WATER POLLUTION CONTROL RESEARCH SERIES 17O7OEKN12/69
Feasibility of Hydrolysis of Sludge
using Low Pressure Sfeam with SO2
as a Hydrolytic Adjunct and Utilization
of the Resulting Hydrolysate
US. DEPARTMENT OF THE INTERIOR FEDERAL WATER POLLUTION CONTROL ADMlNlSTRATl
-------
WATER POLLUTION CONTROL RESEARCH SERIES
The Water Pollution Control Research Reports describe
the results and progress in the control and abatement
of pollution of our Nation's waters. They provide a
central source of information on the research, develop-
ment , and demonstration activities of the Federal Water
Pollution Control Administration, Department of the
Interior, through in-house research and grants and
contracts with Federal, State, and local agencies,
research institutions, and industrial organizations.
Water Pollution Control Research Reports will be
distributed to requesters as supplies permit. Requests
should be sent to the Planning and. Resources Office,
Office of Research and Development', Federal Water
Pollution Control Administration, Department of the
Interior, Washington, D.C. 20242.
-------
FEASIBILITY OF HYDROLYSIS OF SLUDGE USING LOW PRESSURE
STEAM WITH S02 AS A HYDROLYTIC ADJUNCT AND UTILIZATION
OF THE RESULTING HYDROLYSATE
by
Foster D. Snell, Inc.
Hanover Road
Florham Park, New Jersey 07932
for the
FEDERAL WATER POLLUTION CONTROL ADMINISTRATION
DEPARTMENT OF THE INTERIOR
Contract Number 14-12-188
December 1969
-------
FWPCA Review Notice
This report has been reviewed by the Federal
Water Pollution Control Administration and
approved for publication. Approval does not
signify that the contents necessarily reflect
the views and policies of the Federal Water
Pollution Control Administration, nor does
mention of trade names or commercial products
constitute endorsement or recommendation for
use.
-------
TABLE OF CONTENTS
Page
Number
FOREWORD
ABSTRACT
INTRODUCTION 1
GENERAL BACKGROUND 1
THE HYDROLYTIC PROCESS STUDY 7
ESTABLISHING BASIC PARAMETERS OF HYDROLYSIS 7
FILTRATION STUDIES 10
THE HYDROLYTIC EFFECT 13
ANALYSIS AND PROCEDURES FOR PREPARATION OF
ORGANIC MOLASSES 14
DISCUSSION 17
THE NUTRITIONAL STUDY
EXPERIMENTAL PROCEDURE 20
OUTLINE OF THE TEST METHOD 23
RESULTS AND DISCUSSION 26
-------
ABSTRACT
Sulfurous acid, when used as a hydrolytic adjunct in
the treatment of activated sludge, will increase soluble
solids, improve filtration efficiencies and reduce moisture
content of the resultant filter cake. Use of a 0.5% sul-
furous acid, and arbitrarily chosen hydrolytic treatment
parameters of sludge at 140 degrees C for one hour, in-
creased the soluble solids content of the activated sludge
by 20%. Concentration of the solubilized extract produced
a molasses-type syrup; 85% of the solids content of the
syrup was organic. By TKN estimates, over 20% of these
solids were proteinaceous. An amino acid analysis indi-
cated it could be of value as an animal feed, and this was
confirmed in rat feeding studies (at a 10% level in the
diets) showing an EFU (Efficiency of Food Utilization) of
42 for the organic molasses as compared to 39 for cane
molasses.
-------
INTRODUCTION
Many disposal experts feel that the limiting operation
in a modern sewage disposal plant is the disposition of
the solids which accumulate as a result of the various
separation procedures. This study was designed to illus-
trate the usefulness of an extract of the residues for
animal feeds. The report is presented in two parts, "The
Hydrolytic Process Study" and "The Nutritional Study,"
following a review of the "General Background" of the study,
GENERAL BACKGROUND
The general concept of waste water disposal, as it
has developed over the years, can best be pictured as a
repeating process of solid-liquid separations and oxida-
tions, with an aim toward producing pure water, inorganic
salts and gases as the ultimate end products. This is
illustrated in Figure 1. Most currently used disposal
processes involve unit operations selected from the var-
ious types used elsewhere in the chemical process industry.
