^N labeled (NH^)2S0^ for two weeks, frozen and chopped to 1.6 mm length using a Hobart T 215 food processor.

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62 The digesters received A. 7 kgfresh weight of the ^^N labeled water hyacinths and an inoculum volume of 2.5, 5 or 10 L. A control digester received 10 L of inoculum and no plant material. The inoculum used for plants with high N content was obtained from an operating continuous -fed upflow digester receiving a feedstock of water hyacinth and domestic sewage sludge in a blend ratio of 3:1 (Chynoweth et al., 1983). The inoculum used for the plants with low N content was obtained from a non-operating continuously-fed tank digester receiving water hyacinth as feedstock. Each digester was buffered with 210 g NaHCO^ and tap water was used to bring each batch digester to 54. 7 kg. Gas production was monitored for 60 days. At the end of the digestion period, each digester was thoroughly mixed and the total contents were emptied into a 60 L tub. The digested materials were passed through a 1.00 mm fiberglass screen into a second 60 L tub to separate the digested biomass sludge from the effluent. The sludge was drained for 7 minutes and transferred into a polyethylene bag and placed directly into a freezer. The effluent was transferred to a water hyacinth production system. The liquid samples from the digester effluents and screened effluents (sludge removed) were analyzed for pH, electrical conductivity (EC), total solids (TS), fixed solids (FS), volatile solids (VS) (APHA, 1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH^-N and NO^-N by steam distillation (Keeney and Nelson, 1982), and chemical oxygen demand (COD) (APHA, 1980). The screened effluent was also filtered through a 0.2 )m membrane filter and analyzed for Ca, K, Na and Mg by atomic absorption and P by an autoanalyzer

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63 The fresh plant material and digested sludge were freeze-dried (Thermovac-T) and analyzed for the following: TS, FS, VS, TKN (Nelson and Sommers, 1973), total carbon (TC) (LECO Induction Furnace 523-300), lignin, cellulose and hemicellulose (Goering and Van Soest, 1970), and ashed mineral constituents (Gaines and Mitchell, 1979). Results and Discussion Characteristics of Inocula Characteristics of the two inocula varied considerably (Table 9). The inoculum used for plants with high N content (high N plants) contained higher levels of TS, VS, NH^-N, TKN, and COD than the inoculum used for plants with low N content (low N plants). Characteristics of inoculum depended on the type of feedstock used for digestion (Stack et al., 1978) as well as digester operating conditions (Hashimoto et al., 1980). The inoculum used for high and low N plants came from digesters with feedstocks of a 3:1 water hyacinth/domestic sewage sludge blend and water hyacinths, respectively. Ammonium accounted for 68 and 92% of the total N of the inoculum from the water hyacinth/sewage sludge and water hyacinth feedstocks, respectively. Ammonium was the primary N source for methanogenic bacteria (Zeikus, 1977). Carbon and Nitrogen Mineralization During Digestion Biogas (CH^ and CO2) production, corrected to standard conditions (0C and 0.1 MPa), is given in Table 10. Gas production essentially ceased after 60 days of digestion. Cumulative biogas production at 60 days for high N plants was approximately 21% less for 2.5 L of inoculum compared to 10 L, Furthermore, ciimulative biogas production at 15 days

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64 Table 9. Characteristics of the inocula used in the batch digesters. mg L % of TS High N plant material 1072 48 1530 U200 6.3 1.75 33.5 66.5 Low N plant material 535 21 562 784 7.7 0.29 83.1 16.9 ^COD = Chemical oxygen demand, TS, FS, and VS = Total, fixed and volatile solids, respectively.

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65 Table 10. Gas production during anaerobic digestion of high and low N water hyacinth plants. Inoculum Cumulative biogas production Total Gas Yields .volume 15 days 30 days 60 days biogas methane --L---Liters •L 8 ;-l VSHigh N plant material 2.5 16 4 40.8 60.3 U O 1 Zi U 5 28. 1 53.5 73.0 0. 23 0. 15 10 34. 5 59.1 75.4 0. 20 0. 13 Low N plant material 2.5 16. 9 45.5 67.9 0. 25 0. 16 5 20. 0 51.6 72.6 0. 27 0. 17 10 14. 7 52.2 67.4 0. 25 0. 16

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was over twice as great for the digester receiving the largest amount of inoculum. However, for low N plants, the amount of inoculiun did not appreciably affect cumulative biogas production during digestion. Cumulative biogas production at 60 days was similar for both high and low N plants. Biogas production at 15 days was generally greater for high N plants. It appeared that N was not a limiting factor for total gas production in either digestion test. Converting 60 day biogas production to biogas or methane yields (L g ^ VS added) is also presented in Table 10. Volatile solids included inputs from water hyacinths and inoculum. The average methane content of the biogas was 63.7 + 5.2 % based on 18 samples. Surprisingly, biogas and methane yields were higher for the low N plants. This was caused by an increase of VS from inoculum used in digesters for high N plants. The inoculum used for high N plants contained 1.75% TS (66.5% VS of TS) (Table 9). The inoculum for low N plants contained 0.29% TS (16.9% VS of TS). Gas production expressed in these units suggested that inoculum volume did not appreciably affect total biogas or methane yields. The average methane yields were O.IA and 0.16 L g ^ VS added for high and low N water hyacinth plants, respectively. The methane yields were lower than those reported for continuously-fed digesters (Hanisak et al., 1980; Chynoweth et al., 1983). Batch digestion (once fed and sealed) would not promote maximiam gas yields as frequent addition of fresh substrate enhances gas production (Price and Cheremisinof f 1981). Shiralipour and Smith (1984) reported that methane production for water hyacinth roots was lower than for shoots and that increasing N in water hyacinth growth media increased methane yields. Water hyacinths typically produced longer roots as water fertility declined (see p. 32).

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67 Shoot: root dry weight ratios of water hyacinth were higher when nutrients were not limiting and decreased significantly when plants grew in nutrient-poor waters (Reddy, 198A). It was assumed in the present study that gas production, both cumulative and yields, would be greater for the high N plants. A mass balance of N is presented in Table 11. The organic N content decreased after anaerobic digestion for each treatment. Mineralization of organic N to NH^-N was the primary N transformation occurring during digestion. The total N recovered was lower for low N water hyacinth plants. The majority of the N was recovered in the effluent as NH^-N. Most of the N placed in digesters was recovered in the effluent, although the proportion of NH^-N of the total N tended to increase (Hashimoto et al., 1980; Field et al., 1984). Approximately 30% of the organic N placed in the digesters was recovered as organic N in the digested sludge for both high and low N plants. The organic N recovered in the screened effluent was 15 and 36% of the added organic N for high and low N plants, respectively. The total organic N recovered as effluent or sludge organic N was 45 and 66% of added organic N for high and low N plants, respectively. Therefore, a high N content of water hyacinth resulted in more mineralization of added organic N. Total ^^N recovered as "^^NH^-N in the screened effluent was 72 + 4% 4 — for high N plants compared to 35 + 9% for low N plants (Table 12). The organic ''^^N recovered in digested sludge accounted for 20 + 5% of the added ^^N from fresh water hyacinth plants regardless of N content. Approximately 11 and 20% of the added ^^N was recovered as organic N in the screened effluent for digested high and low N plants, respectively.

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68 Table 11. Nitrogen balance for the batch digesters. High N plant material Low N plant material __1 9 c ; T t T Lj 1 n T, 2.5 L 5 L 10 L Nitrogen added g Water hyacinth Organic N 10. 39 10. ,39 10. 39 3 24 3. 24 n J Inoculum Organic N 1. 15 2. ,31 4. ,61 0 0/ 0. 14 U Z / Inorganic N 2. ,68 5. ,36 10. ,72 1 34 2. 68 5 35 Total Organic N 11. ,54 12. ,69 14. ,99 3 ,31 3. ,37 J Dl Inorganic N 2. ,68 5, ,36 10. ,72 1. ,34 2. ,68 5. ,35 14. ,22 18. .05 25. ,72 4. ,64 6. ,05 8. ,86 Nitrogen recovered Screened effluent Organic N 1, ,48 2, .41 2. ,02 1, ,26 1. ,31 1. ,09 Inorganic N 8. ,81 11, .60 15, .81 1, ,20 2. ,63 4. ,98 Digested sludge Organic N 2. .86 4, .91 4, ,94 1, .17 0, .85 0, .87 Total Organic N 4, ,34 7, .32 6, .96 2, .43 2, .16 1, .96 Inorganic N 8, .81 11 .60 15, .81 1, .20 2, .63 4, .98 13, .15 18 .91 22, .77 3, .63 4, .79 6, .94 % Recovered 92 105 89 78 79 78 ^Vol\ime (liters) of inocultim.

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69 Table 12. Nitrogen15 balance for the batch digesters. High N plant material Lovi r N plant material 2.5 Lt 5 L 10 L 2.5 L 5 L 10 L 15 5 N Added •-g Water hyacinth Organic N 10. 39 10 .39 10, .39 o • 24 3. 24 3 .24 N Recovered Screened effluent Organic N 1. 18 1 .48 0. .93 0. 84 0. 65 0 .34 Inorganic N 7. 06 7 .78 7. .68 0. ,79 1. 25 1 .34 Digested Sludge 0 .52 Organic N 1. 67 2 .53 2, .21 0. ,87 0. 57 Total 9. 91 11 .79 10 .82 2. ,50 2. 46 2 .20 % Recovered 95 113 104 77 76 68 + Volume (liters) of inoculum.

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70 The low total recovery of N and ^^N for low N plants after anaerobic digestion is difficult to explain. Nitrogen cycling during anaerobic digestion was primarily mineralization of organic N or immobilization of inorganic N. Volatilization of NH^-N may occur but the potential increases as NH^-N concentrations increase or at higher pH values (Freney et al., 1983). Each digester received 210 g NaHCO^ as a buffer and the pH after digestion was similar for all digester effluents. The NH^-N concentrations after digestion were much higher for the high N plants (Table 13). Effluent composition Characteristics of the digester effluents prior to sludge separation are presented in Table 13. Generally, as the rate of inoculum increased, there were increases in EC, NH^-N, TKN, and TS. The COD increased with increasing inoculum volume for digesters with high N plants. Characteristics of the screened effluent (sludge removed) are reported in Table 14. Removing the digested biomass sludge from the digester effluents decreased the EC, NH^-N and TKN. The screened effluent from the low N plants contained more Ca, Mg, K, and Na, and less P than the screened effluent from the high N plants. Characteristics of the fresh plant biomass and digested biomass sludge are given in Table 15. Anaerobic digestion resulted in increases in TC and TKN of sludge compared to fresh plant biomass. The increases in sludge TKN after digestion of low N plants caused a reduction of the C/N ratio from 35 to 16. The changes in TC or TKN of the digested high N plants did not appreciably alter the C/N ratio. The digested sludge

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Table 13. Characteristics of digester effluents before sludge removal Inoculum Digester Effluent Characteristics' volume pH EC COD NH^-N TKN TS VS FS --L-dS ,-1 --mg L % -% of TSHigh N plant material 2.5 7. A A. 5 3030 189 238 0 .A98 AO. 3 59.7 5 7. 6 5.1 A290 210 315 0 .570 38.7 61.3 10 7. A 5.6 5050 29A A06 0 .610 38.9 61.1 Control 7. 7 NA* A170 205 259 0 .535 36.6 63. A Low N plant material 2.5 7. 3 6.6 1690 50 70 0 .AA5 28.8 71.1 5 7. 2 6.9 23A0 70 112 NA 37.3 62.7 10 7. 3 7.8 16A0 112 15A 0 .A88 28.8 71.2 Control 7. 8 NA 109 98 120 0 .315 lA.O 86.0 COD = Chemical oxygen demand, TS, FS and VS = Total, fixed and volatile solids, respectively. NA = Not available.

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o •H 4-1 w •H (U +J a n) u P c 0) r-l w 0) c 0) o C/3 Cm C/2 > CO H z z I z 33 in o in ON 1 00 av ON ON -H in in VO 1 H 4-1 O m o 6^; 1 • • • — o O 00 1 < ro CN ON r*^ 00 in in vO o CN m • • • o o o o t J 00 00 in 1 • • 1 1 t-H — 1 f— 4 1 r— ( 1 •H rt M O o O O o O tu CN 00 f— < vO r— 1 00 Ov n ro VO C3V a < in in in vO vO in in r~ 1 — in 1 in O in o r1

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1 0 CN C 0 vO 0^ I f— 1 0 0 1-H CM CN ; 00 00 CN vO cu vO 00 00 j 1 .—1 ro *-H in 00 0 0 00 o 00 in 00 to VO 0 0 CN CN rH r-H oO 00 CN 0 00 CN O) r— t CN CN VO CN CN CN 1 10 00 0 00 00 vO * ^ CO CN CN CN CN CN vO 00 0 1-H CN 00 • •H 0 fd f 00 r-H 1-H CN vO CO •H rrl tu CN CN CN CN m CN CO VO I CN t— t CN 1-H M I-H CO T3 T3 (U T— CO ;3 QJ 0) QJ 0) OJ OJ 0) X) 4-) P +J X. -P 4-J -p •H CO CO CO CO CO CO CO CO •>> rH 0 CO OJ QJ QJ QJ QJ OJ QJ QJ J3 OJ )-< £03 CkO in bO 00 00 •H CC PQ J-l •H •H •H Ph •H •H •H E-* Q Q Q Q i— 4 •H

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74 had a higher lignin content compared to fresh plant biomass. The increase in lignin was due to the loss of readily decomposable C during anaerobic digestion. Lignin appears to be practically inert to anaerobic digestion (Hashimoto et al., 1980) Generally, there was a decrease in cellulose after digestion. The hemicellulose remained similar for the fresh plant biomass and digested sludge. Anaerobic digestion resulted in losses of K and Mg from fresh plant biomass, but increased the concentration of sludge Ca, Na, Fe, and Zn. Conclusions Cumulative biogas production at 60 days was similar for high (-3A g N kg ^ dry wt tissue) and low (-10 g N kg *) N plants suggesting that long term digestion of water hyacinth was not influenced by initial N content. Effects of inoculiim volume on cumulative biogas production were seen at 15 days for high N plants but not low N plants. Conversion of cumulative biogas production into biogas and methane yields (L g ^ VS added) showed that low N plants produced more biogas and methane than high N plants. This was due to increase of TS (and consequently VS) in digesters of high N plants from the inoculum source, since cximulative gas production was similar for both types of plants. Mineralization of organic to ^^NH^-N accounted for 72 and 35% of added '''^N for high and low N plants, respectively. Approximately 20% of the added ^^N was recovered as organic N in sludge for both types of plants. A low ^^N recovery was observed for low N plants. Increasing inoculum volume increased electrical conductivity, NH^-N, TKN, and TS of the digester effluents. The digested biomass sludge had higher levels of TC, TKN, lignin, Ca, Na, Fe, and Zn, and lower levels

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75 of K and Mg compared to the fresh plant biomass. The C/N ratio of the fresh plant biomass with a low tissue N content decreased from 35 to 16 after digestion. The C/N ratio of the fresh plant biomass with a high tissue N content was the same as the digested biomass sludge (C/N=12). t

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TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTHS An integrated approach of wastewater renovation using aquatic macrophytes with utilization of biomass for energy production is economically appealing. The plant biomass produced in these systems, along with other wastes such as sewage sludge or animal waste could be anaerobically digested to produce methane (Stack et al., 1981; Shiralipour and Smith, 1984). This process generates a waste by-product which must be disposed of, or preferably utilized to reduce the cost of energy production, in an environmentallysafe manner. The waste by-product consists of digested sludge and a large volume of effluent. Integrating wastewater renovation through water hyacinth production provides an internal option for the disposal of effluent generated during conversion of biomass into methane. The effluent composition of anaerobic digesters varied with type of feedstock used in digestion (Stack et al., 1981). Information on chemical composition of effluents from sewage or animal wastes was readily available (Sommers, 1977; Field et al., 198A). However, anaerobic digestion of plant biomass has only recently gained attention in the United States and information on composition or disposal of the effluent was limited (Hanisak et al., 1980; Atalay and Blanchar, 1984). Digester effluents have high concentrations of BOD, NH^-N, K and Na (Atalay and Blanchar, 1984; Field et al., 1984), while the divalent cations and metals were concentrated in the sludge (Sommers, 1977; Field et al. 1984). 76

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77 Water hyacinth-based wastewater treatment systems have already been evaluated for use in treating primary and secondary sewage effluents (Wolverton and McDonald, 1979; Reddy et al., 1985) and anaerobic digester effluent (Hanisak et al., 1980). The potential productivity of water hyacinth in nutrient-enriched waters has led to its selection in alternative methods of wastewater renovation, particularly in areas where growth is not restricted by climatic limitations. Use of water hyacinth for digester effluent treatment is particularly attractive, because of its ability to grow in waters with high elemental concentrations. The biomass produced could be returned to the digester as a feedstock for methane production. Hanisak et al. (1980) determined that 64.5% of (liquid and sludge) N in diluted • effluents from anaerobically digested water hyacinth could be reassimilated by water hyacinths. Diluting the effluent does not address the full potential of water hyacinth to grow under these nutrient and salt enriched conditions. Haller et al. (1974) concluded that water hyacinth will not live in waters containing sustained salt concentrations in excess of 2500 mg L "*". Optimal dilution of these concentrated effluents to obtain maximum water hyacinth yields and nutrient removal was not reported. The objectives of this study were to 1) evaluate water hyacinth productivity in anaerobic digester effluents obtained from digesters receiving different types of water hyacinth as feedstock, and, 2) determine ''"^N recovery by water hyacinth growing in digester effluents from digested ^^N labeled water hyacinth biomass.

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78 Materials and Methods Anaerobic digester effluents were obtained from six 55 L batch digesters containing water hyacinth with a high or low tissue N content as feedstock. Water hyacinths with low (-10 g N kg ^ dry plant tissue) and high (-34 g N kg ^) tissue N content were grown in nutrient-depleted water and sewage effluent, respectively. After removal from their respective growth media, the hyacinths were grown in ^^N labeled (NH^)2S0^ nutrient solution for two weeks, frozen, and chopped to 1.6 mm length using a Hobart T 215 food processor. The water hyacinths were anaerobically digested for four months in 55 L batch digesters. Each digester received A. 7 kg (fresh weight) of the ^^N labeled water hyacinth, 2.5, 5, or 10 L volume of inoculum from anaerobic digesters receiving water hyacinth as feedstock, and were buffered with 210 g NaHCO^. After digestion, the biomass sludge was separated from the effluent by passing the total contents of the digesters through a 1.00 mm fiberglass screen. The screened effluents (sludge removed) were analyzed for total solids (TS), volatile solids (VS), fixed solids (FS) (APHA, 1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH^-N and NO^-N by steam distillation (Keeney and Nelson, 1982), electrical conductivity (EC) (Hach Mini Conductivity Meter) and pH (Orion Model 404 Specific Ion Meter). Samples passed through a 0.2 pm membrane filter were analyzed for Na, K, Mg and Ca by atomic absorption and ortho P colorimetrically after reacting with ammonium molybdate. Six water hyacinth plants were placed in 10 L of undiluted or diluted effluents in containers having a surface area of 0.051 m^. The water hyacinth plants were collected from the University of Florida's

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79 Bivens Arm research reservoirs in Gainesville, Florida. The plants were clipped of dead tissue and acclimated to greenhouse conditions for two weeks prior to treatments. The studies to evaluate the potential of water hyacinth to treat digester effluents were conducted for a period of 22 days in March and September, 1984. The daily maximum greenhouse temperatures ranged from 19 to 37C in March and 27 to 35C in September There were a total of 10 treatments, replicated three times, as described below. Treatments 1 to 3 were diluted effluents from digesters containing high N water hyacinths and inoculum voliames of 2.5, '5, and 10 L, respectively. The dilutions were 1:8, 1:4, and 1:3 effluent : tap water for digester effluents with inoculum volumes of 2.5, 5, and 10 L, respectively. Treatments 4 to 6 were undiluted effluents from digesters containing high N plants and inoculum volumes of 2.5, 5, and 10 L, respectively. Treatments 7 to 9 were undiluted effluents from digesters containing low N plants and inoculum volximes of 2.5, 5, and 10 L, respectively. The final treatment was a modified 10% Hoagland's solution (p. 42) to serve as a control. Characteristics of the inoculum and effluents are given in the Anaerobic Digestion of Water Hyacinth chapter (pp. 60 to 75). Plant samples collected initially and at the conclusion of the experiment were analyzed for dry weights, TKN (Nelson and Sommers, 1973), Na, K, Ca and Mg by atomic absorption and P by an autoanalyzer Water samples were collected at 0, 1, 2, 3, 4, 8, 15, and 22 days and analyzed for NH^-N and NO^-N by steam distillation (Keeney and Nelson, 1982), TKN (Nelson and Sommers, 1975), Ca, Mg, K, Na by atomic absorption and P by an autoanalyzer. Electrical conductivity (Hach Mini

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80 Conductivity Meter), pH (Orion Model 404 Specific Ion Meter) and dissolved O2 (Yellow Springs Instrvunent Model 54 O2 Meter) were measured every other day. The ^^N analyses of plant and water samples were conducted on a Micro Mass 602 spectrometer. Results and Discussion Chemical Composition of the Effluents Characteristics of digester effluents used in the study are given in Table 16. The initial pH of the effluent sources and nutrient medium were similar. The digester effluents had a wide range of EC, NH^-N, TKN and other nutrients. This provided an opportunity to evaluate water hyacinth growth under diverse media conditions. The EC of the nutrient medium and diluted effluents ranged from 0.7 to 2.3 dS m''^. The EC of the undiluted effluents ranged from 4.3 to 6.7 dS m'"*". The NH^-N and TKN concentrations of the undiluted effluents from digested plants with a high N content (high N plants) were greater than those of the undiluted effluents of digested plants with a low N content (low N plants). The NH^-N concentrations ranged from 23 to 104 mg L~-^ for diluted effluents and 24 to 289 mg L"-^ for undiluted effluents, respectively. High Na and K concentrations were noted for undiluted effluents. The highest levels of P were in undiluted effluents of digested high N plants and the highest levels of Ca and Mg were in undiluted effluents of digested low N plants. The critical levels of Na, K, Ca and Mg needed to achieve maximum water hyacinth yields are relatively unknown. The NH^-N, K and Mg concentrations were higher than levels reported for water hyacinths cultured in primary or secondary sewage effluent (Reddy et al., 1985).

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J (30 1 1 1 00 m CS H 1 .—1 .—1 — ( T-H •H 1 o 1 a) 1 g 1 1 I < 00 CN O m 1 CVl .—1 m ro CN 0) 1 •H 1 Vi 1 4J 1 1 1 00 iri +J 00 00 in 1 w c 1 -JCO CN CN r-H 1 c rH .— I rH (11 1 CO a 1 r-l [/) 1 a z 1 1 — ( QJ 1 o >^ cs u (N o ro 3 J2 4H •H <4-l e g a) o e u o o o ^-i o o O 0) CO u IT) < vO VO 4J 1 — 1 CO (A ( r-l rH (0 +J t— 1 r— 1 0) 1 ca C u 4-1 0} •H [ •b iH p >4-l 0 <4H >o oi Q) 00 vO CN 00 IT) CN 1 (U •— I t3 — i CM ro m (U o j T3 4J Q) (0 z 4J ,— 1 1 •H H f — 1 CS ON +j •H >£) o rH 00 (0 z — t — 1 CN CN •H 0) P U CO CO vO m VI ca o r— 1 in .c u tc 00 VD VO P •H c M 0) • iH 1 SO 3 J in 1-1 O 1—1 1 in o in o O o 1 •—1 rH O c > iH M CO m u c rH a z o u C (U 3 rH Mh Mh 0) t3 0) c JO o ro m in in o m ^
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82 Productivity of Water Hyacinths Total dry weight gains of water hyacinths were consistantly less in the undiluted effluents (Fig 15). Complete death of plants was observed in four undiluted effluents. The loss of dry weight for these treatments was probably due to leaching of soluble plant constituents after plant death. The highest dry weight gains were associated with the lowest EC. However, plants survived in undiluted effluents having EC levels of 5.6 and 5.9 dS m"''' (5600 and 5900 pmhos cm'^). These EC levels were equivalent to -2900 and 3200 mg NaCl L ^ and were higher salt concentrations reported for water hyacinth survival (Penfound and Earle, 19A8; Haller et al., 1974). The undiluted effluents had Na levels in excess of 1100 mg L ^ Apparently water hyacinth has a wide range of adaptability to media composition and, therefore, total salt concentration is not a good criterium for determining plant survival. The diluted effluents were excellent media for plant growth and the gains in dry weight were consistently higher compared to the nutrient medium (Fig. 15). The highest dry weight gain was in the diluted effluent having N and P concentrations of 65 and 5.5 mg L ^, respectively, and a N/P ratio of 11.8:1. Sato and Kondo (1981) reported maximum yields of water hyacinth at a N and P concentration of 50 and 13.8 mg L ^, respectively, and a N/P ratio of 3.7:1. Dry weight gains were noted for two of the undiluted effluents. However, tissue damage was noted in all undiluted effluents. Tissue damage in undiluted effluents was observed within 24 hr after study initiation. Two types of leaf tissue damage were observed. Damaged leaves on younger shoots had burnt (brown) tips which curled up

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83 o 00 r-s CM 00 CSJ O CO O lO ro cvj O O O O 't ro O o CVJ O o o o I C^J" NIVO IHOGM Ada • • o o 3 0) n CO 4= C M T3 C 0) •H e bC 4-1 c AJ (1) ,C -H bO l-i •H 4-1 ? C M C O CO V u s

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8A towards the center of the plant.The other leaf tissue damage on young and older leaves was curling of the entire leaf towards the center of the leaf. Stem damage in undiluted effluents was noticeable after 2 days. Damaged stems collasped under slight manual pressure. The destruction of chlorophyll in stems and leaves was widely observed in the undiluted effluents. The extent and spread of tissue damage increased in undiluted effluents with increasing EC and NH^-N concentrations. At the end of one week, all plants in four undiluted effluents were dead. The EC and NH^-N concentrations of these effluents ranged from A. 3 to 6.7 dS m ^ and 87 to 289 mg N L ^, respectively. The shoots began to separate from the roots at the water surface and the submerged roots sank. Root separation also accounted for the negative dry weight gains of plants in undiluted effluents. Three treatments showed occasional visual signs of tissue damage, i.e., the nutrient medium and 2 diluted effluents. Visual signs of plant damage but noticeable gains in plant dry weights were observed in the diluted effluent having an initial NH^-N concentration of lOA mg L and two undiluted effluents having NH^-N concentrations of 2A and A9 mg L ^. All remaining treatments resulted in plant death, apparently due to high EC or high NH^-N concentrations, or a combination of both. Although water hyacinth plants struggled to survive in the undiluted effluents, algal activity was noted in all undiluted effluent treatments. Upon emptying the containers, algae were found attached to the side and bottom surfaces of the plastic containers.

