EVALUATION OF SOIL TEST METHODS AS INDICATORS OF RELEASABLE
PHOSPHORUS IN WETLAND SOILS
A THESIS PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE
UNIVERSITY OF FLORIDA
This document is dedicated to my parents and all other family members, past and present.
I would like to convey my sincere thanks to all who have directly or indirectly
shared their valuable time to bring this piece of work into its present shape. I desire to
express my sincere gratitude to my committee members, Dr. Vimala D. Nair (Chair), Dr.
K. Ramesh Reddy (Cochair) and Dr. Mark W. Clark, for their constant support, guidance
and advice during this research. I would like to acknowledge the funding agency The
United States Environmental Protection Agency's Office of Water, grant numbers
828212 and 830604. I am very grateful to my parents and other family members for
providing me the opportunity to pursue this degree. Special thanks should go to Miss Yu
of Wetland Biogeochemistry Lab for all of her help in laboratory analyses. Last but not
least, I thank all the laboratory staff and my friends, here and there, for their constant help
and support throughout the work.
TABLE OF CONTENTS
A C K N O W L E D G M E N T S ................................................................................................. iv
L IST O F T A B L E S .................................................................... .......................... .. vii
LIST OF FIGURES ...................................... ....... .......... .......... ..... viii
A B ST R A C T ................. .......................................................................................... x
1 IN TRODU CTION ................................................. ...... .................
2 LITER A TU R E REV IEW ............................................................. ....................... 5
Soil Test Extractions for Phosphorus ........................................ ....................... 5
M ehlich 1 (M l) E xtraction ......................................................................... .... 8
M ehlich 2 (M 2) Extraction........................................................ ............. ..8
M ehlich 3 (M 3) Extraction ........................................................ ............. ..9
B ray and K urtz P-1 Extraction ........................................ ......................... 9
Olsen Extraction ............................... ....... ............ ........10
Acid Ammonium Acetate-EDTA Extraction .................................................10
C aC 12 Extraction (0.01 M ) ..................... ............ ........ ...... ..................... 10
Ammonium Bicarbonate DTPA Extraction (AB-DTPA) ..................................10
O xalate E extraction ................................................... .. ...... .............. .. 11
W after E extraction ................. ...... .......... ...... .... ............. .............. .. 11
Relationships among Soil Test Procedures ............... .....................................12
Agronom ic and Environm ental STP..................................... ......... ............... 14
Extraction M ethods for W etland Soils ............................................ ............... 15
3 SOIL TEST PHOSPHORUS AS INDICATORS OF P RELEASE FROM
W E T L A N D SO IL S ......................................................................... .....................2 0
Introdu action ........................................................ ............. ................. 20
Soil Tests Chosen for This Study ............................................. ............... 22
Soil Tests Used by Various Researchers..........................................................23
M materials and M methods ....................................................................... ..................24
Soil Sam pling ......................................................................24
Soil A naly ses............................................................... 25
Soil C haracterization .................................. ................... . ....... ............25
Methods of Different Extraction Processes Employed ..............................27
Statistical A analyses ......... .. ........................ ........ ..... ......................... .. 27
R results and D iscussions........... ...... ...................................................... .... .... ...... 28
C o n c lu sio n s............................................................................................................ 3 6
4 INDICATORS FOR ENVIRONMENTAL PHOSPHORUS EVALUATION ..........38
In tro d u ctio n ...................................... ................................................ . 3 8
M materials and M methods ........................................................................ ..................42
Statistical M eth o d s............ ... ...................... ...................................... .......... .. ..... .. 4 2
R results and D iscussions.......... ..... ......................................................... ... .... ...... 43
C o n c lu sio n s..................................................... ................ . 5 7
5 SUMMARY AND CONCLUSIONS.......................................................................59
L IST O F R E FE R E N C E S ............................................................................. .............. 62
BIOGRAPH ICAL SKETCH ...................................................... 75
LIST OF TABLES
3-1 Soil tests used by different researchers to assess environmentally and/or
bioav ailable P in soils............ ... .................................................... .. .... .... ... .. 23
3-2 Range of selected physical characteristics of the soils in this study ........................28
3-3 Mean, standard error (SE), median, mode, standard deviation (SD) and range for
WSP, M1-P, M3-P and Ox-P for all wetland samples in this study. .....................29
3-4 Correlation coefficients of different soil tests on 307 wetland samples...................31
3-5 Correlation of different extractable P with WSP (by location)............................31
3-6 Comparison of sediment and wetland soils with respect to water soluble P
(W SP) and organic matter (OM ) content............. ................................................ 34
3-7 Correlation coefficients of strongly associated parameters............... ................... 35
3-8 Comparison of upland and wetland soils ...........................................................35
4-1 Correlation (r) of PSR analyses with water soluble P on 311 wetland samples ....43
4-2 Prediction equations for water soluble P with two different sets of independent
v a riab le s .......................................................................... . 4 7
4-3 Threshold values for P release indicators for wetland soils.............. ..................50
4-4 Correlation of Ml analyses and other parameters (by location) ............................54
4-5 PSR com prison (by location)........................................ ........................... 55
4-6 Characteristics of elemental analyses of soils under different Ox-PSR ranges .....56
LIST OF FIGURES
3-1 Map of the locations of the soil samples collected during the Southeastern
Wetland Biogeochemical Survey (SWBS) study (Adapted from Paris, 2004)........25
3-2 Field sampling scheme during the SWBS study (Adapted from Paris, 2004). .......26
3-3 Relationship of M1-P and M3-P for all soils by location (The symbols A
represents center of wetland, B represents edge of wetland and U represents
upland adjacent to the wetlands) [p < 0.0001] ......................................31
3-4 Correlation between WSP and organic matter; p < 0.0001, only wetland soils are
in clu d ed .......................................................... ................ . 3 2
3-5 Correlation between organic matter (OM) and total carbon (TC) content; p <
0.0001, both upland and wetland soils included. ................ ................ ..............33
4-1 Water soluble P (WSP) as a function of M1-PSR for wetland soils with < 12%
total C and > 12% total C ........................................ ........................................ 44
4-2 Water soluble P (WSP) as a function of M3-PSR for wetland soils with < 12%
total C and > 12% total C ............................................................. .....................44
4-3 Water soluble P (WSP) as a function of Ox-PSR for wetland soils with < 12%
total C and > 12% total C ........................................ ........................................ 44
4-4 Water soluble P (WSP) normalized with respect to bulk density as a function of
M1-PSR for soils < 12% total carbon and > 12% total carbon.............................46
4-5 Water soluble P (WSP) normalized with respect to bulk density as a function of
M3-PSR for soils < 12% total carbon and > 12% total carbon.............. ...............46
4-6 Water soluble P (WSP) normalized with respect to bulk density as a function of
Ox-PSR for soils < 12% total carbon and > 12% total carbon..............................46
4-7 Total P as a function of Ox-PSR for all wetland soils, p < 0.0001 .......................48
4-8 Oxalate-P (Ox-P) as a function of total P (TP);p < 0.0001 ................................48
4-9 M1-P as an indicator of wetland soil's P release trend; p < 0.0001 ....................52
4-10 M1-P and Ox-PSR relationship for upland soils (modified from Nair et al.,
2 0 0 4 ) ...................................... .................................................... . 5 2
4-11 Mehlich 3 P as an indicator of P release from wetland soils, p < 0.0001 ................53
Abstract of Thesis Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Master of Science
EVALUATION OF SOIL TEST METHODS AS INDICATORS OF RELEASABLE
PHOSPHORUS IN WETLAND SOILS
Chair: Vimala D. Nair
Cochair: K. Ramesh Reddy
Major Department: Soil and Water Science
Anthropogenic activities in the watershed cause nutrient loading to wetlands which
in turn has led to the direct impact on ecological integrity of this system. Ongoing
research attempts to establish numeric values to all the pollutants, nutrients or chemicals
for a particular type of water body and the designated use of that given water body. There
are no standard numeric values established for different nutrient concentrations in
wetlands due to the complexity, heterogeneity and limited existing information available
on wetlands. Phosphorus (P) is one of the major nutrients which has a direct impact on
wetland ecosystems. Soil tests have been established for upland ecosystems to predict
crop response to fertilizer application and an increasing trend is observed to use these
existing soil tests to predict P release to surrounding water bodies. Threshold values for
various soil tests and related parameters with respect to P have been assigned for upland
soils to evaluate the potential for P loss to a water body. However, there are no similar
threshold values that have been assigned as a measure of water quality degradation in
Total 630 surface soil samples (0-10 cm) were collected for this study from five
southeastern states of the United States (US): Florida, Alabama, Georgia, South Carolina
and Indiana. Surface soil samples were collected from the center of wetlands, the edge of
the wetlands and from the adjacent uplands. All samples were analyzed for pH, total
carbon (TC), bulk density (BD), total phosphorus (TP), Mehlich 1 (Ml) extractable P,
iron (Fe), aluminium (Al), calcium (Ca), magnesium (Mg), Mehlich 3 (M3) extractable P,
Fe, Al, Ca, Mg, Oxalate (Ox) extractable P, Fe, Al, and water extractable P.
Phosphorus saturation ratio (PSR) is a ratio of extractable P to the P sorption
capacity of a soil. The PSR has also been used to predict the P loss potential from a soil.
For this study, the PSRs were calculated using P, Fe and Al concentrations from Ml
(M1-PSR), M3 (M3-PSR) and Ox (Ox-PSR) solutions. The background TP concentration
of soils from the wetlands was approximately 550 mg kg-1 which was calculated from the
75th percentile of the distribution of all the unimpacted wetland soils of this study
according to the United States Environmental Protection Agency (USEPA) guideline.
Based on this reference background concentration of the wetlands surveyed threshold
values for P release to the surrounding water bodies were calculated to be M1-P = 23 mg
kg-1, M3-P = 41 mg kg-1, Ox-PSR =0.073, M1-PSR = 0.076, and M3-PSR = 0.057.
Non-point source pollution from agricultural and other anthropogenic sources have
been identified as the major cause of degradation of water bodies (USEPA, 1996). In
1972, the Federal Water Pollution Control Act (FWPCA) was amended by the Congress,
and the mandates included the designation of water bodies by their uses and
determination of standards by which water bodies would be held to. Congress set laws
that the US Environmental Protection Agency (EPA) and states will be charged to
implement. Standards were set to be protective of designated uses they will be assigned
to and those uses were mainly drinking, fishing, swimming or recreational activities. But
due to complex hydrologic characteristics and poorly defined designated uses standards
for wetlands have yet to be set for many parts of the US (Paris, 2004).
Before 1987, several amendments were passed to FWPCA including one popularly
referred to as the Clean Water Act (CWA). Before 1987 all standards for water bodies
were based on "narrative" criteria like taste bad, smell bad or look bad. In 1987, the
Water Quality Act (WQA) was passed by the Federal government. This act set goals to
develop and implement "numeric" criteria for all pollutants or nutrients for a particular
type of water body and establish designated use of that given water body. However, there
was little progress 10 years after passing the CWA. Therefore, in 1998, USEPA and US
Congress jointly created the Clean Water Action Plan (CWAP). The CWAP clearly
designated four categories of water bodies, lakes and reservoirs, rivers and streams,
estuaries and wetlands. USEPA was chosen to develop and implement nutrient criteria on
a regional basis, not national or state level. The concept of ecoregions was developed in
the 1980's to help USEPA subdivide the US based on climate, geological morphology,
vegetation, soil and agricultural use (Omernik, 1987; McMahon, et al., 2001). Research
was conducted to determine if ecological differences related to nutrients occur within
these ecoregions (McMahon et al., 2001). The overall goal was to ensure that each
ecoregion should have its own numeric nutrient criteria for each of the four water body
types. Numeric criteria for most of the first three water body types have been established.
The last water body, wetland, still does not have a standard numeric criteria assigned.
Development of numeric criteria is pending for wetlands due to the complexity,
heterogeneity and limited existing information. Two approaches have been recommended
by USEPA in this regard: (1) determination of stimulus response relationships between
causal and temporal variable and therefore identification of the threshold beyond which
that response is undesirable and (2) establishment of a value based on reference
conditions to protect the ecological integrity of the class of the wetlands in a particular
location (Paris, 2004).
Recently, biogeochemical surveys and researches have been conducted (Craft and
Casey, 2000; Paris, 2004) but numeric nutrient criteria for phosphorus (P) in wetlands
still needs to be developed. A nutrient criterion of 10 [tg L-1 total P (TP) in the water
column has been made for the Florida Everglades. There are still ongoing debates among
scientists and regulatory authorities regarding the reliability of this value to protect the
ecological integrity of the Everglades (USEPA, 2005).