Over the years, waste disposal experts found that many
of the improvements and sophistications of their processes
actually increased costs particularly in regard to the
disposal of the solid residues. The rising costs of land,
in and around urban centers, has made land disposal costs
prohibitive. At the same time, public clamor for increas-
ingly purer effluent water has spurred the development of
improved treatments which produce even greater quantities
of solid waste.
Current disposal of sewage plant solids has now become
part of an operation called "ultimate disposal." Ultimate
disposal is concerned with the problem (along with others)
of disposal of refractory (resistant to oxidation) organic
residues to typical biological oxidations.
- 1 -
-------
As Gas
Whole Waste In
Reduction
of
Solids
I
Reduction
of
Solubles
Sludge
Production
of
Solids
Sludge
Reduction
of
Solids
This flow diagram represents
the cycles and oscillating
system of unit operations
involved in the basic con-
cepts of disposal.
Reduction
of
Solubles
As Gas
Etc. until pure water
Figure 1
CONCEPTS OF ADVANCED
WASTE TREATMENT
- 2 -
-------
Sale of Effluents
Traditionally, disposal experts have tried to recover
disposal costs by selling the end product of waste disposal.
Products such as Milorganite , a fertilizer composed of dried
activated sludge, have resulted from their efforts. However,
the use of sludge as a fertilizer has decreased because the
fertilizer, which is the result of biological oxidations, is
non-competitive with ammonia as a nitrogen source. Some dis-
posal plants find it impossible to recover freight costs, let
alone process costs, if they attempt to sell sludge as ferti-
lizer. The high costs usually are attributed to the expense
of dehydrating the low solids sludge. To overcome this cost
of dewatering and dehydration, various conditioning processes
have been proposed to increase the rate of filtration and to
decrease the moisture content of the cake after it is formed.
One such conditioning process is known as the Porteus
Process^1'. In this process, direct injection of steam into a
thickened suspension of activated sludge supposedly alters
the colloidal nature of the sludge and allows easier filtra-
tion and increased efficiency of water removal.
Oxidative and Hydrolytic Processes
Another procedure suggested for processing sludge is
the Zimpro Process '^' , involving high pressure, wet air oxida-
tion which converts the organics to C02, and recovery of the
energy evolved during processing. This process is efficient,
but requires expensive equipment to maintain the necessary
high pressures.
Teletzke et al. ' describe a modified wet air oxidation
process utilizing lower pressures. These digestions, as
described, are essentially hydrolytic reactions involving a
minimal of oxidation, and the resultant solutions are re-
cycled to the biological system.
The Use of Sludge for Animal Feeds
The activation of sludge is, in reality, an aerobic
fermentation process in which the organic matter in sewage
is metabolized and assimilated as the nutrient media for
the growth of microorganisms. In a sense, this process is
- 3 -
-------
equivalent to the growth of single-celled proteins often
suggested as feed sources. Dried activated sludge had
been reported to be equal in nutritional quality, as meas-
ured by nitrogen retention in lambs, to soybean meal and
urea-containing rations of similar crude protein content^)
However, the total nitrogen of a sludge-containing ration
had an apparent lower digestibility in rats (52%) and a
moderately low biological value (51%). Research into the
nature of activated sludges has shown that the cell debris
consists largely of polysaccharide polymers, protein, fat
and inorganic complexes^>) .
Acid Hydrolysis as a Means of Predigestion
Hydrolysis of organic materials has historically been
used as a process to make available otherwise undigestible
food ingredients. After hydrolysis, the solubilized mate-
rial is often concentrated to a thick syrup resembling the
concentrated residues from cane sugar manufacture, called
molasses. In the case of wood, citrus, pineapple, sorghum
and many other syrups, the term "molasses" is used because
it best describes the heterogeneous nature of the syrup,
regardless of its original source. Thus, terms are used
such as wood molasses or soy hydrolysate molasses, etc.
Manufacture of organic molasses is a means of ultimate
disposal. Currently, molasses types sell for from two to
five cents per pound. At $40 to $100 per ton, this selling
price leaves a profit opportunity not previously exploited.
Sulfurous Acid a Unique Hydrolytic Agent
The use of dilute sulfurous acid as a hydrolytic and
extraction agent is well known. Sulfurous acid functions
especially well in solubilizing naturally-occurring sub-
stances because it provides both the acidity required and
inhibits the formation of brown addition products usually
referred to as humins. A whole technology has been built
on the use of sulfite for delignification of pulp for the
paper industry^). McKinney''' reported on the efficacy of
sulfurous acid for the preparation of white soya protein
extracts in 1949.