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85 Nitrogen Removal First-order kinetic equations were used to described NH^-N loss from the effluents. An integrated rate equation for a first-order reaction is expressed as: In C / = kt o t where C = initial NhT"N concentration in the effluent, o 4 = final concentration at time = t, k = first order rate constant (days The rate constant is calculated by solving for k: k = In C / 1/t o t The rate constants for diluted effluents ranged from 0.228 to 0.593 day"''^ (Table 17). The rate constants for undiluted effluents ranged from 0.175 to 0.A46 day'''^. The time required for a 50% reduction in initial NH^-N concentration generally increased with decreasing dry weight gains and increasing NH^-N concentrations. The shortest reduction times were associated with rapid plant growth (50% reduction in 1.12 to 3.04 days). Plant assimilation was probably the primary mechanism of NH^-N loss in these treatments. Reddy (1983) reported a 50% reduction of inorganic N from agricultural drainage water in 18 days. The time required for a 50% reduction of NH^-N in effluents resulting in plant death was 1.98 to 3.96 days. Mechanisms of NH^-N loss in treatments resulting in plant death included microbial assimilation and NH^-N volatilization. The pH of the effluents ranged from 7.0 to 7.9 in treatments with actively growing plants but increased from 8.2 to 9.3 in treatments where plant death was observed and algal growth increased (APPENDIX A, Table 27). The potential of NH^-N volatilization increases as NH^-N concentrations increase or at higher pH values (Freney et al., 1983).

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86 Table 17. First-order kinetic descriptions of NH^-N loss with time. Initial NH, -N volume cone. k time' R Inoculum NH -N Reduction „ 1 J. • t -D^ \r\t^ IT r 1 mo I K --L-mg L ^ day ^ days Diluted effluents from high N plants 2.5 23 0.593 1.12 0.722 5 65 0.A49 1.54 0.917 10 104 0.228 3.04 0.951 Undiluted effluents from high N plants 2.5 161 0.232 2.98 0.938 5 212 0.207 3.35 0.947 10 289 0.175 3.96 0.982 Undiluted effluents from low N plants 2.5 24 0.446 1.55 0.850 5 49 0.325 2.13 0.885 10 87 0.350 1.98 0.845 Nutrient medium 20 0.281 2.47 0.973 t Time required for 50% reduction in initial NH^-N concentration

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87 Clock (1968) reported a 75% reduction of NO^-N in 5 days for water hyacinths growing in secondary sewage effluent. A 75% reduction of NH^-N in digester effluents required 2.3 to 6 days for treatments where positive dry weight gains were observed. For treatments resulting in plant death, 4 to 8 days were required to remove 75% of the NhT-N. 4 Nitrogen-15 plant assimilation was observed for all treatments although a low recovery was observed in treatments resulting in plant death (2 to 16%) (Table 18). The ^^N recovery by plants for the other treatments ranged from 36 to 77%. The majority of the ^^N was found in the shoot material for all treatments (54 to 73%). Approximately 75% of the was unaccounted for in treatments resulting in plant death. Microbial assimilation and NH^-N volatilization were probably important NH^-N removal processes in undiluted effluents where plant death was observed. Plant Tissue Chemical Composition Plants survived in the diluted effluents but death was noted for plants in the undiluted effluents from digested high N plants. Mineral constituents from plants growing in these effluents were analyzed to isolate individual cation and P assimilation or loss from living and dead plant tissue. The concentrations of plant tissue (root and shoot fractions) Na, K, P, Ca and Mg are reported in Table 19. The original plant tissue had low concentrations of Na and P in both shoot and root material, but higher concentrations of K, Ca and Mg in the shoots compared to the roots. There were large increases in Na for both shoots and roots of the surviving and dead plants. The root K concentrations increased for surviving plants but decreased for dead plants. The P concentrations

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88 Table 18. Nitrogen15 balance for labeled effluents. Recovered by Inoculiam Available plants N Recovery volume 15N Roots Shoots Plants Water Unaccounted jjjg % of applied ^^NDiluted effluents from high N plants 2.5 225 44 103 66 7 27 5 649 80 199 43 9 48 10 1044 101 272 36 3 61 Undiluted effluents from high N plants 2.5 1610 20 45 4 19 77 5 2120 18 41 3 27 40 10 2890 19 46 2 25 74 Undiluted effluents from low N plants 2.5 236 62 119 77 6 17 5 487 71 124 40 4 56 10 869 63 73 16 6 78

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89 Table 19. Distribution of nutrients in water hyacinth shoots and roots in diluted or undiluted effluents of digested high N plants. Inoculum volijune Na K P Ca Mg --L-.g kg SHOOTS Diluted effluents from high N plants 2.5 13.3 13.3 2.4 16.9 9.2 5 17.5 19.3 3.9 17.8 9.1 10 18.0 23.2 5.0 15.6 8.6 Undiluted effluents from high N plants 2.5 21.0 18.0 3.5 18.2 5.6 5 20.5 17.7 3.9 22.6 6.2 10 19.2 14.5 3.8 19.3 5.7 LSD (0.05) 4.4 7.1 1.3 3.7 1.9 Original shoot tissue 2.8 22.0 2.7 15.4 5.7 ROOTS Diluted effluents from high N plants 2.5 14.5 7.1 2.1 6.1 4.6 5 17.8 8.9 2.6 6.0 4.7 10 16.0 12.4 3.4 9.0 4.2 Undiluted effluents from high N plants 2.5 16.2 3.1 11.2 26.5 4.6 5 16.8 3.3 10.3 23.6 4.3 10 17.2 3.3 10.2 24.6 4.1 LSD (0.05) 3.0 3.7 1.9 3.0 0.7 Original root tissue 4.6 5.5 2.8 6.6 2.6

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90 increased for the dead plant root material but remained similar for the other materials. The Ca content increased for the shoots of all plants and the roots of the dead plant material. The Mg content increased for roots of all plants and shoots of the surviving plants. The net assimilation or loss of plant nutrients from plants in diluted or undiluted effluents from digested high N plants are reported in Table 20. The plants in diluted effluents assimilated large amounts of Na and K, apparently because these nutrients move rapidly with the transpiration stream. The net shoot assimilation of all nutrients was greater than the net root assimilation. The dead plants from undiluted effluents showed a net loss of K but net gains of the other nutrients. Potassium, Na, Ca and Mg were reported as being rapidly lost during the early leaching phase of plant decomposition in fresh water (Boyd, 1970b; Davis and van der Valk, 1978). Most of the Na moved into the shoot region. Generally the net gains of P and Ca were found in dead plant roots compared to shoots of surviving plants in diluted effluents. Final Chemical Composition of the Effluents Characteristics of the digester effluents and nutrient medium after water hyacinth treatment are given in Table 21. For treatments where plant dry weight gains were observed, generally there was large reductions of the elements analyzed in the effluents. For the treatments resulting in plant death, K and Mg increased and the reductions of other elements were of lesser magnitude. The largest reductions of EC (49 + 7% reduction), K (93 + 3%) and P (92 + 3%) were observed in diluted effluents. The highest gains of

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Table 20. Net assimilation or loss of plant nutrients in diluted or undiluted effluents from digested high N plants. Inoculum .volume Na K P Ca Mg --L-mg m day SHOOTS Diluted effluents from high N plants 2.5 2A0 35 19 179 126 5 353 187 57 223 138 10 333 220 69 148 110 Undiluted effluents from high N plants 2.5 190 -23 10 42 4 5 201 -36 15 89 9 10 181 -92 11 35 -4 LSD (0.05) 64 101 17 62 27 ROOTS Diluted effluents from high N plants 2.5 217 80 14 53 59 5 273 111 22 50 61 10 227 155 32 94 48 Undiluted effluents from high N plants 2.5 40 -26 31 73 2 5 50 -29 32 72 1 10 70 -29 42 103 3 LSD (0.05) 57 50 27 60 14

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3 P O r-l o o c > 00 1 1 (Nl 00 00 rH a> — 1 u 1 1 1 .-H •— 1 rH 1 1 1 1 n -4n in ON rH a. 1 • > • • • • 1 o O o o 1 .—1 1 1 m 1 to +J 1 -P c 1— 1 c (0 1 1-1 ;^ iH ON 6 o o 1-1 V4 m (0 m o 00 +J CN C3N 4-1 c C m OJ o 0) 3 00 00 3 iH iH m m CN <• in in d rH lO in 00 V V V CN vO 00 < -3c CN m
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93 plant dry weight were also noted for these effluents. AmmoniiMi was >1.0 mg L '^ and TKN was reduced 83 to 97% in these effluents after 22 days. Sodium reduction was in the range of 34 to 52%. Calcium and Mg decreased for the 2.5 and 5 L inoculum diluted effluents but increased for the 10 L inoculum diluted effluent. Intermediate reductions of EC (25 +3%), K (58 + 6%) and P (69 + 7%) were observed in undiluted effluents from digested low N plants. Ammonium was < 1.0 mg L ^ and TKN was reduced 90 to 93% in these effluents after 22 days. Sodium decreased 54 to 63% after 22 days. The largest reductions of Ca (82 + 4%) and Mg (43 + 8%) were found in these effluents. Plant death was noted in all undiluted effluents from digested high N plants. These treatments had the lowest reductions of EC (5 + 3%) and P (26 + 10%). Plants were removed after 10 days following complete death of aerial tissue and separation of root masses. The death of plant tissue resulted in an increase of K and Mg in the effluents. Sodium was reduced 18 to 23%. Calcium reduction was in the range of 52 to 70%. The loss of NH^-N (83 to 86% reduction) was probably due to NH^ volatilization and microbial assimilation. Conclusions The highest dry weight gains were for plants growing in diluted effluents with EC levels ranging from 0.7 to 2.3 dS m"'^ and Nflt-N 4 concentrations of 23 to 104 mg L ^ Plants survived and grew in two undiluted effluents with EC levels of 5.6 and 5.9 dS m'^ and NH^-N 4 concentrations of 24 and 49 mg l''''. All other EC and NhT-N combinations 4 of undiluted effluents resulted in plant death.

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94 A first-order kinetic equation was used to describe NH^-N loss with time. Rate constants for diluted effluents ranged from 0.228 to 0.593 day ^. Rate constants for undiluted effluents ranged from 0.175 to 0.446 day ^. The time required for a 50% reduction of NH^-N was 1.12 to 3.04 days for treatments with positive water hyacinth dry weight gains. A 50% reduction of NH^-N in treatments resulting in plant death required 1.98 to 3.96 days. Plant assimilation was one of the primary mechanisms of NH^-N loss in the systems with actively growing plants. Microbial assimilation and NH^'N volatilization were probably important mechanisms of NH^-N removal for treatments resulting in plant death. Plant assimilation accounted for a 36 to 77% recovery of effluent ^^N for surviving plants. Only 2 to 16% of the ^^N was recovered in dead plant tissue. Approximately 75% of the ^^N was unaccounted for in effluents resulting in plant death. Surviving plants assimilated large amounts of Na, K, Ca and Mg while dead plants lost K and had small gains of Ca and P. Sodium accumulated in dead plant tissue. Death was attributed to an indiscriminate salt injury and/or NH^-N toxicity. The largest reductions of EC, K and P were observed in diluted effluents. The highest plant dry weight gains were also found in these effluents. Potassium and Mg increased in effluents where plant death i was noted.

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DECOMPOSITION OF FRESH AND ANAEROBICALLY DIGESTED PLANT BIOMASS IN SOIL Anaerobic digestion of plant biomass, sewage sludge, or animal wastes generates a waste by-product which must be disposed of, or preferably utilized, in an environmentally-safe manner. Disposal of the digested sludge by land application is one option often considered (Miller, 1974; Terry et al., 1979; Atalay and Blanchar, 1984). The digested biomass sludge differs chemically from the fresh plant biomass. A consequence of anaerobic digestion is a reduction of the more readily decomposable C of the plant tissue during production of CH^ and CO2. Many sludges contained relatively large amounts of Ca, Mg, P and Zn and lower contents of soluble elements such as K (Sommers, 1977) Miller (1974) concluded that anaerobically digested sewage sludge was rather resistant to further decomposition with a maximum of 20% of the added C was evolved as CO2 during a 6-month incubation. Tester et al. (1977) reported 16% of added C from composted sewage sludge was evolved as CO^ during 54 days of incubation. Hsieh et al. (1981a) showed that activated sludge had a much higher C mineralization rate comparjiad to digested sludge due to a larger portion of active organic C. Epstein et al. (1978) determined that the percentage of added N mineralized from digested sludge remained essentially constant irrespective of application rate. However, Ryan et al. (1973) and Stark 95

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96 and Clapp (1980) observed an enhanced rate of N mineralization with increasing rate of sewage sludge application. Nitrogen mineralization potential was found to be 30 and 38% of organic N in activated and digested sludge, respectively, during 60 days of incubation (Hsieh et al. 1981b). Much of the available information dealt with land application of anaerobically digested sewage sludge (Miller, 197A; Terry et al., 1979), and limited data was reported on the decomposition of sludge obtained from the anaerobic digestion of plant biomass (Atalay and Blanchar, 1984). The objective of this study was to evaluate the decomposition and N mineralization rates of anaerobically digested plant biomass added to' soil. Four materials were evaluated: fresh plant biomass with a low or high tissue N content, and their respective anerobically digested residues Materials and Methods Surface (0-15 cm depth) soil samples of a Kendrick fine sand (Arenic paleudult) were collected at the Agronomy Farm, University of Florida in Gainesville, Florida. The soil was air-dried and passed through a 2 mm sieve. The soil had a particle size distribution of 92.9% sand, A. 6% silt, and 2.5% clay. The CEC was 3.44 cmol(+) kg'^ soil with a base saturation of 47%. Water hyacinths with low (-10 g N kg ^ dry plant tissue) and high (-34 g N kg ^) tissue N content were grown in nutrient-depleted water and sewage effluent, respectively. After removal from their respective growth media, the hyacinths were grown in ^^N labeled (NH,)^SO, nutrient 4 2 4 solution for two weeks, frozen, and chopped to 1.6 mm length using a Hobart T 215 food processor.

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97 The water hyacinths were anaerobically digested for four months in 55 L batch digesters. Each digester received 4.7 kg (fresh weight) of the labeled water hyacinth, 5 L of an inoculum from anaerobic digesters receiving water hyacinth as feedstock, and were buffered with 210 g NaHCO^. After digestion, the biomass sludge was separated from the effluent by passing the total contents of the digester through a 1.00 mm fiberglass screen. Samples of the fresh water hyacinth and anaerobically digested water hyacinth sludge were freeze-dried (Thermovac T) and ground through a 0.84 mm screen of a Wiley Mill. The freeze-dried materials were characterized for lignin, cellulose and hemicellulose (Goering and Van Soest, 1970), ashed mineral constituents (Gaines and Mitchell, 1979), total solids (TS), volatile solids (VS), total C (TO) (LECO Induction Furnace 523-300), and total Kjeldahl N (TKN) (Nelson and Sommers, 1973). Carbon/nitrogen (C/N) ratios of the four residues were calculated from percentage TC and TKN. Fifty gram soil samples were preincubated for 5 days at a water content adjusted to 0.01 MPa before addition of the residues. The freeze-dried materials were added to the soil at a rate of 5 g (dry wt) kg ^ soil (10 Mg ha ^) and incubated for 90 days at 27C. Water content was adjusted to 0.01 MPa every 15 days. Ambient laboratory air, with CO. and NH^-N removed by 3 M NaOH and 4 M H^SO^ traps, respectively, was pumped through the incubation flask at a rate of 50 ml min ^. The evolved from soil samples was collected in 0.1 M NaOH traps and determined by titration with acid after reacting with saturated BaCl2. The percentage C evolved with time was calculated by subtracting C evolved as COof the control soil (no organic C amendment) from the

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various treatments and dividing by the amount of C added for each residue. Soil samples were analyzed at 0, 30, 60, and 90 days for 2 M KCl-extractable NH^-N and NO^-N by steam distillation (Keeney and Nelson, 1982), TKN (Nelson and Sommers, 1972), Mehlich I extractable Ca, K, Mg, Na, Fe, Zn, and P (Melich, 1953), organic C by the Walkley-Black method (Nelson and Sommers, 1982), and pH. The ''^^N analyses were conducted on a Micro Mass 602 spectrometer. Results and Discussion Plant Residue Characterization Characteristics of the fresh plant biomass and anaerobically digested biomass sludge are presented in Table 22. The TC and TKN concentrations of the digested sludges were higher than their respective fresh plant materials. Total C was not significantly different for low and high N fresh plant biomass or digested sludge. The C/N ratio of the fresh plant biomass with a low N content (low N plant biomass) decreased from 35 to 16 after digestion. The C/N ratio of fresh plant biomass with a high N content (high N plant biomass) did not change during digestion. Lignin content was significantly higher in digested sludges due to loss of readily-decomposable C during anaerobic digestion. The low N fresh plant biomass contained approximately twice as much lignin as the high N fresh plant biomass. Moore and Bjorndal (1984, Unpublished results, Univ. of Florida, Gainesville) concluded that water hyacinth roots, in general, have higher lignin content than shoots. The lignin content of the low N fresh plant biomass was approximately double that of

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Table 22. Characteristics of the fresh and digested plant biomass Chemical Low N plant biomass High N plant biomass constituent Fresh Digested Fresh Digested -1 kg ^ of biomass-g Volatile solids 839 b^ 866 a 837 b 825 c Ash 161 b 134 c 166 b 173 a Total carbon 373 b 425 a 385 b 446 a Lignin 83 b 145 a 43 c 130 a Cellulose 266 a 180 b 167 b 206 b Hemicellulose 247 a 243 a 183 b 180 b Total nitrogen 10.6 c 26.6 b 34.0 a 39.3 a Calcium 21.0 b 24.2 a 17.6 c 18.4 c Potassium 22.2 a 2.8 b 23.5 a 3.8 b Magnesium 6.7 a 2.2 a 3.2 b 2.2 c Sodium 10.9 c 17.4 b 8.0 d 26.0 a Iron 1.9 c 7.2 a 1.6 c 4.7 b Zinc 0.7 c 2.0 a 0.5 d 0.9 b C/N ratio 35 a 16 b 12 c 12 c Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

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100 the high N fresh biomass due to a larger rooting mass associated with plants grown in nutrient-depleted water. The low N fresh and digested plant biomass contained more hemicellulose than high N materials. Cellulose content decreased for low N fresh plant biomass after digestion, but increased for high N fresh plant biomass. Anaerobic digestion resulted in a large loss of K from fresh plant biomass. Potassium is relatively soluble and is used for translocation of anions via the zylem and phloem, enzyme activation, and stomatal movements. Additional K may be required to enhance the microbial degradation of digested biomass sludges when added to soil. The increase in Na concentration in the digested sludges was due to the addition of NaHCO^ buffer to stabilize digester pH. The digestion process resulted in increases in the relative amounts of Ca, Na, Fe, and Zn in the sludge compared to fresh plant biomass. Carbon and Nitrogen Mineralization Carbon evolution from the four materials, reported as mg C evolved as CO2 per g residue C added, is shown in Fig. 16. The soils with fresh plant biomass additions always released more CO^ compared to that from digested biomass sludges. The two sludges showed similar evolution rates during the first 40 days of incubation despite their difference in TKN. After AO days, the CO2 evolution of high N digested sludge increased and continued to increase for the remaining incubation period. This suggests a possible lag period during which the microbial population is adjusting to those species which tolerate high levels of Na. However, there were no significant differences in CO^ evolution from the two sludges throughout the incubation period (Duncan's multiple range test). Overall C decomposition cannot be described by simple kinetic equations. However, first-order kinetics describe plant residue or

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500 400300o 200residue 100 1 500 400 O 300 E 200 High N Fresh Biomass I I I I 00 300 Low N Fresh Biomass I I I I High N Digested Biomass 0 10 20 30 40 50 60 70 80 90 TIME (days) Figure 16. Carbon evolution from soil applied fresh and digested plant biomass.

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animal waste decomposition if the overall decomposition sequence is presented as occurring in stages (Gilmour et al., 1977; Hunt, 1977; Reddy et al., 1980). Each stage is thought to represent the sequential ease of C constituent decomposition, i.e. soluble sugars and starch, cellulose and hemicellulose, and lignin. The rate equation for a first-order reaction is expressed as -dC,/dt = k.C. i 11 where subscript i refers to a particular stage of C decomposition. The integrated first-order rate equation is Ct. = C.exp(-k.t) 1 1 1 where = C at beginning of a decomposition stage, Ct^ = C remaining at end of a decomposition stage at time = t, = first-order rate constant. Therefore, a rate constant can be calculated for each decomposition stage of an organic C material. A graphical representation of the stages and their respective rate constants of the materials used in this study are shown in Fig. 17. Decomposition of fresh plant biomass required a three stage first-order kinetic description. Rate constants for the first stage (soluble sugars and starch) were 0.0441 and 0.0222 day"^ for high and low N plant biomass, respectively. This stage of decomposition was essentially completed in 4 days. A longer time was required during the second stage (cellulose and hemicellulose) of decomposition for low N plant material due to a higher content of these C constituents (Table 22). The rate constants for the final stage (lignin) of decomposition were low for both materials.

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103 < :e iij cr o 00 90 ;\ M = 0.0441 80 = 0.0119 70 k3 = 0.0046 60 50 High N -1 1 Fresh Biomass — 1 1 1 L 1 1 — 1 1 — ^ 100 90 80 70 60 100 90 80 LlI 70 O cr LU 100 CL 90 k2 = 0.0137 Low N Digested Biomass k3 = 0.0018 = 0.0222 k2 = 0.0057 k3 = 0.0035 Low N Fresh Biomass • t ko = 0.0097 k3 = 0.0026 High 1 N Digested Biomass 1 1 1 1 J 1 1 0 10 20 30 40 50 60 70 80 90 TIME (days) Figure 17. Decomposition stages and rate constants of fresh and digested plant biomass added to soil

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104 Decomposition of digested biomass sludges was adequately described in two stages. The conversion of soluble sugars and starch to CH^ and during anaerobic digestion eliminated the first stage of soluble C degradation for the digested sludges. Therefore, the first stage of sludge decomposition corresponded to the second stage (cellulose and hemicellulose) of fresh plant biomass. The longer second stage corresponded to the final stage (lignin) of fresh plant biomass. Soil NO^-N concentrations during incubation are presented in Table 23. Soil NH^-N concentrations (APPENDIX B, Table 29) were < 2 mg kg'^ during the incubation period. Initial NO^-N concentrations were similar for all treatments. No NO^-N accumulated in the soil amended with low N fresh plant biomass until 60 days had elapsed. The concentrations of soil NO^-N were similar for both digested biomass sludges throughout the incubation despite the higher TKN concentration of the high N sludge. Although the C/N ratios of high N fresh biomass or digested biomass sludge were the same, the amount of NO^-N that accumulated in the soil was more than double for fresh biomass compared to digested biomass sludge. A summary of C and ^^N mineralization during the incubation period is presented in Table 2A. Approximately 8% of the applied N was recovered as ^^NO^-N at 90 days for both sludges. Tester et al. (1977) reported 6% of composted sewage sludge N mineralized to NO^-N in 54 days. In contrast, 3 and 33% of applied N was recovered as ^^NO~-N for fresh plant biomass with a low and high N content, respectively. Ogwada (1983) incorporated shredded ^^N labeled water hyacinth with a C/N ratio of 35 into a Myakka fine sand at a rate of 20 Mg dry wt ha"'''. At 60 days, 33% of ^^N from the soilincorporated water hyacinth was

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105 Table 23. Soil NO^-N concentration from added fresh and digested plant biomass. Low N plant biomass HiRh N plant biomass Day Control Fresh Digested Fresh Digested jng i^g ^ soil 0 9.1 a-t 8.2 ab 7.9 abc 8.4 ab 6.6 c 30 11.0 c 0.9 d 20.0 b 51.4 a 17.2 b 60 13.2 c 1.0 d 24. A b 70.3 a 24.8 b 90 1A.5 c A. 8 d 28. A b 68.8 a 30.3 b ^Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test. I

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106 Table 24. Carbon and ^^N mineralization from added fresh and digested plant biomass. Low N plant biomass High N plant biomass Fresh Digested Fresh Digested % of added C or N Day 30 C 22.4 b t 11.2 c 34. 2 a 9.8c ^^N ND 4.7 b 24.8 a 3.8 b Day 60 C 32.5 b 14.9 c 41.6 a 16.1 c ^^N ND 6.2 b 34.3 a 6.0 b Day 90 C 39.0 b 19.1 c 49.9 a 23.1 c ^^N 3.3 c 7.7 b 33.3 a 7.7 b Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test. ND = not detectable.

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107 mineralized and 20% of the initial ^^N was recovered in sorhgumsudangrass plants. After 90 days of incubation, approximately 20% of the added C of digested biomass sludges had evolved as compared to 39 and 50% of fresh plant biomass with a low and high N content, respectively. More than half of the C evolved from fresh plant biomass occurred within the first 10 days. The percentage of C evolved with time from anaerobically digested biomass sludge is similar to results of Miller (1974) and Tester et al. (1977) for anaerobically digested and composted sewage sludge. Miller (1974) reported a maximum of 20% of the added C was evolved as during a 6-month incubation period. Tester et al. (1977) reported that 16% of the added C was evolved as CO^ from composted sewage sludge during 54 days of incubation. However, Terry et al. (1979) found a total of 26 to 42% of C was evolved as from anaerobically digested sewage sludge during 130 days of incubation. Carbon/nitrogen ratio is commonly used as a guideline for predicting the relative decomposability or mineralization potential of organic materials added to soil (Reddy et al., 1980). The high C/N ratio of low N fresh plant biomass (C/N = 35) resulted in immobilization of inorganic N. The low C/N ratio of high N fresh plant biomass (C/N = 12) resulted in rapid mineralization of organic N. However, C/N ratios may have limited applicability for predicting N transformations of digested biomass sludges. Both high N fresh biomass and digested biomass sludge had C/N ratios of 12, but only 8% of the applied N was recovered as ^^NO^-N from digested sludge compared to 33% from fresh plant biomass.

