Group Title: Citrus Station mimeo report - University of Florida Citrus Experiment Station ; CES 64-6
Title: Progress in the 2,3- butylene glycol fermentation of citrus wastes
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 Material Information
Title: Progress in the 2,3- butylene glycol fermentation of citrus wastes
Series Title: Citrus Station mimeo report
Physical Description: 5 leaves : ; 28 cm.
Language: English
Creator: Long, Sterling Krueger, 1927-
Patrick, Roger
Citrus Experiment Station (Lake Alfred, Fla.)
Florida Citrus Commission
Publisher: Florida Citrus Experiment Station :
Florida Citrus Commission
Place of Publication: Lake Alfred FL
Publication Date: 1963
Subject: Citrus fruit industry -- By-products -- Florida   ( lcsh )
Citrus fruit industry -- Waste disposal -- Florida   ( lcsh )
Genre: government publication (state, provincial, terriorial, dependent)   ( marcgt )
bibliography   ( marcgt )
non-fiction   ( marcgt )
Bibliography: Includes bibliographical references (leaf 5).
Statement of Responsibility: Sterling K. Long and Roger Patrick.
General Note: Caption title.
General Note: "October 8, 1963."
 Record Information
Bibliographic ID: UF00072419
Volume ID: VID00001
Source Institution: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: oclc - 75957555

Full Text

Citrus Station Mimeo Report CES 64-6
October 8, 1963

Progress in the 2,3-Butylene Glycol Fermentation of Citrus Wastes

Sterling K. Long and Roger Patrick
University of Florida
Citrus Experiment Station
Lake Alfred, Florida

The laboratory-scale fermentation production of 2,3-butylene glycol from
citrus wastes was discussed at an earlier Processors' Meeting and will be
mentioned only briefly in this presentation. Further study has carried the pro-
ject through a pilot plant-scale operation and a method for subsequent extraction
of the glycol on a large volume basis.

The earlier paper on butylene glycol indicated that press liquor and dilute
molasses were satisfactory for the production of this alcohol. Glycol was readily
obtained using either of two bacterial cultures, Aerobacter aerogenes 199 or
Bacillus polymyxa B510, although Aerobacter consumed more sugar and produced more
glycol than the latter species. Subsequent work has established the superiority
of Aerobacter in all respects.

Type of Fermentation Substrate

The early laboratory and pilot fermentations indicated that molasses adjusted
to 200 Brix was more desirable than press liquor due to the higher sugar content
and, consequently, the higher glycol concentration obtained. Molasses alone was
found to be inadequate to support the fermentation since it was deficient in
naturally-occurring nitrogen. This deficiency was readily satisfied by addition
of 0.h% urea.

Only slight differences in the fermentability of molasses obtained during the
various seasons was observed. It is presumed that the plentifulness of fruit and
quantity production of molasses resulted in a "mixed" molasses obtained from a
number of varieties with, very likely, considerable mixing in the storage tanks.
Under these conditions it is apparent that it was practically impossible to ob-
tain meaningful comparative results on the various types of molasses. However
during the past year, due to shortages of fruit and subsequent interruptions in
molasses production, it was possible to obtain reasonably "pure" molasses from
the seasonal varieties. The results of the evaluation of such molasses indicate
that not all are equally suitable for this fermentation when used in an unmixed
condition. Each type will be discussed individually using the following terms:

Early Molasses produced from all early season varieties.
Midseason Molasses produced from mid-season varieties.
Valencia Molasses produced from late season varieties. May be
regarded as relatively "pure" Valencia molasses.
Grapefruit Molasses produced from grapefruit only.

Florida Citrus Experiment Station
and Florida Citrus Commission,
Lake Alfred Florida.
400-10/8/63 SKL

A. Early This molasses required little supplementation to give
good yields of butylene glycol. Good fermentability was obtained by
addition of O.4% urea. Control of the pH of the fermentation at the
optimum of 6.2 was not difficult and was accomplished by periodic
addition of 1ON NaOH.

B. Midseason This was the most satisfactory type of molasses.
Butylene glycol production was good and supplementation with O.h% urea
satisfied the requirements for nitrogen. The pH of the fermentation
was easily controlled.

C. Valencia Fermentations using this type of molasses were more
difficult to control than with the previous types. Supplementation
with 0.h% urea and yeast extract was required. When only urea was used
poor butylene glycol yields were obtained. Control of pH was also con-
siderably more difficult, with rather large quantities of 1ON NaOH re-
quired to maintain the optimum. Excessive quantities of yeast extract
promoted foaming to an undesirable degree.