- 4 -
-------
Baker et al.(8) undertook a careful study of the sulfurous
acid hydrolysis of casein, taking special note of its effect
on the tryptophan content of the humin-free extract. Results
showed that SC>2 hydrolysis protects the tryptophan present in
hydrolysates.
In general, the advantage of sulfurous acid as a hydro-
lytic agent can only be fully realized in treatments of het-
erogeneous natural substances containing both polysaccharides
and proteins. The presence of S02 during hydrolysis of products
containing both of the latter products prevents or minimizes
the formation of the black (or brown) humins.
Wet-Rendering Used in Packing Houses
Historically, in the packing house industry, a process
often used to digest inedible animal residues is a wet-
rendering process, whereby hair, feathers, meat scraps,
bone, etc. are cooked at relatively low pressures (60 Ibs.
steam) for digestion and fat recovery and the preparation
of specialty protein foods(9). in many cases, sulfite is
added to the digestion tank in the form of SC>2.
The meat-packing industry does not use S02~rendering
as a means of disposal but as a highly efficient procedure
for recovery of industrial fat and protein and for protein
hydrolysis as exemplified in the manufacture of culture
media universally used in commercial fermentations, such as
Wilson's B-407 Peptone.
The primary intent of this research program is to apply
the techniques of hydrolysis and solubilization of meat scrap
to the solubilization of activated sludge.
Preliminary Work
On May 11, 1967, Mulbarger^10^ observed a preliminary
demonstration experiment on the use of this concept for
sludge hydrolysis. Mulbarger!s summary of the report on
this single experiment, utilizing only one type of sludge,
is as follows:
"A preliminary trial at Wilson and Company using
activated sludge from the Southwest Plant of Metro-
politan Chicago showed that sulfurous acid hydrolysis
- 5 -
-------
is comparable to anaerobic digestion with regard to
solids reduction. Dried activated sludge was re-
suspended in a 1.25 percent SC>2 solution to give a
10 percent solids slurry. The slurry was pressure
cooked at 60 psi for four hours.
Hydrolyzed sludge solids were qualitatively
easier to filter and offer the added benefit of
being completely sterile. The decanted fluid con-
tained approximately 45 percent of the initial COD.
Lime treatment of the decant for pH adjustment
removed approximately 43 percent of the soluble
COD, 33 percent of the soluble TKN, and essentially
all of the phosphorus. The final decant could then
be recycled to the secondary biological system."
In a program involving the use of dilute sulfurous
acid as a hydrolytic adjunct in treatment of activated
sludge, we not only produced a syrup high in both sugar
and nitrogenous substances but found that the resultant
hydrolysate could be filtered more readily than a compar-
able hydrolysate made by heat treatment only.
The purpose of the present report is to describe the
experiments performed during this study and is presented in
two sections:
1. Studies on the hydrolysis of activated sludge
with heat and sulfurous acid, and
2. A nutritional study on an extract obtained from
the sulfurous acid hydrolysis of activated
sludge.
- 6 -
-------
THE HYDROLYTIC PROCESS STUDY
The objectives of this part of the program were:
to establish the basic parameters for hydrolysis
and solids separation
to characterize the organic molasses
to prepare sufficient organic molasses for the
feeding study.
By definition, organic molasses is the concentrated extract
of activated sludge produced by hydrolysis with sulfurous
acid.
Establishing Basic Parameters of Hydrolysis
A preliminary experiment was performed to establish
hydrolytic parameters. For this purpose, dried activated
sludge from the Chicago Sanitary District was weighed into
glass jars. The samples of sludge were introduced into
pint-size, Ball-brand Mason jars sealed with rubber gaskets
and wire-clamped glass lids and autoclaved in a horizontal
pressure device capable of maintaining up to 200 psi.
Figure 2 is a photograph of the autoclave and jars used.
The contents of the sample jars were adjusted to contain
200 ml of a mixture of sludge, water and various sulfur
dioxide concentrations (added as a solution of 3.9% 802)
Each jar contained 6% dried sludge. This latter concentra-
tion was chosen because previous multiple tests indicated
that the activated sludge from the Bergen County Sewerage
Authority Plant at Little Ferry, New Jersey (the model plant
for this study) averaged 4 to 5% solids over a period of
several months.