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108 Other Soil Parameters Soil pH increased by one and two pH units after addition of anaerobically digested biomass sludge with a low and high N content, respectively (Table 25). The addition of fresh plant biomass did not appreciately alter the initial soil pH. During the 90 day incubation, soil pH generally decreased with all residue additions, except for low N fresh plant biomass. The Na content of the digested biomass sludges explains, in part, the increase in the initial soil pH. Atalay and Blanchar (1984) found that addition of anaerobically digested plant residue to soil increased pH from 5.5 to 7.6. They attributed the pH increase to a limestone buffer used during anaerobic digestion. The addition of fresh or anaerobically-digested water hyacinth increased Mehlich I extractable soil constituents after 90 days of incubation (Table 26). Mehlich I extractable soil constituents at 0, 30 and 60 days of incubation are presented in APPENDIX B, Tables 30 to 32. The increases were a direct reflection of the mineral composition of the respective residue (Table 22). There was a large increase in soil K with fresh plant biomass additions compared to digested biomass sludges. All treatments resulted in large increases of soil Na and Ca. Parra and Hortenstine (1976) concluded that fresh water hyacinths contained appreciable amounts of soluble salts and that crops susceptible to salt injury should not be planted immediately after soil additions.

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Table 25. Soil pH (1:2 w/v) from added fresh and digested plant biomass. Low N plant biomass High N plant biomass Day Control Fresh Digested Fresh Digested 0 5.4A d + 5.38 d 6.49 b 5.73 c 7.34 a 30 5.48 c 5.99 b 6.27 a 4.90 d 6.21 a 60 4.99 c 5.96 ab 6.03 a 4.30 d 5.77 b 90 5.42 b 5.94 a 6.03 a 4.72 c 6.02 a Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

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110 Table 26. Mehlich I extractable constituents at Day 90 from added fresh and digested plant biomass. Chemical constituent Control Low N plant biomass High N plant biomass Fresh Digested Fresh Digested Calcivun Potassivim Magnesium Sodium Iron Zinc 252 e 25 e 24 d 3 e lA d 3.3 e 361 ab 115 b 59 a 53 c 14 d 5.7 d -mg kg ^ soil373 a 40 d 35 c 72 b 20 a 11.3 a 327 d 119 a 39 b 41 d 15 c 6.0 c 348 be 47 c 35 c 117 a 18 b 7.2 b Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

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Ill Conclusions After 90 days of incubation, approximately 20% of the added C of the digested biomass sludges had evolved as CO^ compared to 39 and 50% of the fresh plant biomass with a low and high N content, respectively. Decomposition of fresh plant biomas followed a three stage first-order kinetic description. Decomposition of digested sludge was adequately described by two stage first-order kinetics. Mineralization of organic N to "^^NO^-N accounted for approximately 8% of applied N for both digested biomass sludges at the end of 90 days. Nitrogen mineralization accounted for 3 and 33% of applied N for fresh plant biomass with a low and high N content, respectively. The soil pH increased after addition of digested biomass sludge, but was not appreciately altered after addition of fresh plant biomass. The Na content of digested sludges was attributed as the primary factor for increasing soil pH. Increases in Mehlich I extractable soil constituents were a direct reflection of the mineral composition of fresh or digested plant biomass. The high Na concentration of digested biomass sludge suggests pretreating the sludge to remove some of the salts and selecting salt tolerant plants if the sludge is used as a soil amendment.

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MASS BALANCE OF NITROGEN IN AN INTEGRATED "BIOMASS FOR ENERGY SYSTEM A series of experiments were conducted to evaluate N cycling in three components of an integrated "biomass for energy" system, i.e. water hyacinth production, anaerobic digestion of hyacinth biomass, and recycling of digester effluent and sludge. Two types of water hyacinth biomass production systems were evaluated; 1) plants growing in nutrient-enriched waters representing wastewater treatment systems (sewage and digester effluents, animal wastes or agricultural drainage water), and plants growing in nutrient-limited systems representing natural waters receiving low external N inputs. Nutrient -Enriched Systems Nitrogen cycling in an integrated system for water hyacinths growing in nutrient -enriched systems is summarized in Fig. 18. Water Hyacinth Production -2 -1 A wide range of productivity (5 to 6A g dry wt m day ) has been recorded for plants growing in nutripnt-enriched waters (Boyd, 1976; Hanisak et al., 1980; Reddy and DeBusk, 1983; Reddy et al., 1985). Plants growing in nutrient-enriched media generally have an N content of 30 to AO g N kg '^ of dry tissue (Boyd, 1976; Wolverton and McDonald, 1979; Reddy et al., 1985). In this study the tissue N content of plants growing in sewage effluent was 35 g N kg of dry biomass. 112

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113 O o F CO o n_ o o IjlJ Q 00 CVJ 11:0 p o CD CO ro C£> CD ^ CD ro I 'J? p o o Z o> CD • • rO — OJ — 7I\ X o CVJ o o LU O Q ZD _J CO CO CO < o CVJ o "c CD cn UJ o o o ro cn cn CO •43 c: >> cn J*: c: cn OJ in ro CVJ w o CD _v/ CD ro V-/ o. o> rO CO 0) c 60 •ri CO 15 0 c J-i 60 rc CO P. 4-J (U CO cd IJ to Q) 0) 4J 03 CO 01 01 (U .c 60 4-) •H CO c -o IS 0) Vj 0 J-l CO •H 0 P. ^ M-1 0 c U E (U OJ cfl 4-> cn C CO CO OJ to 0) e ti 3 z i-> c •H 60 cn (U 6
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114 Although plant assimilation is a major process for N removal in ponds of water hyacinths, other N transformations may occur in the root zone, water column and underlying sediment. These processes accoxint for much of the loss of N from the system. For example, a dense mat of water hyacinth could potentially enhance denitrif ication of NO^-N present in the water column (Boyd, 1970; Reddy, 1981). Total N reduction in water hyacinth systems was reported in a range from 65 to 94% of various wastewaters (Sheffield, 1967; Clock, 1968; Hanisak et al., 1980; Reddy et al., 1982). Total fertilizer N recovered by plants in this study ranged from 50% in a field study to 90% in controlled-greenhouse studies. Results indicated that detritus accounted from 3 to 14% of the total biomass produced in water hyacinth ponds. Up to 28% of the total plant N was found in detritus, which could potentially be deposited at the sedimentwater interface. Anaerobic Digestion of Hyacinth Biomass Anaerobic digestion of biomass with high tissue N and low C/N ratio resulted in mineralization of 70% of plant organic N to NH^-N. Total N recovered for digested high N plants was -100% of the added N which agreed with results by Hashimoto et al. (1980) and Field et al. (1984). After digestion, 80% of the plant organic N was recovered in the effluent and 20% was recovered in the sludge. Waste By-Product Recycling Undiluted and diluted effluents were used as nutrient sources to produce additional water hyacinth biomass and for potential N recovery. Plants growing in undiluted effluents did not survive the combination of high salt and NH^-N concentrations. However, diluting the digester effluents provided a media which stimulated water hyacinth growth.

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115 Plant assimilation of effluent N recycled 38% of the initial N placed in the digester. Nitrogen loss from this component of the integrated system accounted for A2% of the N placed in the digester. Possible N loss mechanisms during recycling of digester effluent include algal assimilation and NH^-N volatilization. The digested sludge contained 20% of the initial N placed in the digester. The sludge N could potentially be recovered during crop production after land application. The sludge was fairly resistant to decomposition in soil and only 20% of the sludge C was evolved as CO^ in 90 days of incubation. A low decomposition rate of sludge applied to soil was attributed to loss of the more readily-decomposable C constituents during anaerobic digestion. The decomposition rates observed in this study for digested biomass sludge applied to soil were found to be similar to those reported for digested sewage sludge (Miller, 197A; Tester et al., 1977). Nitrogen mineralization of sludge organic N during 90 days of decomposition in soil accounted for only 2% of the initial N placed in the digester. Fresh water hyacinth biomass was also added to soil to compare its decomposition to that of digested sludge. Approximately 50% of the biomass C was evolved as during 90 days of incubation. Nitrogen mineralization of biomass organic N accounted for 34% of the initial plant N placed in the soil. Mixing the fresh water hyacinth biomass with anaerobically digested sludge may enhance the decomposition of the sludge. Total N recovery by sludge and effluent recycling in the integrated "biomass for energy" system was 60% of the initial N placed in the digester for high N plants. The remaining 40% was lost from the system during effluent recycling in a water hyacinth production system.

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116 Nutrient-Limited Systems Nitrogen cycling in an integrated system for water hyacinths growing in nutrientlimited systems is summarized in Fig. 19. Water Hyacinth Production -2 -1 Growth rates of 2 to 29 g dry wt m day have been reported for plants in natural waters of central and south Florida (Yount and Grossman, 1970; DeBusk et al., 1981). Plants growing in nutrientlimited systems generally have a low N content. In this study, tissue N content of plants grown in tap water without nutrients was 10 g N kg ^ of dry biomass. In addition to a low N content, the shoot: root dry weight ratio decreased as nutrient availability decreased in the water media (Reddy, 1984). The shoot: root dry weight ratio of plants with a low N content was 1.6 compared to 4.2 of plants with a high N content. Anaerobic Digestion of Plant Biomass Shiralipour and Smith (1984) concluded that increasing N in the growth medi\im used for water hyacinth production increased methane yields during anaerobic digestion. Surprisingly, gas yields were higher for plants with a low N content compared to plants with a high N content. However, the gas yield differences were attributed to characteristics of inoculum used during digestion. Only 75% of the initial N placed in the digester was recovered in the effluent and sludge for low N plants. The low total N recovery is difficult to explain. Only 35% of the plant organic N placed in the digester was mineralized to NH^-N compared to 70% mineralization for high N plants. Approximately 40% of the initial N placed in the digester remained as organic N after digestion. After digestion, 54% of the initial plant organic N was recovered in the effluent while 20% was recovered in the digested sludge.

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117 O CO CsJ ^ O o Z C7) 0> 00 CVJ d o w o CL o o Q ro o I 2 O o z X o C\J o o 0> in ro ro z: o z •e 1 u "c o -i • to CB l-i (U x: CO 4J (U CO )-i CO 0) ^ CO CD 2 c tJ 60 •H OJ 4-1 O 00 •H r-l c •H tH rH CO o 1 •H 4-1 CJ C •H (U C c •H •H (U bO 4J (U O 3 I-i C 4J 4-1 •H C z •H o 3 60

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Waste By-Product Recycling The undiluted effluent from digested plants with a low N content was recycled directly to a water hyacinth production system. Plants survived in undiluted effluents due to lower NH^-N concentrations. Approximately 21% of the N placed in the digester was reassimilated by water hyacinths during effluent recycling. Nitrogen loss during digestion and effluent recycling accounted for 52% of the initial N placed in the digester. The digested sludge contained 20% of the initial N placed in the digester. Similar decomposition rates were noted for both digested sludges applied to soil despite their differences of initial N content (27 and 39 g N kg"''^ dry sludge). The mineralization of sludge organic N accounted for 2% of the initial N placed in the digester. The fresh water hyacinth biomass decomposed rapidly in soil but only 3% of the applied N was mineralized to NO^-N. Total N recovery by sludge and effluent recycling was 48% of the initial N placed in the digester for low N plants. The remaining N was lost from the system during anaerobic digestion and effluent recycling.

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CONCLUSIONS The three components of the integrated "biomass for energy" system were 1) the water hyacinth biomass production system; 2) anaerobic digestion of water hyacinth biomass, and 3) waste recycling of digested biomass sludge and effluent. Nitrogen cycling was investigated for each component of the integrated system. Water Hyacinth Productivity and Detritus Production Productivity of water hyacinth was influenced by ambient air temperature, solar radiation, and nutrient compostion of the medium. The highest net productivity occurred during spring and summer and over 75% of the biomass produced was recorded during this time period. The detritus production exceeded net biomass production during winter regardless of water fertility. Seasonal yields of water hyacinth ranged from 1.9 to 23.1 Mg (dry wt) ha ^ and -0.2 to 10.2 Mg ha ^ for plants growing in eutrophic lake water with and without added nutrient, respectively. Detritus comprised 3 to 14% of the total biomass and detritus production was not significant between reservoirs or seasons. Although detritus production was similar for both reservoirs, fertilization resulted in significant increases in detritus N content. Nitrogen loading to the reservoirs from detritus was 148 and 92 kg N ha ^ yr ^ for plants grown in eutrophic lake water with and without added nutrients, respectively. 119

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120 Approximately 50% of the fertilizer N was recovered by plants. Total N assimilated by water hyacinth (live plants and detritus) was 720 and 325 kg N ha ^ yr ^ for plants grown in eutrophic lake water with and without added nutrients, respectively. Detritus and Nitrogen Transf romations Detritus had no apparent effect on rate of N loss in water with water hyacinth plant cover due to rapid plant assimilation. However, N loss in water without plant cover was more rapid with detritus additions. Both sediment and detritus appeared to be potential N sources for plant assimilation. Total N recovered by water hyacinth ranged from 57 to 72% for added ^\o~-N and 70 to 89% for added ^^NH^-N. Less than 10% of added ^^NH"!"-N 3 A 4 was immobilized by detritus in water with plant cover. However, up to 35% of the added ^^NH^-N was associated with detritus in water without A plant cover. This suggests that during periods of low water hyacinth productivity, i.e. winter, detritus is an important sink for added N. Increasing amounts of detritus generally decreased dissolved 2 concentrations of water. The pH of water without plant cover generally was lower when detritus was added. Anaerobic Digestion of Water Hyacinth Initial water hyacinth N content and volume of inoculum did not affect long term (60 days) biogas production. At 15 days, biogas production was generally greater for plants with a high N content. Inoculum volume showed little effect of biogas production for low N plants throughout incubation. However, a larger volvime of inoculum

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121 increased biogas production for plants with a high N content after 15 days of digestion. Removing the biomass sludge from digester effluents decreased the electrical conductivity, NH^-N and TKN of the effluent. Anaerobic digestion resulted in a loss of K and Mg from fresh plant biomass but increased lignin, total C, TKN, Ca, Na, Fe and Zn of the digested sludge. Generally cellulose decreased during digestion and hemicellulose remained unchanged. Mineralization of organic N to NH^-N was the primary N transformation occurring during anaerobic digestion. Approximately 20% of the organic N placed in the digesters was recovered as sludge organic N. Net mineralization of organic N to NH^-N was 70 and 35% of the added organic N of plants with a high and low N content, respectively. Digester Effluent Recyling The initial electrical conductivities and NH^-N concentrations of the digester effluents ranged from 0.7 to 6.7 dS m ^ and 23 to 289 mg N L ^. The highest water hyacinth dry weight gains were associated with the lowest electrical conductivities. However, plants survived in effluents having electrical conductivities of 5.6 and 5.9 dS m ^. Plant death was observed in four undiluted effluents. Additional information would be needed to establish optimum dilution of anaerobic digester effluents to promote maximum biomass yields. First-order kinetic equations were used to describe NH^-N loss from effluent with k values ranging from 0.175 to 0.593 day ^. Generally the highest k values were associated with rapid plant growth. Plant ^^N assimilation was observed for all effluents although plant death

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122 resulted in low recoveries (2 to 16%). The N recovery for other effluents ranged from 36 to 77%. The majority of the was recovered in shoot material. Plants grown in diluted effluents assimilated large amounts of Na and K and net shoot assimilation of all nutrients was greater than net root assimilation. Dead plants showed a net loss of K but net gains of other nutrients. Digested Sludge Recycling Decomposition of fresh and anaerobically digested water hyacinth biomass added to soil was evaluated by CO^ evolution and ''^^N mineralization. Approximately 39 and 50% of the added C was evolved as in 90 days for fresh plant biomass with a low and high tissue N content, respectively. Approximately 20% of the added C was evolved as for both digested sludges in 90 days. Decomposition of fresh plant biomass required a three stage first-order kinetic description of C loss compared to a two stage first-order description for digested sludge. Apparently the loss of readily-decomposable C during anaerobic digestion eliminated the first stage of decomposition in soil. Only 8% of the applied ^^N was mineralized to NO^-N for digested sludges despite their differences of initial N content. In contrast, 3 and 33% of the applied ''^^N was recovered as NO^-N for fresh biomass with a low and high N content, respectively. The C/N ratio of digested sludge was not a predictive guide for N mineralization potential. Mixing the fresh biomass with digested sludge may enhance the decomposition of the sludge in soil.

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APPENDIX A DIGESTER EFFLUENT CHARACTERISTICS DURING WATER HYACINTH TREATMENT

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cN in lo r-~ m 00 CX5 00 00 C!0 00 ^ < • • z 00 0^ IT) o in in r>. I — m o 00 00 0\ 00 CO cs < • • Z oo o\ in 00 n < vo r>. tN 00 in 00 00 00 (N < • • z 00 CTi a, c o u c 0) 3 13 0) 3 vO 00 •<} — 00 w p c r-{ o z (U m 00 m 00 00 oo o m o 00 00 00 o m 00 00 00 <3vD <• 00 00 00 m (N 00 00 00 <• CNI (N 00 oo oo m (Ni cN 00 00 00 tn P c (U m V4-I Q) xi (U 00 T-H rn 00 o\ CJ\ 00 r4 m 00 C7\ C7 < < < z z z 3vo 00 r-~ vo vo vo r-. I — in in in 0) 3 I O i-H O O c > I I I in in • in o CM ^ • m o CM 124

PAGE 135

125 1 ro 00 ro 1—4 CN 1 [ (N r-> ro rsi 1—1 c^ o >300 lO ON ON CO ro ro ro ro 1— 1 NO CO 00 LO ON lO ro o 1 X) Q) e c •H 4-1 3 OV VO o VO < z < z o IT) m VO ov VO

PAGE 136

APPENDIX B SOIL CHARACTERISTICS FROM ADDED FRESH AND ANAEROBICALLY DIGESTED PLANT BIOMASS

PAGE 137

Table 29. Soil anraionium concentrations from added fresh and digested plant biomass. Low N plant biomass High N plant biomass Day. Control Fresh Digested Fresh Digested 8 kg 0 1.2 bt 2.2 b 1.7 b 25.1 a 1.9 b 30 1.2 a 1.4 a 1.8 a 1.8 a 1.3 a 60 1.1 a 1.2 a 1.7 a 1.0 a 1.0 a 90 1.0 a 1.0 a 1.0 a 1.0 a 1.0 d ^Values with same letter within rows are not si gnificantly different 0.05 level by Duncan's Multiple Range Test. 127

PAGE 138

128 Table 30. Mehlich I extractable constituents at day 0 from added fresh and digested plant biomass. Chemical Low N plant biomass High N plant biomass constituent Control Fresh Digested Fresh Digested mg kg Calcium 257 c"^ 377 a 344 ab 332 b 351 ab Potassium 27 d 128 a 44 c 127 a 53 b Magnesivim 25 d 55 a 34 c 37 b 33 c Sodium 7 e 60 c 79 b 56 d 133 a Iron 18 d 25 b 35 a 21 c 26 b Zinc 4.1 d 7.3 b 13.3 a 6.1 c 7.2 b ^Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

PAGE 139

129 Table 31. Mehlich I extractable constituents at day 30 from added fresh and digested plant biomass. Chemical Low N plant biomass High N plant biomass constituent Control Fresh Digested Fresh Digested mg kg Calcium 245 ct 396 a 383 a 340 b 379 a Potassium 39 c 139 a 55 c 124 b 49 c Magnesium 26 c 68 a 39 b 39 b 34 be Sodium 3 e 57 c 75 b 41 d 114 a Iron 17 c 18 b 25 a 15 d 18 b Zinc 4. 1 d 6.1 c 11 9 a 6 1 c 7.7 Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

PAGE 140

130 Table 32. Mehlich I extractable constituents at day 60 from added fresh and digested plant biomass. Chemical Low N plant biomass High N plant biomass constituent Control Fresh Digested Fresh Digested mg kg Calcium 276 c 359 b 393 a 343 b 357 b Potassium 27 d 113 b 44 c 123 a 48 c Magnesium 27 d 56 a 39 b 40 b 34 c Sodium 4 e 55 c 74 b 38 d 119 a Iron 16 c 16 c 23 a 16 c 18 b Zinc A. ,5 d 6. ,5 be 12. ,8 a 6. ,0 c 7. ,3 Values with same letter within rows are not significantly different at 0.05 level by Duncan's Multiple Range Test.

PAGE 141

BIBLIOGRAPHY Alexander, M. 1977. Introduction to Soil Microbiology. 2nd Edition. John Wiley and Sons. New York, NY. A.P.H.A. 1980. Standard Methods for the Examination of Water and Wastewater. 20th Edition. Amer. Publ. Health Assoc. Washington, DC. Almazan, G. and C. E. Boyd. 1978. Effects of nitrogen level on rates of 0^ consumption during decay of aquatic plants. Aquat. Bot. 5:119-126. Atalay, A., and R. W. Blanchar. 1984. Evaluation of methane generator sludge as a soil amendment. J. Environ. Qual. 13:341-344. Berg, B., B. Wessen, and G. Ekbohm. 1982. Nitrogen level and decomposition in Scots pine needle litter. Oikos 38:291-296. Bock, J. H. 1969. Productivity of the water hyacinth Eichhornia crassipes (Mart.) Solms. Ecol. 50:460-464. Bouldin, D. R., R. L. Johnson, C. Burda, and C. W. Kao. 1974. Losses of inorganic nitrogen from aquatic systems. J. Environ. Qual. 3:107-114. Boyd, C. E. 1970a. Vascular aquatic plants for mineral nutrient removal from polluted waters. Econ. Bot. 24:95-103. Boyd, C. E. 1970b. Losses of mineral nutrients during decomposition of Typha latifolia Arch. Hydrobiol. 66:511-517. Boyd, C. E. 1976. Accumulation of dry matter, nitrogen, and phosphorus by cultivated water hyacinths. Econ. Bot. 30:51-56. Broadbent, F. E., and W. V. Bartholomew. 1948. The effect of quantity i of plant material added to soil on its rate of decomposition. Soil Sci. Soc. Am. Proc. 13:271-274. Bryant, M. P. 1979. Microbial methane production: Theoretical aspects. J. Animal Sci. 48:193-201. Carpenter, S. R. 1980. Enrichment of Lake Wingra, Wisconsin, by submersed macrophyte decay. Ecol. 61:1145-1155. Carpenter, S. R., and M. S. Adams. 1979. Effects of nutrients and temperature on decomposition of Myriophyllum spicatum L. in a hard-water eutrophic lake. Limnol. Oceanogr. 24:520-528. 131

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132 Chynoweth, D. P., D. A. Dolene, B. Schwegler, and K. R. Reddy. 1983. Wastewater reclamation and methane production using water hyacinth and anaerobic digestion. Presented at 10th Energy Technology Conference. Washington, DC. Clock, R. M. 1968. Removal of nitrogen and phosphorus from secondary sewage treatment effluent. Ph.D. dissertation. Univ. of Florida. Cooley, T. N. D. F. Martin, W. C. Dunden, Jr., and B. D. Perkins. 1979. A preliminary study of metal distribution in three water hyacinth biotypes. Water Res. 13:343-348. Cornwell, D. A., J. Zoltek, Jr., C. D. Patrinely, T. deS. Furman, and J. I. Kim. 1977. Nutrient removal by water hyacinth. J. Water Poll. Contr. Fed. 49:57-65. Davis, C. B., and A. G. van der Valk. 1978. The decomposition of standing and fallen litter of Typha glauca and Scirpus f luviatilis Can. J. Bot. 56:662-675. DeBusk, T. A., J. H. Ryther, M. D. Hanisak, and L. D. Williams. 1981. Effects of seasonality and plant density on the productivity of some freshwater macrophytes. J. Environ. Qual. 10:133-142. DeBusk, T. A., L. D. Williams, and J. H. Ryther. 1983. Removal of nitrogen and phosphorus from wastewater in a water hyacinth-based treatment system. J. Environ. Qual. 12:257-262. De La Cruz, A. A., and B. C. Gabriel. 1974. Caloric, elemental, and nutritive changes in decomposing Juncus roemerianus leaves. Ecol. 55:882-886. Dinges, R. 1978. Upgrading stabilization pond effluent by water hyacinth culture. J. Water Poll. Contr. Fed. 50:833-845. Dunigan, E. P., R. A. Phelan, and Z. H. Shamsuddin. 1975. Use of water hyacinths to remove N and P from eutrophic waters. Hyacinth Contr. J. 13:59-61. Engler, R. M. and W. H. Patrick, Jr. 1974. Nitrate removal from floodwater overlying flooded soils and sediments. J. Environ. Qual. 3:409-413. Epstein, E. J. M. Taylor, and R. L. Chaney. 1976. Effects of sewage sludge and sludge compost applied to soil on some soil physical and chemical properties. J. Environ. Qual. 5:422-426. Epstein, E. D. B. Keane, J. J. Meisinger, and J. 0. Legg. 1978. Mineralization of nitrogen from sewage sludge and sludge compost. J. Environ. Qual. 7:217-221. Fenchel, T. 1970. Studies on the decomposition of organic detritus derived from the turtle grass Thalassia testudinum Limnol. Oceanogr. 15:14-20.

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133 Fenchel, T. M., and B. B. Jorgensen. 1977. Detritus food chains of aquatic ecosystems: The role of bacteria. In Alexander, M. (ed.). Advances in Microbial Ecology. Volume. 1. Plenum Press, New York. Field, J. A., J. S. Caldwell, S. Jeyanayagam, R. B. Reneau, Jr., W. Kroontje, and E. R. Collins, Jr. 1984. Fertilizer recovery from anaerobic digesters. Transactions of the ASAE 27:1871-1881. Franco, A. A., and D. N. Munns. 1982. Plant assimilation and nitrogen cycling. Plant Soil 67:1-13. Freney, J. R. J. R. Simpson, and 0. T. Denmead. 1983. Volatilization of ammonia. In Freney, J. R. and J. R. Simpson (eds.). Gaseous Loss of Nitrogen From Plant-Soil Systems. Developments in Plant and Soil Sciences. Vol. 9. Martinus Nijhoff/Dr. W. Junk Publishers, The Hague. Gaines, T. P., and G. A. Mitchell. 1979. Chemical Methods for Soil and Plant Analysis. Agron. Handbook No. 1. Univ. of Georgia. Coastal Plains Experiment Station. Gambrell, R. P., and W. H. Patrick, Jr. 1978. Chemical and microbiological properties of anaerobic soils and sediments. In Hook, D. D., and R. M. M. Crawford (eds.). Plant Life in Anaerobic Environments. Ann Arbor Science Publishers, MI. Gilmour, C. M. E. F. Broadbent, and S. M. Beck. 1977. Effects of waste applications on soil carbon and nitrogen cycles. In Elliot, L. F., and F. J. Stevenson (eds.). Soils for Management of Organic Waste and Waste Waters. Soil Sci. Soc. Am. Madison, WI. Godshalk, G. L., and R. G. Wetzel. 1978a. Decomposition of aquatic angiosperms. I. Dissolved components. Aquat. Bot. 5:281-300. Godshalk, G. L., and R. G. Wetzel. 1978b. Decomposition of aquatic angiosperms. II. Particulate components. Aquat. Bot. 5:301-327. Gosselink, J. G., and C. J. Kirby. 197A. Decomposition of salt marsh grass, Spartina alternif lora Loisel. Limnol. Oceanogr. 19:825-831. Goering, H. K. and P. J. Van Soest. 1970. Forage Fiber Analyses. (Apparatus, Reagents, Procedures, and Some Applications). ARS-USDA. Agriculture Handbook No. 379. Washington, DC. Haller, W. T. and D. L. Sutton. 1973. Effect of pH and high P concentration on growth of water hyacinths. Hyacinth Contr. J. 11:59-61. Haller, W. T. D. L. Sutton, and W. C. Barlowe. 1974. Effects of salinity on growth of several aquatic macrophytes. Ecol. 55:891-894.