Four integral parts of a wetland are water, litter, vegetation and soil of which
sorption or binding characteristics of soil and associated materials like organic matter and
leaf litter could be most important parameter to establish a numeric value for P in
wetlands. Therefore, soils may be good indicator strata to use in assessing the nutrient
condition of a wetland.
Soil testing for available P has been used to predict the fertility status of the soil,
plant availability of the nutrient and crop response to applied P by fertilizer or manures
(Gartley and Sims, 1994; Sotomayor-Ramirez et al., 2004). Recently, there were a
number of studies using soil tests conducted to predict the risk of P release to the
surrounding water bodies by leaching or surface runoff (Sotomayor-Ramirez et al., 2004).
As past researches have shown that there exists high positive statistically significant
correlation between different soil test P (STP) and P in surface runoff or leachable P or
simply water soluble P (WSP) (Mc Dowell and Sharpley, 2001; Pautler and Sims, 2000;
Cox and Hendricks, 2000; Torrent and Delgado, 2001; Bundy et al., 2001, Sotomayor-
Ramirez et al., 2004) our first objective was to find a soil test which could predict the
release of P from wetland soils to the water column or surrounding waterbodies.
This research attempts to identify a soil test for phosphorus (STP) or a phosphorus
(P) related parameter as an indicator of P release from wetland soils.
The hypotheses for this study were:
* HI: A STP or a P related parameter would be an appropriate indicator of P release
from wetland soils, irrespective of location or type of wetland.
* H2: STPs or P related parameters used as P release indicators for runoff or leaching
predictions for upland soils would be good indicators for P release from wetland
soils as well.
The objectives of this study were to:
* Identify a routine STP that could be used as an indicators) of P release to the water
column in wetland soils, irrespective of location or type of wetland.
* Determine the relationship between the identified indicators) and other wetland
soil parameters such as organic carbon (OC), pH, bulk density (BD) and metal
concentrations extracted by various soil extraction processes.
* Assign threshold values to the identified indicators for protection of water quality
of a wetland from undesirable P concentrations taking TP concentrations of the 75th
percentile distribution of all unimpacted wetland soils in this study based on
Soil Test Extractions for Phosphorus
Soil test methods have been developed to predict crop response under different
fertilizer recommendations in various best management practice (BMP) programs
(Sotomayor-Ramirez et al., 2004). Chemical extraction methods are successfully used to
evaluate soil nutritional status in relation to crop response (Nelson et al., 1953). All soil
testing procedures can be grouped into two broad categories: chemical extraction
methods and biological methods. Chemical methods have been again grouped into four
categories: (a) water, (b) carbon dioxide saturated water, (c) acids, bases, salts and
buffered solutions, and (d) electrolysis and ion exchangers (Nelson et al., 1953).
Biological methods can be grouped into two categories: (a) higher plants, and (b)
microbiological. The method involving higher plants have been reviewed in detail by
Vandecaveye (1948). The methods of the category higher plants involve field plot
technique, pot method, seedling method, Fried and Dean's (1952) standard method and
plant analysis (Nelson et al., 1953). The oldest was the field plot technique where
comparative yields or uptake of plant nutrients from treated and untreated portions of soil
usually reflect the measure of plant nutrient status (Nelson et al., 1953). Several
researchers (Bray, 1948; Vandecaveye, 1948; Jenny et al., 1950) discussed details of the
pot method originally developed by Mitscherlich (1930). In this method a particular crop
yield with and without phosphorus is compared. The percentage (%) yield of that crop
without P is expressed as the maximum yield and yield with P is used as a measure of soil
P level. Later Jenny et al. (1950) modified this pot method. Neubauer and Schneider
(1923) proposed seedling method based on the amount of P taken up by rye from 100 g.
of soil in 17 days. This method has been extensively used as a reference method for
calibrating chemical methods, mainly in Germany where it became "official method" one
time (Nelson et al., 1953). A detail discussion of this method was reported by
Fried and Dean (1952) proposed a method involving a mathematical expression as
follows: A = [B (1-y)]/y where, A = amount of nutrient available in the soil, B = amount
of nutrient in the standard which should be prepared by using a tagged 32P known
solution, and y = proportion of the nutrient in the plant derived from the standard (Nelson
et al., 1953). Their concept was that if a plant has two sources of a nutrient it could
absorb nutrients from both sources in direct proportions to the amounts available. They
proposed that the quantity of the available nutrient can be determined in terms of a
standard provided that the proportion of the nutrient from the standard can be determined
and using the mathematical relationship shown above. Several researchers (Krantz et al.,
1948; Ulrich, 1948) reviewed in detail the method of plant analysis used as an indicator
of P status of soils. This is a method using plant tissue test and is more rapid method than
the other "higher plants" methods described earlier and it was reported that plant should
be used as a good indicator of the soil environment as it could integrate all the associated
factors (Nelson et al., 1953). On the other hand, microbiological methods use
microorganisms as a measure of soil P and these methods were reviewed elsewhere
(Vandecaveye 1948; Nelson et al., 1953). Aspergillus niger (A. niger) was reported as the
first biological indicator for P and potassium (K) in soils by Butkewitsch (1909). Several
researchers (Pantanelli 1924; Benecke and Soding, 1928) suggested the use of other
microorganisms like A. oryzae, A. flavus, Cladosporium herbarum and Stichococus in
combination with A. niger to test P level in soils. The A. Niger method was developed
extensively by Niklas et al. (1930). Some researchers (Smith et al., 1932; Sekera and
Schober, 1934) reported the presence of organic acids like citric or tannic in the medium
and suggested that the similar information can be obtained by simply extracting with
these organic acids. Soil-plaque method using Azotobacter involves the use of natural soil
(non-sterilized) and first proposed by Winogradsky and Ziemiecka (1927). As this
organism can not tolerate excess acidity this biological method can not be applicable for
the soils having pH below 5.0 (Nelson et al., 1953). In calcareous soil this method was
extensively studied by Hockensmith et al. (1933). In order to overcome this difficulty
Mehlich et al. (1934) proposed the use of Cunninghamella blakesleeana because this
organism could grow in a wide range of soil pH (4 to 9) and was reported that it could
utilize various forms of organic P (Mehlich, 1939). It was reported elsewhere that
Cunninghamella method is better compared to other methods involving plants (Reuszer,
1936; Mooers, 1938; Long, 1947).
Many attempts have been made to describe the types and mechanisms of P-
containing materials present in soil. According to Russel and Russel (1950), the main P
containing materials are apatite (both hydroxide and fluoride), calcium and magnesium
phosphates, iron and aluminium phosphates and organic phosphates. Different soil
physical and chemical properties are responsible for the "available" P determined by the
various extraction processes for the determination of soil test phosphorus (STP). The
main soil factors for P availability are pH, type and amount of organic matter, and type of
colloids and clays (Brady and Weil, 2002).
Mehlich 1 (Ml) Extraction
This extraction method has been developed by Mehlich and his coworkers
(Mehlich, 1953; Nelson et al., 1953). They have used and compared 12 different reagents
0.05 M hydrochloric acid, 0.0125 M sulfuric acid, 0.05 M perchloric acid, 1% lactic acid,
and mixtures of salts or acids of different strengths to test the ability of bringing P into
the soil solution. All extraction procedures were tested on North Carolina soils. The
mixture of 0.05 M HC1 and 0.0125 M H2S04 gave maximum crop response to extractable
P for North Carolina soils (Nelson et al., 1953) and therefore mixture o 0.05 M HC1 and
0.0125 M H2S04 is used as Ml extracting solution..
Mehlich 2 (M2) Extraction
This method developed by Mehlich in 1978 before discovering Mehlich 3
extraction process (Mehlich, 1978a, 1978b). This Mehlich 2 (M2) extractant is composed
of 0.2 M NH4C1, 0.2 M CH3COOH, 0.015 M NH4F, and 0.012 M HC1 at approximately
pH 2.5. This soil test was well correlated with P uptake by Millet in six Ultisols and also
reported to be highly correlated with other soil tests like Ml, Bray 1, Olsen with soils
having a wide range of soil properties (Mehlich, 1978b). Mehlich 2 method was a better
extractant over Ml because it was reported that Ml was not applicable for calcareous or
acid soils containing recently added rock phosphate (Barnes and Kamprath, 1975) while
M2 was applicable across wide range of soil pH due to the presence of ammonium
fluoride and acetic acid in its extracting solution (Mehlich 1978b).
Mehlich 3 (M3) Extraction
This method was the one of the last attempts by Mehlich and his coworkers
regarding all the extraction processes he developed (Mehlich, 1984). It is the
modification of M2 extraction which included Cu among the extractable nutrients,
applicable for wide range of soil pH. The substitution of nitrate (NO3-) by chloride (C1-)
with addition of ethylene di-amine tetra acetic acid (EDTA) accomplished the reduction
of M2's corrosive properties (Mehlich, 1984). The M3 solution is composed of 0.2 M
CH3COOH, 0.25 M NH4N03, 0.015 M NH4F, 0.013 M HNO3, and 0.001 M EDTA. It is
now well accepted that M3 was developed for improving the laboratory efficiency of soil
testing and for providing a "universal extractant" which could be used for a wide range of
soils (Sims, 1989).
Bray and Kurtz P-1 Extraction
The Bray and Kurtz P-l method was developed by Roger H. Bray and Touby Kurtz
in 1945. This extraction has been shown to be well correlated with crop yield on most
acid or neutral soils. This soil test is not suitable for calcareous soils or soils with high pH
(pH > 6.8) or high base saturation. This soil is widely used in the Midwestern and North
Central United States (Bray and Kurtz, 1945; Frank et al., 1998). The extracting solution
is a mixture of 0.025 M HC1 and 0.03 M NH4F and the resulting solution pH about 2.6.
The fluoride ions in this extract enhance P release from aluminium phosphates by
decreasing Al activity in solution by formation of various aluminium fluoride complexes.
Fluoride is effective at suppressing the readsorption of P by soil colloids. The acidic
nature of the extraction favors dissolution reaction.
In 1954, Sterling R. Olsen and coworkers developed Olsen P or sodium bicarbonate
soil test for P to predict crop response on calcareous soils (Olsen et al., 1954). This soil
test is mainly used in the north central and western United States. This extraction is best
suited for calcareous soils but was reported to be effective in acidic soils as well (Fixen
and Grove, 1990). The extracting solution contains 0.5 M NaHCO3 with pH 8.5. This
method uses different anions like HCO3-, CO32- and OH- to bring P in solution. In
calcareous soils Ca2+ is precipitated as CaCO3 and Fe and Al as Fe and Al oxy-
hydroxides, thus enhance P solubility.
Acid Ammonium Acetate-EDTA Extraction
This extraction process was developed in 1994 at Agricultural Research Center of
Finland which has been globally used in soil testing and pollution studies and proved
useful in testing non alkaline soils (Sippola, 1994). The solution is made of 0.5 M with
respect to both ammonium acetate and acetic acid with pH 4.65 and sodium salt of
ethylene di-amine tetra acetic acid (Na2EDTA) is added to the final concentration of 0.02
CaCl2 Extraction (0.01 M)
This process was developed for soil fertility and environmental quality testing in
Hungary recently. The extracting solution of 0.01 M CaC12 was tested as an alternative
conventional extracting procedure (Jaszberenyi et al., 1994).
Ammonium Bicarbonate DTPA Extraction (AB-DTPA)
This extraction process is often known as Soltanpour method and this process can
be used to extract NO3-, P, K, Zn, Fe, Cu and Mn from alkaline soils (Soltanpour and
Schwab, 1977). The reagent is the mixture of 1.0 M NH4HCO3 and 0.005 M (Di-ethylene
tri-amine penta acetic acid (DTPA). Later Soltanpour and Workman modified this
process by omitting carbon black (Soltanpour and Workman, 1979; Soltanpour 1985).