The extent of the solubiiization (or hydrolysis) was
measured by filtering the reacted slurry and evaporating
50 ml of the clear solution to dryness and weighing the
residue. The results of this experiment are shown in
Table 1.
- 7 -
-------
IQ
°
o
n
o
I
i/i
in
n
Q_
o_
o°
TJ
o
n
ro
-------
Table 1
Grams of Solids in 50 ml of Hydrolysis Liquor
Prepared under Various Conditions
S02 Concentration
No S02 Added
2.5% S02 Added
1.2% S02 Added
.63% SO 2 Added
.33% S02 Added
Note; Total
Results: Almost
Solids in 50 ml
1 hour @
100 psi
1.47
1.38
1.21
1.18
available solids in
50% of the solids
6 hours @
15 psi
0.51
1.41
0.96
0.68
0.64
1 hour @
15 psi
0.21
0.71
0.48
0.48
0.44
50 ml would be 3 grams .
from dried
activated
sludge can be solubilized while simple cooking
solubilizes only a small percentage.
Operating Conditions
All sludge samples (unless otherwise noted) were treated
for one hour under 30 Ib. steam pressure (140 degrees C).
Where sulfur dioxide was used, final concentration of SO2 was
0.5% based on total mixture.
All tests were conducted on fresh activated sludge
obtained from the aforementioned Bergen County Plant. It is
a thickened suspension of sludge of the type normally shipped
from this plant in barges for disposal. Bergen County thickens
its sludge by natural settling; no chemicals are used.
The average solids content of the thickened sludge was
determined by evaporation to dryness. At the same time, the
soluble and insoluble solids were determined by filtration.
The liquid portion was colloidal in nature and resembled
blood serum.
- 9 -
-------
Analysis of the sludge in repeated samples resulted
in the following average specification:
pH 6.8
moisture 91%
insoluble solids 4.18%
soluble solids 4.82%.
Filtration Studies
Separating the solids from the thickened sludge was
both time-consuming and tedious, requiring as long as
24 hours of filtration to obtain a dry cake. Hence, it
was decided to treat the thickened sludge directly and
measure the effect of hydrolysis by comparing the soluble
solids content of the filtrate before and after treatment.
The values so obtained, when compared with the total solids
content of the original sludge, give a true measure of the
amount of solubilization of the colloids.
Test samples consisting of 500 gm of thickened sludge,
having an average soluble solids content of 4.18% by weight,
were sealed in pint-sized Mason jars and placed in the auto-
clave previously described. The samples were divided into
three groups of 25 samples each, and subjected to treatments
A, B and C below.
Treatment
Designation Conditions
A. untreated raw activated sludge
B. raw activated sludge autoclaved
for one hour under 30 Ib. steam
pressure in accordance with arbi-
trary parameters, and
C. raw activated sludge containing
0.5% by weight of sulfur dioxide
added as 4.6% sulfurous acid sub-
jected to the same heat treatment
as B.
- 10 -
-------
After cooling to room temperature, 100 ml of each sample
was filtered through Whatman No. 1 analytical filter paper
using a battery of six Buchner funnels with a common vacuum
manifold. A photograph of the filter used in this study is
shown in Figure 3. The average filtration rate was deter-
mined by measuring the volume of liquid collected in 40
seconds and is summarized in Table 2.
Table 2
Filtration Rate of Treated and Untreated Sludge
Treatment Volume of Filtrate Collected (ml)
A 15
B 40
C 90
The filtration rate of activated sludge is increased
almost three-fold by heat treatment alone; when sulfur di-
oxide is added to the activated sludge before heat treat-
ment, the filtration rate is increased six-fold.
A second series of tests were performed to measure the
effects of heat and SC>2 treatment on the absolute rate of
dewatering. The same procedure was used as in the previous
experiment, except that the rate of filtration was deter-
mined by noting the length of time required to reach a dry
cake. In this series of experiments, 200 ml samples of
sludge, treated and untreated, were filtered and both the
volume of the filtrate collected and the time required to
achieve a dry cake were measured. Results are summarized
in Table 3.
- 11 -
-------
10
c
5
OJ
o
3
(D
1/1
n
Q-
c
Q_
UQ
(D
o
-------
Table 3
Absolute Rate of Dewatering
(200 ml sample)
Time to
Treatment Filtrate Collected (ml) Achieve a Dry Cake (min)
A 147 420
B 150 4
C 175 2
The foregoing illustrates that filtration time is
reduced drastically by heat treatment alone or heat plus
sulfur dioxide. Also, the amount of moisture retained in
the filter cake is reduced.