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134 Hanisak, M. D., L. D. Williams, and J. H. Ryther. 1980. Recycling the nutrients in residues from methane digesters of aquatic macrophytes for new biomass production. Resource Recovery Conser. 4:313-323. Hargrave, B. T. 1972. Aerobic decomposition of sediment and detritus as a function of particle surface area and organic content. Limnol. Oceanogr. 17:583-596. Harrison, P. G. and K. H. Mann. 1975. Detritus formation from eelgrass ( Zostera marina L.): The relative effects of fragmentation, leaching, and decay. Limnol. Oceanogr. 20:924-934. Hashimoto, A. G. Y. R. Chen, V. H. Varel, and R. L. Prior. 1980. Anaerobic fermentation of agricultural residues. In Shuler, M. Utilization and Recycle of Agricultural Wastes and Residues. ORG Press, Inc., Boca Raton, FL. Hill, B. H. 1979. Uptake and release of nutrients by aquatic macrophytes. Aquat. Bot. 7:87-93. House, D. 1981. The Biogas Handbook. Peace Press, Inc., San Francisco, CA. Howard-Williams, C., S. Pickmere, and J. Davies. 1983. Decay rates and nitrogen dynamics of decomposing watercress ( Nasturtium officinale R.Br.). Hydrobiol. 99:207-214. Howeler, R. H. 1972. The oxygen status of lake sediments. J. Environ. Qual. 1:366-371. Hsieh, Y. P., L. A. Douglas, and H. L. Motto. 1981a. Modeling sewage sludge decomposition in soil: I. Organic carbon transformations. J. Environ. Qual. 10:54-59. Hsieh, Y. P., L. A. Douglas, and H. L. Motto. 1981b. Modeling sewage sludge decomposition in soil: II. Nitrogen transformations. J. Environ. Qual. 10:59-64. Hughes, W. L. (ed.). 1981. Supplement Energy for Rural Development. Renewable Resources and Alternative Technologies for Developing Gountries. Natl. Acad. Press, Washington, DC. Hunt, H. W. 1977. A simulation model for decomposition in grasslands. Ecol. 58:469-484. Hunter, R. D. 1976. Changes in carbon and nitrogen content during decomposition of three macrophytes in freshwater and marine environments. Hydrobiol. 51:119-128. Jenkinson, D. S. 1965. Studies on the decomposition of plant material in soils. I. Losses of carbon from G labelled ryegrass incubated with soil in the field. J. Soil Sci. 16:104-115.

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135 Jenkinson, D. S. 1971. Studies on the decomposition of C labelled organic matter in soil. Soil Sci. 111:64-70. Keeney, D. R. 1973. The nitrogen cycle in sediment -water, systems. J. Environ. Qual. 2:15-29. Keeney, D. R., and D. W. Nelson. 1982. Nitrogen-Inorganic Forms. In Page, A. L. (ed.). Methods of Soil Analysis. Part 2. Second Edition. Agronomy 9. ASA, Madison, WI. Kirkby, E. A., and K. Mengel. 1967. The ionic balance in different tissues of the tomato plant in relation to nitrate, urea, or ammonium nutrition. Plant Physiol. A2:6-14. Knipling, E. B. S. H. West, and W. T. Haller. 1970. Growth characteristics, yield potential, and nutritive content of water hyacinths. Soil Crop Sci. Soc. Fla. 30:51-63. Mah, R. A., D. M. Ward, L. Baresi, and T. L. Glass. 1977. Biogenesis of methane. Ann. Rev. Microbiol. 31:309-341. Hanson, J. G. and B. E. Manson. 1958. Water hyacinth reproduces by seed in New Zealand. New Zealand J. Agric. 96:191. McDonald, R. C, and B. C. Wolverton. 1980. Comparative study of wastewater lagoon with and without water hyacinth. Econ. Bot. 34:101-110. Mehlich, A. 1953. Determination of P, Ca, Mg, K, Na, and NH^. North Carolina Soil Test Division (mimeo. 1953), Raleigh, NC. Melillo, J. M., J. D. Aber, and J. F. Muratore. 1982. Nitrogen and lignin control of hardwood leaf litter decomposition dynamics. Ecol. 63:621-626. Mengel, K. 1974. Plant ionic status. In Carson, E. W. (ed.). The Plant Root and its Environment. Univ. Press of Virginia, Charlottesville, VA. Miller, R. D., and D. D. Johnson. 1964. The effect of soil moisture tension on carbon dioxide evolution, nitrification, and nitrogen mineralization. Soil Sci. Soc. Am, Proc. 28:644-647. Miller, R. H. 1974. Factors affecting the decomposition of an anaerobically digested sewage sludge in soil. J. Environ. Qual. 3:376-380. Moore, J., and K. Bjorndal. 1984. Unpublished results. Univ. of Florida, Gainesville, FL. Nelson, D. W. 1982. Gaseous losses of nitrogen other than through denitrif ication. In Stevenson, F. J. (ed.). Nitrogen in Agricultural Soils. Agronomy 22. ASA, Madison, WI.

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136 Nelson, D. W. and L. E. Sommers. 1972. A simple digestion procedure for estimation of total nitrogen in soils and sediments. J. Environ. Qual. 1:A23-A25. Nelson, D. W. and L. E. Sommers. 1973. Determination of total nitrogen in plant materials. Agron. J. 65:109-112. Nelson, D. W. and L. E. Sommers. 1975. Determination of total nitrogen in natural waters. J. Environ. Qual. 4:465-468. Nelson, D. W., and L. E. Sommers. 1982. Total carbon, organic carbon, and organic matter.' In Page, A. L. (ed.). Methods of Soil Analysis. Part. 2. Second Edition. Agronomy 9. ASA, Madison, WI. Nichols, D. S., and D. R. Keeney. 1973. Nitrogen and phosphorus release from decaying water milfoil. Hydrobiol. 42:509-525. Nyhan, J. W. 1975. Decomposition of carbon-14 labeled plant materials in a grassland soil under field conditions. Soil Sci. Soc. Am. Proc. 39:643-648. Nyhan, J. W. 1976. Influence of soil temperature and water tension on the decomposition rate of carbon-14 labeled herbage. Soil Sci. 121:288-293. Odiom, W. E., and M. A. Heywood. 1978. Decomposition of intertidal freshwater marsh plants. In Good, R. E. D. F. Whigham, and R. J. Simpson (eds.). Freshwater Wetlands. Ecololgical Processes and Management Potential. Academic Press, New York. Ogwada, R. A. 1983. Growth, nutrient uptake, and nutrient regeneration by selected aquatic macrophytes. M.S. Thesis. Univ. of Florida. Ogwada, R. A., K. R. Reddy, and D. A. Graetz. 1984. Effects of aeration and temperature on nutrient regeneration from selected aquatic macrophytes. J. Environ. Qual. 13:239-243. Orchard, V. A., and F. J. Cook. 1983. Relationship between soil respiration and soil moisture. Soil Biol. Biochem. 15:447-453. Otsuki, A., and R. G. Wetzel. 1974. Release of dissolved organic matter by autoanalysis of a submerged macrophyte, Scriptus subterminalis Limnol. Oceanogr. 19:842-845. Parra, J. V., and C. C. Hortenstine. 1976. Response by pearl millet to soil incorporation of water hyacinths. J. Aquat. Plant Manage. 14:75-79. Patterson, D. T., and S. 0. Duke. 1979. Effect of growth irradiance on the maximum photosynthetic capacity of water hyacinth ( Eichhornia crassipes ) Plant Cell Physiol. 20:117-184.

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137 Peevey, W. J., and A. G. Norman. 19A8. Influence of composition of plant materials on properties of the decomposed residues. Soil Sci. 65:209-226. Penfound, W. T. and T. T. Earle. 19A8. The biology of the water hyacinth. Ecol. Monogr. 18:447-472. Ponnamperuma F. N. 1972. The chemistry of submerged soils. Adv. Agron. 24:29-96. Price, E. C, and P. N. Cheremisinof f 1981. Biogas production and utilization. Ann Arbor Science Publishers, Inc., Ann Arbor, MI. Puriveth, P. 1980. Decomposition of emergent macrophytes in a Wisconsin marsh. Hydrobiol. 72:231-242. Rai, D. N., and J. D. Munshi. 1979. The influence of thick floating vegetation (water hyacinth: Eichhornia crassipes ) on the physico-chemical environment of a freshwater wetland. Hydrobiol. 62:65-69. Raven, J. A., and F. A. Smith. 1976. Nitrogen assimilation and transport in vascular land plants in relation to intracellular pH regulation. New Phytol. 76:415-431. Reddy, K. R. 1981. Diel variations in physico-chemical parameters of water in selected aquatic systems. Hydrobiol. 85:201-207. Reddy, K. R. 1983. Fate of nitrogen and phosphorus in a waste-water retention reservoir containing aquatic macrophytes. J. Environ. Qual. 12:137-141. Reddy, K. R. 1984. Water hyacinth ( Eichhornia crassipes ) biomass product ion in Florida. Biomass 6:167-181. Reddy, K. R., and L. 0. Bagnall. 1981. Biomass production of aquatic plants used in agricultural drainage water treatment. In 1981 International Gas Res. Conf. Proc. Govt. Inst. Inc., Rockville, MD. Reddy, K. R. and W. F. DeBusk. 1984. Growth characteristics of aquatic macrophytes cultured in nutrient-enriched water: I. Water hyacinth, water lettuce, and pennywort. Econ. Bot. 38:229-239. Reddy, K. R., and D. A. Graetz. 1981. Use of shallow reservoir and flooded soil systems for wastewater treatment: Nitrogen and phosphorus transformations. J. Environ. Qual. 10:113-119. Reddy, K. R., F. M. Hueston, and T. McKim. 1985. Biomass production and nutrient removal potential of water hyacinth cultured in sewage effluent. J. Solar Energy Eng. 107:128-135.

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138 Reddy, K. R., R. Khaleel, and M. R. Overcash. 1980. Carbon transformations in the land areas receiving organic wastes in relation to nonpoint source pollution: A conceptual model. J. Environ. Qual. 9:A3A-AA2. Reddy, K. R., and P. D. Sacco. 1981. Decomposition of water hyacinth in agricultural drainage water. J. Environ. Qual. 10:228-233. Reddy, K. R. P. D. Sacco, D. A. Graetz, K. L. Campbell, and L. R. Sinclair. 1982. Water treatment by aquatic ecosystems: Nutrient removal by reservoirs and flooded fields. Environ. Manage. 6:261-271. Reddy, K. R., and D. L. Sutton. 198A. Water hyacinths for water quality improvement and biomass production. J. Environ. Qual. 13:1-8. Reddy, K. R., D. L. Sutton, and G. Bowes. 1983. Freshwater aquatic plant biomass production in Florida. Soil Crop Sci. Soc. Fla. A2:28-A0. Reddy, K. R. and J. C. Tucker. 1983. Productivity and nutrient uptake of water hyacinth, Eichhornia crassipes I. Effect of nitrogen source. Econ. Bot. 37:237-2A7. Rogers, H. H. and D. E. Davis. 1972. Nutrient removal by water hyacinth. Weed Sci. 20:A23-A27. Ryan, J. A., D. R. Keeney, and L. M. Walsh. 1973. Nitrogen transformations and availability of an anaerobically digested sewage sludge in soil. J. Environ. Qual. 2:A89-A92. Sain, P., and F. E. Broadbent. 1977. Decomposition of rice straw in soils as affected by some management factors. J. Environ. Qual. 6:96-100. Sato, H., and T. Kondo. 1981. Biomass production of waterhyacinth and its ability to remove inorganic minerals from water. I. Effect of the concentration of culture solution on the rates of plant growth and nutrient uptake. Japan. J. Ecol. 31:257-267. Scarsbrook, E., and D. E. Davis. 1971. Effect of sewage effluent on growth of five vascular aquatic species. Hyacinth Contr. J. 9:26-30. Schreiner, S. D. 1980. Effects of water hyacinth on the physio-chemistry of a south Georgia pond. J. Aquat. Plant Manage. 18:9-12. Schwegler, B. R. and T. W. McKim. 1981. Reedy Creek Improvement District Water Hyacinths Program. Report to EPA. Grant S805655. Nov. 1981.

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i 139 Sheffield, C. W. 1967. Water hyacinth for nutrient removal. Hyacinth Contr. J. 6:27-30. 14 Shields, J. A., and E. A. Paul. 1973. Decomposition of • C-labelled plant material under field conditions. Can. J. Soil Sci. 53:297-306. Shir^lipour, A., and P. H. Smith. 198A. Conversion of biomass into methane gas. Biomass 6:85-92. Sinha, M. K., D. P. Sinha, and H. Sinha. 1977. Organic matter transformations in sbils: V. Kinetics of carbon and nitrogen mineralization in soils amended with different organic materials. Plant Soil 46:579-590. Sommers, L. E. 1977. Chemical composition of sewage sludges and analysis of their potential use as fertilizer. J. Environ. Qual. 6:225-232. Sompongse, D. 1982. The role of wetland soils in nitrogen and phosphorus removal from agricultural drainage water. Ph.D. dissertation. Univ. of Florida. Stack, C, P. Lichtenberger, and J. Martin. 1981. Economic and environmental consequences of anaerobic digestion of animal wastes. In Preprints of the 1981 International Gas Research Conference. Stark, S. A., and C. E. Clapp. 1980. Residual nitrogen availability from soils treated with sewage sludge in a field experiment. J. Environ. Qual. 9:505-512. Steward, K. K. 1970. Nutrient removal potentials of various aquatic plants. Hyacinth Contr. J. 8:34-35. Tenny, F. G., and S. A. Waksman. 1929. Composition of natural organic materials and their decomposition in the soil. IV. The nature and rapidity of decomposition on the various organic complexes in different plant materials, under aerobic conditions. Soil Sci. 28:55-84. Terry, R. E. D. W. Nelson, and L. E. Sommers. 1979. Carbon cycling during sewage sludge decomposition in soils. Soil Sci. Soc. Am. J. 43:494-499. Terry, R. E., D. W. Nelson, and L. E. Sommers. 1981. Nitrogen transformations in sewage sludge-amended soils as affected by soil environmental factors. Soil Sci. Soc. Am. J. 45:506-513. Tester, C. F., L. J. Sikora, J. M. Taylor, and J. F. Parr. 1977. Decomposition of sewage sludge in soil: I. Carbon and nitrogen transformations. J. Environ. Qual. 6:459-463.

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140 Toerien, D. F., and W. H. J. Hattingh. 1969. Anaerobic digestion. I. The microbiology of anaerobic digestion. Water Res. 3:385-416. Wolverton, B. C, and R. C. McDonald. 1975a. Water hyacinths and alligator weeds for removal of lead and mercury from polluted waters. NASA Tech. Memo. No. TM-X-72723. Natl. Space Technol. Lab. Louis, MS. Wolverton, B. C, and R. C. McDonald. 1975b. Water hyacinths and alligator weeds for removal of silver, cobalt, and strontium from polluted waters. NASA Tech. Memo. No. TM-X-72727. Natl. Space Technol. Lab., Louis, MS. Wolverton, B. C, and R. C. McDonald. 1979. Energy from aquatic plant wastewater treatment systems. NASA/NSTL. Tech. Memorandum TM-X-72733. Natl. Space Technol. Lab., Louis, MS. Wolverton, B. C, and R. C. McDonald. 1981. Energy from vascular plant wastewater treatment systems. Econ. Bot. 35:224-232. Yount, J. L., and R. Grossman. 1970. Eutrophication control by plant harvesting. J. Water Poll. Contr. Fed. 42:173-183. Zeikus, J. G. 1977. The biology of methanogenic bacteria. Bacteriol. Rev. 41:514-541.

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BIOGRAPHICAL SKETCH Kevin Moorhead was born in Findlay, Ohio, on May 22, 1956, and spent the first 18 years of his life there. After high school graduation and a summer in California, Kevin moved to Swannanoa, North Carolina, and began his college career at Warren Wilson College. He graduated in December, 1978, with a Bachelor of Arts degree in biology. Kevin moved to Pintlala, Alabama, and worked for Alaga-Whitf ield food company in Montgomery, Alabama, for 7 months. In August, 1979, he moved to Columbus, Ohio, to begin a graduate program in agronomy at the Ohio State University. Kevin received his Master of Science degree in December, 1981, with an emphasis in soil fertility, under the direction of the late Dr. E. 0. McLean. Since January, 1982, he has pursued a Ph.D. degree at the University of Florida in the Department of Soil Science. He is presently a candidate for the degree of Doctor of Philosophy with an emphasis in soil biochemistry. Kevin has 7 sisters, a brother looking for Sasquatch, and (at this point) 3 nieces and 1 nephew. lAl

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-I I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. D. A. Graetz, Chairma^^^^' Associate Professor of Soil Science I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. K. R. Reddy, CocKairman Professor of Soil Science I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Bow^' essor of Botany I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. G. A. Fiskell Professor of Soil Science

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I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. R. A. -illordstedt Associate Professor of Agricultural Engineering This dissertation was submitted to the Graduate Faculty of the College of Agriculture and to the Graduate School and was accepted as partial fulfillment of the requirements for the degree of Doctor of Philosophy. May 1986 Dean, C^'lege of Agricvfiture Dean, Graduate School


63
The fresh plant material and digested sludge were freeze-dried
(Thermovac-T) and analyzed for the following: TS, FS, VS, TKN (Nelson
and Sommers, 1973), total carbon (TC) (LECO Induction Furnace 523-300),
lignin, cellulose and hemicellulose (Goering and Van Soest, 1970), and
ashed mineral constituents (Gaines and Mitchell, 1979).
Results and Discussion
Characteristics of Inocula
Characteristics of the two inocula varied considerably (Table 9).
The inoculum used for plants with high N content (high N plants)
contained higher levels of TS, VS, NH+-N, TKN, and COD than the inoculum
used for plants with low N content (low N plants). Characteristics of
inoculum depended on the type of feedstock used for digestion (Stack et
al., 1978) as well as digester operating conditions (Hashimoto et al.,
1980). The inoculum used for high and low N plants came from digesters
with feedstocks of a 3:1 water hyacinth/domestic sewage sludge blend and
water hyacinths, respectively. Ammonium accounted for 68 and 92% of the
total N of the inoculum from the water hyacinth/sewage sludge and water
hyacinth feedstocks, respectively. Ammonium was the primary N source for
methanogenic bacteria (Zeikus, 1977).
Carbon and Nitrogen Mineralization During Digestion
Biogas (CH^ and CC^) production, corrected to standard conditions
(0C and 0.1 MPa), is given in Table 10. Gas production essentially
ceased after 60 days of digestion. Cumulative biogas production at 60
days for high N plants was approximately 21% less for 2.5 L of inoculum
compared to 10 L. Furthermore, cumulative biogas production at 15 days


DECOMPOSITION OF FRESH AND ANAEROBICALLY
DIGESTED PLANT BIOMASS IN SOIL
Anaerobic digestion of plant biomass, sewage sludge, or animal
wastes generates a waste by-product which must be disposed of, or
preferably utilized, in an environmentally-safe manner. Disposal of the
digested sludge by land application is one option often considered
(Miller, 1974; Terry et al., 1979; Atalay and Blanchar, 1984). The
digested biomass sludge differs chemically from the fresh plant biomass.
A consequence of anaerobic digestion is a reduction of the more readily
decomposable C of the plant tissue during production of CH^ and CC^.
Many sludges contained relatively large amounts of Ca, Mg, P and Zn and
lower contents of soluble elements such as K (Sommers, 1977)
Miller (1974) concluded that anaerobically digested sewage sludge
was rather resistant to further decomposition with a maximum of 20% of
the added C was evolved as CO^ during a 6-month incubation. Tester et
al. (1977) reported 16% of added C from composted sewage sludge was
evolved as CO^ during 54 days of incubation. Hsieh et al. (1981a) showed
that activated sludge had a much higher C mineralization rate compared to
digested sludge due to a larger portion of active organic C.
Epstein et al. (1978) determined that the percentage of added N
mineralized from digested sludge remained essentially constant
irrespective of application rate. However, Ryan et al. (1973) and Stark
95


31
Shoot and root lengths were similar for plants in both reservoirs
during autumn and winter (Table 2). During spring, the water hyacinth
root lengths were shorter and shoot lengths longer in the fertilized
reservoir compared to the plants in the control reservoir. An
interesting development in plant morphology was the dislodging of
practically the entire root system from plants in the fertilized
reservoir beginning in March after daily temperatures began to increase.
The majority of plants were typified by a small root system compared to
plants in the control reservoir. Some root dislodging was noticed in
the control reservoir during spring and summer but was not as
characteristically uniform as in the fertilized reservoir. Root lengths
in the fertilized reservoir began to increase during summer, but the
shoot lengths were double those in the control reservoir.
Under nutrient-limiting conditions, water hyacinths produce a large
volume of root material presumably to increase their nutrient absorption
capacity. With nutrient-enriched media, water hyacinth use more
photosynthetic energy in shoot production. Cornwell et al. (1977)
measured shoot lengths in excess of 1 m in wastewater media. Penfound
and Earle (1948) recorded root lengths of 0.1 to 1 m. Maximum shoot
length recorded during this study was 55 cm during summer for fertilized
plants compared to 28 cm during summer for control plants.
i
The plant tissue N content is shown in Fig. 8. Fertilization
resulted in increases in shoot, root, and detritus N content compared to
plants in the control reservoir. Maximum tissue N content for
fertilized plants occurred during winter when plant productivity was
low. The increase in plant productivity in spring and summer diluted
the N content of the tissue although total N assimilation by the plants


16
(Hughes, 1981). Bioconversion of biomass with a higher C/N ratio was
limited by N. A lower initial C/N ratio resulted in mineralization of
organic N during digestion. The C/N ratio of the digester effluent was
lower than the C/N ratio of the fresh slurry because of the release of C
as CO^ and CH^ (House, 1981).
Anaerobic digestion of plant biomass resulted in high
concentrations of NH*-N in the digester (Hashimoto et al., 1980; Field
et al., 1984). Ammonium was toxic to methogens at concentrations > 3.0
g L-l, regardless of pH (Hashimoto et al., 1980). Losses of NH^-N
through volatilization should be low in digesters operating at the
optimum pH of 6.7 to 7.2 unless NH^-N concentrations are high.
Waste By-Product Recycling
The final component of the integrated "biomass for energy" system
was recycling of the waste by-product generated from the anaerobic
digestion of plant biomass. The waste by-product from the anaerobic
digester was screened to separate the digested biomass sludge from the
effluent. The effluent was recycled in the water hyacinth biomass
production system discussed earlier. Methane digester effluent contains
high levels of NH+-N (> 200 mg L ^) which may inhibit plant growth.
Dilution of the effluent is required before use in a water hyacinth
production system. Optimum dilution of the effluent for maximum water
hyacinth yields has not been established. Nitrogen cycling in a water
hyacinth production system was presented earlier.
The sludge was added to soil as an organic amendment. A
consequence of anaerobic digestion was a reduction of the readily
decomposable C of the plant tissue during production of CH^ and CO^-


11
microbial-controlled degradation (Boyd, 1970b; Hunter, 1976; Godshalk
and Wetzel, 1978a; Howard-Williams et al., 1983).
Otsuki and Wetzel (1974) reported a rapid leaching loss of
dissolved organic matter regardless of conditions of aerobiosis or
whether plants were fresh or freeze-dried. Hill (1979) concluded that
rapid leaching of soluble material accounted for a 21 to 60% dry weight
loss of aquatic macrophytes during the first 8 days of incubation.
Leaching was established as the major process in the decomposition of
eelgrass and total loss of organic matter by leaching accounted for 82%
of dried leaves and 65% of fresh leaves (Harrison and Mann, 1975).
Leaching rates appeared to be independent of temperature (Carpenter,
1980).
Potassium, Na, Mg, and Ca have all been reported as being rapidly
lost during the early leaching phase of plant decomposition (Boyd,
1970b; Davis and van der Valk, 1978; Puriveth, 1980). Carpenter (1980)
found that the higher the initial P concentration, the more rapid was P
leaching.
The second stage of decomposition is attributed to biological
processes. Microbial-controlled decomposition was influenced by
temperature (Carpenter, 1980; Puriveth, 1980), pH (Sompongse, 1982),
available 0^ (Godshalk and Wetzel, 1978a), and available nutrients
(Carpenter and Adams, 1979; Puriveth, 1980). Godshalk and Wetzel
(1978a) found that the presence of O2, regardless of temperatures of 10
or 25 C, permitted rapid degradation of dissolved and particulate
organic matter. Decomposition of water hyacinth was found to be faster
under aerobic than anaerobic conditions (Reddy and Sacco, 1981).
However, Sompongse (1982) determined that aeration did not have a


Figure 18. Nitrogen cycling in an integrated system for water hyacinths growing
in nutrient-enriched systems. Numbers in parentheses are percentages
of the initial 35 g N placed in the anaerobic digester.
113


I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
R. A. dtfordstedt
Associate Professor of Agricultural
Engineering
This dissertation was submitted to the Graduate Faculty of the College
of Agriculture and to the Graduate School and was accepted as partial
fulfillment of the requirements for the degree of Doctor of Philosophy.
May 1986
[c vZ/ty
Dean, College of Agrictfftc
ture
Dean, Graduate School


NITROGEN CYCLING IN AN INTEGRATED
"BIOMASS FOR ENERGY" SYSTEM
By
KEVIN KEITH MOORHEAD
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS
FOR THE DEGREE OF DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA
1986


74
had a higher lignin content compared to fresh plant biomass. The
increase in lignin was due to the loss of readily decomposable C during
anaerobic digestion. Lignin appears to be practically inert to
anaerobic digestion (Hashimoto et al., 1980) Generally, there was a
decrease in cellulose after digestion. The hemicellulose remained
similar for the fresh plant biomass and digested sludge. Anaerobic
digestion resulted in losses of K and Mg from fresh plant biomass, but
increased the concentration of sludge Ca, Na, Fe, and Zn.
Conclusions
Cumulative biogas production at 60 days was similar for high (-34 g
N kg ^ dry wt tissue) and low (-10 g N kg ^) N plants suggesting that
long term digestion of water hyacinth was not influenced by initial N
content. Effects of inoculum volume on cumulative biogas production
were seen at 15 days for high N plants but not low N plants. Conversion
of cumulative biogas production into biogas and methane yields (L g ^ VS
added) showed that low N plants produced more biogas and methane than
high N plants. This was due to increase of TS (and consequently VS) in
digesters of high N plants from the inoculum source, since cumulative gas
production was similar for both types of plants.
Mineralization of organic to ^NH^-N accounted for 72 and 35% of
15 I
added N for high and low N plants, respectively. Approximately 20% of
15
the added N was recovered as organic N in sludge for both types of
plants. A low recovery was observed for low N plants.
Increasing inoculum volume increased electrical conductivity, NH+-N,
TKN, and TS of the digester effluents. The digested biomass sludge had
higher levels of TC, TKN, lignin, Ca, Na, Fe, and Zn, and lower levels


80
Conductivity Meter), pH (Orion Model 404 Specific Ion Meter) and
dissolved 0^ (Yellow Springs Instrument Model 54 O2 Meter) were measured
every other day. The analyses of plant and water samples were
conducted on a Micro Mass 602 spectrometer.
Results and Discussion
Chemical Composition of the Effluents
Characteristics of digester effluents used in the study are given
in Table 16. The initial pH of the effluent sources and nutrient medium
were similar. The digester effluents had a wide range of EC, NH^-N, TKN
and other nutrients. This provided an opportunity to evaluate water
hyacinth growth under diverse media conditions. The EC of the nutrient
medium and diluted effluents ranged from 0.7 to 2.3 dS m ^. The EC of
the undiluted effluents ranged from 4.3 to 6.7 dS m ^.
The NH+-N and TKN concentrations of the undiluted effluents from
4
digested plants with a high N content (high N plants) were greater than
those of the undiluted effluents of digested plants with a low N content
(low N plants). The NH^-N concentrations ranged from 23 to 104 rag L ^
for diluted effluents and 24 to 289 mg L ^ for undiluted effluents,
respectively.
High Na and K concentrations were noted for undiluted effluents.
The highest levels of P were in undiluted effluents of digested high N
plants and the highest levels of Ca and Mg were in undiluted effluents
of digested low N plants. The critical levels of Na, K, Ca and Mg
needed to achieve maximum water hyacinth yields are relatively unknown.
The NH*-N, K and Mg concentrations were higher than levels reported for
water hyacinths cultured in primary or secondary sewage effluent (Reddy
et al., 1985).