D. Grapefruit This molasses required both O.h% urea and even
greater quantities of yeast extract than Valencia for good production
of glycol. Control of pH was very difficult, especially when an ex-
cessive quantity of yeast extract was used.

E. Mixture of Valencia and Grapefruit This combination was more
satisfactory than Grapefruit molasses alone although O.h% urea and
yeast extract were required. Apparently the low nitrogen concentration
of Grapefruit molasses was partially compensated for by addition of the
Valencia molasses since less yeast extract was required than when Grape-
fruit alone was used.

The following supplements may be regarded as typical requirements for good
glycol production using the various types of molasses in a 20 gallon pilot plant

Type Molasses Nitrogen Supplements
Early O.h% (13 oz.) urea
Midseason 0.h% (13 oz.) urea
Valencia 0.h% (13 oz.) urea +
0.013% (10 g.) yeast extract
Grapefruit 0.h% (13 oz.) urea +
0.065% (50 oz.) yeast extract
Valencia-Grapefruit 0.4% (13 oz.) urea +
0.013% (10 g.) yeast extract

Florida Citrus Experiment Station
and Florida Citrus Commission,
Lake Alfred, Florida.
400-10/8/63 SKL


Pilot-Plant Fermentation

A 30 gallon "Cavitator" (Yeomans Brothers, Melrose Park, Ill.) was obtained
for use as a pilot fermentor. The Cavitator was equipped with a highly efficient
impeller, a foam disperser, cooling coil, and an air metering device. The fer-
mentor was constructed of Lucite with a stainless steel top and internal mechanism.
Since heat could not be employed for sterilization, the fermentation chamber was
filled with a hypochlorite solution before use, followed by thorough rinsing with
sterile water.

The fermentation substrate was prepared for use by diluting molasses with
water to 200 Brix, followed by pasteurization in a stainless steel feed tank
equipped with a heating and cooling coil. The Cavitator was filled with the
molasses solution and urea and yeast extract were added as needed. After ade-
quate mixing, the pH was adjusted to the optimum of 6.2 with 1CN NaOH and the
culture added. The fermentation was run at 800F. while under aeration at 0.2 cfm.
The pH was maintained at 6.2 by periodic adjustment with O1N NaOH. Although the
fermentor was equipped with an automatic foam disperser, it was found to be
inadequate to handle the large amounts of foam produced. Satisfactory control
was obtained by regular dosing with Dow-Corning Antifoam B.

The course of the fermentation was followed by sampling and analysis for
remaining sugar and 2,3-butylene glycol content.

Under optimum fermentation conditions, Aerobacter aerogenes 199 cultures
produced maximum butylene glycol concentrations of 3.0-.0 %O with a residual
sugar of approximately 2.5 % within a period of 48-72 hours.

Criteria for Healthy and Completed Fermentations

In both A. aerogenes and B. polymyxa fermentations, repetition of a given
sequence of events was found to be a reliable criterion for predicting the prob-
able course of the fermentation, as well as the completion of glycol production.

A. Decreasing pH. The active stage of a "healthy" fermentation was
characterized by a steady decrease in pH which necessitated frequent addition
of 1ON NaOH to maintain the optimum of 6.2. The period of greatest activity in
this respect was during the first 30 hours of fermentation. After this period,
healthy fermentations required no further adjustment of pH.

Increasing pH of the fermentation beer, in conjunction with other factors
indicated below, was uniformly reliable in indicating the cessation of glycol
production, due either to sub-optimal conditions or normal completion of the
fermentation. This pH phenomenon was, however, the earliest indication of the
condition of the fermentation.

B. Simultaneous decrease in glycol concentration and apparent increase
in sugar concentration. Other analytical procedures which were of value in de-
termining the state of the fermentation were the colorimetric methods employed

Florida Citrus Experiment Station
and Florida Citrus Commission,
Lake Alfred, Florida.
400-10/8/63 SKL

for determining total sugar (1) and 2,3-butylene glycol content (2). Naturally,
these procedures are useful in the usual manner in following the course of the
fermentation by observing decreasing sugar and increasing glycol levels, however,
these methods can also be used to determine fairly accurately the termination of
glycol production.