The Hydrolytic Effect
It is difficult to measure accurately the real solubil-
izing effect of treatment with S02 and heat because of the
difficulty in adequately dewatering the untreated sludge.
This difficulty was overcome by carrying the filtration to
completion and measuring the solids content of the filtrate
Table 4
Effects of Treatment on Soluble Solids Content of Filtrate
Sample Treatment Soluble Solids Content (%)
A No Treatment 4.82
B Heat Alone 7.34
C Heat + .5% S02, 8.84
Results in this case indicate that heat treatment alone
increases the soluble solids content by nearly 90% and an
additional 20% increase is obtained by addition of SC>2.
- 13 -
-------
Analysis and Procedures for Preparation of Organic Molasses
A 10-gallon sample of thickened activated sludge was
collected from the outfall of Bergen County, New Jersey dis-
posal plant. Figure 4 is a photograph of the apparatus used.
The sludge had approximately 9% solids, half of which was in-
soluble. This creamy slurry was charged into a closed jacketed,
20-gallon stainless steel vessel equipped with a mechanical
agitator. Sulfur dioxide gas was added until the final con-
centration was equivalent to 0.5% l^SO-j. This mixture had an
initial pH of 1.6. Steam introduced through an internal coil
and the jacket brought the pressure and temperature up to an
equivalent of 30 Ib. and 135 degrees C respectively. The
reaction mixture was held at this temperature and pressure
for three hours, then cooled to 80 degrees C and the mixture
filtered. Filtration was very rapid, producing a cake with
above 40% solids and a light yellow liquor smelling slightly
of SC>2 with approximately 8% dissolved solids, and a pH of
3.00.
This liquor was concentrated in a Rodney-Hunt evaporator
to a syrup with 60% solids. Chemical analysis of the solids
gave approximately 18% ash and 82% organics. Repeated pro-
duction runs produced molasses of slightly varying analysis
but usually within the range of the example shown.
The ash was high in iron (2.7%) and calcium (1.8%). The
sodium, potassium and magnesium content was typical of that
found in natural products. It was low in phosphorus (0.005%)
and high in sulfur, as would be expected because of sulfite
residues.
The organics analyzed close to 3.5% nitrogen, which, when
calculated as proteinaceous (6.25X) material, would be equiva-
lent to almost 20% protein. However, when the protein content
was reconciled against the total sugar analysis (40%) , it
became obvious that a portion of the nitrogen was in the form
of amino-sugars or other non-protein nitrogen.
In subsequent runs, it appeared that when the total solids
of the activated sludge was high in soluble solids; the sugar
content of the molasses was high. On the other hand, when the
soluble solids content was low, the sugar content was also low.
- 14 -
-------
10
c
*1
n
Xk.
a
n
c
10
n
-------
An amino acid analysis on the organic molasses yielded:
Amino Acid Percentage
1. Arginine 0.79
2. Cystine 0.17
3. Glycine 1.88
4. Histidine 0.37
5. Isoleucine 1.08
6. Leucine 1.01
7. Lysine 0.87
8. Methionine 0.36
9. Phenylalanine 0.74
10. Threonine 1.13
11. Valine 1.37
- 16 -
-------
DISCUSSION
Based upon our experience in the laboratory, and on
batch-pilot runs, and relating this experience to produc-
tion of the meat by-product, Peptone, and past experience
in the sugar field, we have prepared preliminary cost fac-
tors for the production of organic molasses. These factors
do not include the process improvements in dewatering --
certainly a notable factor.
Essentially three operations are required:
(1) Digestion of sludge
(2) Filtration or separation of solids
(3) Concentration of extract to molasses.
The principal expense involved will be in the cooking
and concentration steps.
The cost of filtration will be a minor factor when com-
pared to cooking and evaporation.
At this time, we feel the cost of digesting 25,000 Ibs.
of solids in stainless steel tanks, using direct steam (and
SOo) addition and yielding of syrup containing 20% solids,
will cost $160.69 (this does not include the cost of the SO?)
Evaporation to molasses (50% solids) will cost an additional
$244.67. Costs are itemized in Table 5.