22
chosen because N is often identified as a limiting factor for plant
growth and is used to establish loading rates in the disposal of solid
and liquid waste.
Plant uptake was established as a major N removal process during
water hyacinth biomass production (Reddy and Sutton, 198A). However,
the role of water hyacinth detritus as a N source or sink has not been
established. Methane yields during anaerobic digestion of water
hyacinth were enhanced with increasing N content (Shiralipour and Smith,
1984). However, N mineralization rates were not investigated. Limited
information was available on disposal or utilization of digester
effluent or sludge from anaerobically digested plant biomass (Hanisak et
al., 1980; Atalay and Blanchar, 1984). The overall objective of this
research was to integrate the three components of biomass production,
anaerobic digestion of biomass, and digester waste recycling with
respect to N cycling.


59
attributed to increasing heterotrophic respiration due to increasing
amounts of C. Although this general relationship existed for all
treatments, plant cover and sediment layer appeared to have more of a
regulatory role in dissolved 0^ dynamics than detritus.
Water pH was constant in water having plant cover and sediment.
The decreasing pH of water with plant cover and no sediment was
attributed to NH^-N assimilation by plants in exchange for H+. The pH
of open water was generally lower as the rate of detritus increased.
Detritus had no apparent effect on rate of N loss in water with
water hyacinths. However, N loss was more rapid in open water as the
rate of detritus increased.
Total plant ^NH^-N uptake exceeded ^N0^-N uptake. Both sediment
and detritus appeared to be a potential N source for water hyacinths.
Total recovered by water hyacinths ranged from 57 to 72% for added
15N0"-N and 70 to 89% for added 15NH+-N.
3 4
Less than 10% of the added ^NH^-N was immobilized by detritus in
water with plant cover. However, in water without plant cover, up to
35% of the added ^NH*-N was associated with detritus. This suggests
that during periods of low water hyacinth productivity, typical in cold
weather conditions, detritus is an important sink for added N.


LIST OF FIGURES
FIGURE PAGE
1. Integrated water hyacinth aquaculture system
of biomass production, bioconversion to methane
and digester waste recycling 2
2. A generalized diagram of a water hyacinth plant 5
3. Nitrogen cycling in a water hyacinth production sytem ... 9
4. Nitrogen cycling during anaerobic digestion 15
5. Nitrogen cycling in soil treated with plant residues. ... 18
6. Weekly averages of daily temperatures and solar
radiation 27
7. Monthly averages of daily primary productivity and
detritus production 28
8. Seasonal plant tissue nitrogen content 33
9. Dissolved 0 in sediment-water-plant systems with
added nitrate 45
10. Dissolved 0^ in sediment-water-plant systems with
added ammonium 46
11. The pH of sediment-water-plant systems with
added nitrate 48
12. The pH of sediment-water-plant systems with
added ammonium 49
13. Nitrogen loss from sediment-water-plant systems
with added nitrate 50
14. Nitrogen loss from sediment-water-plant systems
with added ammonium 51
15. Dry weight gains of water hyacinths in digester
effluents and nutrient medium 83
vii


44
Water pH (Orion Model 404 Specific Ion Meter), dissolved 02 (Yellow
Springs Instrument Model 54 0^ meter) and temperature were measured
every other day. Electrical conductivity (Hach Mini Conductivity Meter)
was measured weekly.
Results and Discussion
Effect of Detritus on Water Dissolved 02
The dissolved 0^ concentrations of water with added ^NO^-N and
15 +
NH^-N are shown in Figs. 9 and 10, respectively. Dissolved
concentrations remained < 5 mg L ^ in water having plant cover but lower
dissolved O2 concentrations were recorded as the rate of detritus
increased. This reflected increasing microbial 0^ demand for
respiratory functions with increasing C source (Fenchel and Jorgensen,
1977). For water without plant cover (open water), the dissolved 0^
concentrations were scattered more with time. The dissolved 0^
measurements were taken between 2:30 and 3:30 pm and should represent
near maximum concentrations on a diurnal basis (Howeler, 1972). The
increased dissolved 0^ concentrations of open water were due to an
increased rate of photosynthesis by algae during the day compared to
respiration (Reddy, 1981).
Generally the effect of decreasing dissolved 0^ concentrations with
increasing detritus was seen for open water with or without sediment.
Dissolved 0^ concentrations were generally lower in open water with
sediment compared to open water without sediment. Nichols and Keeney
(1973) reported lower dissolved concentrations for sediment-water
systems than water only. Although detritus appeared to have a role in
C>2 dynamics in water, plant cover was the primary regulator.


BIOGRAPHICAL SKETCH
Kevin Moorhead was born in Findlay, Ohio, on May 22, 1956, and spent
the first 18 years of his life there. After high school graduation and a
summer in California, Kevin moved to Swannanoa, North Carolina, and began
his college career at Warren Wilson College. He graduated in December,
1978, with a Bachelor of Arts degree in biology. Kevin moved to Pintlala,
Alabama, and worked for Alaga-Whitfield food company in Montgomery,
Alabama, for 7 months. In August, 1979, he moved to Columbus, Ohio, to
begin a graduate program in agronomy at the Ohio State University. Kevin
received his Master of Science degree in December, 1981, with an emphasis
in soil fertility, under the direction of the late Dr. E. 0. McLean.
Since January, 1982, he has pursued a Ph.D. degree at the University of
Florida in the Department of Soil Science. He is presently a candidate
for the degree of Doctor of Philosophy with an emphasis in soil
biochemistry. Kevin has 7 sisters, a brother looking for Sasquatch, and
(at this point) 3 nieces and 1 nephew.
141


Table 17. First-order kinetic descriptions of NH^-N
loss with time.
Inoculum
Initial
nh4-n
Reduction
volume
cone.
k
time^
R2
--L--
T-1
mg L
day ^
days
Diluted effluents from high N plants
2.5
23
0.593
1.12
0.722
5
65
0.449
1.54
0.917
10
104
0.228
3.04
0.951
Undiluted effluents from high N plants
2.5
161
0.232
2.98
0.938
5
212
0.207
3.35
0.947
10
289
0.175
3.96
0.982
Undiluted effluents from
low N plants
2.5
24
0.446
1.55
0.850
5
49
0.325
2.13
0.885
10
87
0.350
1.98
0.845
Nutrient medium
20
0.281
2.47
0.973
^ Time required for 50% reduction in initial NH^-N
concentration.


133
Fenchel, T. M., and B. B. Jorgensen. 1977. Detritus food chains of
aquatic ecosystems: The role of bacteria. In Alexander, M. (ed.).
Advances in Microbial Ecology. Volume. 1. Plenum Press, New York.
Field, J. A., J. S. Caldwell, S. Jeyanayagam, R. B. Reneau, Jr., W.
Kroontje, and E. R. Collins, Jr. 1984. Fertilizer recovery from
anaerobic digesters. Transactions of the ASAE 27:1871-1881.
Franco, A. A., and D. N. Munns. 1982. Plant assimilation and nitrogen
cycling. Plant Soil 67:1-13.
Freney, J. R., J. R. Simpson, and 0. T. Denmead. 1983. Volatilization
of ammonia. In Freney, J. R., and J. R. Simpson (eds.). Gaseous
Loss of Nitrogen From Plant-Soil Systems. Developments in Plant
and Soil Sciences. Vol. 9. Martinus Nijhoff/Dr. W. Junk
Publishers, The Hague.
Gaines, T. P., and G. A. Mitchell. 1979. Chemical Methods for Soil and
Plant Analysis. Agron. Handbook No. 1. Univ. of Georgia. Coastal
Plains Experiment Station.
Gambrell, R. P., and W. H. Patrick, Jr. 1978. Chemical and
microbiological properties of anaerobic soils and sediments. In
Hook, D. D., and R. M. M. Crawford (eds.). Plant Life in Anaerobic
Environments. Ann Arbor Science Publishers, MI.
Gilmour, C. M., E. F. Broadbent, and S. M. Beck. 1977. Effects of
waste applications on soil carbon and nitrogen cycles. In Elliot, L.
F., and F. J. Stevenson (eds.). Soils for Management of Organic
Waste and Waste Waters. Soil Sci. Soc. Am. Madison, WI.
Godshalk, G. L., and R. G. Wetzel. 1978a. Decomposition of aquatic
angiosperms. I. Dissolved components. Aquat. Bot. 5:281-300.
Godshalk, G. L., and R. G. Wetzel. 1978b. Decomposition of aquatic
angiosperms. II. Particulate components. Aquat. Bot. 5:301-327.
Gosselink, J. G., and C. J. Kirby. 1974. Decomposition of salt marsh
grass, Spartina alterniflora Loisel. Limnol. Oceanogr. 19:825-831.
Goering, H. K., and P. J. Van Soest. 1970. Forage Fiber Analyses.
(Apparatus, Reagents, Procedures, and Some Applications).
ARS-USDA. Agriculture Handbook No. 379. Washington, DC.
Haller, W. T., and D. L. Sutton. 1973. Effect of pH and high P
concentration on growth of water hyacinths. Hyacinth Contr. J.
11:59-61.
Haller, W. T., D. L. Sutton, and W. C. Barlowe. 1974. Effects of
salinity on growth of several aquatic macrophytes. Ecol.
55:891-894.


47
Effect of Detritus on Water pH
The pH of water with added ^NO^-N or ^NH+-N is shown in Figs. 11
and 12, respectively. A fairly constant pH of 7.0 was noted in water
having plant cover and sediment regardless of detritus additions. The
pH decreased in water with plant cover but without sediment. The
decreasing pH was noted immediately for added ^NH^-N and after 20 days
for added ^NO^-N. The decrease in pH was less as the detritus rate
increased.
The immediate pH decrease in water with plants and added NH^-N was
probably due to production of H+ during plant NH^-N assimilation (Raven
and Smith, 1976). The H+'generated is actively exuded, partly in
exchange for cations (Franco and Munns, 1982). Plant NO^-N assimilation
occurs by exchange with another anion or by simultaneous cation
assimilation to maintain ion equilibrium (Kirkby and Mengel, 1967;
Mengel, 1974). The decreasing pH in water with plant cover but without
sediment suggested that the underlying sediment had a buffering role in
pH regulation.
The pH of water without plant cover were generally higher and more
variable than water with plant cover. Reddy (1981) reported high
mid-day pH values in ponds where algal activity was high. Bouldin et
al. (1974) found high pH values (> 8.5) for ponds containing submersed
macrophytes during sunlight hours. The pH of open water was generally
lower as the rate of detritus increased.
Effect of Detritus on Nitrogen Loss
Nitrogen loss from water with added ^N0_-N or ^NhI'-N is shown in
3 4
Figs. 13 and 14, respectively. Sediment or detritus had no apparent


Figure 1. Integrated water hyacinth aquaculture system of biomass
production, bioconversion to methane and digester waste
recycling.


Table 21. Characteristics of the digester effluents and nutrient medium after
water hyacinth treatment.
Inoculum
volume pH EC NH*-N TKN Na K P Ca Mg
--L-- dS mg L
Diluted effluents from high N plants
2.5
7.2
0.4
< 1
4.3
80
2
0.3
14
11
5
7.5
0.7
< 1
6.0
173
2
0.4
16
12
10
7.5
1.2
< 1
3.8
297
4
0.3
19
18
Undiluted effluents
from hiRh
N plants
2.5
8.7
4.2
22.2
39.4
873
103
10.5
11
28
5
8.6
4.4
31.9
80.2
903
134
9.9
9
24
10
8.4
4.9
49.9
87.9
950
163
7.1
11
22
Undiluted effluents
from low
N plants
2.5
8.8
4.2
< 1
4.3
450
72
1.2
23
24
5
9.2
4.6
< 1
5.1
523
98
1.2
10
33
10
9.3
4.8
< 1
8.6
547
159
1.3
10
38
Nutrient
Medium
3.5
0.7
< 1
1.8
10
1
0.5
42
17


36
immobilization of N as plant detritus was 3 and 33% of standing crop
assimilation for harvested and non-harvested water hyacinth plants,
respectively. However, they did not include plant detritus trapped
within the water hyacinth mat.
The annual N assimilation by water hyacinth is low compared to N
removal rates reported for plants growing in nutrient-enriched waters.
Reddy et al. (1985) found annual N removal rates of 1726 and 1193 kg N
ha ^ yr ^ for water hyacinths growing in primary and secondary sewage
effluent, respectively. Rogers and Davis (1972) concluded that water
hyacinths could remove 2500 kg N ha ^ yr ^ if maximum growth could be
sustained. Sato and Kondo (1981) measured a maximum removal rate of
4782 kg N ha ^ yr ^ for plants growing in a nutrient medium. The low
annual N assimilation reported in the present study was due to low rates
of fertilization.
Plant uptake played a major role in removing N in both the
reservoirs (Table 4). A large portion of lake water N was present as
organic N, which was not readily available to plants. In both
reservoirs, plants derived N from mineralization of lake water organic
N, N release from underlying sediments, and mineralization of organic N
in detritus. In the fertilized reservoirs, plants also derived N from
the fertilizer N applied. Nitrogen assimiliation by water hyacinth from
the added fertilizer was calculated as follows: (Total N assimilation
by plants in the fertilized reservoir Total N assimilation by plants
in the control reservoir / Total fertilizer N added) 100.
About 51% of the added fertilizer N was taken up by the plants in
the fertilized reservoir, and the remaining 49% may have been lost
through denitrification. Reddy et al. (1982) observed a reduction of 78


25
20
I 5
I 0
5
0
20
I 5
10
5
0
Fertilized
TIME (months)
. Monthly averages of daily primary productivity and
detritus production.


98
various treatments and dividing by the amount of C added for each
residue.
Soil samples were analyzed at 0, 30, 60, and 90 days for 2 M
KCl-extractable NH+-N and NO^-N by steam distillation (Keeney and Nelson,
1982), TKN (Nelson and Sommers, 1972), Mehlich I extractable Ca, K, Mg,
Na, Fe, Zn, and P (Melich, 1953), organic C by the Walkley-Black method
(Nelson and Sommers, 1982), and pH. The ^N analyses were conducted on
a Micro Mass 602 spectrometer.
Results and Discussion
Plant Residue Characterization
Characteristics of the fresh plant biomass and anaerobically
digested biomass sludge are presented in Table 22. The TC and TKN
concentrations of the digested sludges were higher than their respective
fresh plant materials. Total C was not significantly different for low
and high N fresh plant biomass or digested sludge. The C/N ratio of the
fresh plant biomass with a low N content (low N plant biomass) decreased
from 35 to 16 after digestion. The C/N ratio of fresh plant biomass
with a high N content (high N plant biomass) did not change during
digestion.
Lignin content was significantly higher in digested sludges due to
loss of readily-decomposable C during anaerobic digestion. The low N
fresh plant biomass contained approximately twice as much lignin as the
high N fresh plant biomass. Moore and Bjorndal (1984, Unpublished
results, Univ. of Florida, Gainesville) concluded that water hyacinth
roots, in general, have higher lignin content than shoots. The lignin
content of the low N fresh plant biomass was approximately double that of


110
Table 26. Mehlich I extractable constituents at Day 90 from added
fresh and digested plant biomass.
Chemical
constituent
Control
Low N plant biomass
High N plant
biomass
Fresh
Digested
Fresh
Digested
, -1
mg Kg soil
Calcium
252 e f
361 ab
373 a
327 d
348 be
Potassium
25 e
115 b
40 d
119 a
47 c
Magnesium
24 d
59 a
35 c
39 b
35 c
Sodium
3 e
53 c
72 b
41 d
117 a
Iron
14 d
14 d
20 a
15 c
18 b
Zinc
3.3 e
5.7 d
11.3 a
6.0 c
7.2 1
^Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.


116
Nutrient-Limited Systems
Nitrogen cycling in an integrated system for water hyacinths
growing in nutrient-limited systems is summarized in Fig. 19.
Water Hyacinth Production
-2 -1
Growth rates of 2 to 29 g dry wt m day have been reported for
plants in natural waters of central and south Florida (Yount and
Crossman, 1970; DeBusk et al., 1981). Plants growing in nutrient-
limited systems generally have a low N content. In this study, tissue N
content of plants grown in tap water without nutrients was 10 g N kg ^
of dry biomass. In addition to a low N content, the shoot:root dry
weight ratio decreased as nutrient availability decreased in the water
media (Reddy, 1984). The shoot:root dry weight ratio of plants with a
low N content was 1.6 compared to 4.2 of plants with a high N content.
Anaerobic Digestion of Plant Biomass
Shiralipour and Smith (1984) concluded that increasing N in the
growth medium used for water hyacinth production increased methane
yields during anaerobic digestion. Surprisingly, gas yields were higher
for plants with a low N content compared to plants with a high N
content. However, the gas yield differences were attributed to
characteristics of inoculum used during digestion. Only 75% of the
initial N placed in the digester was recovered in the effluent and
sludge for low N plants. The low total N recovery is difficult to
explain. Only 35% of the plant organic N placed in the digester was
mineralized to NH+-N compared to 70% mineralization for high N plants.
Approximately 40% of the initial N placed in the digester remained as
organic N after digestion. After digestion, 54% of the initial plant
organic N was recovered in the effluent while 20% was recovered in the
digested sludge.


21
Jenkinson (1965, 1971), using different plants and soils,
determined that the proportion of added plant C retained in the soil
under different climatic conditions was remarkably similar over time.
Generally, one-third of the added plant C remained after one year,
falling to one-fifth after 5 years.
Atalay and Blanchar (1984) determined that anaerobically digested
biomass sludge decomposed rapidly in soil as evidenced by nearly 40% of
the C added evolved as CO^ during 100 days of decomposition. However,
Miller (1974) concluded that anaerobic digested sewage sludge was
resistant to further decomposition with a maximum of 20% of the added C
evolved as C0 during a 6-month incubation. Terry et al. (1979) found
that 26 to 42% of anaerobically digested sewage sludge C was evolved as
CO^ during incubation. Generally, the majority of the CO2 produced in
incubation studies was evolved in the first 30 days (Miller, 1974; Terry
et al., 1979; Ataway and Blanchar, 1984).
Other soil properties influenced by plant biomass additions
included increasing water-holding capacity, CEC, and electrical
conductivity (Stark and Clapp, 1980; Atalay and Blanchar, 1984).
Epstein et al. (1976) found levels of salinity and chloride in sewage
sludge applied to soils increased to a level which may affect
salt-sensitive plants.
Conclusions
Although information is available on N cycling for each component
of the system, no attempt has been made to follow N transformations in
an integrated "biomass for energy" system. Evaluation of N cycling was


42
hyacinths. Specifically, the objectives were 1) to determine the
regulatory function of detritus on dissolved 0^ and pH of water and 2)
15 -
to determine the influence of detritus on the fate of N0^ -N and
^NH+-N in sediment- water-plant systems.
Materials and Methods
Two greenhouse studies were conducted to evaluate the effect of
detritus on the fate of labeled N0--N or NH+-N in water with and
3 4
without water hyacinth plants. Treatments evaluated were: 1) with and
without underlying sediment, 2) with and without water hyacinth plant
cover, and, 3) three rates of added water hyacinth detritus. There were
\
24 tanks in each study having dimensions of 50 cm 50 cm 25 cm depth.
Twelve of the 24 tanks contained a 2.5 cm sediment layer (1.875 kg
soil). The sediment was a Lauderhill organic soil (Lithic medisaprists)
collected at the Central Florida REC research farm in Zellwood, Florida.
The soil was air-dried and passed through a 2 mm sieve. Fifty liters of
tap water were added to sediment tanks to obtain a 20 cm water depth.
The greenhouse studies were initiated after sediment/water
equilibration of 1 week. A nutrient medium (a modified 10% Hoagland's
solution) was added to all tanks to obtain nutrient concentrations of:
15NH+-N or 15N0-N = 20.0 mg L_1; K = 23.5 mg L_1; PC^-P = 3.1 mg L_1j
Ca = 20.0 mg L ; Mg = 4.8 mg L SO^-S = 6.4 mg L ^; Fe = 0.6 mg L ^
and micronutrients. Micronutrients were applied through commercially
available liquid fertilizer (Nutrispray-Sunniland, Chase and Co.,
Sanford, Florida) to obtain concentrations of 0.2 mg Cu L ^; 1.5 mg Mn
L ^; 0.04 mg B L ^; and 0.02 mg Mo L ^.


115
Plant assimilation of effluent N recycled 38% of the initial N
placed in the digester. Nitrogen loss from this component of the
integrated system accounted for 42% of the N placed in the digester.
Possible N loss mechanisms during recycling of digester effluent include
algal assimilation and NH^-N volatilization.
The digested sludge contained 20% of the initial N placed in the
digester. The sludge N could potentially be recovered during crop
production after land application. The sludge was fairly resistant to
decomposition in soil and only 20% of the sludge C was evolved as CO^ in
90 days of incubation. A low decomposition rate of sludge applied to
soil was attributed to loss of the more readily-decomposable C
constituents during anaerobic digestion. The decomposition rates
observed in this study for digested biomass sludge applied to soil were
found to be similar to those reported for digested sewage sludge
(Miller, 1974; Tester et al., 1977). Nitrogen mineralization of sludge
organic N during 90 days of decomposition in soil accounted for only 2%
of the initial N placed in the digester.
Fresh water hyacinth biomass was also added to soil to compare its
decomposition to that of digested sludge. Approximately 50% of the
biomass C was evolved as CO^ during 90 days of incubation. Nitrogen
mineralization of biomass organic N accounted for 34% of the initial
plant N placed in the soil. Mixing the fresh water hyacinth biomass
with anaerobically digested sludge may enhance the decomposition of the
sludge.
Total N recovery by sludge and effluent recycling in the integrated
"biomass for energy" system was 60% of the initial N placed in the
digester for high N plants. The remaining 40% was lost from the system
during effluent recycling in a water hyacinth production system.