McKeague and Day (1966) first proposed the oxalate extraction method. They used
acidified ammonium oxalate solution as the extracting reagent to test the usefulness of
the extractant for comparing acid-ammonium oxalate extractable Fe and Al with
dithionite-citrate-bicarbonate extractable Fe and Al for distinguishing Bf horizons of
Canadian soils having wide range of properties. They proposed a solution pH about 3.0
with equilibrium reaction time of four hours. They noted that under these conditions
oxalate extraction could dissolve much of the Fe and Al from amorphous materials but
very small portion from crystalline oxides. The dithionite extraction dissolved large
proportion of Fe oxides from amorphous as well as crystalline materials but was less
effective for the extraction of amorphous forms of Al. So, they concluded that oxalate
extraction gives an approximation of amorphous Al and Fe oxides whereas dithionite
extraction dissolves crystalline forms of Fe and Al in soil. They also reported that oxalate
extractable materials have a major influence on some of the soil properties; high amounts
of oxalate extractable Fe and Al are associated with soils having high pH dependent
charge and high P sorbing capacity. Now-a-days oxalate extraction for P, Fe and Al is
carried on using oxalic acid, ammonium oxalate mixture at pH 3.0 with shaking/reaction
time of four hours in dark with 1:40 (0.5 g. soil and 20 mL solution) soil to solution ratio.
Luscombe et al. (1979) were the one group of initial researchers who used distilled
water as a soil extractant to predict plant available P in soil. In their glasshouse
experiment they found a very good correlation between water extractable P and dry
matter yield of perennial ryegrass grown in three contrasting soil types. They concluded
that this method has potential as a soil test to determine P status of soils which received
In order to determine environmentally available P many studies examined methods
that best correlate water extractable soil P levels to surface runoff P or leachable P
(Sharpley, 1995; Pote et al., 1996). Pote et al. (1996) found excellent correlation between
water extractable P and dissolved reactive P (DRP) in surface runoff. As various
extractants are either more acid or more alkaline in nature than the actual soil solution,
they receive criticism that a portion of extractable P is actually of low availability.
Therefore, it was suggested that using distilled water or 0.01 M CaC12 could overcome
this criticism (Pote et al., 1995). The usual water extraction process involves 2 g. of soil
and 20 mL of distilled water (or 1:10 soil to solution ratio) with two hours shaking time.
Relationships among Soil Test Procedures
Pautler and Sims (2000) analyzed quantitative relationships between agronomic
STP values and other P measurements proposed as indicative of the potential for
groundwater contamination of P. They tried to assess the soil P test that would be most
easily adopted by routine soil testing laboratories to predict environmental and agronomic
P recommendations. Pautler and Sims (2000) conducted their study on 127 Delaware and
Dutch soils and found that STP (in this study M1-P) was significantly correlated with
total P (r=0.57), oxalate extracted P (Pox) (r=0.84), Fe oxide strip P (r=0.84) and dilute
salt (0.01 M CaC12) extractable P (r=0.71). Their data suggested that STP could be used
to predict the concentrations of Pox and desorbable P (strip P) but less reliable as an
indicator of total or soluble P. Therefore M1-P alone may not be able to predict P loss
from soil to water due to other factors such as hydrology, topography, nutrient and
Pote et al. (1996) found that various STPs were most useful for predicting
dissolved reactive P (DRP) and bioavailable P (BAP) in runoff from a Captina silt loam
(fine-silty, siliceous, mesic Typic Fragiudult) soil. The STP was extracted by six methods
which were M3, Bray and Kurtz P-l, Olsen, distilled water, Fe oxide paper and acidified
ammonium oxalate. They found that the STP correlated best to runoff P when the soil
was extracted with distilled water, acidified ammonium oxalate or Fe oxide paper strips
and the form of P (DRP or BAP) in runoff had very little correlation with STP.
Delgado and Torrent (2001) reported that the release of TP to the freshwater body
was not only dependent on the amount of P that was present in the source but also on the
soil properties, such as P sorbing capacity, the degree of P saturation (DPS) and the form
of P present in the soil or sediment. For agronomic fertilizer recommendation the soil
capacity factor is also a very important factor to be considered (Quintero et al., 2003).
Delgado and Torrent (1997, 2001) suggested that an extraction method using goethite
would be able to predict the total plant available P (TPAP) as well as P released from a
soil. Maftoun et al. (2003) believed that the Olsen method would predict P availability for
rice under both reduced and oxidized calcareous soil conditions better than most other
Various researchers (Sharpley, 1995; Pote et al., 1999a, 1999b; Pierson et al., 2001;
Schroeder et al., 2004) tried to establish relationships between STP and DRP in surface
runoff and suggested that soils with high STP values could indicate significant amount of
P to runoff as dissolved or as particulate bound P. It is very difficult to assign a specific
range of "universally acceptable" STP value beyond which the surface runoff P will be of
environmental concern (Sharpley, 1995; Schroeder et al., 2004) because it was reported
that the relationship between STP and the surface runoff P would be soil and time
specific (Sharpley, 1995; Pierson et al., 2001). Gartley and Sims (1994) reported that
Bray and Kurtz P-l, Ml, M3, Morgan, modified Morgan and Olsen P are the most
commonly used routine STP. Schroeder et al. (2004) conducted a study to establish
relationships between STP and DRP and TP in surface runoff considering all the other
effects like soil types, time of application of manures and soil depths. They used three
extractions: M3 (Mehlich, 1984), Fe203 paper (Myers et al., 1997), deionized water (Pote
et al., 1996) and oxalate extractable P, Fe and Al. They reported that sampling depth had
no significant effect on the relationship between STP and P in runoff (Schroeder et al.,
Agronomic and Environmental STP
Maguire and Sims (2002b) conducted a thorough study in order to establish
relationships between agronomical and environmental soil test P (STP). They chose five
different soil types with a wide range in STP. Intact soil columns were collected for soil
column studies. The leachates were analyzed for DRP and the soils were analyzed for
water extractable phosphorus (WEP) or water soluble phosphorus (WSP), 0.01 M CaC12
P (CaC12-P), FeO-P and Ml and M3 extractable P, Al and Fe. They detected a change
point while comparing relationships between DRP in leachate and P in all of the soil tests
used. That change point is important because below that change point leachate DRP
increased very slowly per unit increase in STP and above which leachate DRP increased
rapidly. Ml and M3 which are mixtures of strong acids, bases, completing agents and
chemicals are generally function by dissolution and desorption reactions which assess the
fertility status of soils (Mehlich, 1984; Sims, 2000; Maguire and Sims, 2002b). However,
soil P tests like FeO-P, WSP, and CaC12-P could measure soluble and easily desorbable P
in soils (Sharpley et al., 1996; Sims et al., 2000; Sims and Coale, 2002, Maguire and
Sims, 2002b) and thus designed as environmental soil tests.
Hooda et al. (2000) compared five different STPs (Olsen, M3, Acidified
ammonium oxalate-oxalic acid, Fe203-coated paper strip and distilled water) and the
amount of P release in water and suggested that the acidified ammonium oxalate was the
least effective while the water extraction correlated best with the amount of P desorbed.
According to Ebeling et al. (2003), it is possible to assess the status of soil P and the
environmentally available P by simply running distilled deionized (DDI) water extraction
or the routine agronomic soil test instead of more expensive environmental soil tests due
to the strong correlations they found between agronomic and environmental soil tests.
Extraction Methods for Wetland Soils
Phosphorus could be present in wetland in various forms such as: dissolved
inorganic P (DIP), dissolved organic P (DOP), particulate inorganic P (PIP) and
particulate organic P (POP) in which dissolved inorganic P fraction could be considered
as bioavailable (Reddy et al., 1999). Sequential extraction methods have been extensively
used to understand P fractions in wetland soils. The early fractionation schemes grouped
soil P into: P sorbed onto the surface of P-retaining components as orthophosphate ions
(nonoccluded P), P present within the matrices of P retaining compounds (occluded P)
and P present in phosphate minerals like apatite (Chang and Jackson, 1957; Petersen and
Corey, 1966; Williams et al., 1971a, 1971b; Reddy et al., 1999). Chang and Jackson
(1957) method of sequential extraction involved 0.5 N NH4F (Al bound P), 0.1 N NaOH
(Fe bound P), 0.5 N H2SO4 (Ca and Mg bound P), Na2S204-citrate (reductant-soluble Fe-
P), 0.5 N NH4F (occluded Al bound P) and 0.1 N NaOH (occluded Fe-Al bound P).
Another more detailed fractionation scheme was proposed by van Eck (1982) with
methods for extracting six P pools as: 0.5 M NaCl (exchangeable P), 1 M NH4C1 (labile
organic P and carbonate bound P), 0.1 M NaOH (Fe and Al bound P), 0.5 M HC1 (Ca
bound P) and resistant organic P. The form of P removed by the first or second extracting
solution in the fractionation scheme is usually considered as bioavailable (Gunatilaka,
1988; Psenner et al., 1988; Reddy et al., 1999) and usual bioavailable extracts are 1 M
NH4C1 (Hieltjes and Lijklema, 1980; van Eck, 1982), 1 M KC1 (Reddy et al., 1995), 0.1
M NaOH (Wildung et al., 1977; Hieltjes and Lijklema, 1980), citrate-bicarbonate-
dithionite (Williams et al., 1971a, 1971b) and nitrolotriacetic acid (NTA) (Gunatilaka,
Wetlands and sediments usually contain high organic matter (OM). Various
physical, chemical, biological and environmental factors control the breakdown of OM
which in turn determines the storage of nutrients in organic pool (Reddy et al., 1999). In
wetlands, the maximum amount of P storage could be in organic form (Stevenson, 1982;
Reddy et al., 1999). Most of the wetland studies, however, focus on inorganic P forms
which are considered bioavailable due to the slow rate of decomposition of OM. Less P is
released from the organic P pool under anaerobic conditions in wetland soils (Reddy et
al., 1999). Due to abundance of OM and various degrees of availability of organic P in
wetland soils, mineralization of organic P could play an important role in contributing to
the bioavailable P (Hedley and Stewart, 1982; Condron et al., 1985).
Reddy et al. (1995) characterized wetland soils and sediments from Lake
Okeechobee watershed using a modified fractionation scheme of Hieltjes and Lijklema
(1980) where they replaced 1 M NH4C1 by 1 M KC1. They characterized stream
sediments and wetland soils with respect to various forms of P. They had low P bound
with Ca and Mg (0.5 M HC1 extractable) but relatively higher portion of P was organic
(0.1 M NaOH extractable) which was reported to be hydrolyzed to transfer into
bioavailable form. Reddy et al. (1995) concluded that wetland soils adjacent to the
streams had higher capacity to retain P and wetland-sediment system functions as sink for
P under most conditions except high rainfall periods. Olila et al. (1995) also used P
fractionation scheme of Hieltjes and Lijklema (1980) to compare various P fractions
between some lakes in Florida and lakes from other country reported in literature. They
reported that readily available P (NH4C1 extractable P) is similar in concentration in
selected lakes of Florida, Hungary, The Netherlands, and Sweden.
Qiu and McComb (2000) used a fractionation scheme (Hieltjes and Lijklema,
1980) to characterize seven wetland sediment soils of Western Australia. The method
separates inorganic P into three fractions: loosely bound P by 1 M NH4C1 extractable P,
Fe and Al bound P by 0.1 M NaOH extractable P and Ca bound P by 0.5 M HCI
extractable P. According to several researchers (Hieltjes and Lijklema, 1980; Bostrom, et
al., 1988) loosely bound P and Fe and Al bound P could be regarded as bioavailable but
Ca bound P may not be directly bioavailable. They determined sediment organic P by the
difference between ashed and non-ashed sediments extracted with HC1. They reported
percentage (%) of their various P fractions to total P (TP) and showed that labile P or
NH4C1 extractable P was generally accounted for 10% of the TP. The same fractionation
scheme developed by Hieltjes and Lijklema (1980), was used by Pulatsu et al. (2003) to
determine the changes in concentrations and seasonal variations of P and Fe in pondwater
and porewater in the littoral zone of a pond in Central Turkey. They reported that the
highest value of concentration of P in porewater, pondwater and sediment were found in
April and October and suggested that it would be easier to control P concentration in
spring and autumn than in summer. Hogan et al. (2004) compared P retention capacity of
two types of wetlands on Kent Island, Maryland, USA. They used a modification of
sequential extraction method of Paludan and Jensen (1995) to identify nine inorganic
(Pi)and organic P (Po) fractions which were: water soluble (H20-Pi and Po), bicarbonate-
dithionite (BD-Pi and Po), sodium hydroxide P (NaOH-Pi and Po), humic acid P (HA-P),
hydrochloric acid P (HC1-Pi) and residual P (Res-P). They also analyzed soils for 0.5 M
NaHCO3-extractable or labile or available P and CHC13-labile P by the method of Hedley
et al. (1982) and microbial biomass P was determined by taking the difference between
NaHCO3 extractable and CHC13 extractable P fractions.