- 17 -
-------
Table 5
Economics for Production of Organic Molasses
(25,000 Ibs. of Solids)
Digestion Cost
(1) 35,000 Ib. Steam $ 45.50
(2) Water .69
(3) Electricity 1.50
(4) Labor (16 man hours 80.00
8 $5.00)
(5) Depreciation and use 33.00
$160.69
Evaporation
(1) 90,000 Ib. Steam $117.00
(2) Water 2.07
(3) Electricity 5.10
(4) Labor (17 man hours) 85.00
(5) Equipment and depreciation 35.50
$244.67
Total Cost $405.36
This brings our production costs to around .015 cents
per pound of organic molasses.
A flow diagram of a possible plant for production of
organic molasses is shown in Figure 5.
- 18 -
-------
RAW SEWAGE
SCREENING
DEGRITTING
ALTERNATE SO?
DIGESTION
SYSTEM
f
JL
SEDIMENTATION
[Primary
Sludge
oj
iH
o
>1
o
Si
0)
&>
t)
^
iH
en
AERATION
SETTLING
(1)
&l
s
H
cn
M
-------
EXPERIMENTAL PROCEDURE
A preliminary experiment was performed to evaluate the
utility of organic molasses in animal feeds. From the design
of the experiment/ it was anticipated that one could deter-
mine maximum levels that can be used in animal feeds before
adverse reactions are produced in the animal.
The diets for each group supposedly contained sufficient
protein, fat, carbohydrate, vitamins and salts to permit ade-
quate growth. The diet of Group I contained 25% cornstarch.
In Group II, the cornstarch was replaced by an equivalent
quantity (on a dry weight basis) of the wet organic "molasses."
In Group III, the cornstarch was replaced by an equivalent
weight of non-digestible fiber. The study was designed to
determine whether the organic molasses could replace the energy
provided by the cornstarch. Assuming that the organic molasses
could not be utilized for energy, the growth would then be
equivalent to that of Group III. If depressive effects were
present, the growth would be less than that of Group III.
Measurements were made of food intake, weight gain and
major organ weights. The data are presented in Table 6.
- 20 -
-------
Table 6 Organic Molasses 17 Day Mean Data
Food Body Weight Organ Weights (gm) Organ Weight: Body Weight Ratio (%)
Group Intake Gain Testes Heart Liver Kidney Lung TestesHeartLiverKidneyLung
(gm) (gm)
I 238.2 80.3 1.12 0.40 5.56 1.09 0.84 0.87 0.31 4.33 0.85 0.65
i II 206.3 51.1 0.94 0.43 6.27 1.41 1.39 0.99 0.45 6.58 1.48 1.46
tsj
I
, III 211.2 55.8 ' 0.98 0.37 5.25 1.01 0.67 0.87 0.33 4.69 0.90 0.60
-------
At the start of the experiment, the animals were fed
ad libitum. Within the first three days, it was obvious
that the two control groups were eating considerably more
than the group receiving the organic molasses. This was
obviously due to the fact that the organic molasses made
the diet very wet. Rats do not like wet diets. Conse-
quently, in order to avoid interpreting data from groups
receiving different quantities of food, Groups I and III
were pair-fed with Group II.
The results indicate that Group II (organic molasses),
grew less than Group III (25% less carbohydrate) and Group I,
probably an indication of the effect of the mushy diet. Con-
sequently, it could not be determined to what extent the
organic molasses was being utilized for energy. In comparing
the pair-fed groups, the food efficiency of Group II appeared
to be less than those of the other groups. In addition, the
organ/body weight rations for kidneys, livers and lungs of
Group II were considerably greater than for the other two
groups, which we interpret to be a reflection of the high
mineral content of the molasses.
In summary, groups of rats fed organic molasses as part
of their diet ate less food, had decreased body weights and
greater organ weight/body weight ratios for several signi-
ficant organs.
At this time, it appears that 25% organic molasses was
excessive. The high moisture and high salt content were
responsible for poor food utilization.
A second set of experiments was set up using less organic
molasses (5% and 10%) and compared in the same trials with
cane molasses at the same levels.
- 22 -
-------
OUTLINE OF TEST METHOD
Seventy male, albino, Sprague-Dawley derived weanling
rats were distributed evenly among seven experimental groups
as follows:
Groups Diet
! Ad lib sucrose control
2- 5% organic molasses, sucrose
replaced on dry weight basis
3. 10% organic molasses, sucrose
replaced on dry weight basis
4« 5% cane molasses, sucrose
replaced on dry weight basis
5« 10% cane molasses, sucrose
replaced on dry weight basis
6. Sucrose control, pair fed with
5% replacement groups
7. Sucrose control, pair fed with
10% replacement groups.