Table 28. Effluent dissolved 0^ concentration during water hyacinth treatment.
Inoculum
volume
0
2
4
6
8
10
12
14
16
18
20
22
T --
r-1
-mg 02
Li
Diluted i
effluents
from high N
plants
2.5
6.2
4.6
5.4
5.2
4.9
4.7
5.3
5.6
3.9
3.8
3.7
3.3
5
5.4
4.4
3.2
4.0
4.0
4.1
4.5
4.6
3.2
3.5
3.4
2.8
10
4.4
3.9
3.6
2.8
3.5
1.8
4.6
4.3
3.0
3.2
3.7
3.3
Undiluted
effluents
from
ilgh.
N plants
2.5
1.8
2.6
2.9
0.6
0.5
1.0
2.6
2.3
2.9
2.9
3.9
3.1
5
1.7
2.9
2.1
1.0
0.4
0.8
2.9
2.5
2.8
2.5
3.8
2.1
10
0.4
2.1
1.4
0.7
0.2
0.3
2.2
1.3
1.1
1.3
1.5
1.0
Undiluted
effluents
from
low N plants
2.5
4.1
4.5
3.1
2.7
5.1
NA1"
NA
8.0
7.0
8.0
5.9
7.2
5
4.1
4.7
3.7
2.1
3.8
NA
NA
11.9
12.1
11.4
6.9
9.1
10
3.9
4.8
3.5
1.1
1.4
NA
NA
9.1
11.0
NA
NA
NA
Nutrient
medium
7.1
6.9
6.3
5.7
7.0
NA
NA
6.7
7.0
7.2
6.9
6.4
NA = Not available.
125


6
The elongated internodes were designated as stolons (Penfound and Earle,
1948).
The relatively rapid rate of colonization by water hyacinth is due
primarily to vegetative reproduction (stolon and offset production).
The plants reproduce sexually during warmer months until freezing
terminates anthesis. The developing fruits containing the seeds are
cast off onto the mat or into the water. They sink in water and remain
in a viable condition for several years. Manson and Manson (1958)
reported that each plant could produce 5000 to 6000 seeds which remained
viable for at least 5 years.
The geographical distribution of water hyacinth is'regulated by
temperature and salt concentration in the water. When average minimum
temperature reached 10C, productivity of water hyacinth approached zero
(Reddy and Bagnall, 1981). Optimum growth was found in a temperature
range of 25 to 30C (Bock, 1969; Knipling et al., 1970). Water hyacinth
is basically a freshwater plant and will die in waters with sustained
salt concentrations in excess of 2500 mg L (Haller et al., 1974).
Water hyacinth growth is regulated by the nutrient composition of
the water medium, temperature, solar radiation, and plant density.
Water hyacinth potentially could be grown in nutrient-enriched waters
such as sewage effluents, agricultural runoff and drainage effluents,
methane digester effluents, and runoff from animal waste operations.
Nitrogen is present as NH*-N, NO^-N, and organic N in water media
avaiable for water hyacinth production. Organic N often predominates in
most water media and is not readily available for plant assimilation.
Water hyacinths are efficient users of inorganic N and plant
assimilation is one of the major processes of N removal in hyacinth
ponds.


118
Waste By-Product Recycling
The undiluted effluent from digested plants with a low N content
was recycled directly to a water hyacinth production system. Plants
survived in undiluted effluents due to lower NH+-N concentrations.
4
Approximately 21% of the N placed in the digester was reassimilated by
water hyacinths during effluent recycling. Nitrogen loss during
digestion and effluent recycling accounted for 52% of the initial N
placed in the digester.
The digested sludge contained 20% of the initial N placed in the
digester. Similar decomposition rates were noted for both digested
sludges applied to soil despite their differences of initial N content
(27 and 39 g N kg ^ dry sludge). The mineralization of sludge organic N
accounted for 2% of the initial N placed in the digester.
The fresh water hyacinth biomass decomposed rapidly in soil but
only 3% of the applied N was mineralized to NO^'N. Total N recovery by
sludge and effluent recycling was 48% of the initial N placed in the
digester for low N plants. The remaining N was lost from the system
during anaerobic digestion and effluent recycling.


122
resulted in low recoveries (2 to 16%). The N recovery for other
15
effluents ranged from 36 to 77%. The majority of the was recovered
in shoot material.
Plants grown in diluted effluents assimilated large amounts of Na
and K and net shoot assimilation of all nutrients was greater than net
root assimilation. Dead plants showed a net loss of K but net gains of
other nutrients.
Digested Sludge Recycling
Decomposition of fresh and anaerobically digested water hyacinth
biomass added to soil was evaluated by CO^ evolution and
mineralization. Approximately 39 and 50% of the added C was evolved as
CO^ in 90 days for fresh plant biomass with a low and high tissue N
content, respectively. Approximately 20% of the added C was evolved as
CO^ for both digested sludges in 90 days. Decomposition of fresh plant
biomass required a three stage first-order kinetic description of C loss
compared to a two stage first-order description for digested sludge.
Apparently the loss of readily-decomposable C during anaerobic digestion
eliminated the first stage of decomposition in soil.
Only 8% of the applied was mineralized to NO^-N for digested
sludges despite their differences of initial N content. In contrast, 3
and 33% of the applied was recovered as NO^-N for fresh biomass with
a low and high N content, respectively. The C/N ratio of digested
sludge was not a predictive guide for N mineralization potential.
Mixing the fresh biomass with digested sludge may enhance the
decomposition of the sludge in soil.


BIBLIOGRAPHY
Alexander, M. 1977. Introduction to Soil Microbiology. 2nd Edition.
John Wiley and Sons. New York, NY.
A.P.H.A. 1980. Standard Methods for the Examination of Water and
Wastewater. 20th Edition. Amer. Publ. Health Assoc. Washington,
DC.
Almazan, G., and C. E. Boyd. 1978. Effects of nitrogen level on rates
of 0 consumption during decay of aquatic plants. Aquat. Bot.
5:119-126.
Atalay, A., and R. W. Blanchar. 1984. Evaluation of methane generator
sludge as a soil amendment. J. Environ. Qual. 13:341-344.
Berg, B., B. Wessen, and G. Ekbohm. 1982. Nitrogen level and
decomposition in Scots pine needle litter. Oikos 38:291-296.
Bock, J. H. 1969. Productivity of the water hyacinth Eichhornia
crassipes (Mart.) Solms. Ecol. 50:460-464.
Bouldin, D. R., R. L. Johnson, C. Burda, and C. W. Kao. 1974. Losses
of inorganic nitrogen from aquatic systems. J. Environ. Qual.
3:107-114.
Boyd, C. E. 1970a. Vascular aquatic plants for mineral nutrient
removal from polluted waters. Econ. Bot. 24:95-103.
Boyd, C. E. 1970b. Losses of mineral nutrients during decomposition of
Typha latifolia. Arch. Hydrobiol. 66:511-517.
Boyd, C. E. 1976. Accumulation of dry matter, nitrogen, and phosphorus
by cultivated water hyacinths. Econ. Bot. 30:51-56.
Broadbent, F. E., and W. V. Bartholomew. 1948. The effect of quantity
i of plant material added to soil on its rate of decomposition. Soil
Sci. Soc. Am. Proc. 13:271-274.
Bryant, M. P. 1979. Microbial methane production: Theoretical
aspects. J. Animal Sci. 48:193-201.
Carpenter, S. R. 1980. Enrichment of Lake Wingra, Wisconsin, by
submersed macrophyte decay. Ecol. 61:1145-1155.
Carpenter, S. R., and M. S. Adams. 1979. Effects of nutrients and
temperature on decomposition of Myriophyllum spicatum L. in a
hard-water eutrophic lake. Limnol. Oceanogr. 24:520-528.
131


67
Shoot:root dry weight ratios of water hyacinth were higher when nutrients
were not limiting and decreased significantly when plants grew in
nutrient-poor waters (Reddy, 1984). It was assumed in the present study
that gas production, both cumulative and yields, would be greater for the
high N plants.
A mass balance of N is presented in Table 11. The organic N
content decreased after anaerobic digestion for each treatment.
Mineralization of organic N to NH+-N was the primary N transformation
occurring during digestion. The total N recovered was lower for low N
water hyacinth plants. The majority of the N was recovered in the
effluent as NH+-N. Most of the N placed in digesters was recovered in
the effluent, although the proportion of NH^-N of the total N tended to
increase (Hashimoto et al., 1980; Field et al., 1984).
Approximately 30% of the organic N placed in the digesters was
recovered as organic N in the digested sludge for both high and low N
plants. The organic N recovered in the screened effluent was 15 and 36%
of the added organic N for high and low N plants, respectively. The
total organic N recovered as effluent or sludge organic N was 45 and 66%
of added organic N for high and low N plants, respectively. Therefore,
a high N content of water hyacinth resulted in more mineralization of
added organic N.
Total recovered as ^Nh1"-N in the screened effluent was 72 + 4%
4
for high N plants compared to 35 + 9% for low N plants (Table 12). The
organic recovered in digested sludge accounted for 20 + 5% of the
added from fresh water hyacinth plants regardless of N content.
Approximately 11 and 20% of the added was recovered as organic N in
the screened effluent for digested high and low N plants, respectively.


71
Table 13. Characteristics of digester effluents before sludge removal.
Inoculum
volume
Digester
Effluent
Characteristics^
pH
EC
COD
NH*-N
4
TKN
TS
VS
FS
-i
--L--
dS
mg L
%
-% of
TS-
High N plant material
2.5
7.4
4.5
3030
189
238
0.498
40.3
59.7
5
7.6
5.1
4290
210
315
0.570
38.7
61.3
10
7.4
5.6
5050
294
406
0.610
38.9
61.1
Control
7.7
NA
4170
205
259
0.535
36.6
63.4
Low N plant material
2.5
7.3
6.6
1690
50
70
0.445
28.8
71.1
5
7.2
6.9
2340
70
112
NA
37.3
62.7
10
7.3
7.8
1640
112
154
0.488
28.8
71.2
Control
7.8
NA
109
98
120
0.315
14.0
86.0
^ COD = Chemical
oxygen
demand,
TS, FS
and VS
- Total
, fixed
and
volatile solids, respectively.
*NA = Not available.


88
Table 18. Nitrogen-15 balance for labeled effluents.
Recovered by
Inoculum Available plants N Recovery
volume 15N Roots Shoots Plants Water Unaccounted
L
mg
% of applied
Diluted effluents from high N plants
2.5
225
44
103
66
7
27
5
649
80
199
43
9
48
10
1044
101
272
36
3
61
Undiluted effluents from high N plants
2.5
1610
20
45
4
19
77
5
2120
18
41
3
27
40
10
2890
19
46
2
25
74
Undiluted <
affluents from
low N plants
2.5
236
62
119
77
6
17
5
487
71
124
40
4
56
10
869
63
73
16
6
78


93
plant dry weight were also noted for these effluents. Ammonium was >1.0
mg L ^ and TKN was reduced 83 to 97% in these effluents after 22 days.
Sodium reduction was in the range of 34 to 52%. Calcium and Mg
decreased for the 2.5 and 5 L inoculum diluted effluents but increased
for the 10 L inoculum diluted effluent.
Intermediate reductions of EC (25 + 3%), K (58 + 6%) and P (69 +
7%) were observed in undiluted effluents from digested low N plants.
Ammonium was < 1.0 mg L ^ and TKN was reduced 90 to 93% in these
effluents after 22 days. Sodium decreased 54 to 63% after 22 days. The
largest reductions of Ca (82 + 4%) and Mg (43 + 8%) were found in these
effluents.
Plant death was noted in all undiluted effluents from digested high
N plants. These treatments had the lowest reductions of EC (5 + 3%) and
P (26 + 10%). Plants were removed after 10 days following complete
death of aerial tissue and separation of root masses. The death of
plant tissue resulted in an increase of K and Mg in the effluents.
Sodium was reduced 18 to 23%. Calcium reduction was in the range of 52
to 70%. The loss of NH^-N (83 to 86% reduction) was probably due to NH^
volatilization and microbial assimilation.
Conclusions
The highest dry weight gains were for plants growing in diluted
effluents with EC levels ranging from 0.7 to 2.3 dS m ^ and NH")"-N
4
concentrations of 23 to 104 mg L Plants survived and grew in two
undiluted effluents with EC levels of 5.6 and 5.9 dS m'1 and Nh|-N
A
concentrations of 24 and 49 mg L_1. All other EC and NH+-N combinations
A
of undiluted effluents resulted in plant death.


TIME (days)
Figure 11. The pH of sediment-water-plant systems with added nitrate.


66
was over twice as great for the digester receiving the largest amount of
inoculum. However, for low N plants, the amount of inoculum did not
appreciably affect cumulative biogas production during digestion.
Cumulative biogas production at 60 days was similar for both high
and low N plants. Biogas production at 15 days was generally greater
for high N plants. It appeared that N was not a limiting factor for
total gas production in either digestion test.
Converting 60 day biogas production to biogas or methane yields (L
g ^ VS added) is also presented in Table 10. Volatile solids included
inputs from water hyacinths and inoculum. The average methane content
of the biogas was 63.7 + 5.2 % based on 18 samples. Surprisingly, biogas
and methane yields were higher for the low N plants. This was caused by
an increase of VS from inoculum used in digesters for high N plants. The
inoculum used for high N plants contained 1.75% TS (66.5% VS of TS)
(Table 9). The inoculum for low N plants contained 0.29% TS (16.9% VS
of TS). Gas production expressed in these units suggested that inoculum
volume did not appreciably affect total biogas or methane yields.
The average methane yields were 0.14 and 0.16 L g ^ VS added for
high and low N water hyacinth plants, respectively. The methane yields
were lower than those reported for continuously-fed digesters (Hanisak et
al., 1980; Chynoweth et al., 1983). Batch digestion (once fed and
t
sealed) would not promote maximum gas yields as frequent addition of
fresh substrate enhances gas production (Price and Cheremisinoff, 1981).
Shiralipour and Smith (1984) reported that methane production for
water hyacinth roots was lower than for shoots and that increasing N in
water hyacinth growth media increased methane yields. Water hyacinths
typically produced longer roots as water fertility declined (see p. 32).


17
Anaerobically digested sludge was considered to be stable and resist
further decomposition (Sommers, 1977).
Nitrogen Cycling in Soil Treated with Plant Residues
The land application of fresh or anaerobically digested plant
biomass has significant implications on N cycling. Nitrogen
transformations occurring after residue additions include 1)
mineralization/immobilization; 2) microbial or plant assimilation; 3)
nitrification; 4) denitrification; and 5) NH^-N volatilization (Fig. 5).
Since most of the N in fresh or digested plant biomass is in
organic forms, the rate of mineralization becomes the rate limiting step
for all transformations that follow. Mineralization or immobilization
depends on the initial N concentration of the plant biomass as well as
the composition of the C constituents. A low N content or a wide C/N
ratio was associated with slow decomposition and rates of decomposition
were proportional to lignin content (Alexander, 1977). A wide C/N ratio
(> 30:1) favored N immobilization whereas a narrower ratio (< 20:1)
resulted in N mineralization (Alexander, 1977).
Mineralization of N from anaerobically digested sewage sludges was
reported to be affected by the rate of application (Ryan et al., 1973;
Stark and Clapp, 1980). However, Epstein et al. (1978) found that
irrespective of the amount of material (sewage sludge and sludge
|
compost) applied, the percentage of added N mineralized remained
essentially constant.
The mineralization of NH+-N from organic N is accompanied by
microbial assimilation or plant uptake. In aerobic environments the
NH^-N was quickly converted to NC^-N (nitrification) which could also be


PERCENT C REMAINING
TIME (days)
Figure 17. Decomposition stages and rate constants
of fresh and digested plant biomass added
to soil


89
Table 19. Distribution of nutrients in water hyacinth shoots
and roots in diluted or undiluted effluents of
digested high N plants.
.Inoculum
volume Na K P Ca Mg
--L-- g kg
SHOOTS
Diluted effluents from high N plants
2.5
13.3
13.3
2.4
16.9
9.2
5
17.5
19.3
3.9
17.8
9.1
10
18.0
23.2
5.0
15.6
8.6
Undiluted effluents
from high N plants
2.5
21.0
18.0
3.5
18.2
5.6
5
20.5
17.7
3.9
22.6
6.2
10
19.2
14.5
3.8
19.3
5.7
LSD (0.05)
4.4
7.1
1.3
3.7
1.9
Original shoot tissue
2.8
22.0
2.7
15.4
5.7
ROOTS
Diluted effluents
from high N
plants
2.5
14.5
7.1
2.1
6.1
4.6
5
17.8
8.9
2.6
6.0
4.7
10
16.0
12.4
3.4
9.0
4.2
Undiluted effluents
from high N plants
2.5
16.2
3.1
11.2
26.5
4.6
5
16.8
3.3
10.3
23.6
4.3
10
17.2
3.3
10.2
24.6
4.1
LSD (0.05)
3.0
3.7
1.9
3.0
0.7
Original root tissue
4.6
5.5
2.8
6.6
2.6


128
Table 30. Mehlich I extractable constituents at day 0 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant biomass
constituent
Control
Fresh
Digested
Fresh
Digested
- -mg kg
Calcium
257 cf
377 a
344 ab
332 b
351 ab
Potassium
27 d
128 a
44 c
127 a
53 b
Magnesium
25 d
55 a
34 c
37 b
33 c
Sodium
7 e
60 c
79 b
56 d
133 a
Iron
18 d
25 b
35 a
21 c
26 b
Zinc
4.1 d
7.3 b
13.3 a
6.1 c
7.2 b
^Values with same letter within rows are not significantly different at
0.05 level by Duncan's Multiple Range Test.


Organic
Carbon
Organic
Nitrogen
/
t
/ /
tv////
\//
Organic
Carbon
/
/
Organic
Nitrogen
2. ,
/ y
/
t /
N TRANSFORMATIONS
1. Rant Uptake
2. Mineralization /
Immobilization
3. Nitrification
4. Denitrification
5. NH^ Volatilization
Figure 3. Nitrogen cycling in a water hyacinth production system.
Nitrogen transformations investigated during this study
are indicated with larger arrows.


19
assimilated by microbes or plants (Ryan et al., 1973). A high organic
loading rate may result in 0^ depletion during decomposition which
promotes the denitrification process (Epstein et al., 1978; Hsieh et
al., 1981b).
Decomposition of Plant Residues in Soil
Decomposition of plant residues in soil occurs in two stages. The
first stage was attributed to loss of the easily decomposable labile
fraction which was followed by the second stage of slow decomposition of
a resistant residual fraction (Shields and Paul, 1973; Reddy et al.,
1980). Both stages were thought to be controlled by two simultaneously
occurring superimposed first-order kinetic reactions (Sinha et al.,
1977).
Fresh and anaerobically digested plant biomass differ widely in
their chemical composition. Anaerobic digestion converts most of the
easily-decomposable plant C constituents into CH^ and CC^. The digested
biomass sludge has a higher lignin content and is more resistant to
decomposition. There is little information available on decomposition
of anaerobically digested plant biomass added to soil. However, the
rates and the factors which influence decomposition of fresh plant
biomass added to soil have been well-established.
Tenny and Waksman (1929) concluded that water-soluble organic
substances were first to be decomposed in the soil, followed by
hemicellulose and at the same time, or immediately after, cellulose.
Lignin was very resistant to decomposition and may even delay the
disintegration of cellulose or hemicellulose because of the structural
proximity of these C constituents in the cell wall (Tenny and Waksman,
1929; Peevey and Norman, 1948; Berg et al., 1982).


10
8
6
4
2
10
8
6
4
2
v-
Planta
S#d¡mnt
1 .1 1.1 i
) 4 8
-Tt+-~r v '
i i I -i, l I ,I.J
12 16 20 24 28
TIME (days)
12. The pH of sediment-water-plant systems with added ammonium.


8
The potential productivity and nutrient removal capacities of water
hyacinth has led to its selection as a biomass feedstock for methane
generation while providing a means for treatment of nutrient-enriched
waters. Extensive research, both in laboratory and field applications,
was conducted on the use of water hyacinth in wastewater treatment
during the past 20 years (Sheffield, 1967; Boyd, 1970a; Steward, 1970;
Scarsbrook and Davis, 1970; Rogers and Davis, 1971; Dunigan et al.,
1975; Cornwell et al., 1977; McDonald and Woverton, 1980; Reddy et al.,
1982; DeBusk et al., 1983). Water hyacinth was shown to be effective in
removing N, P and other nutrients, and reducing biological oxygen demand
and total suspended solids.' Water hyacinth was also shown to readily
absorb and concentrate heavy metals (Wolverton and McDonald, 1975a,b;
Cooley et al., 1978).
Nitrogen Cycling in the Water Hyacinth Production System
Nitrogen transformations occurring in a water hyacinth production
system include 1) plant uptake; 2) mineralization/immobilization; 3)
nitrification; 4) denitrification; and 5) NH^-N volatilization (Fig. 3).
Plant uptake is one of the major processes for N removal from water
hyacinth-based wastewater systems. Plant uptake is directly related to
the growth rate and the nutrient composition of the water. Water
hyacinth was more efficient in utilizing NH+-N than NO^-N when both
forms were supplied in equal proportions (Reddy and Tucker, 1983).
A dense cover of floating water hyacinths will regulate dissolved
0^, temperature and pH of water which influences several N
transformations. Generally, diel fluctuations of these water parameters
were reported to be lower in areas covered with water hyacinths compared
to open areas (Rai and Munshi, 1979; McDonald and Wolverton, 1980;
Reddy, 1981).


27
Figure 6. Weekly averages of daily temperatures and solar
radiation.


140
Toerien, D. F., and W. H. J. Hattingh. 1969. Anaerobic digestion. I.
The microbiology of anaerobic digestion. Water Res. 3:385-416.
Wolverton, B. C., and R. C. McDonald. 1975a. Water hyacinths and
alligator weeds for removal of lead and mercury from polluted
waters. NASA Tech. Memo. No. TM-X-72723. Natl. Space Technol.
Lab., Louis, MS.
Wolverton, B. C., and R. C. McDonald. 1975b. Water hyacinths and
alligator weeds for removal of silver, cobalt, and strontium from
polluted waters. NASA Tech. Memo. No. TM-X-72727. Natl. Space
Technol. Lab., Louis, MS.
Wolverton, B. C., and R. C. McDonald. 1979. Energy from aquatic plant
wastewater treatment systems. NASA/NSTL. Tech. Memorandum
TM-X-72733. Natl. Space Technol. Lab., Louis, MS.
Wolverton, B. C., and R. C. McDonald. 1981. Energy from vascular plant
wastewater treatment systems. Econ. Bot. 35:224-232.
Yount, J. L,, and R. Crossman. 1970. Eutrophication control by plant
harvesting. J. Water Poll. Contr. Fed. 42:173-183.
Zeikus, J. G. 1977. The biology of methanogenic bacteria. Bacteriol.
Rev. 41:514-541.


57
Table 8. Mass
balance
of added
^NH^-N in sediment-water
4
-plant systems.
-
Plant
Sediment:^
Unaccounted
Treatment
or algae
Detritus
Org Inorg
Water
Total
For
/o IVU v w 1. y ui
PLANTS
Without sediment
0 mg C L_1
82.2
7.3
2.3
91.8
8.2
100 mg C L_1
69.6
3.6
5.8
79.0
21.0
400 mg C L_1
80.7
7.1
3.8
91.6
8.4
With sediment
0 mg C L-1
89.1
ND §
4.3 2.6
1.0
97.0
3.0
100 mg C L"1
84.5
1.7
3.0 2.9
1.5
93.6
6.4
400 mg C L_1
82.8
3.9
3.3 3.5
1.5
95.0
5.0
NO PLANTS
Without sediment
0 mg C L_1
ND
ND
9.6
9.6
90.4
100 mg C L_1
ND
14.7
17.5
32.2
67.8
400 mg C L"1
6.1
34.7
5.2
46.0
54.0
With sediment
0 mg C L_1
14.9
ND
10.2 7.1
2.3
34.5
65.5
100 mg C L'1
15.0
5.9
11.0 8.8
5.8
46.5
53.5
400 mg C L_1
11.1
26.2
3.9 6.9
3.8
51. 9
48.1
t

§
Carbon source was plant detritus.
Org, Inorg = organic and inorganic N,
ND = Not detectable.
respectively.


7
Water hyacinth adapted to light intensity and full sunlight elicted
the greatest photosynthetic rate (Patterson and Duke, 1979). Optimum
plant density to obtain maximum biomass yield varied with season and
available plant nutrients in the water (Reddy and Sutton, 1984). DeBusk
et al. (1981) and Reddy et al. (1983) established that optimum plant
density for achieving maximum growth cultured in wastewaters was in the
-2
range of 15 to 35 kg wet wt m
Water hyacinth productivity has been evaluated in natural and
-2 -1
nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day
were reported for plants growing in natural waters of central and south
Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide-range
-2 -1
of productivity (5 to 42 g dry wt m day ) was recorded for plants
cultured in nutrient-enriched waters (Schwegler and Kim, 1981; Hanisak
et al., 1980). Reddy and DeBusk (1984) obtained an average of 52 and a
-2 -1
maximum of 64 g dry wt m day for water hyacinths grown in
nutrient-nonlimiting conditions.
The effectiveness of water hyacinth in removing inorganic N was
reported for several nutrient-enriched wastewaters. Sheffield (1967)
and Clock (1968) reported a 75 to 94% reduction of inorganic N from
secondary sewage effluent in systems containing water hyacinths. Reddy
et al. (1982) observed a 78 to 81% reduction of inorganic N from organic
soil drainage water containing water hyacinths. Hanisak et al. (1980)
concluded that 65% of N in digester effluents could be assimilated when
water hyacinths were grown in diluted effluents. Boyd (1976) calculated
average rates of N and P removal were 3.4 and 0.43 kg ha1 day1 in
fertilized fish ponds. Rogers and Davis (1972) concluded that water
hyacinth removal capacities were less effective with increasing nutrient
concentrations.


135
Jenkinson, D. S. 1971. Studies on the decomposition of C labelled
organic matter in soil. Soil Sci. 111:64-70.
Keeney, D. R. 1973. The nitrogen cycle in sediment-water- systems.
J. Environ. Qual. 2:15-29.
Keeney, D. R., and D. W. Nelson. 1982. Nitrogen-Inorganic Forms. In
Page, A. L. (ed.). Methods of Soil Analysis. Part 2. Second
Edition. Agronomy 9. ASA, Madison, WI.
Kirkby, E. A., and K. Mengel. 1967. The ionic balance in different
tissues of the tomato plant in relation to nitrate, urea, or
ammonium nutrition. Plant Physiol. 42:6-14.
Knipling, E. B., S. H. West, and W. T. Haller. 1970. Growth
characteristics, yield potential, and nutritive content of water
hyacinths. Soil Crop Sci. Soc. Fla. 30:51-63.
Mah, R. A., D. M. Ward, L. Baresi, and T. L. Glass. 1977. Biogenesis
of methane. Ann. Rev. Microbiol. 31:309-341.
Manson, J. G., and B. E. Manson. 1958. Water hyacinth reproduces by
seed in New Zealand. New Zealand J. Agrie. 96:191.
McDonald, R. C., and B. C. Wolverton. 1980. Comparative study of
wastewater lagoon with and without water hyacinth. Econ. Bot.
34:101-110.
Mehlich, A. 1953. Determination of P, Ca, Mg, K, Na, and NH^. North
Carolina Soil Test Division (mimeo. 1953), Raleigh, NC.
Melillo, J. M., J. D. Aber, and J. F. Muratore. 1982. Nitrogen and
lignin control of hardwood leaf litter decomposition dynamics.
Ecol. 63:621-626.
Mengel, K. 1974. Plant ionic status. In Carson, E. W. (ed.). The
Plant Root and its Environment. Univ. Press of Virginia,
Charlottesville, VA.
Miller, R. D., and D. D. Johnson. 1964. The effect of soil moisture
tension on carbon dioxide evolution, nitrification, and nitrogen
mineralization. Soil Sci. Soc. Am, Proc. 28:644-647.
Miller, R. H. 1974. Factors affecting the decomposition of an
anaerobically digested sewage sludge in soil. J. Environ. Qual.
3:376-380.
Moore, J., and K. Bjorndal. 1984. Unpublished results. Univ. of
Florida, Gainesville, FL.
Nelson, D. W. 1982. Gaseous losses of nitrogen other than through
denitrification. In Stevenson, F. J. (ed.). Nitrogen in
Agricultural Soils. Agronomy 22. ASA, Madison, WI.