Bruland and Richardson (2004) determined relationships between P sorption, soil
properties and patterns of spatial variability to quantify and determined the soil properties
that best explained the variability of P sorption of two forested riparian wetlands in North
Carolina. They used the phosphorus sorption index (PSI) to quantify the P sorption
capacity of the soils. The PSI was determined by shaking a sterilized soil sample with a
solution of 130 mg P L-1 for 24 hours and by taking the difference in concentration of
inorganic P between initial (130 mg L1) and final concentration representing the amount
of P sorbed. They reported that two forested riparian wetlands with similar vegetation and
hydrology had substantially different mean P sorption capacity due to the differences in
% clay and oxalate extractable Fe and Al. They also reported that oxalate extractable Al
was the best predictor of PSI at each site and P sorption dynamics could vary from site to
site depending on the spatial distributions of soil properties of that particular site.
SOIL TEST PHOSPHORUS AS INDICATORS OF P RELEASE FROM WETLAND
In the upland ecosystem, several soil chemical extraction methods have been used
to determine the availability of phosphorus (P) and other plant available nutrients present
in soil (Sims et al., 2000). A known weight of soil samples is equilibrated with a known
volume of an extractant for a fixed time. The filtrate contains the dissolved plant
nutrients. This filtrate is analyzed for P and other elements using standard methods of
nutrient analyses. These soil tests for P (STP) have been extensively evaluated for upland
soils and have been related to response variables such as plant bioavailability,
leachability and surface runoff (Maguire and Sims, 2002b). There is a growing interest to
apply these methods to saturated and inundated soils to indicate potential thresholds of
nutrient impairment in natural systems. However, wetland soils have very different
characteristics, most notably a significant increase in organic matter, compared to upland
soils. Because of these differences in soil characteristics, questions regarding the
usefulness and relationship between extraction methods and their environmental
implications under flooded or wet soil conditions remain.
Different soil extraction methods for P have been evolved to estimate various forms
of P pool availability. The methods include Mehlich 1 (Ml) (Mehlich 1953), Mehlich 3
(M3) (Mehlich 1984), total inorganic P (1.0 M HC1 extractable) (Reddy et al., 1998), 0.5
M NaHCO3 method (Olsen et al., 1954), water extraction (Kuo, 1996), neutral salt (KC1)
extraction, oxalate extraction (McKeague and Day, 1966; Sheldrick, 1984), Bray and
Kurtz P-l (Bray and Kurtz, 1945) and total soil P (TP) by ashing (Jackson, 1958;
These soil test extractants are used for different types of soils. Bray and Kurtz
(1945) suggested HC1 and NH4F in combination to remove easily acid soluble Al- and
Fe-phosphates P forms. The Ml soil test was developed by Mehlich and his coworkers in
1953 (Mehlich 1953; Nelson et al., 1953) and introduced a combination of 0.05 M HCI
and 0.025 M H2S04 (Ml, also known as the dilute double acid or North Carolina
extractant) to extract P from soils in the north-central region of the U.S. Sulfate ions in
this acid solution can dissolve Al, Fe and Ca phosphates in addition to P adsorbed on
colloidal surfaces in soils. Ml extractant is suitable for acid soils (pH<6.5) and is the soil
test currently used in Florida. Water extraction can remove only dissolved forms of P but
is not suitable for adsorbed, completed or mineral forms of P. The M3 soil test was
developed by Mehlich (1984) as an improved multi element extractant for P, K, Ca, Al,
Cu, Fe, Mn and Zn. The extracting solution of M3 contains CH3COOH, NH4N03, NH4F,
HN03 and EDTA. The chelating reagent EDTA is used to remove the micronutrients
from the soil sample. This multi-element extractant (M3) is suitable for removing P and
other elements in acid and neutral soils. Olsen et al. (1954) introduced 0.5 M NaHCO3
solution at a pH of 8.5 to extract P from calcareous, alkaline, and neutral soils. Total
inorganic P can be extracted by using a 1.0 M HC1 solution (Reddy et al., 1998).
In this research soil extraction methods developed for upland soils have been
evaluated to determine their applicability for predicting release of P to surrounding water
bodies in wetland soils.
Soil Tests Chosen for This Study
The literature review suggests that researchers (Table 3-1) have been using
different soil tests to understand the fate and behavior of environmentally available P.
The most common soil tests chosen to serve the purpose were water extraction, Ml, M3,
oxalate extraction, Bray P-l and Olsen method. Among these extraction methods three
most commonly used methods in the United States (US) are Olsen's NaHCO3, Bray P-l
and Ml (Wolf and Baker, 1985). For this study four different soil test methods (for P)
chosen were Ml, M3, oxalate extraction and water extraction.
Mehlich 1 was developed for acid soils and is easily determined in private and
public analytical laboratories and is the most often used STP in Florida and therefore
chosen as one of the soil tests for evaluation in this study (Nair et al., 2004; Maguire and
Sims, 2002b). Mehlich 3 is a popular extractant in the Southeastern US and is applicable
over a wide range of pH and therefore selected as an STP for evaluation (Mehlich, 1984;
Khiari et al., 2000; Sims and Coale, 2002; Maguire and Sims, 2002a, 2002b). Water
extraction has been used by several workers throughout the world and therefore selected
as a possible indicator of P release from a wetland (McDowell et al., 2001; Sardi and
Fuleky, 2002; Maguire and Sims, 2002b; Elrashidi et al., 2003; Ebeling et al., 2003;
Kleinman et al., 2003; Schroeder et al., 2004; Indiati and Neri, 2004). Oxalate extractable
P, though not used often as an STP, is widely employed in order to calculate the Degree
of P Saturation (DPS) (Pote et al., 1996; Pautler and Sims, 2000; Nair and Harris, 2004;
Nair et al., 2004). According to many researchers oxalate extractable iron (Ox-Fe) and
aluminium (Ox-Al) are important measures of amorphous and poorly crystalline forms of
iron oxides. Oxalate iron is particularly important for evaluation of the P sorption
capacity of a soil especially when the soils are flooded (Khalid et al., 1977; Rhue and
Harris, 1999) as in the case of wetlands. Therefore, oxalate extraction is important in the
research in order to characterize soils in terms ofP sorption capacity and thus included in
Soil Tests Used by Various Researchers
Various researchers used different types of soil tests in order to assess fertility
status of soil in agricultural practices as well as to predict P availability to surrounding
waterbodies. Table 3-1 addresses various soil extraction methods used by some
researchers in last two decades.
Table 3-1. Soil tests used by different researchers to assess environmentally and/or
bioavailable P in soils
Number Soil tests conducted Reference
1. Hot water, Ammonium lactate. Sardi and Fuleky, 2002.
2. M3, Fe203 paper cycle, DI water. Schroeder et al., 2004.
3. NaOH, Bicarbonate, Anion exchange resin (AER), Delgado and Torrent, 2001.
Mixed resin, Iron paper strip, TP.
4. Olsen, Colwell, Brayl, Soltanpour, M3, Morgan, Maftoun et al., 2003.
Sr-citrate and Resin.
5. WSP, Bray P, Olsen P, AER. Elrashidi et al., 2003.
6. DI water, M3, Bray-Kurtz P, Oxalate, AER, Ebeling et al., 2003.
Bioavailable P (BAP) method.
7. Bray P, Olsen P, AEM P, Aslyng P. Quintero et al., 2003.
8. Morgan, Bray 1, Bray 2, Ml, M2, M3, Olsen. Michaelson and Ping, 1986.
9. M3, Olsen, Oxalate. Kleinman and Sharpley, 2002.
10. Ml, M3, Bray P, 1 N ammonium acetate, 0.1 N Gartley et al., 2002.
11. Bray 1, M3, Olsen, Fe strip, Resin capsule, Resin Kleinman et al., 2001.
membrane, Water, CaCl2, Modified Hedley
12. 1 M KC1, Ml, Pi test (Fe oxide coated strip paper). Sarkar and O'Connor, 2001.
13. Ammonium acetate, Bicarbonate, DTPA, KC1, Ion Sherrod et al., 2002.
14. WSP, CaC12- P, FeO-P, Ml, M3. Maguire and Sims, 2002b.
15. Olsen, Bray 1, Mehlich 3, 0.01 M CaC12. Sotomayor-Ramirez et al.,
16. Olsen, Bray 1, Ml, M3. Wolf and Baker, 1985.
17. Ml, M3, AE R, Acidified ammonium acetate. Zones and Piha, 1989.
18. Bray 1, Ml, Olsen. Sharply et al., 1984.
19. Olsen, CaC12- P, M3. McDowell et al., 2001.
20. M3, Bray 1, Ammonium bicarbonate, DTPA, Hanlon and Johnson, 1984.
Table 3-1. Continued
Number Soil tests conducted Reference
21. Bray 1, Ml, Fe Strip, Resin strip, 0.01 M CaCl2. Leal et al., 1994.
22. Bray 1, Bray 2, Ml, Olsen, Truog. Tchuenteu, 1994.
23. Morgan's solution, Oxalate extractable P (Ox-P). Kleinman, et al., 1999.
24. M3, Olsen-P, water, 0.01 M CaCl2. McDowell and Sharpley,
25. Water, M3, Acid ammonium oxalate. Kleinman et al., 2003.
26. 0.03 M NH4F and 0.1 M HC1 (Bray 2). Laverdiere and Karam, 1984.
27. Olsen, Bray 1, Bray 2, M3, Egner-P, CaCl2-P, Indiati and Neri, 2004.
Water, Fe-strip, AERM (resin-P) and 1 anion
28. Bray 1, Olsen, Ml, M2, resin-P, NaOH, NaHCO3, Cajuste et al., 1994.
Materials and Methods
Surface soil (0 to 10 cm) samples were collected from the center (A), edge (B) of
the wetland and adjacent upland (U) areas of the Southeastern United States in 2003 and
2004 (February 2003 to March 2004 and June 2004 onwards, respectively). Soil samples
were collected from three ecoregions: Southeastern Forested Plain (IX), Southern Coastal
Plain (XII), and Eastern Coastal Plain (XIV). The first set of soil samples were collected
from five southeastern states Florida (FL), Alabama (AL), Georgia (GA), South Carolina
(SC), and Indiana (IN) representing different ecoregions for each state : FL (IX, XII), AL
(IX), GA (IX), SC (IX, XII, XIV), IN (IX). The total number of soil samples collected
was 630 of which 331 were impacted and 299 were unimpacted soils. A total of 415 soil
samples were collected in the first stage of sampling (first set) and 215 soil samples were
collected during the second stage (second set) of which all were impacted soils. The
different types of the wetlands were riverine swamp, riverine marsh, non-riverine marsh
and non-riverine swamp. A map of the areas surveyed and a field sampling scheme are
given in Figures 3-1 and 3-2 (adapted from Paris, 2004).
Impacted sites = 94
A Least impacted = 115
Eastern Coasal Plain (XIV) \
Southern Costal Plain (XII)
Southeastern Forested Plain (IX)
Figure 3-1. Map of the locations of the soil samples collected during the Southeastern
Wetland Biogeochemical Survey (SWBS) study (Adapted from Paris, 2004).
Soil pH was determined by using 1: 2 soil to solution ratio on air-dried soils
(Hanlon, 1994; Thomas, 1996). Other parameters were determined as follows: soil bulk
density by drying a known weight of wet soil for 72 hours at 70C (Grossman and
Reinsch, 2002), loss of ignition (LOI) to measure organic matter content (Nelson and
Sommers, 1996), total carbon (TC) (Nelson and Sommers, 1996), and total P (Jackson,
1958; Anderson, 1976; USEPA, 1993).
B) Small non-rlverlne
C) Large non-riverine
Figure 3-2. Field sampling scheme during the SWBS study (Adapted from Paris, 2004).
The composite soil collected from each site was thoroughly mixed and pH and bulk
density were determined. The soils were air-dried, ground and passed through 100 mess
sieve. Soil samples were analyzed for P by Mehlich 1 (Ml), Mehlich 3 (M3), water
extraction and oxalate extraction. Water soluble extracts were analyzed for P
colorimetrically by molybdate blue method (Murphy and Riley, 1962) using a Technicon
II colorimetric auto analyzer using USEPA Method 365.1 (USEPA, 1993). All other
extracts for P and other extractable metals were analyzed by inductively coupled plasma
atomic emission spectroscopy (ICP-AES).
Methods of Different Extraction Processes Employed
The following four methods of extraction were employed:
Water Extractable Phosphorus (Kuo, 1996)
* Weigh 2 g dry soil into labeled 50 mL polypropylene centrifuge tubes.
* Add 20 mL of distilled deionized water (DDI) into tubes & place in a reciprocating
shaker at low speed for 1 hour.
* Centrifuge for 10 min at 6000 rpm.