During the course of these fermentation studies it was observed that, at a
given stage in the fermentation cycle, glycol production leveled off and remained
at the same concentration for a period of several hours. This was followed by a
slow but steady decrease in glycol concentration. Simultaneously with cessation
of glycol production, sugar utilization ceased. Then as the glycol concentration
began to decrease, the sugar concentration of the fermentation beer began to show
an apparent increase. This trend was consistently observed with both types of

In summary, increasing pH, with simultaneously decreasing glycol content and
increasing sugar content, indicated the approach of the conclusion of glycol pro-
duction and signified the point at which the fermentation should be discontinued.

Steam-Stripping of Glycol from Fermentation Beer

Recovery of the butylene glycol from the fermentation beer by steam-stripping
was attempted using two types of beer. A comparison was made of glycol removal by
this method from single-strength or unconcentrated beer and from beer which had
been previously concentrated by evaporation in an Atkins Model B pilot plant
evaporator (3) under 29-30 inches vacuum at 700F.

The process of steam-stripping involves the passage of steam under high
pressure, yielding a temperature approaching or exceeding the boiling point of
2,3-butylene glycol, through a thin film of the fermentation beer with subse-
quent condensation of the effluent glycol vapor-laden steam.

Single-strength or concentrated fermentation beer was pumped, by means of a
proportioning pump, through a pre-heater at 800-900F. and introduced as a spray
into the top of a 2h foot jacketed vertical column. The beer then traveled the
length of the column as a thin film over the column packing. The packing con-
sisted of layers of 1.9 inch diameter steel rings, with alternate layers offset
to provide maximum baffling.

The column was brought to a steam pressure of 100-120 p.s.i.g. (3370-350F.)
prior to introduction of the beer and was maintained in this pressure range
throughout operation. The steam was introduced into the base of the column and
passed upward through the falling film of beer, vaporizing the glycol. This
steam-glycol mixture was removed through the top of the column and passed into
the column jacket where partial cooling and condensation occurred. The jacket,
heated in this manner, also served to prevent premature condensation within the
main column. The partially condensed vapors passed through a valve at the base
of the jacket, which was used to control the rate of passage of the vapors, and
then through a water-cooled condensing coil. Stripped beer or sludge collected
in the base of the column and was discharged periodically through a valve as

Florida Citrus Experiment Station
and Florida Citrus Commission,
Lake Alfred, Florida.
0OO-10/8/63 SKL


Samples of butylene glycol for analysis were obtained by rectification of
representative samples of condensate, accomplished by concentration in a labora-
tory-scale rotary evaporator, and followed by distillation and redistillation.
In actual practice such rectification would be done in a plant-scale fraction-
ating column.

Use of this column solved the previous difficulties experienced in glycol
recovery by solvent extraction or direct distillation. The column remained free
of coagula and viscous sludge during all periods of operation. The column also
remained free of burned sugar deposits. Clean-up problems were simplified by
backflushing with steam and water.

Although optimum feed and discharge rates have not been established, con-
siderable basic information has been obtained. A number of feed rates of beer
into the column have been tested. As might be expected, the more concentrated
the beer, the more rapid the feed rate permitted. With single strength beer,
satisfactory recovery was obtained at a feed rate of 1.0 gal./hr., while with
4-fold concentrated beer, 7.0 gal/hr. gave excellent results. Pre-heating of the
beer to 80-90F. was found to be necessary for maximum glycol removal. Main-
taining steam pressure within the column at 100-120 p.s.i.g. was satisfactory for
rapid vaporization of the glycol. The proper rate of passage of the condensate
through the column has not yet been definitely established.

Literature Cited

1. Morris, D. L. 1948. Analytical Methods for Bacterial Fermentations, 2nd
rev., pp. 33-35. Nat. Res. Council Canada, No. 2952, Saskatoon, Sask.

2. Desnuelle, P. and Naudet, M. 19L5. Analytical Methods for Bacterial
Fermentations, 2nd rev., pp. 37-38. Nat. Res. Council Canada, No. 2952,
Saskatoon, Sask.

3. Atkins, C. D., Wenzel, F. W., and Moore, E. L. 1951. An evaporator of
improved design for the concentration of citrus juices. Proc. Florida
State Hort. Soc. 64, 188-191.

Florida Citrus Experiment Station
and Florida Citrus Commission,
Lake Alfred, Florida.
o00-10/8/63 SKL

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