132
Chynoweth, D. P., D. A. Dolene, B. Schwegler, and K. R. Reddy. 1983.
Wastewater reclamation and methane production using water hyacinth
and anaerobic digestion. Presented at 10th Energy Technology
Conference. Washington, DC.
Clock, R. M. 1968. Removal of nitrogen and phosphorus from secondary
sewage treatment effluent. Ph.D. dissertation. Univ. of Florida.
Cooley, T. N., D. F. Martin, W. C. Dunden, Jr., and B. D. Perkins.
1979. A preliminary study of metal distribution in three water
hyacinth biotypes. Water Res. 13:343-348.
Cornwell, D. A., J. Zoltek, Jr., C. D. Patrinely, T. deS. Furman, and J.
I. Kim. 1977. Nutrient removal by water hyacinth. J. Water Poll.
Contr. Fed. 49:57-65.
Davis, C. B., and A. G. van der Valk. 1978. The decomposition of
standing and fallen litter of Typha glauca and Scirpus fluviatilis.
Can. J. Bot. 56:662-675.
DeBusk, T. A., J. H. Ryther, M. D. Hanisak, and L. D. Williams. 1981.
Effects of seasonality and plant density on the productivity of
some freshwater macrophytes. J. Environ. Qual. 10:133-142.
DeBusk, T. A., L. D. Williams, and J. H. Ryther. 1983. Removal of
nitrogen and phosphorus from wastewater in a water hyacinth-based
treatment system. J. Environ. Qual. 12:257-262.
De La Cruz, A. A., and B. C. Gabriel. 1974. Caloric, elemental, and
nutritive changes in decomposing Juncus roemerianus leaves. Ecol.
55:882-886.
Dinges, R. 1978. Upgrading stabilization pond effluent by water
hyacinth culture. J. Water Poll. Contr. Fed. 50:833-845.
Dunigan, E. P., R. A. Phelan, and Z. H. Shamsuddin. 1975. Use of water
hyacinths to remove N and P from eutrophic waters. Hyacinth Contr.
J. 13:59-61.
Engler, R. M., and W. H. Patrick, Jr. 1974. Nitrate removal from
floodwater overlying flooded soils and sediments. J. Environ.
Qual. 3:409-413.
Epstein, E., J. M. Taylor, and R. L. Chaney. 1976. Effects of sewage
sludge and sludge compost applied to soil on some soil physical and
chemical properties. J. Environ. Qual. 5:422-426.
Epstein, E., D. B. Keane, J. J. Meisinger, and J. 0. Legg. 1978.
Mineralization of nitrogen from sewage sludge and sludge compost.
J. Environ. Qual. 7:217-221.
Fenchel, T. 1970. Studies on the decomposition of organic detritus
derived from the turtle grass Thalassia testudinum. Limnol.
Oceanogr. 15:14-20.


38
to 81% of the agricultural drainage effluent NO^-N and NH^-N in 3.6 days
in a reservoir containing water hyacinths. DeBusk et al. (1983)
calculated that 45% of the N removed from wastewater was immobilized in
water hyacinth standing crop and detritus. About 30% of the fertilizer
N was added as NO^-N, which could be potentially lost due to
denitrification. Since water hyacinth plants prefer NH+-N over NO^-N
(Reddy and Tucker, 1983), the majority of the plant N uptake probably
came from NH*-N added through fertilizer. The role of underlying
sediment in the immobilization/mineralization, and denitrification of N
from these systems needs further investigation.
Total N recovery in the fertilized reservoir was about 90%, and
plant uptake represented about 71% of total N inputs. In the control
reservoir, total N recovery was higher than the N inputs. Plants
removed 325 kg N ha ^, as compared to 238 kg N ha ^ added. Release of N
from sediment or mineralization of N during decomposition of detritus
may account for the higher N recovery compared to total N inputs.
Ogwada (1983) found a yearly average of 150 + 34 kg KCl-extractable
inorganic N ha ^ sediment using monthly sediment N concentrations of the
same reservoirs.
Conclusions
Primary productivity of water hyacinths was influenced by ambient
air temperature, solar radiation, and nutrient composition of the
culture medium. Net detritus production (total detritus detritus lost
through decomposition) was relatively constant throughout the year and
represented 3.5 to 14.0% of the total standing crop. Detritus plant
tissue of the fertilized reservoir contained higher tissue N, compared


Table 29. Soil ammonium concentrations from added fresh and digested
plant biomass.
Day
Control
Low N plant biomass
High N plant biomass
Fresh
Digested
Fresh
Digested
-1
0
1.2 b t
2.2 b
1.7 b
25.1 a
1.9 b
30
1.2 a
1.4 a
1.8 a
1.8 a
1.3 a
60
1.1 a
1.2 a
1.7 a
1.0 a
1.0 a
90
1.0 a
1.0 a
1.0 a
1.0 a
1.0 d
^ Values
with same letter
within rows
are not significantly different at
0.05 level by Duncan's Multiple Range Test.
127


139
Sheffield, C. W. 1967. Water hyacinth for nutrient removal. Hyacinth
Contr. J. 6:27-30.
14
Shields, J. A., and E. A. Paul. 1973. Decomposition of C-labelled
plant material under field conditions. Can. J. Soil Sci.
53:297-306.
Shir^lipour, A., and P. H. Smith. 1984. Conversion of biomass into
methane gas. Biomass 6:85-92.
Sinha, M. K., D. P. Sinha, and H. Sinha. 1977. Organic matter
transformations in sils: V. Kinetics of carbon and nitrogen
mineralization in soils amended with different organic materials.
Plant Soil 46:579-590.
Sommers, L. E. 1977. Chemical composition of sewage sludges and
analysis of their potential use as fertilizer. J. Environ. Qual.
6:225-232.
Sompongse, D. 1982. The role of wetland soils in nitrogen and
phosphorus removal from agricultural drainage water. Ph.D.
dissertation. Univ. of Florida.
Stack, C., P. Lichtenberger, and J. Martin. 1981. Economic and
environmental consequences of anaerobic digestion of animal wastes.
In Preprints of the 1981 International Gas Research Conference.
Stark, S. A., and C. E. Clapp. 1980. Residual nitrogen availability
from soils treated with sewage sludge in a field experiment. J.
Environ. Qual. 9:505-512.
Steward, K. K. 1970. Nutrient removal potentials of various aquatic
plants. Hyacinth Contr. J. 8:34-35.
Tenny, F. G., and S. A. Waksman. 1929. Composition of natural organic
materials and their decomposition in the soil. IV. The nature and
rapidity of decomposition on the various organic complexes in
different plant materials, under aerobic conditions. Soil Sci.
28:55-84.
Terry, R. E., D. W. Nelson, and L. E. Sommers. 1979. Carbon cycling
during sewage sludge decomposition in soils. Soil Sci. Soc. Am. J.
43:494-499.
Terry, R. E., D. W. Nelson, and L. E. Sommers. 1981. Nitrogen
transformations in sewage sludge-amended soils as affected by soil
environmental factors. Soil Sci. Soc. Am. J. 45:506-513.
Tester, C. F., L. J. Sikora, J. M. Taylor, and J. F. Parr. 1977.
Decomposition of sewage sludge in soil: I. Carbon and nitrogen
transformations. J. Environ. Qual. 6:459-463.


APPENDIX A
DIGESTER EFFLUENT CHARACTERISTICS DURING WATER HYACINTH TREATMENT


62
The digesters received A.7 kg- fresh weight of the N labeled water
hyacinths and an inoculum volume of 2.5, 5 or 10 L. A control digester
received 10 L of inoculum and no plant material. The inoculum used for
plants with high N content was obtained from an operating continuous-fed
upflow digester receiving a feedstock of water hyacinth and domestic
sewage sludge in a blend ratio of 3:1 (Chynoweth et al., 1983). The
inoculum used for the plants with low N content was obtained from a
non-operating continuously-fed tank digester receiving water hyacinth as
feedstock. Each digester was buffered with 210 g NaHCO^ and tap water
was used to bring each batch digester to 54.7 kg.
Gas production was monitored for 60 days. At the end of the
digestion period, each digester was thoroughly mixed and the total
contents were emptied into a 60 L tub. The digested materials were
passed through a 1.00 mm fiberglass screen into a second 60 L tub to
separate the digested biomass sludge from the effluent. The sludge was
drained for 7 minutes and transferred into a polyethylene bag and placed
directly into a freezer. The effluent was transferred to a water
hyacinth production system.
The liquid samples from the digester effluents and screened
effluents (sludge removed) were analyzed for pH, electrical conductivity
(EC), total solids (TS), fixed solids (FS), volatile solids (VS) (APHA,
1980), total Kjeldahl N (TKN) (Nelson and Sommers, 1975), NH^-N and NO^-N
by steam distillation (Keeney and Nelson, 1982), and chemical oxygen
demand (COD) (APHA, 1980). The screened effluent was also filtered
through a 0.2 jum membrane filter and analyzed for Ca, K, Na and Mg by
atomic absorption and P by an autoanalyzer.


Table 16. Initial characteristics of the digester effluents and nutrient medium.
Inoculum
volume
pH
EC
NH+-N
4
TKN
Na
K
P
Ca
Mg
T-1
i_j
QO
mg u
Diluted effluents from high N plants
2.5
7.8
0.7
23
26
166
20
2.8
24
17
5
7.7
1.6
65
72
360
44
5.5
18
18
10
7.7
2.3
104
124
450
62
4.7
13
13
Undiluted effluents from high N plants
2.5
7.6
4.3
161
188
1140
82
12.8
32
13
5
7.6
4.7
212
256
1160
80
12.8
30
12
10
7.6
5.3
289
326
1160
123
11.5
23
14
Undiluted effluents from low N plants
2.5
7.5
5.6
24
45
1220
195
3.1
147
50
5
7.4
5.9
49
72
1240
245
4.6
53
53
10
7.5
6.7
87
111
1200
325
4.5
61
63
Nutrient medium
7.5
0.7
20
21
11
24
3.6
20
5


EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER HYACINTH SYSTEMS
Plant detritus (dead and decaying plant debris) is an integral part
of water hyacinth mats and comprises 3 to 14% of the total biomass (see
p. 39). It is usually derived from natural aging of plants, biological
or chemical control, and frost damage. The addition of detritus to an
aquatic system influenced several C and N transformations (Fenchel and
Jorgenson, 1977).
Nitrogen is present as NH*-N, NO^-N, and organic N in water media
available for water hyacinth production. Organic N predominates in most
water media and is not readily available for plant assimilation. Water
hyacinths were efficient users of inorganic N and plant assimilation was
a major process of N removal in aquatic systems containing water
hyacinth (Reddy and Sutton, 1984). Other N transformations in aquatic
systems resulting in removal of NO^-N or NH+-N include microbial
assimilation, nitrification/denitrification, and NH^-N volatilization
(Keeney, 1973; Bouldin et al., 1974). Addition of detritus
significantly alters the rates of these processes.
Mineralization or immobilization of N occurs during decomposition
of detritus in water and sediment. Decomposition of detritus and
subsequent N release was found to be related to C/N ratio, initial N and
fiber contents (De La Cruz and Gabriel, 1974; Godshalk and Wetzel,
1978b; Odum and Heywood, 1978; Ogwada et al., 1984).
40


96
and Clapp (1980) observed an enhanced rate of N mineralization with
increasing rate of sewage sludge application. Nitrogen mineralization
potential was found to be 30 and 38% of organic N in activated and
digested sludge, respectively, during 60 days of incubation (Hsieh et al.
1981b).
Much of the available information dealt with land application of
anaerobically digested sewage sludge (Miller, 1974; Terry et al., 1979),
and limited data was reported on the decomposition of sludge obtained
from the anaerobic digestion of plant biomass (Atalay and Blanchar,
1984). The objective of this study was to evaluate the decomposition and
N mineralization rates of anaerobically digested plant biomass added to
soil. Four materials were evaluated: fresh plant biomass with a low or
high tissue N content, and their respective anerobically digested
residues.
Materials and Methods
Surface (0-15 cm depth) soil samples of a Kendrick fine sand
(Arenic paleudult) were collected at the Agronomy Farm, University of
Florida in Gainesville, Florida. The soil was air-dried and passed
through a 2 mm sieve. The soil had a particle size distribution of
92.9% sand, 4.6% silt, and 2.5% clay. The CEC was 3.44 cmol(+) kg *
soil with a base saturation of 47%.
Water hyacinths with low (-10 g N kg 1 dry plant tissue) and high
(-34 g N kg ^) tissue N content were grown in nutrient-depleted water and
sewage effluent, respectively. After removal from their respective
growth media, the hyacinths were grown in 15N labeled (NH,)S0. nutrient
4 2 4
solution for two weeks, frozen, and chopped to 1.6 mm length using a
Hobart T 215 food processor.


Figure 16. Carbon evolution from soil applied fresh
and digested plant biomass.


82
Productivity of Water Hyacinths
Total dry weight gains of water hyacinths were consistently less in
the undiluted effluents (Fig 15). Complete death of plants was observed
in four undiluted effluents. The loss of dry weight for these
treatments was probably due to leaching of soluble plant constituents
after plant death.
The highest dry weight gains were associated with the lowest EC.
However, plants survived in undiluted effluents having EC levels of 5.6
and 5.9 dS m ^ (5600 and 5900 pmhos cm ^). These EC levels were
equivalent to -2900 and 3200 mg NaCl L ^ and were higher salt
concentrations reported for water hyacinth survival (Penfound and Earle,
1948; Haller et al., 1974). The undiluted effluents had Na levels in
excess of 1100 mg L ^. Apparently water hyacinth has a wide range of
adaptability to media composition and, therefore, total salt
concentration is not a good criterium for determining plant survival.
The diluted effluents were excellent media for plant growth and the
gains in dry weight were consistently higher compared to the nutrient
medium (Fig. 15). The highest dry weight gain was in the diluted
effluent having N and P concentrations of 65 and 5.5 mg L
respectively, and a N/P ratio of 11.8:1. Sato and Kondo (1981) reported
maximum yields of water hyacinth at a N and P concentration of 50 and
13.8 mg L 1, respectively, and a N/P ratio of 3.7:1. Dry weight gains
were noted for two of the undiluted effluents. However, tissue damage
was noted in all undiluted effluents.
Tissue damage in undiluted effluents was observed within 24 hr
after study initiation. Two types of leaf tissue damage were observed.
Damaged leaves on younger shoots had burnt (brown) tips which curled up


~ 20.0
CD
7.5
CD I 5.0
12.5
10.0
h-
7.5
f
¡Z
<
_J
CL
5.0
2.5
Fertilized Control Fertilized Control
AUTUMN WINTER
S R D S R D
Fertilized Control
SPRING
Figure 8. Seasonal plant tissue nitrogen content.
Fertilized Control
SUMMER
u>
OJ


Table 2. Seasonal water hyacinth shoot and root lengths.
Fertilized reservoir Control reservoir
Season Shoots Roots Shoots Roots
cm
Autumn (78)
39.6
21.5
35.7
22.9
Winter (90)
25.2
14.2
23.3
20.2
Spring (84)
26.6
9.9
18.8
14.8
Summer (88)
54.9
21.3
27.8
27.1
Test of significance *
Shoot
length
Root length
Season
**
**
Month (Season)
NS
**
Reservoir
*
**
t Number of days in Season.
Significant at 0.05 (*) or 0.01 (**) level, or not significant
(NS).


NITRATE NITROGEN (mg
TIME (days)
Figure 13. Nitrogen loss from sediment-water-plant systems with added nitrate.


APPENDIX B
SOIL CHARACTERISTICS FROM ADDED FRESH AND
ANAEROBICALLY DIGESTED PLANT BIOMASS


I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
A. ^Graet'z ,'^Chadm^^^
Associate Professor of Soil Science
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
K. R. Reddy, CocKairman
Professor of Soil Science
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
J. G. A. Fiskell
Professor of Soil Science


100
the high N fresh biomass due to a larger rooting mass associated with
plants grown in nutrient-depleted water. The low N fresh and digested
plant biomass contained more hemicellulose than high N materials.
Cellulose content decreased for low N fresh plant biomass after
digestion, but increased for high N fresh plant biomass.
Anaerobic digestion resulted in a large loss of K from fresh plant
biomass. Potassium is relatively soluble and is used for translocation
of anions via the zylem and phloem, enzyme activation, and stomatal
movements. Additional K may be required to enhance the microbial
degradation of digested biomass sludges when added to soil. The increase
in Na concentration in the digested sludges was due to the addition of
NaHCO^ buffer to stabilize digester pH. The digestion process resulted
in increases in the relative amounts of Ca, Na, Fe, and Zn in the sludge
compared to fresh plant biomass.
Carbon and Nitrogen Mineralization
Carbon evolution from the four materials, reported as mg C evolved
as CO^ per g residue C added, is shown in Fig. 16. The soils with fresh
plant biomass additions always released more CO^ compared to that from
digested biomass sludges. The two sludges showed similar evolution rates
during the first 40 days of incubation despite their difference in TKN.
After 40 days, the CO^ evolution of high N digested sludge increased and
i
continued to increase for the remaining incubation period. This suggests
a possible lag period during which the microbial population is adjusting
to those species which tolerate high levels of Na. However, there were
no significant differences in CO^ evolution from the two sludges
throughout the incubation period (Duncan's multiple range test).
Overall C decomposition cannot be described by simple kinetic
equations. However, first-order kinetics describe plant residue or


TABLE OF CONTENTS
PaSe
ACKNOWLEDGEMENTS . ii
LIST OF TABLES v
LIST OF FIGURES vii
ABSTRACT ix
INTRODUCTION 1
LITERATURE REVIEW A
Water Hyacinth Biomass Production A
Anaerobic Digestion 13
Waste By-Product Recycling 16
Conclusions 21
WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION 23
Materials and Methods 2A
Results and Discussion 26
Conclusions 38
EFFECT OF DETRITUS ON NITROGEN TRANSFORMATIONS IN WATER
HYACINTH SYSTEMS AO
Materials and Methods A2
Results and Discussion AA
Conclusions 58
ANAEROBIC DIGESTION OF WATER HYACINTH 60
Materials and Methods 61
Results and Discussion 63
Conclusions 7A
TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTH 76
Materials and Methods 78
Results and Discussion 80
Conclusions 93
iii


124
Table 27. Effluent pH during water hyacinth treatment.
Inoculum
volume
Day
0
2
4
6
8
10
12
14
16
18
20
'22
T
--pH
Diluted effluents from high N plants
2.5
7.8
7.7
7.7
7.6
7.5
7.3
7.5
7.2
7.4
7.3
7.5
7.2
5
7.7
7.7
7.9
7.8
7.8
7.8
7.6
7.4
7.6
7.4
7.5
7.5
10
7.7
8.0
8.1
7.9
7.9
7.6
7.7
7.4
7.8
7.6
7.7
7.5
Undiluted
effluents from high N plants
2.5
7.6
8.1
8.3
8.4
8.7
8.4
8.0
8.0
8.3
8.2
8.3
8.3
5
7.6
8.0
8.2
8.2
8.3
8.6
8.7
8.3
8.8
8.8
9.0
8.8
10
7.6
8.0
8.2
8.2
8.2
8.4
8.3
8.0
8.3
8.5
8.8
8.8
Undiluted
effluents from
low N
plants
2.5
7.5
8.1
8.3
8.4
8.5
NAt
NA
8.8
8.8
8.7
8.8
8.8
5
7.4
8.2
8.3
8.5
8.6
NA
NA
9.2
9.1
9.2
9.2
9.1
10
7.5
8.1
8.3
8.5
8.6
NA
NA
9.3
9.3
NA
NA
NA
Nutrient medium
7.5
6.4
4.8
3.9
3.6
NA
NA
3.3
3.5
3.4
3.5
3.5
t NA = Not available.


26
Sommers, 1973). Solar radiation and high and low daily temperatures
were recorded. The results were statistically analyzed for a randomized
block design with the fertilized and control reservoirs as treatments.
Results and Discussion
The weekly averages of daily maximum and minimum air temperatures
and solar radiation are shown in Fig. 6. Maximum temperatures ranged
from 21.9C during January to March and 36.5C during July to September.
Minimum temperatures for these time periods were 8.2C and 20.3C,
respectively. Maximum and minimum temperatures for the rest of the year
were similar (high= 30C, low= 13.5C). Maximum and minimum solar
radiation occurred from April to September and from November to March,
respectively.
The monthly averages of daily primary productivity and detritus
production of water hyacinth are presented in Fig. 7. Maximum daily
water hyacinth productivity during this study was observed in August for
_2
the fertilized reservoir (28.3 g dry wt m day ) compared to June for
the control reservoir (14.7 g dry wt m day ). Detritus production
remained fairly consistent with time for both reservoirs. Detritus
production in the fertilized reservoir increased noticeably in September
when plant productivity began to decline. The average daily detritus
-2 -1
production in the fertilized reservoir was 3.7 g dry wt m day
-2 -1
compared to 3.5 g dry wt m day in the control reservoir. DeBusk et
al. (1983) found that detritus production occurred at a relatively
constant rate regardless of harvested or nonharvested conditions.
Monthly data for the plant parameters have been summarized by
seasons: 1) autumn (October, November, and December); 2) winter


129
Table 31. Mehlich I extractable constituents at day 30 from added
fresh and digested plant biomass.
Chemical
Low N plant biomass
High N plant
biomass
constituent
Control
Fresh
Digested
Fresh Digested
-l
- mg kg
Calcium
245 ct
396 a
383 a
340 b
379 a
Potassium
39 c
139 a
55 c
124 b
49 c
Magnesium
26 c
68 a
39 b
39 b
34 be
Sodium
3 e
57 c
75 b
41 d
114 a
Iron
17 c
18 b
25 a
15 d
18 b
Zinc
4.1 d
6.1 c
11.9 a
6.1 c
7.7 b
^Values with same letter within rows are not significantly different at
0.05 level by Duncan's Multiple Range Test.


68
Table 11. Nitrogen balance for the batch digesters.
High N plant material Low N plant material
2.5 L+ 5 L 10 L 2.5 L 5 L 10 L
Nitrogen added
Water hyacinth
Organic N
Inoculum
Organic N
Inorganic N
Total
Organic N
Inorganic N
Nitrogen recovered
Screened effluent
Organic N
Inorganic N
Digested sludge
Organic N
Total
Organic N
Inorganic N
g
10.39
10.39
10.39
1.15
2.31
4.61
2.68
5.36
10.72
11.54
12.69
14.99
2.68
5.36
10.72
14.22
18.05
25.72
1.48
2.41
2.02
8.81
11.60
15.81
2.86
4.91
4.94
4.34
7.32
6.96
8.81
11.60
15.81
13.15
18.91
22.77
3.24
3.24
3.24
0.07
0.14
0.27
1.34
2.68
5.35
3.31
3.37
3.51
1.34
2.68
5.35
4.64
6.05
8.86
1.26
1.31
1.09
1.20
2.63
4.98
1.17
0.85
0.87
2.43
2.16
1.96
1.20
2.63
4.98
3.63
4.79
6.94
% Recovered 92 105 89 78 79 78
t
Volume (liters) of inoculum.


94
A first-order kinetic equation was used to describe NH^-N loss with
time. Rate constants for diluted effluents ranged from 0.228 to 0.593
day 1. Rate constants for undiluted effluents ranged from 0.175 to
0.446 day ^. The time required for a 50% reduction of NH+-N was 1.12 to
3.04 days for treatments with positive water hyacinth dry weight gains.
A 50% reduction of NH*-N in treatments resulting in plant death required
1.98 to 3.96 days. Plant assimilation was one of the primary mechanisms
of NH^-N loss in the systems with actively growing plants. Microbial
assimilation and NH^-N volatilization were probably important mechanisms
of NH+-N removal for treatments resulting in plant death.
Plant assimilation accounted for a 36 to 77% recovery of effluent
for surviving plants. Only 2 to 16% of the was recovered in
dead plant tissue. Approximately 75% of the was unaccounted for in
effluents resulting in plant death.
Surviving plants assimilated large amounts of Na, K, Ca and Mg
while dead plants lost K and had small gains of Ca and P. Sodium
accumulated in dead plant tissue. Death was attributed to an
indiscriminate salt injury and/or NH^-N toxicity.
The largest reductions of EC, K and P were observed in diluted
effluents. The highest plant dry weight gains were also found in these
effluents. Potassium and Mg increased in effluents where plant death
was noted.


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24
DeBusk et al. (1983) measured detritus production in harvested and
nonharvested water hyacinth based sewage treatment systems. Detritus
-2 -1
production in both systems averaged 2 g dry wt m day More than 80%
of the detritus consisted of root material. The bulk of the standing
crop detritus remained trapped in the floating plant mat. However, this
study did not reveal the potential of detritus as a nutrient input to
the water hyacinth ponds.
The objectives of this study were to 1) measure productivity and
detritus (shoot and root) production of water hyacinths grown in
eutrophic lake water with and without added nutrients and 2) determine
the potential of detritus as a nutrient source or sink to the ponds.
Materials and Methods
The study was conducted in two reservoirs located at the Central
Florida Research and Education Center research farm near Lake Apopka in
Zellwood, Florida. The reservoirs were constructed with 2.0 m high
levees of a Lauderhill organic soil (Lithic medisaprists) and with
bottoms composed of calcareous clay. The water depth was 60 cm and the
dimensions of the reservoirs were 7.6 m by 61 m (total surface area of
2
465 m ). Both reservoirs were filled with water from nearby Lake
Apopka, and were sectioned into four equal areas for replication and
stocked with water hyacinths.
2
A total of eight 0.25 m cages (Vexar mesh screen connected to 5 cm
diameter PVC pipe) were stocked with water hyacinth at an initial
_2
density of 16 kg (fresh wt) m The cages were placed within the four


90
increased for the dead plant root material but remained similar for the
other materials. The Ca content increased for the shoots of all plants
and the roots of the dead plant material. The Mg content increased for
roots of all plants and shoots of the surviving plants.
The net assimilation or loss of plant nutrients from plants in
diluted or undiluted effluents from digested high N plants are reported
in Table 20. The plants in diluted effluents assimilated large amounts
of Na and K, apparently because these nutrients move rapidly with the
transpiration stream. The net shoot assimilation of all nutrients was
greater than the net root assimilation.
The dead plants from undiluted effluents showed a net loss of K but
net gains of the other nutrients. Potassium, Na, Ca and Mg were
reported as being rapidly lost during the early leaching phase of plant
decomposition in fresh water (Boyd, 1970b; Davis and van der Valk,
1978). Most of the Na moved into the shoot region. Generally the net
gains of P and Ca were found in dead plant roots compared to shoots of
surviving plants in diluted effluents.
Final Chemical Composition of the Effluents
Characteristics of the digester effluents and nutrient medium after
water hyacinth treatment are given in Table 21. For treatments where
plant dry weight gains were observed, generally there was large
reductions of the elements analyzed in the effluents. For the
treatments resulting in plant death, K and Mg increased and the
reductions of other elements were of lesser magnitude.
The largest reductions of EC (49 + 7% reduction), K (93 + 3%) and P
(92 + 3%) were observed in diluted effluents. The highest gains of


AMMONIUM NITROGEN Cmg LT
TIME (days)
Figure 14. Nitrogen loss from sediment-water-plant systems with added ammonium.