* Filter the supernatant through 0.45gm filters into 20 mL HPDE scintillation vials.
Mehlich 1 Extraction (Mehlich 1953, Nelson et al., 1953)
* Weigh out 5 g. of soil into a 100 mL centrifuge tube.
* Add 20 mL of the double acid extractant solution and shake for 5 mins.
* Immediately filter through a no. 41 filter paper.
Mehlich 3 Extraction (Mehlich 1984)
* Weigh 3 g. of soil into a 100 mL centrifuge tube.
* Add 25 mL of extracting solution to each tube and shake for 5 mins.
* Immediately filter through a no. 42 filter paper.
Oxalate Extraction (McKeague and Day, 1966; Sheldrick, 1984)
* Weigh out 0.5 g. soil.
* Add 20 mL of oxalate reagent.
* Shake in dark for 4 hours.
* Centrifuge at 6000 rpm for 10 mins.
* Filter with 0.45 [m filter paper with the lights off
Simple linear regression analyses, correlation of all parameters and different plots
were performed with the Data Analysis Tool Pack in Excel 2003 (Microsoft Office XP
Professional, 2003). Stepwise regression analyses were run on combinations of different
sets of dependent and independent variables by using SAS (SAS Institute, 2001).
Regression and correlation analyses ran with JMP version 4.0 (SAS Institute, 2000) and
Minitab version 14.0 (Minitab, 2004) software packages.
Results and Discussions
Several studies found that STP is related to the concentration of soluble P as
leachable P, surface or subsurface drainage or as dissolved reactive P (DRP) (McDowell
et al., 2001) by two different linear relationships having statistically significant different
slopes. In this study, soil tests like Ml, M3 and oxalate will be compared to water
extractable P. The soils studied in this research have a wide range of soil physical
characteristics (Table 3-2).
Table 3-2. Range of selected physical characteristics of the soils in this study
Soil characteristics First set of samples Second set of samples
pH 3.29 to 7.53 3.0 to 7.26
Bulk density (g cm-3) 0.02 to 2.4 0.05 to 1.6
Loss on ignition (%) 0.5 to 98.2 1.4 to 93.8
Soil test for P using different extraction processes is a well-established agricultural
practice worldwide by which the soils under investigation could be characterized as P
deficient or the vice-versa (Sims et al., 2000). Research on environmental soil P testing
has been going on for more than a decade but most soil scientists agree that these
chemical soil tests can determine the concentration of soluble, biologically available and
potentially desorbable P in soils, but they provide little information on the transport
processes and management practices which have strong influence on the movement of P
through runoff or by leaching (Sims et al., 2000). There is a trend in establishing
relationships of water soluble P or CaC12 soluble P (CaC12-P) as surface runoff dissolve
reactive P (DRP) or leachate P against different STPs like M1-P, M3-P, Ox-P, Olsen P,
bioavailable (BAP) (measured by 0.1 M NaOH), oxide strip P etc. (Dorich et al., 1985;
Sallade and Sims, 1997; Sims et al., 2000; McDowell and Sharpley, 2001; McDowell et
al., 2001; Maguire and Sims, 2002b). In our study we have included M1-P, M3-P, Ox-P
and WSP as four different soil tests because these four are the most common, convenient
and general soil tests for P used in parts of United States (US) and elsewhere in the
A wide range of extractable soil P was observed for each of the soil tests used in
this study (Table 3-3). The variability of four extraction methods used in this study
(irrespective of location and other soil properties associated) is given in Table 3-3. This
table shows that: (1) the standard deviation (SD) and range of concentration of P of each
soil test show the variability and wide range of the physical and chemical characteristics
of the samples in this study. This wide range and variability of STPs indicates that the
location and other physical and chemical properties have profound influence on the
samples under investigation; and (2) the P concentration of a wetland soil varies as Ox-P
> M3-P > M1-P > WSP which is the same trend as reported elsewhere for upland soils
(Maguire and Sims, 2002b; Nair et al., 2004).
Table 3-3. Mean, standard error (SE), median, mode, standard deviation (SD) and range
for WSP, M1-P, M3-P and Ox-P for all wetland samples in this study.
Criteria WSP (mg kg-1) M1-P (mg kg1) M3-P (mg kg1) Ox-P (mg kg-1)
Mean 8.4 38.5 43.5 286
SE 1.2 5.8 2.8 21
Median 0.99 17 30.3 199
Mode 0.2 21.6 34.4 290
SD 21 101.3 49.6 359
Range 0 to 236 0.8 to 922 0 to 330 14 to 3222
Mehlich 1 or Mehlich 3 were developed to assess fertility status of soils and
therefore regarded as agronomic soil tests (Mehlich, 1984; Sims, 2000; Maguire and
Sims, 2002b). However soil P tests such as WSP or Ox-P were developed more as
environmental soil tests (Sharpley et al., 1996; Sims et al., 2000, Sims and Coale, 2002;
Maguire and Sims, 2002b). Correlation of different STP (Table 3-4) is similar but weaker
than upland soils from findings of some other researchers (Sims, 1989; Sallade and Sims,
1997; Maguire and Sims, 2002b). There is significant (p < 0.0001) strong correlation
between different soil test P except WSP (Table 3-4) which is attributed to high organic
matter (OM) content with mean LOI 19.4 %, range (0.5% to 98.2%) (Table 3-2). All
samples were divided into three groups by location, as center of wetland (A), edge of
wetland (B) and upland (U). The correlation of M1-P and M3-P for all the soils improves
significantly when soils were separated based on the location (Figure 3-3). The
correlation between Ml and M3 extracted phosphorus for 513 samples was less strong
for center of wetland (A) (R2 = 0.46***) and moderate for edge of wetland (B) (R2
0.55***) but strong for adjacent uplands (U) (R2 = 0.81***) (Figure 3-3).
Water soluble P varies with OM (Figure 3-4). This finding supports the conclusion
of Maguire and Sims (2002b) that acidic, high OM soils behaved differently from the
other soils in their study. From these two initial findings it could be concluded that the
relationships among soil tests are site specific and likely dependent on the soil organic
The correlation of WSP is highly significant (p < 0.0001) with respect to M3-P
irrespective of location of the samples. Mehlich 3 is a strong extractant compared to other
soil test Ps (STP) and M3 would work over wide range of soil pH (Maguire and Sims,
2002b). Therefore, M3-P could be used as a predictor of wetland's bioavailable P
Table 3-4. Correlation coefficients of different soil tests on 307 wetland samples
Correlation matrix WSP M1-P M3-P
M1-P 0.096 _
M3-P 0.265*** 0.678***
Ox-P 0.043 0.664*** 0.624***
Table 3-5. Correlation of different extractable P with WSP (by location)
Correlation (r) with A (n = 146) B (n = 148) U (n = 114)
M1-P 0.106 0.077 0.273*
M3-P 0.342*** 0.297*** 0.255*
Ox-P 0.012 0.072 0.068
TP 0.085 0.082 0.036
, *,p < 0.05, ***,p < 0.0001; A, center of wetland, B, edge of wetland, U, upland
y = 0.95x + 14.0
R= 0.46 (A), n = 196
0 50 100
M1-P (mg kg1)
* Center of wetland (A) Edge of wetland (B)
y = 1.30x + 8.1
S R2= 0.56 (B), n = 195
R= 0.81 (U), n = 122
Figure 3-3. Relationship of M1-P and M3-P for all soils by location (The symbols A
represents center of wetland, B represents edge of wetland and U represents
upland adjacent to the wetlands) [p < 0.0001].
Literature review suggests that for upland soils in most of the cases the relationship
between M1-P and M3-P gives a strong correlation (Maguire and Sims, 2002b). The
important findings from Figure 3-3 is that the simple regression coefficient (R2) and the
slope of the regression line both increase in the order from center of wetland to the
upland through edge of the wetland (Figure 3-3) which means that the location of the soil
does have a significant importance on the P extracted by different procedures.
WSP is related (R2 = 0.56***) to the soil organic matter (Figure 3-4, Tables 3-6,
and 3-7). WSP is also well correlated to the TC (R2 = 0.88***) as indicated by the TC
and OM relationship (Figure 3-5 and Table 3-7).
y = 0.0097x2 0.22x + 4.4
R2 = 0.56
Sn n=409 ** *
0 20 40 60 80 100
Figure 3-4. Correlation between WSP and organic matter; p < 0.0001, only wetland soils
60y = 0.49x- 1.24
R2 = 0.88
40 4 n = 594
0 * -
0 20 40 60 80 100 120
Figure 3-5. Correlation between organic matter (OM) and total carbon (TC) content; p <
0.0001, both upland and wetland soils included.
The role of OM in P sorption-desorption characteristics is still not clear (Afif et al.,
1995) but it has been reported to be linked to a decrease in P sorption (Barrow, 1989).
McDowell and Sharpley (2001) proposed that OM could occupy sorption sites and
therefore their grassland soils with larger amounts of OM increased the desorption
potential of loosely bound P compared to their arable soils with lower OM.
Mandal (1963) showed that when an easily decomposable organic matter source is
incorporated or included in waterlogged soils then the soluble P release is increased in the
waterlogged soils and eventually Ca bound P is decreased. The possible reason was
attributed to the solubilizing effects of CO2, or the organic acids produced as a result of
microbial decomposition of Ca bound P (Mandal, 1963; Sallade and Sims, 1997). Mandal
(1963) suggested that the large amount of CO2 has had the solubilizing effect on
insoluble tri-calcium phosphates; in other words this solubilizing effect could convert
least soluble tri-calcium phosphates to more soluble di-calcium phosphates.
WSP is strongly associated with loss of ignition (LOI) which is the measure of
organic matter content of the sample (Figure 3-4, 3-5). The amount of WSP in our case is
generally high with a wide range (mean 8.4 mg kg1, standard deviation 21; Table 3-3) in
concentration. Figure 3-4 shows that 56% variability of WSP could be explained by OM
alone. From these findings it may be hypothesized that high amount of WSP and
relationship with OM indicate that water extractable P is loosely sorbed on the large
surface area of OM which was reported elsewhere (Sallade and Sims, 1997; McDowell
and Sharpley, 2001).
Table 3-6. Comparison of sediment and wetland soils with respect to water soluble P
(WSP) and organic matter (OM) content
Parameter Source R2 References
Soluble P vs. Sediments 0.462 Sallade and Sims, 1997
Organic Matter Wetlands 0.562*** Mukherjee, 2005 (This study)
***,p < 0.0001
A study of the potential for P release from sediments to drainage water was
determined by incubating sediments under flooded conditions (Sallade and Sims, 1997).
Phosphorus released after 21 days was well correlated to DPS (r= 0.75 ***), calculated as
the ratio of bioavailable P to the P sorption index (PSI). The initial amount of P in
solution was well correlated to the organic matter content as is the situation in our studies
(Table 3-6). They attributed a strong correlation between WSP and organic matter to
loosely sorbed P being retained by organic matter (Sallade and Sims, 1997). Correlation
coefficients (r) of different STPs and other physical parameters irrespective of location of
the samples assuming linear relationships are given in Table 3-7.
Table 3-7. Correlation coefficients of strongly associated parameters
All Two Years Data Set
Parameters Associated Correlation Coefficient (r)
WSP versus OMt 0.719
WSP versus TC 0.749
M1-P versus M3-P- 0.647
M1-P versus TP 0.764
M1-P versus Ox-P 0.69
M3-P versus TP 0.661
M3-P versus Ox-P 0.653
Ox-P vs. TP 0.935
LOI versus % TC 0.921
p < 0.0001; t, WSP is better related to OM as an exponential equation (Figure 3-4); +,
M1-P versus M3-P is dependent on the location of sampling (Figure 3-4). The correlation
coefficient in this table is for all locations considered together.
A brief comparison of upland, sediment and wetland on the basis of M1-P as the
function of different extractable P is given in Table 3-8. This table shows the difference
of different soil extractions conducted on uplands, sediment and wetlands. The wetland in
the third column of Table 3-8 refers to the wetland soils in this study. The correlation
coefficient (r) of WSP and M1-P is very poor (0.1) in the case of wetland, moderate
(0.53) for sediment and good (0.71) for upland (Table 3-8).
Table 3-8. Comparison of upland and wetland soils
M1-P Upland Sediment Wetland
Reference (i) (ii) (iii)
TP 0.57 ND 0.54
Ox-P 0.84 ND 0.43
Fe-Strip P 0.84 ND ND
0.01 M CaC12 P/
WSP 0.71 0.53 0.01
(i) Pautler and Sims, 2000
(ii) Sallade and Sims, 1997
(iii) Mukherjee, 2005 (This study)
ND, not detected.
Wetland soils are a more complicated ecosystem compared to uplands with P
release often associated with anaerobic conditions, particularly when the Fe concentration
of the soil is high. A better indicator of P release could be from P flux experiments but
such evaluations would not be practical for quick determinations. Water extraction does
not appear to be a good indicator of P release from wetland soils. The WSP values
reported here were on air-dried basis. It is likely that WSP done on wet samples could
have a better relationship with other STPs like Ml and M3. Relationship of Ml and M3
is moderate in the cases of center of wetland and edge of wetland but strong for uplands.
Therefore, soil sampling location has strong influence on these extraction methods.
For upland soils, it has been found that the P leaching or runoff properties of a soil
from different agricultural fields have varying soil chemical and physical properties
which could affect the retention and transport of P (Sims et al., 2000; Delgado and
Torrent, 2001; Maguire and Sims, 2002b). Sharpley and Tunney (2000) discussed the
importance of addressing concerns for four areas of research, namely, soil P testing for
environmental risk assessment, pathways of P transport, best management of practice
(BMP) development and implementation and strategic initiatives to manage P. STP alone
can quantify the concentration of bioavailable, soluble and potentially desorbable P in
soils but fails to provide information on the transport processes involved (Sims et al.,
Potential P release STP indicators for wetland soils from this study include M1-P
and M3-P. However, soil testing alone may not be sufficient as a predictor of P release
from wetland soils since STP does not take into account for the P retention capacity of a
soil. Therefore the comparison of STP in conjugation with different soil chemical and
physical properties and P losses to water are considered in the next chapter.
INDICATORS FOR ENVIRONMENTAL PHOSPHORUS EVALUATION
There is a critical need to establish a practical indicator to assess the ability of P
movement from a particular upland site to surface or ground waters by surface runoff,
subsurface drainage or leaching (Maguire and Sims, 2002b; Nair et al., 2004). The
concept of Degree of Phosphorus Saturation (DPS) for sandy soils was first introduced by
scientists of Netherlands (van der Zee et al., 1987; van der Zee et al., 1988; Breeuwsma
and Silva, 1992) and now it is widely accepted across the world as an indicator of P
release from a soil (Maguire and Sims, 2002b; Nair and Harris, 2004). However, there is
no alternative indicator for P release from wetland soils that has been identified so far.
A method of determining environmentally available P by using DPS or phosphorus
saturation ratio (PSR) against WSP or CaC12-P, M1-P or M3-P by several researchers in
upland soils (van der Zee et al., 1987; Breeuwsma and Silva, 1992; Sallade and Sims,
1997; McDowell et al., 2001; Maguire and Sims, 2002b; Kleinman et al., 2003; Nair et
al., 2004; Nair and Harris, 2004) has been used in this study in order to determine
environmentally available P in wetlands.
The iron and aluminium bound P (Fe-P and Al-P, respectively) values are taken
into consideration in the Phosphorus Saturation Ratio (PSR) and DPS calculations (Nair
et al., 2004).
The equations of PSR and DPS used are given below:
PSR = (P)/(Fe + Al), (Nair et al., 2004) [Equation 1];
DPS = [P/a*(Fe + Al)]*100; where a = 0.5, (Nair et al., 2004) [Equation 2];
Where, P, Fe, and Al are Ml, M3 or Oxalate extractable and expressed in mmoles
in Equations 1 and 2.
In DPS calculations there is an empirical factor, a, which researchers use with
different values based on the soil types (Pautler and Sims, 2000; Nair et al., 2004). In
order to avoid complexity, and as PSR will give similar information to DPS, PSR will be
used in this study.
From the previous chapter, we note that P release from wetland soils is strongly
influenced by organic matter. Therefore, there will be some concern on the use of (Fe +
Al) as a surrogate for P sorption in wetland soils. Oxidation reduction changes could play
an important part in P solubility and sorption mechanisms (Reddy et al., 1998). Further,
hydrated Fe oxides associated with Al and organic matter in gel complexes has been
shown to control inorganic P sorption in lake sediments (McCallister and Logan, 1978).
However, Reddy et al. (1998) found that P retention by stream sediments and wetland
soils was strongly correlated with Fe and Al, and that adding total organic C to predictive
equations improved the variability only slightly, ie., from 87% to 92%. For almost all
practical purposes it should be possible to use (Fe + Al) as an indicator of the P sorption
capacity of wetland soils.
Some soil tests could be predictors of soluble P, desorbable P and DPS (Pautler and
Sims, 2000). Our results (previous chapter) also show the ability of some soil tests as
predictors of soluble P in wetland soils. Degree of P saturation from oxalate extraction
(Ox-DPS) has been suggested as an indicator of P leaching from agricultural soils
(Breeuwsma et al., 1995; Hooda et al., 2000; Maguire et al., 2001; Nair et al., 2002a,
2002b; Nair et al., 2004; Nair and Harris, 2004). Pautler and Sims (2000) have shown
that the agricultural soils will meet the adequate crop P supply and environmental
protection both at the same time for those soils below an Ox-DPS threshold. But as the
oxalate extraction is not a routine soil test and as Ox-DPS values are not widely
available; Khiari et al. (2000) reported that M3 P/A1 ratio could be useful to predict both
the crop P requirement as well as environmental P threshold above which excess P will
Maguire and Sims (2002a) conducted a study where they compared M3 P
saturation ratio (M3-PSR) with Ox-DPS and they evaluated M3-PSR for predicting
agronomic and environmental soil P saturation thresholds. They collected soils from five
soil series having wide ranges of physical and chemical properties measured. Soils were
extracted by M3 and oxalate methods and analyzed for P, Fe and Al. They calculated
M3-PSRs in two different ways. M3-PSRs were calculated as molar ratios of M3
extractable P/[A1 + Fe] (ratio I) and P/A1 (ratio II). They found that M3-PSRs (ratio I and
ratio II) were well correlated to each other and also to Ox-DPS and all of these P
saturation measurements showed a change point above which the concentration of P in
the column leachate increased rapidly. They also reported that the coefficient of
determination indicated that the M3-PSR ratio I was better than Ox-DPS at predicting
losses in the column leachate. Maguire and Sims (2002b) showed that M3-PSR was well
correlated to the DPS based on an acid ammonium oxalate extraction which they utilized
to suggest a potential risk for P loss from soils to surface and eventually groundwater.
They have also indicated a 0.10 to 0.15 range of M3-PSR to represent a risk of P loss to
surface and groundwater.
As it was reported elsewhere that traditional routine soil tests for P alone cannot
predict the P movement due to difference in soil's physical and chemical properties (Sims
et al., 2000; Delgado and Torrent, 2001; Maguire and Sims, 2002b), so Maguire and Sims
(2002b) employed three different environmental soil P tests (WSP, CaCl2-P and FeO-P)
and three agronomic soil P tests (M1-P, M3-P and M3-PSR) to predict dissolved reactive
phosphorus (DRP) concentrations in leachate and compare environmental and agronomic
soil P tests. They reported that the environmental soil P tests were slightly better to
predict DRP concentrations than that of the agronomic soil P tests but M3-PSR was as
good as any of the environmental soil tests to predict the same.
An intensive study by Sims et al. (2000) showed that there was statistically
significant linear relationships between M3 [P/(A1 + Fe)] and [M3 (P/A1)] ratios and
DPSox and suggested that the soil testing laboratories could use M3 test after developing
environmental interpretations related to P saturation. They also compared the M3 [(P/(Al
+ Fe))] ratio with DRP in runoff from soils ranging in STP from low to excessive P using
a rainfall simulation box study method and dissolved reactive P (DRP) in leachate from
undisturbed column. They found in both the cases that there was a distinct "change point"
for M3 [(P/(A1 + Fe))] ratio and above that value DRP in runoff or column leachates
increased rapidly. So, they suggested that split line models using M3 [(P/ (Al + Fe))] ratio
is the better predictor of DRP in runoff and column leachate than M3 alone.
The M3-PSR is a combination of agronomic STP and as well as environmental STP
(Khiari et al., 2000; Sims and Coale, 2002; Maguire and Sims 2002a, 2002b). For upland
soils, WSP and DPS have been used as indicators of P loss to surface and ground waters
(van der Zee et al., 1988; Leinweber et al., 1999; Maguire and Sims, 2002a; 2002b). Both
WSP and DPS have been cited as better P release indicators than agronomic soil P tests
(Sims et al., 2000). Schroeder et al. (2004) calculated DPS and reported that the strongest
correlations existed with total P and DRP with DPS. Attempt was made to use the
information from upland studies to wetland conditions in this study. The overall objective
for this study was to find at least a soil test or soil extraction method which could predict
P loss from wetlands to surrounding water bodies and attempt to establish threshold point
beyond which the wetland soils could be assigned as problematic.
Materials and Methods
Soil tests for P including Ml, M3 and oxalate extractions were carried out. In
addition to P determination, Fe, Al, Ca and Mg were performed on the Ml and M3
extracts, and Fe and Al in the Oxalate extracts. Water extraction for P and TP by ignition
ash method also conducted on the same soils. The details of these soil tests are given in
the previous chapter.
Based on Equation 1 the different PSR were analyzed using following equations:
PSR using Mehlich 1 P, Fe and Al (M1-PSR):
M1-PSR = (M1-P)/(M1-Fe+M1-Al) [Equation 3]
PSR using Mehlich 3 P, Fe and Al (M3-PSR):
M3-PSR = (M3-P)/(M3-Fe+M3-Al) [Equation 4]
PSR using Oxalate P, Fe and Al (Ox-PSR):
Ox-PSR = (Ox-P)/(Ox-Fe+Ox-Al) [Equation 5]
In the above equations 3, 4 and 5 P, Fe and Al are expressed as mmol kg1.
Simple regression analyses were run by using Excell 2003 (Microsoft Office XP
Professional, 2003). Other regression and correlation analyses computed with JMP
version 4.0 (SAS Institute, 2000) and MINITAB version 14.0 (Minitab, 2004) software
packages. Stepwise regression analyses ran with different sets (sets 1 and 2) of
independent variables pH, BD, LOI, Ml, M3 and Ox-extractable P and metals, and Ml-
PSR, M3-PSR and Ox-PSR to predict variability of WSP by using SAS (SAS Institute,
Results and Discussions
As WSP gives change point with DPS or PSR analyses in upland soils, the same
relationships have been used for wetland conditions and the correlations (r) of WSP of
311 wetland soils against different PSR analyses (Table 4-1) show that WSP is
significantly well correlated with M3-PSR in these wetland soils. M3-PSR was shown to
be better in predicting P losses by leaching than M1-PSR or Ox-PSR which supports the
findings of other researchers under upland conditions (Khiari et al., 2000; Sims and
Coale, 2002; Maguire and Sims, 2002b). Stepwise regression analysis (Table 4-2) for
WSP and correlation of WSP with M3-P (Table 3-5) also agree with these findings.
Table 4-1. Correlation (r) of PSR analyses with water soluble P on 311 wetland samples
Correlation [r] WSP M1-PSR M3-PSR
M1-PSR 0.398 -
M3-PSR 0.627 0.631
Ox-PSR 0.197 0.416 0.584
Significant atp < 0.0001 level
The influence of organic carbon in predicting WSP for the entire two data sets was
studied by dividing into two different categories: 0 to 12 % TC, and greater than (>) 12 %
TC. The WSP and PSR relationships for wetland soils are presented in Figures 4-1, 4-2
Figures 4-1, 4-2 and 4-3 show that at approximately 0.1 M1-PSR, 0.1 M3-PSR and
0.1 Ox-PSR (for < 12% TC) WSP becomes suddenly elevated. The change point is not
S 20 .
% 12% TC, n = 236
12% TC, n = 48
0 0.05 0.1 0.15 0.2 0.25
A (1) 0% 12% TC O (2) > 12% TC
1. Water soluble P (WSP) as a function
12% total C and > 12% total C.
(1) 0% 12% TC, n =
1 (2) > 12% TC, n =49
of M1-PSR for wetland soils with <
0 0.05 0.1 0.15 0.2 0.25 0.3
A (1) 0% 12% TC (2) > 12% TC
2. Water soluble P (WSP) as a function of M3-PSR for wetland soils with <
12% total C and > 12% total C.
1) 0% 12% TC, n = 287
2) > 12% TC, n = 56
* Fi A"
AA + Au
0 0.05 0.1 0.15 0.2 0.25 0.3 0.35
A (1) 0% 12% TC m (2) > 12% TC
Figure 4-3. Water soluble P (WSP) as a function of Ox-PSR for wetland soils with <
12% total C and > 12% total C.
that sharp as predicted for upland soils elsewhere (Nair et al., 2004). For wetland soils
with TC > 12% this change point seems to be probably lower and about 0.05 PSR range.