WATER HYACINTH BIOMASS AND DETRITUS PRODUCTION
Water hyacinth is one of the most productive aquatic macrophytes
found throughout the tropical and subtropical regions of the world. The
plant has been used extensively for treatment of nutrient-enriched
waters and currently there are a number of wastewater treatment systems
in the U. S. utilizing water hyacinths for secondary and tertiary
treatment (Cornwell et al.,1977; Dinges, 1978; Wolverton and McDonald,
1979; Reddy et al., 1985).
Water hyacinth productivity has been evaluated in natural and
-2 -1
nutrient-enriched waters. Growth rates of 2 to 29 g dry wt m day
were reported for plants growing in natural waters of central and south
Florida (Yount and Crossman, 1970; DeBusk et al., 1981). A wide range
-2 -1
of productivity (5 to 42 g dry wt m day ) was recorded for plants
cultured in nutrient-enriched waters (Hanisak et al., 1980; Reddy and
Bagnall, 1981; Reddy, 1984). Maximum growth rates provided an average
-2 -1
of 52 and a maximum of 64 g dry wt m day for plants cultured under
nutrient-nonlimiting conditions (Reddy and DeBusk, 1984).
Plant detritus (dead and decaying plant debris) is an integral part
of water hyacinth mats. Detritus is usually derived from natural aging
of plants, biological or chemical control, and frost damage.
Decomposition of detritus releases nutrients which can be subsequently
utilized by water hyacinths. Information on water hyacinth productivity
was extensive (Reddy et al., 1983), but research on detritus production
and its role as a nutrient sink or source was limited.
23


56
15
Table 7. Mass balance of added NO^-N in sediment-waterwplant systems.
Plant
Sediment $
Unaccounted
Treatment or
algae
org
inorg
Water
Total
For
15,
PLANTS
Without
sediment
0
mg
C L_lt
65.2


ND §
65.2
34.8
100
mg
c l'1
67.5


ND
67.5
32.5
400
mg
C L_1
60.6


ND
60.6
39.4
With
sediment
0
mg
C L-1
72.0
3.2
0.4
ND
75.6
24.4
100
mg
C L-1
65.6
4.4
0.4
ND
70.4
29.6
400
mg
CL_1
56.7
3.6
0.6
ND
59.9
40.1
NO PLANTS
Without
sediment
0
mg
C L_1
ND


6.2
6.2
93.8
100
mg
c l"1
6.8


ND
6.8
93.2
400
mg
C L_1
8.2


ND
8.2
91.8
With
sediment
0
mg
c l1
ND
11.4
0.4
5.1
16.9
83.1
100
mg
cl"1
8.1
10.1
0.4
2.0
20.6
79.4
400
mg
C L-1
8.4
4.9
0.5
0.5
14.3
85.7
^Carbon source was plant detritus.
^Org, Inorg = organic and inorganic N, respectively.
§ND = Not detectable.


Table 3. Seasonal nitrogen uptake by water hyacinth and detritus.
Fertilized
reservoir
Control reservoir
Season Water hyacinth Detritus
Water hyacinth
Detritus
1. ~ XT T ~ ^
Autumn (78) ^ 127.8
28.2
30.9
14.6
Winter (90) 33.9
42.4
-1.2
23.7
Spring (84) 167.7
37.5
104.3
29.1
Summer (88) 242.4
39.8
98.7
24.4
Test of significance t
Water hyacinth
Detritus
Season
**
NS
Month (season)
NS
*
Reservoir
**
**
^ Number of days in season.
Significant at 0.05 (*) or 0.01 (**) level or not significant (NS).


41
A dense cover of floating water hyacinth depleted dissolved 0^ of
the underlying water, thus creating anaerobic conditions (Boyd, 1970;
McDonald and Wolverton, 1980; Reddy, 1981). Decomposition of plant
detritus also consumed 0^ (Nichols and Keeney, 1973; Rai and Munshi,
1979). Anaerobic conditions may restrict nitrification and favor
denitrification, which may proceed within the water hyacinth mat, in the
water column, or in the underlying sediment. Detritus also provides
energy source for denitrification. Denitrification occurred primarily
in the underlying sediment and the rate depended on NO^-N diffusion from
the water column to the sediment (Engler and Patrick, 1974; Reddy and
Graetz, 1981).
Volatilization becomes increasingly important as the water pH
increases. The partial pressure of NH^-N in equilibrium with a solution
of NH^-N increased rapidly in a pH range of 8.5 to 10.0 (Bouldin et al.,
1974). A pH of 7.0 in water occurred in areas covered with water
hyacinth with little diel variation (McDonald and Wolverton, 1980;
Reddy, 1981) which suggests that NH^-N volatilization is minimal in
areas covered with plants.
The relative role of N assimilation by water hyacinth on total N
removal from reservoirs was investigated by Reddy (1983). Approximately
40% of added ^NH^-N or ^no^-N was assimilated by plants. Less than
15 I
10% of the added N was found in the surface sediment layer. Over 40%
of the added was unaccounted for.
Information on the role of detritus in aquatic systems on
immobilization or mineralization of inorganic N is limited. The overall
objective of this study was to determine the effect of detritus on
selected N transformations in water columns with and without water


LIST OF TABLES
TABLE PAGE
1. Seasonal water hyacinth yield and detritus production ... 30
2. Seasonal water hyacinth shoot and root lengths 32
3. Seasonal nitrogen uptake by water hyacinth and detritus . 35
A. Nitrogen balance for the two reservoirs 37
5. Total plant N and ^NO^N assimilation 53
6. Total plant N and ^NH+-N assimilation 5A
7. Mass balance of added ^NO^-N in sediment-water-plant
systems 56
8. Mass balance of added ^NH*-N in sediment-water-plant
systems 57
9. Characteristics of the inoculum used in the batch
digesters 64
10. Gas production during anaerobic digestion of high and
low N water hyacinth plants 65
11. Nitrogen balance for the batch digesters 68
12. Nitrogen-15 balance for the batch digesters 69
13. Characteristics of digester effluents before sludge
removal 71
14. Characteristics of screened effluents (sludge removed)
after digestion 72
15. Characteristics of fresh and digested biomass residues. . 73
16. Initial characteristics of the digester effluents
and nutrient medium 81
17. First-order kinetic descriptions of NH+-N loss with time. 86
18. Nitrogen-15 balance for labeled effluents 88
v


108
Other Soil Parameters
Soil pH increased by one and two pH units after addition of
anaerobically digested biomass sludge with a low and high N content,
respectively (Table 25). The addition of fresh plant biomass did not
appreciately alter the initial soil pH. During the 90 day incubation,
soil pH generally decreased with all residue additions, except for low N
fresh plant biomass.
The Na content of the digested biomass sludges explains, in part,
the increase in the initial soil pH. Atalay and Blanchar (1984) found
that addition of anaerobically digested plant residue to soil increased
pH from 5.5 to 7.6. They attributed the pH increase to a limestone
*,
buffer used during anaerobic digestion.
The addition of fresh or anaerobically-digested water hyacinth
increased Mehlich I extractable soil constituents after 90 days of
incubation (Table 26). Mehlich I extractable soil constituents at 0, 30
and 60 days of incubation are presented in APPENDIX B, Tables 30 to 32.
The increases were a direct reflection of the mineral composition of the
respective residue (Table 22). There was a large increase in soil K with
fresh plant biomass additions compared to digested biomass sludges. All
treatments resulted in large increases of soil Na and Ca. Parra and
Hortenstine (1976) concluded that fresh water hyacinths contained
appreciable amounts of soluble salts and that crops susceptible to salt
injury should not be planted immediately after soil additions.


Table 10. Gas production during anaerobic digestion of high
and low N water hyacinth plants.
Inoculum Cumulative biogas production Total Gas Yields
.volume 15 days 30 days 60 days biogas methane
--L-- Liters L g-1 VS
High N plant material
2.5 16.4 40.8 60.3 0.21 0.14
5 28.1 53.5 73.0 0.23 0.15
10 34.5 59.1 75.4 0.20 0.13
Low N plant material
2.5 16.9 45.5 67.9 0.25 0.16
5 20.0 51.6 72.6 0.27 0.17
14.7 52.2 67.4 0.25 0.16
10


5
Figure 2. A generalized diagram of a water hyacinth plant.
The major morphological structures are
adventitious roots (AR); root hairs (RA); rhizome
(RH); stolon (ST); detritus tissue (DT) attached
to the plant; float (F); leaf isthmus (IS);, leaf
petiole (PT); peduncle (PD); spathe (SP); leaf
lamina (LA); inflorescence (IN).


64
Table 9. Characteristics of the inocula used in the batch digesters.
Inocula characteristics t
NH+-N
4
no3-n
TKN COD
pH TS
FS
VS
T-1
-% of
TS---
mg L
High N plant
material
1072
48
1530 1-4200
6.3 1.75
33.5
66.5
Low N plant
material
535
21
562 784
7.7 0.29
83.1
16.9
^ COD = Chemical
oxygen demand,
TS, FS, and VS
= Total,
fixed and
volatile solids, respectively.


99
Table 22. Characteristics of the fresh and digested plant biomass
Chemical Low N plant biomass High N plant biomass
constituent Fresh Digested Fresh Digested
g kg of biomass
Volatile solids
839 bf
866 a
837 b
825 c
Ash
161 b
134 c
166 b
173 a
Total carbon
373 b
425 a
385 b
446 a
Lignin
83 b
145 a
43 c
130 a
Cellulose
266 a
180 b
167 b
206 b
Hemicellulose
247 a
243 a
183 b
180 b
Total nitrogen
10.6 c
26.6 b
34.0 a
39.3 a
Calcium
21.0 b
24.2 a
17.6 c
18.4 c
Potassium
22.2 a
2.8 b
23.5 a
3.8 b
Magnesium
6.7 a
2.2 a
3.2 b
2.2 c
Sodium
10.9 c
17.4 b
8.0 d
26.0 a
Iron
1.9 c
7.2 a
1.6 c
4.7 b
Zinc
0.7 c
2.0 a
0.5 d
0.9 b
C/N ratio
t
35 a
16 b
12 c
12 c
'Values with same letter within rows are not significantly different
at 0.05 level by Duncan's Multiple Range Test.


77
Water hyacinth-based wastewater treatment systems have already been
evaluated for use in treating primary and secondary sewage effluents
(Wolverton and McDonald, 1979; Reddy et al., 1985) and anaerobic
digester effluent (Hanisak et al., 1980). The potential productivity of
water hyacinth in nutrient-enriched waters has led to its selection in
alternative methods of wastewater renovation, particularly in areas
where growth is not restricted by climatic limitations.
Use of water hyacinth for digester effluent treatment is
particularly attractive, because of its ability to grow in waters with
high elemental concentrations. The biomass produced could be returned
to the digester as a feedstock for methane production. Hanisak et al.
(1980) determined that 64.5% of (liquid and sludge) N in diluted
effluents from anaerobically digested water hyacinth could be
reassimilated by water hyacinths. Diluting the effluent does not
address the full potential of water hyacinth to grow under these
nutrient and salt enriched conditions. Haller et al. (1974) concluded
that water hyacinth will not live in waters containing sustained salt
concentrations in excess of 2500 mg L ^. Optimal dilution of these
concentrated effluents to obtain maximum water hyacinth yields and
nutrient removal was not reported.
The objectives of this study were to 1) evaluate water hyacinth
productivity in anaerobic digester effluents obtained from digesters
receiving different types of water hyacinth as feedstock, and, 2)
determine recovery by water hyacinth growing in digester effluents
from digested labeled water hyacinth biomass.


Table 14. Characteristics of screened effluents (sludge removed) after digestion.
Inoculum Screened Effluent Characteristics^
volume pH EC NH^-N TKN Ca Mg K Na P TS VS FS
-L- dS mg L_1 % --% of TS
High N plant material
2.5 7.6 4.3 161 188 32 13 82 1140 12.8 0.357 32.4 67.6
5 7.6 4.7 212 256 30 12 80 1160 12.8 0.342 41.5 58.5
10 7.6 5.3 289 326 23 14 123 1160 11.5 0.369 41.0 59.0
Low N plant material
2.5 7.5 5.6 22 45 147 50 195 1220 3.1 0.367 30.3 69.7
5 7.4 5.9 48 72 53 53 245 1240 4.6 0.408 30.5 69.5
10 7.5 6.7 91 111 61 63 325 1200 4.5 0.425 28.1 71.9
-j-TS, VS, FS = Total, volatile and fixed solids, respectively.


TREATMENT OF ANAEROBIC DIGESTER EFFLUENTS USING WATER HYACINTHS
An integrated approach of wastewater renovation using aquatic
macrophytes with utilization of biomass for energy production is
economically appealing. The plant biomass produced in these systems,
along with other wastes such as sewage sludge or animal waste could be
anaerobically digested to produce methane (Stack et al., 1981;
Shiralipour and Smith, 1984). This process generates a waste by-product
which must be disposed of, or preferably utilized to reduce the cost of
energy production, in an environmentally-safe manner. The waste
by-product consists of digested sludge and a large volume of effluent.
Integrating wastewater renovation through water hyacinth production
provides an internal option for the disposal of effluent generated
during conversion of biomass into methane.
The effluent composition of anaerobic digesters varied with type of
feedstock used in digestion (Stack et al., 1981). Information on
chemical composition of effluents from sewage or animal wastes was
readily available (Sommers, 1977; Field et al., 1984). However,
anaerobic digestion of plant biomass has only recently gained attention
in the United States and information on composition or disposal of the
effluent was limited (Hanisak et al., 1980; Atalay and Blanchar, 1984).
Digester effluents have high concentrations of BOD, NH+-N, K and Na
(Atalay and Blanchar, 1984; Field et al., 1984), while the divalent
cations and metals were concentrated in the sludge (Sommers, 1977; Field
et al., 1984).
76


34
was much greater during this time period (Table 3). Plant tissue N
content remained nearly consistent with time in the control reservoir
and root tissue N content generally exceeded that of the shoot tissue
(Fig. 8).
Seasonal N assimilation by water hyacinth ranged from 34 to 242 kg
N ha ^ for plants in the fertilizer reservoir and from <0 to 104 kg N
ha ^ for plants in the control reservoir (Table 3). There were
significant differences between seasons and reservoirs in water hyacinth
N assimilation. The detritus N content was significantly greater for
fertilized than control plants, but there were no significant
differences in detritus N content between seasons.
Data on mass balance of N in both reservoirs are shown in Table 4.
Nitrogen input from the lake was 238 kg N ha ^ with 89% of the N in the
organic fraction. Total amount of fertilizer applied during the study
period was 781 kg N ha ^, with NH^-N, NO^-N, and organic N representing
55, 30, and 15% of total fertilizer applied, respectively.
The total N assimilated by water hyacinth (live plants and
detritus) was 720 and 325 kg ha ^ yr ^ for fertilized and control
reservoirs, respectively (Table 4). Annual net N loading by detritus
was 148 and 92 kg ha ^ for fertilized and control reservoirs,
respectively (Table 4). Maximum detritus N loading occurred during
i
winter for the fertilized reservoir and during spring for the control
reservoir. This corresponded to the time of root dislodging from plants
in the two reservoirs.
The annual net N immobilized by detritus represented 21 and 28% of
the total N removed by water hyacinth in the fertilized and control
reservoirs, respectively. DeBusk et al. (1983) concluded that


Table 24. Carbon and N mineralization from added
fresh and digested plant biomass.
Low N plant
biomass
High N
plant
biomass
Fresh
Digested
Fresh
Digested
/o OX aflQGQ
l. o tr
N
Day
30
c
22.4 b t
11.2 c
34.2
a
9.8c
15n
ND *
4.7 b
24.8
a
3.8 b
Day
60
c
32.5 b
14.9 c
41.6
a
16.1 c
15n
ND
6.2 b
34.3
a
6.0 b
Day
90
c
39.0 b
19.1 c
49.9
a
23.1 c
15n
3.3 c
7.7 b
33.3
a
7.7 b
t

Values with same letter within rows are not
significantly different at 0.05 level by Duncan
Multiple Range Test.
ND = not detectable.


TABLE
PAGE
19. Distribution of nutrients in water hyacinth shoots
and roots in diluted and undiluted effluents of
digested high N plants 89
20. Net assimilation or loss of plant nutrients in diluted
or undiluted effluents from digested high N plants 91
21. Characteristics of the digester effluents and nutrient
medium after water hyacinth treatment 92
22. Characteristics of the fresh and digested plant biomass . 99
23. Soil NO^-N concentration from added fresh and digested
plant biomass 105
24. Carbon and mineralization from added fresh and
digested plant biomass 106
25. Soil pH (1:2 w/v) from added fresh and digested plant
biomass 109
26. Mehlich I extractable constituents at Day 90 from
added fresh and digested plant biomass 110
27. Effluent pH during water hyacinth treatment 124
28. Effluent dissolved 0^ concentration during water
hyacinth treatment 125
29. Soil ammonium concentrations from added fresh and
digested plant biomass 127
30. Mehlich I extractable constituents at Day 0 from
added fresh and digested plant biomass 128
31. Mehlich I extractable constituents at Day 30 from
added fresh and digested plant biomass 129
32. Mehlich I extractable constituents at Day 60 from
added fresh and digested plant biomass 130


102
animal waste decomposition if the overall decomposition sequence is
presented as occurring in stages (Gilmour et al., 1977; Hunt, 1977;
Reddy et al., 1980). Each stage is thought to represent the sequential
ease of C constituent decomposition, i.e. soluble sugars and starch,
cellulose and hemicellulose, and lignin.
The rate equation for a first-order reaction is expressed as
-dC./dt = k.C.
i li
where subscript i refers to a particular stage of C decomposition. The
integrated first-order rate equation is
Ct. = C.exp(-k.t)
i i r i
where C. = C at beginning of a decomposition stage,
Ct^ = C remaining at end of a decomposition stage at time = t,
k. = first-order rate constant.
i
Therefore, a rate constant can be calculated for each decomposition
stage of an organic C material.
A graphical representation of the stages and their respective rate
constants of the materials used in this study are shown in Fig. 17.
Decomposition of fresh plant biomass required a three stage first-order
kinetic description. Rate constants for the first stage (soluble sugars
and starch) were 0.0441 and 0.0222 day ^ for high and low N plant
biomass, respectively. This stage of decomposition was essentially
completed in 4 days. A longer time was required during the second stage
(cellulose and hemicellulose) of decomposition for low N plant material
due to a higher content of these C constituents (Table 22). The rate
constants for the final stage (lignin) of decomposition were low for
both materials.


CONCLUSIONS
The three components of the integrated "biomass for energy" system
were 1) the water hyacinth biomass production system; 2) anaerobic
digestion of water hyacinth biomass, and 3) waste recycling of digested
biomass sludge and effluent. Nitrogen cycling was investigated for each
component of the integrated system.
Water Hyacinth Productivity and Detritus Production
Productivity of water hyacinth was influenced by ambient air
temperature, solar radiation, and nutrient compostion of the medium.
The highest net productivity occurred during spring and summer and over
75% of the biomass produced was recorded during this time period. The
detritus production exceeded net biomass production during winter
regardless of water fertility.
Seasonal yields of water hyacinth ranged from 1.9 to 23.1 Mg (dry
wt) ha ^ and -0.2 to 10.2 Mg ha ^ for plants growing in eutrophic lake
water with and without added nutrient, respectively. Detritus comprised
3 to 14% of the total biomass and detritus production was not
significant between reservoirs or seasons. Although detritus production
was similar for both reservoirs, fertilization resulted in significant
increases in detritus N content. Nitrogen loading to the reservoirs
from detritus was 148 and 92 kg N ha 1 yr 1 for plants grown in
eutrophic lake water with and without added nutrients, respectively.
119


TIME (days)
Figure 9. Dissolved 0^ in sediment-water-plant systems with added nitrate.
Ln


43
Water hyacinth detritus (shoot and root material) was added at the
rates of 0, 100, and 400 mg C L *. The detritus was chopped manually to
lengths of -2 cm. The detritus for treatments with added ^NO^-N was
collected from a natural water hyacinth stand in Zellwood, Florida and
had an initial N content of 5.6 mg g 1 dry tissue. The detritus for
15 +
treatments with added NH^-N was collected from a water hyacinth stand
located in a wastewater stabilization pond at the University of Florida
wastewater treatment plant in Gainesville, Florida and had an initial N
content of 23.1 mg g dry tissue.
_2
Water hyacinths, at an initial density of 10 kg (fresh wt) m ,
were placed in 12 of 24 tanks. The plants were collected from the
University of Florida's Bivens Arm research reservoirs in Gainesville,
Florida. The plants were clipped of dead tissue and rinsed with tap
water prior to placement in tanks.
The disappearance of added inorganic N was determined by collecting
water samples at 0, 1, 2, 3, 4, 8, 15, 28 days and measuring NH+-N,
NO^-N, and total Kjeldahl N (TKN). The changes in water hyacinth fresh
weight were measured weekly. Plant samples and detritus were analyzed
for TKN. The sediment was characterized for organic and inorganic N
prior to and at the conclusion of each study. Fifty grams (dry wt) of
moist sediment samples were extracted with 2 M KC1 and analyzed for
NH^-N and NO^-N. Sediment samples were air-dried, ground by mortar and
pestle, and analyzed for TKN. The inorganic N for all samples was
determined by steam distillation (Keeney and Nelson, 1982). The TKN of
water, plant, and sediment samples were determined by micro-Kjeldahl
procedures (Nelson and Sommers, 1972; 1973; 1975). The analyses on
water, sediment, plant and detritus samples were conducted using a Micro
Mass 602 spectrometer.


INTRODUCTION
Several types of aquatic plants are widely distributed in
freshwater lakes and streams. These plants assimilate nutrients and
produce biomass, which could potentially be used for beneficial
purposes. Water hyacinth (Eichhornia crassipes [Mart] Solms) is one of
the dominant aquatic plants distributed throughout the tropical and
subtropical regions of the world. This freshwater macrophyte has
already been evaluated for use in treating nutrient-enriched waters such
as sewage effluent (Cornwell et al., 1977; Wolverton and McDonald, 1979;
Reddy et al., 1985), agricultural drainage water (Reddy and Bagnall,
1981; Reddy et al., 1982), anaerobic digester effluent (Hanisak et al.,
1980), and fertilized fish ponds (Boyd, 1976). The characteristics that
make this plant grow rapidly in polluted waters make it an ideal
candidate for large-scale nutrient removal and water purification (Reddy
and Sutton, 1984).
An integrated aquaculture system has been developed using water
hyacinth for water treatment and for total resource recovery. The
components of an integrated aquaculture system are schematically
illustrated in Fig. 1. Water hyacinth plants have been used for
wastewater treatment while the biomass produced was harvested
periodically and processed through anaerobic digestion to produce
methane. The process produced a waste by-product which must be disposed
of, or preferably utilized, in an environmentally-safe manner.
1


(23 to 289 mg N L ^). Biomass yields were maximum at electrical
conductivities of < 2.5 dS m 1 and ^NH^-N concentrations of < 100 mg N
L_1.
Addition of water hyacinth biomass to soil resulted in
decomposition of 39 to 50% of added C for fresh plant biomass and 19 to
15.
23% of added C for digested biomass sludge. Only 8% of added N in
V
digested sludges was mineralized to ^N0_-N despite differences in
initial N content (27 and 39 g N kg 1 dry sludge). In contrast, 3 and
33% of added ^ N in fresh biomass with low and high N content,
respectively, was recovered as ^NO^-N.
Total recovery after anaerobic digestion ranged from 70 to 100%
of the initial plant biomass Land application of digester sludge
resulted in the mineralization of 2% of initial biomass into plant
available form. Use of water hyacinth for digester effluent treatment
resulted in recycling of 21 to 38% of the initial biomass ^N. Total N
recovery by sludge and effluent recycling in the integrated "biomass for
energy" system was 48 to 60% of the initial plant biomass The
remaining was lost from the system during anaerobic digestion and
effluent recycling.
x


13
weight loss under aerated conditions in sediment-water systems than in
water only. They attributed this difference to an additional supply of
N from the sediments.
Anaerobic Digestion
The second component of the integrated "biomass for energy" system
was anaerobic digestion of plant biomass for methane production.
Anaerobic digestion is a biological process in which organic matter, in
the absence of oxygen, is converted to methane and carbon dioxide
(Toerien and Hattingh, 1969).
During the process of anaerobic digestion, waste organic C was
stabilized by the nearly complete microbial fermentation of
carbohydrates resulting in a reduction of volatile solids (Miller,
1974). Anaerobically digested sewage sludges were considered more
stable to microbial degradation than were aerobically digested sludges
(Sommers, 1977).
Processes which regulated anaerobic digestion include hydrolysis of
polymers, the dissimilation of starting subtrates to the level of acetic
acid, and the conversion of acetic acid to CH^ and CO^ (Mah et al.,
1977). Factors which influenced anaerobic digestion include pH and
temperature changes. All methanogens were reported to be strict
i
anaerobes with an optimum pH of 6.7 to 7.4 (Bryant, 1979). The optimum
temperature range was 30 to 35C (House, 1981).
Water hyacinth biomass could be anaerobically digested to produce
methane. Hanisak et al. (1980) found average methane yields of 0.24 L
g volatile solids (VS) of shredded water hyacinth in 162 L digesters.
Chyoweth et al. (1983) reported methane yields of 0.19 and 0.28 L g ^ VS