These evaluations are based on the WSP concentrations as determined on air-dried soils.
The wetland soils with high OM (> 12% TC) also exhibit high WSP because P is loosely
bound on the large surface area of OM and thus when the PSR is low WSP is high.
The WSP versus PSR graphs (Figure 4-1, 4-2 and 4-3) show two different TC
ranges where there is a need to assign a threshold value or change point above which PSR
values suggest environmentally problematic situations. For soils having < 12% TC, the
threshold PSR is likely higher than for the soils having > 12% TC. The change point for
WSP-PSR relationships for Florida upland soils were 0.1 for WSP and M1-PSR, 0.1 for
WSP and Ox-PSR and 0.08 for WSP and M3-P (Nair et al., 2004). For those wetland
soils where TC is less than 12%, these change points may be reasonable as well.
Stepwise regression equations (Equations 6, 7, and 8) for WSP with independent
variables LOI, pH, BD, and Ml, M3, and Ox-extractable P, and metals indicate BD and
LOI as variables contributing to the prediction equation for Ox-PSR < 0.10 and Ox-PSR
> 0.10.These equations show the influence of BD and LOI as a measure of OM to predict
For Ox-PSR < 0.10:
WSP = 3 1.4 pH + 2.5 BD* + 0.4 LOI*** 0.01 M1-Al*** (R2 = 0.4089),
WSP = (9.9) 1.9 pH*** + 0.48 LOI*** 0.0002 Ox-Fe* 0.003 Ox-Al* (R2
0.4768), [Equation 7]
For Ox-PSR > 0.10:
WSP = (4.2) + 0.38 LOI* + 0.05 M1-Ca*** 0.19 M1-Mg*** (R2 = 0.5658).
We therefore normalized the WSP concentration (i.e. from mg kg-1 to ptg cm-3) to
take bulk density into consideration in our WSP concentration. The calculation was to
<12% TC o >12% TC
10 lP D
0 0.1 0.2 0.3 0.4 0.5
Figure 4-4. Water soluble P (WSP) normalized with respect to bulk density as a function
of M1-PSR for soils < 12% total carbon and > 12% total carbon
* <12% TC O >12% TC
I. .s 'A
Figure 4-5. Water soluble P (WSP) normalized with respect to bulk density as a function
of M3-PSR for soils < 12% total carbon and > 12% total carbon
* < 12% TC o >12% TC
0.1 0.2 0.3
Figure 4-6. Water soluble P (WSP) normalized with respect to bulk density as a function
of Ox-PSR for soils < 12% total carbon and > 12% total carbon
simply multiply BD and the concentration of WSP of a particular soil to normalize WSP
from weight basis to volume basis with the unit [tg cm-3. The resulting plots are presented
in Figures 4-4, 4-5 and 4-6. It is difficult to confirm if this "normalization" allows better
interpretation of the data. The change point is still not as sharp as in various studies of
upland soils (McDowell et al., 2001; Maguire and Sims, 2002b; Nair et al., 2004).
Mehlich 3 phosphorus saturation ratio (M3-PSR) had a strong influence in
predicting WSP in wetland soils (Table 4-2). Similar finding for upland soils are
described elsewhere (Khiari et al., 2000; Sims and Coale, 2002; Maguire and Sims,
2002b; Nair et al., 2004).
Table 4-2. Prediction equations for water soluble P with two different sets of
Set number Prediction Equations
1 WSP = (- 7.8) + 0.37 LOI*** + 160.04 M3-PSR*** [R2 = 0.5221]
2 WSP = (- 9.5) + 3.3 BD + 0.35 LOI*** + 0.04 Ox-Ca 0.0009 Ox-Al +
10.3 M3-PSR*** [R2 = 0.4333]
***, significant atp < 0.0001
Total P (TP) also gave a significant regression coefficient (R2 = 0.60, p < 0.0001)
against Ox-PSR with a polynomial second order curve (Figure 4-7). Total P is not
currently used as one of the STPs or as an indicator of environmentally available P.
However some researchers (Pautler and Sims, 2000) have shown that a percent of easily
desorbable P could be calculated by using the ratio of Ox-P: TP or FeO strip P: TP and
could be used as an indicator of P release to the surrounding water bodies. Lookman et al.
(1995) reported that the all form of oxalate extractable P should be regarded as reversibly
adsorbed or reversibly fixed P and Fe-oxide P represents as "fast desorbing pool" of soil
P. Pautler and Sims (2000) calculated Fe-oxide strip P to Ox-P as the ratio in Y axis with
R2 = 0.49. They suggested that the impacted soils with low concentrations of [Ox-Fe +
Ox-Al] could be susceptible to P leaching.
y = 7655x2 + 2721x + 310
R2 = 0.60
n = 413
%/ I - I I I I I I
0 0.1 0.2 0.3 0.4 0.5 0.6
Figure 4-7. Total P as a function of Ox-PSR for all wetland soils, p < 0.0001
y = 0.50x- 12
R2 = 0.86***
0 1000 2000 3000 4000 5000 6000 7000
TP (ng kg1)
Figure 4-8. Oxalate-P (Ox-P) as a function of total P (TP);p < 0.0001
In our study, approximately 50% (R2 = 0.86, p < 0.0001, Figure 4-8) of TP is oxalate
extractable, suggesting that half of the TP is reversibly adsorbed, and likely to be released
to the water column.
One of the major questions to be addressed is the upper, environmentally based
critical limit for soil test P such as M1-P and M3-P for determining the water quality in a
wetland. Total P (TP) is a potential P release indicator for a wetland soil, thus there could
be an upper limit set for environmentally sound wetland conditions. The background
concentration of total P (TP) of all the center of wetlands of this study is approximately
550 mg kg-1 calculated from the 75th percentile of the distribution of all the unimpacted
wetland soils of this study according to the United States Environmental Protection
Agency (USEPA) guideline (USEPA, 2000). Based on this reference background
concentration of TP = 550 mg kg-1 the threshold values for Ox-PSR will be 0.073
according to the second degree equation of Figure 4-7. The other threshold values were
calculated based on the background TP = 550 mg kg-1 of all the wetlands surveyed in this
study from statistically significant linear or curvilinear relationships of different STPs
and PSRs (Table 4-3) and those are M1-P = 23 mg kg-1, M3-P = 41 mg kg-1, Ox-PSR
=0.073, M1-PSR = 0.076, and M3-PSR = 0.057 (Table 4-3).
The historical background TP concentration in the Water Conservation Area 2A
(WCA-2A) in the northern Everglades, Florida, has been estimated as 500 mg kg-1
(DeBusk et al., 2001). Using satellite image of vegetation distribution and soil TP data
(DeBusk et al., 1994), Wu et al. (1997) predicted cattail invasion into the indigenous
sawgrass marsh at a TP concentration of 650 mg kg-'.The threshold values based on
background concentration of TP = 550 mg kg-1 for our wetland soils irrespective of types
of wetlands are given in Table 4-3.
Table 4-3. Threshold values for P release indicators for wetland soils
Variables Background Concentration Reference
TP 550 (mg kg1) Figure 4-7
M1-P 23 (mg kg-) Figure 4-9
M3-P 41 (mg kg') Figure 4-11
Ox-PSR 0.073 Figure 4-7
Gartley and Sims (1994) rated fertility status based on 8980 upland soils collected
from four regions of the US (northeast, midwest, mid-Atlantic, and southeast) with Ml
soil test. According to them the categories identified were : (i) 0 to 6 mg kg-1 M1-P as
very low, (ii) 7-15 mg kg-1 M1-P as low, (iii) 16-30 mg kg-1 M1-P as medium, (iv) 31-60
mg kg-1 M1-P as high and (v) > 61 mg kg-1 M1-P as excessive. It was also reported that
the soils having M1-P > 30 mg kg-1 should require more intensive management. Florida's
suggested threshold M1-P is 30 mg kg-1 (Nair et al., 2004) also supports the findings of
Gartley and Sims (1994). The calculated threshold value of M1-P (33 mg kg-1, Table 4-3)
for the wetland soils fall in the high risk category designed for upland soils.
Several researchers (Sims et al., 2001; Sims and Coale, 2002) proposed four agri-
environmental M3-P limits in order to optimize crop production and as well as to ensure
the minimization of non point source P pollution: (i) < 50 mg kg-1 P, where profitable
crop response is expected, (ii) 50 to 100 mg kg-1 P, where P should be considered
adequate, (iii) 101 to 150 mg kg-1 P, where no P application is recommended and (iv) >
150 mg kg-1 P, high risk of P losses by erosion, runoff or leaching (Maguire and Sims,
2002b). The numbers (ii) and (iii) are considered as intermediate between agronomicc"
and "environmental" categories (Maguire and Sims, 2002b). Maguire and Sims (2002b)
found change points for M3-P as 181 mg kg-', M1-P as 81 mg kg-1, WSP as 8.6 mg kg-1
and M3-PSR as 0.2. These agronomic and environmental categories were suggested for
upland soils. However, for the wetland soils in this study, there is no change point for
M3-P and Ox-PSR relationship. The relationship is linear (R2 = 0.68, p < 0.0001, Figure
Soil test P is a common determination in Florida in both private and public
laboratories (Nair et al., 2004). Also for upland soils it is well established that M1-P and
M3-P are strongly correlated to neutral salt solution, leachate P, soluble reactive P (SRP),
dissolve reactive P (DRP) or in rainfall simulation which all are practically water solution
(Sims et al., 2000; Pautler and Sims, 2000; McDowell and Sharpley, 2001; McDowell et
al., 2001; Maguire and Sims, 2002a, 2002b; Sotomayor-Ramirez et al., 2004). Therefore,
there is an increasing trend to relate STP to PSR analyses for predictive purpose (Pautler
and Sims, 2000; Nair and Harris, 2004; Nair et al., 2004). Since Mehlich 1-P is Florida's
soil test P, M1-P to PSR relationship was studied for wetland soils. A plot of M1-P
against Ox-PSR gave a second order polynomial curve with strong significant correlation
(R2 = 0.85***) for all the wetland soils (Figure 4-9). Similar relationships have been
drawn by the split point technique to detect the change point in upland soils (Pautler and
Sims, 2000; Nair et al., 2004, Figure 4-10) for M1-P and Ox-DPS. In our case there is a
lack of data points showing a linear increase in STP (Figure 4-9), the better fitted second
degree curvilinear line has been drawn.
While Ox-P/(Ox-Fe + Ox-Al) (where all the elements are in molar ratios) is the
normal procedure used in PSR or DPS calculations for upland soils (van der Zee et al.,
1988; Leinweber et al., 1999; Maguire and Sims, 2002a, 2002b), we cannot conclude that
the same calculations would be ideal for wetland soils as well. However, PSR
0 0.1 0.2 0.3 0.4 0.5 0.6
Figure 4-9. M1-P as an indicator of wetland soil's P release trend; p < 0.0001
y = 290x 7.3
R2 = 0.42***
y= 1522x 259
R2 = 0.77*** 00
0 O* 0
0.10 'o o ^^
0.0 0.1 0.2 0.3
0.4 0.5 0.6
Figure 4-10. MI-P and Ox-PSR relationship for upland soils (modified from Nair et al.,
y=2216x2 -0.69x+ 11
R2 = 0.85
n = 357
calculations are operationally defined, and the use of the PSR (as calculated above) as a
potential indicator for wetland soils cannot be ruled out.
Pautler and Sims (2000) calculated Ox-DPS with the a factor as 0.68 and
concluded that M1-P versus Ox-DPS could give the change point as approximately 25%
for 50 mg kg-1 of M1-P which should be equal to 0.17 Ox-PSR for Delaware and Dutch
soils. They also mentioned that 50 mg kg-1 of M1-P should be rated "agronomically
excessive" and thus of a serious environmental concern.
350 y = 504x + 4.4
R2 = 0.68
250 n = 357 .
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7
Figure 4-11. Mehlich 3 P as an indicator of P release from wetland soils, p < 0.0001
Sallade and Sims (1997) found that M1-P was significantly correlated to P release
(r = 0.52*** at 35C) in a sediment/water flux study. This finding supports our
suggestion for the use of M -P as a wetland P release indicator. The researchers noted
that sediments with similar BAP and PSI values did have different PSRs and concluded
that the PSR could be used to target ditch sediments with higher potential to release P to
Table 4-4. Correlation of Ml analyses and other parameters (by location)
Only Center of Wetland (A)
Correlation [r] M1-P M1-Fe M1-AI M1-PSR
M3-P 0.697*** -0.146 0.264* 0.21*
Ox-P 0.828*** -0.047 0.404*** 0.189*
WSP 0.106 -0.226* -0.052 0.128
TP 0.865*** -0.08 0.363*** 0.226*
% LOI -0.007 -0.078 0.216* 0.082
% TC -0.007 -0.072 0.166* 0.079
pH 0.075 0.066 0.011 0.2*
MC -0.106 0.248* 0.151 0.032
BD -0.279* -0.148 -0.134 -0.152
M3-PSR 0.52*** -0.178* -0.084 0.406***
Ox-PSR 0.861*** -0.219* 0.184* 0.293***
Only Edge of Wetland (B)
Correlation [r] M1-P M1-Fe MI-AI M1-PSR
M3-P 0.656*** -0.153 0.224* 0.54***
Ox-P 0.516*** -0.005 0.439*** 0.37***
WSP 0.077 -0.241* -0.152 0.123
TP 0.685*** -0.04 0.363*** 0.573***
% LOI -0.026 -0.108 0.144 -0.024
% TC -0.029 -0.121 0.084 -0.017
pH 0.229* 0.017 0.105 0.187*
MC -0.066 0.206* 0.091 -0.062
BD -0.13 -0.091 -0.128 -0.09
M3-PSR 0.62*** -0.197* -0.107 0.604***
Ox-PSR 0.847*** -0.198* 0.095 0.791***
Only Uland (U)
Correlation [r] M1-P M1-Fe MI-AI M1-PSR
M3-P 0.901*** -0.024 0.384*** 0.308*
Ox-P 0.68*** 0.013 0.435*** 0.305*
WSP 0.273* -0.168 -0.216 0.278*
TP 0.567*** -0.058 0.311* 0.381***
% LOI -0.095 0.278* 0.195* -0.015
% TC 0.212* 0.172 0.237* 0.051
pH 0.282* -0.116 0.191* 0.208*
MC -0.132 0.316* 0.063 -0.049
BD -0.08 -0.238* -0.181 0.128
M3-PSR 0.519*** -0.164 -0.18 0.584***
Ox-PSR 0.422*** -0.262* -0.143 0.432***
*,p < 0.05; ***, p < 0.001
The various Ml parameter relationships to other soil parameters are presented in
Table 4-4. Significant relations are presented in detail at various places in this text.
Table 4-5. PSR comparison (by location)
Location Comparison R2 n (outlier) Equation
Ml versus M3 0.2591 201 y = 0.0536x + 0.0519
Center of M1 versus Ox 0.7425 173 (3) y = 0.5147x + 0.0337
wetland (A) M3 versus Ox 0.437 176 (1) y = 1.0586x + 0.0171
Ml versus M3 0.2376 204 y = 0.0341x + 0.0472
Edge of M1 versus Ox 0.6138 180 y = 0.2591x + 0.0446
wetland (B) M3 versus Ox 0.6174 180 y = 1.304x + 0.0012
Ml versus M3 0.7733 120 (1) y = 0.6367x + 0.0111
Ml versus Ox 0.6192 118(1) y = 0.8189x + 0.0202
Upland (U) M3 versus Ox 0.6942 120 y = 1.1388x + 0.0123
, p < 0.0001; Number in the parenthesis are outliers, not included in the equations
Table 4-5 shows a strong correlation between M1-PSR and Ox-PSR and M3-PSR
and Ox-PSR for center of wetland and edge of wetland and M1-PSR, M3-PSR and Ox-
PSR all are significantly correlated for upland soils which agree with the findings ofNair
et al. (2004). This indicates that all these methods of calculation could be appropriate for
use as potential P release indicators for wetland soils. Using the equations in Table 4-5,
the threshold M1-PSR for P release for the wetland soils (center) is calculated as 0.129
and the same for M3-PSR is 0.078. The background concentration of TP of all the center
of wetland soils of this study is approximately 550 mg kg-1 which was calculated from
the 75th percentile of the distribution of all the unimpacted wetland soils of this study
according to United States Environmental Protection Agency (USEPA) guideline
(USEPA, 2000). Based on this reference background concentration of the wetlands
surveyed threshold values for P release to the surrounding water bodies will be M1-P =
23 mg kg-1, M3-P = 41 mg kg-1, Ox-PSR =0.073, M1-PSR = 0.076, and M3-PSR = 0.057
(Table 4-3). Note that these values were estimated at this time and more information is
needed for setting threshold values for these parameters.
An attempt was made to understand the reasons for slope differences for the M1-P
and Ox-PSR relationship in two different arbitrarily chosen Ox-PSR ranges, less than
0.10 and greater than 0.10 Ox-PSR. The slope difference between two Ox-PSR range is
attributed due to the lower value of M1-P in combination with higher values of Ox-Fe
and Ox-Al for the soils having Ox-PSR less than 0.10 than that of the soils having Ox-
PSR greater than 0.10 (Table 4-6).
Table 4-6. Characteristics of elemental analyses of soils under different Ox-PSR ranges
n = 487 (Ox- WSP M1-P M3-P Ox-P TP Ox-Fe Ox-Al
PSR < 0.1) mg kg-
Mean 4 14 21 206 424 5067 1721
Standard Error 0.6 0.8 0.9 8 14.0 279 63
Median 0.6 9.4 19 156 376 2734 1368
Deviation 12 17 21 176 310 6170 1400
Variance 153 277 420 30807 96046 38071273 1959860
Skewness 5.2 2.8 1.7 1.7 1.2 2.8 3.3
0to Oto Oto 11.4 to 75 to 153.1 to
Range 98.6 128.9 138.5 1174.9 Oto 1769 49806 13519
n = 92 (Ox-PSR WSP M1-P M3-P Ox-P TP Ox-Fe Ox-Al
> 0.1) mg kg-
Mean 14.4 88 89 476 920 1317 1636
Standard Error 2.8 18.3 8.7 61 115 137 220
Median 7.7 28 61 293 595 887 1140
Deviation 27 175 83 586 1107 1317 2106
Variance 728 30670 6918 343280 1225004 1734563 4433590
Skewness 6.4 3.4 1.2 2.7 2.5 1.7 3.8
Oto Oto Oto 0 to
Range 236 923 330 0 to 3225 0 to 6119 0 to 6712 15440
Stepwise regression equations (Equations 6, 7, and 8) suggest that WSP is related to the
organic matter, bulk density and to some extent to the extractable metals in two Ox-PSR
Mean values for WSP, M1-P and Ox-P are all lower when Ox-PSR < 0.10
compared to the P concentrations at > 0.10 Ox-PSR (Table 4-6). Here also 0.10 Ox-PSR
has been chosen as the threshold value. It is noted that the concentrations of Ox-Fe and
Ox-Al are lower at the higher Ox-PSR range. The high P concentrations coupled with the
low retention capacity of the soils results in a sudden P release beyond this threshold
Our initial findings comparing different soil tests with water soluble P suggests that
P is sorbed loosely on the organic matter and that is the primary reason for high amounts
of WSP in wetland soils. There is a lack of data on wetland soils to establish a numeric
value but our wetland soils showed a wide range of WSP with a mean concentration of
8.4 mg kg-1 (Table 3-3). Stepwise regressions suggest that organic matter is the cause of
high P concentrations in water which is similar to the findings of other researchers
(Mandal, 1963; Kuykendall et al., 1999; McDowell and Sharpley, 2001). The poor
correlation of different STP and water extraction (Table 3-5) is attributed to the wetland
soils having high organic matter. WSP determinations on wetland soils were performed
on air-dried soils. It appears that this procedure may not be the most appropriate method
as a wetland P release indicator.
The background TP concentration of all the wetlands of this study is approximately
550 mg kg-1 which was calculated from the 75th percentile of the distribution of all the
unimpacted wetland soils of this study according to United States Environmental
Protection Agency (USEPA) guideline (USEPA, 2000). Based on this reference
background concentration of the wetlands surveyed threshold values for P release to the
surrounding water bodies will be M1-P = 23 mg kg-1, M3-P = 41 mg kg-1, Ox-PSR
=0.073, M1-PSR = 0.076, and M3-PSR = 0.057.
SUMMARY AND CONCLUSIONS
The selection of P release indicators for wetland soils was made from a wide range
of soils representing four types of wetlands: non-riverine marsh, non-riverine swamp,
riverine marsh and riverine swamp from three ecoregions (IX, XII and XIV) surveyed.
Samples were collected from two zones of the wetlands: center of the wetland and edge
of the wetland and one zone was selected from the upland adjacent to each wetland.
Phosphorus concentration of the selected wetlands ranged from 0 to 236 mg kg-1 WSP, 0
to 922 mg kg- M1-P, 0 to 330.2 mg kg- M3-P and 14 to 3222 mg kg- Ox-P with mean
values of 8.4 mg kg- WSP, 38.5 mg kg- M1-P, 43.5 mg kg- M3-P and 286 mg kg- Ox-
Relationships between STP concentrations are dependent on the location of the
sampling, i.e. center of wetland, edge of wetland and adjacent upland. Apparently,
different extraction procedures remove different forms of P from the soil. While the
relationship between Ml and M3 was highly significant for upland soils (R2 = 0.81***) it
less significant for edge of wetlands (R2 = 0.43***) and for center of wetlands (R2
WSP is exponentially related to the OM in the soil. Stepwise multiple regression
analyses for prediction of WSP indicated OM as the most important variable with little
contribution from Ox-Fe or Ox-Al. The only other significant variable in WSP prediction
was M3-PSR which was similar to the findings of several researchers working on upland
soils. Neither M1-PSR nor Ox-PSR appeared a variable in WSP prediction. This finding
can once again be attributed to the forms of P extracted by M3 which appears to be
different than P extracted by Ml and oxalate reagents. No definite mechanism for
extractions by Ml, M3 and oxalate can be made at this time from the information
generated from this study.
Relationships of WSP and PSR indicate a dependence on TC irrespective of the
method used in PSR calculations. For soils with > 12% TC, the P release is higher at a
lower PSR compared to soils with < 12% TC. Again, it may be premature to set threshold
PSR values for wetland soils based on the WSP information available at this time. The
pattern of change of M1-P with Ox-PSR is similar for wetland and upland soils. Either
curvilinear trends or split line model appear appropriate, confirming the use ofM1-P or
Ox-PSR as potential P release indicators for wetland soils irrespective of the type or
location of the wetland.
The relationship of M3-P with Ox-PSR is linear and the soil test could also be a
potential P release indicator. Total P is exponentially related to the PSR and could also be
used as an indicator to identify environmentally problematic wetland soils. While linear
relationships have been predicted for calculations of Ox-PSR, M1-PSR and M3-PSR for
upland soils, the best relationship for the center of a wetland is between M1-P and Ox-
PSR (R2 = 0.79).
Ml, M3, TP and PSR could all be potential P release indicators for wetland soils.
For Florida, the use of Ml, TP or PSR calculated from Ml parameters may be most
practical. WSP conducted on dry soils is not an indicator of P release from a wetland soil.
Additional research is necessary to determine if WSP extraction conducted under field
moisture conditions along with flux studies could be an indicator of P release.
The background TP concentration of all the wetlands of this study is approximately
550 mg kg-1 which was calculated from the 75th percentile of the distribution of all the
unimpacted wetland soils of this study according to United States Environmental
Protection Agency (USEPA) guidelines. Based on this reference background TP
concentration of the wetlands surveyed, threshold values for P release to the surrounding
water bodies will be M1-P = 23 mg kg-1, M3-P = 41 mg kg-1, Ox-PSR =0.073, M1-PSR =
0.076, and M3-PSR = 0.057.
The PSR may be a preferred indicator for P release from a wetland if based on a
reference wetland condition. Soil test P does not take into account the retentive capacity
of a soil due to additional Fe and Al particles that would be relocated in the wetland from
adjacent agricultural areas. Higher STP values compared to a reference wetland need not
necessarily indicate a poorer wetland condition. It is emphasized, however, that all
threshold values established in the current study, may require modifications when
additional research information on wetland water quality criteria are generated.
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I am Atanu Mukherjee. I was born in Calcutta (now Kolkata) in Eastern India.
From my childhood I had affection for environmental studies and also various processes
which control nature. I received my bachelor's degree in B.Sc. from the University of
Calcutta in 1998 with chemistry honors. I received an M.Sc from the University of
Calcutta in agricultural chemistry and soil science in 2000. I came to the United States in
2003 to pursue higher studies related to the environment.