All seven synthetic diets were basically the same
(Table 7) with substitutions being made only so that part
of the carbohydrate contribution of diets 2, 3, 4, and 5
were replaced by either 5 or 10 percent (dry weight basis)
organic molasses or cane molasses. Adjustment was also
made for water content of the diets. On a dry-weight basis,
the basal diet contained 20% casein, 5% fat, 58% carbohydrate,
1.15% vitamin mix, 4.0% minerals and 10% non-nutritive fiber.
Due to the high water content of the organic molasses, the
moisture contents of the diets were equalized. Since this
resulted in a rather fluid consistency to the diets, the
high level of fiber (10%) was used to improve the consis-
tency and increase the palatability of the diets.
- 23 -
-------
The recipes for the synthetic rat diets contained the
following materials: 90% vitamin-free test casein #160040,
General Biochemicals, Chagrin Falls, Ohio; Commercial Cotton-
seed Oil, Hunt-Wesson Foods, Fullerton, California; Cane
Sugar (Sucrose), American Sugar Company, New York, New York;
Vitamin Fortification mix #40060, General Biochemicals,
Chagrin Falls, Ohio*; U.S.P. XVII Salt Mix #170890 (reference
U.S. Pharmacopeia XVII, pg. 862), General Biochemicals,
Chagrin Falls, Ohio**; Non-Nutritive Fiber (cellulose type)
#160390, General Biochemicals, Chagrin Falls, Ohio; Cane
Molasses, R. J. Reynolds Foods, Inc., New York (diluted with
distilled water to 40% solids); organic molasses (40%) solids.
Rats were obtained from the KG Corporation, Parsippany,
New Jersey, breeders of research animals, divided into seven
of equal average weight (mean weight, 50 gm) and individually
housed in galvanized rat cages in a temperature controlled
environment. Water was available at all times. Diets were
prepared bi-weekly, to supply the same caloric intake for all.
Control and cane molasses groups were pair-fed with organic
molasses groups to eliminate variable food intake and the ad-
verse possible effect on palatability on efficiency of food
utilization. A control group was fed ad lib to indicate sig-
nificant differences, if any, on acceptability of sugar and
molasses diets. Body weight and daily food consumptions were
recorded for all during the 21-day feeding study. Animals
were observed daily for normal or abnormal activity, appetite,
health, and fecal character.
* The vitamin mix added at 1% of the diet provided:
(in mg/kg of diet) vitamin A, 39,6825; vitamin D2,
40492, alpha tocapherol acetate, 485.0090; ascorbic
acid, 1017.5200; 1-inositol, 110.2290, choline di-
hydrogen citrate, 3715.1230; menadione, 49.6030;
p-aminobenzoic acid, 110.2290; niacin, 99.2060;
ribovlavin, 22.0450, pyridoxine HCL, 22.0450;
thiamine HCL 22.0450; calcium pantothenate, 66.1376;
biotin, 0.44092; folic acid, 1.98412; vitamin Bi2/
29.7619.
** The mineral mix added at 3.5% of the diet (4.0% on a
dry weight basis) contained: (in percent composition)
Calcium Carbonate, 38.140; Cobalt Chloride, 0.0023;
Cupric Sulfate, 0.0477; Ferrous Sulfate, 2.700;
Magnesium Sulfate, 5.730; Manganese Sulfate, 0.401;
Potassium Iodide, 0.079; Potassium Phosphate Monobasic,
38.900; Sodium Chloride, 13.930, Zine Sulfate, 0.0548.
- 24 -
-------
After 21 days on test, all animals were sacrificed
using chloroform anesthesia and necropsies were performed.
Adrenals, heart, liver, kidneys and lung were weighed and
saved in 10% buffered formalin for possible future histo-
logical examination. In addition, any gross pathology was
noted and lesions were saved in formalin. Calculations
were made of 21-day body weight gain, food intake and effi-
ciency of food utilization (Table 8) as well as organ-body
weight ratios for the weighed organs (Table 9). Three rats
from Groups 1, 3 and 5 were frozen after necropsy for body
water determination if examinations of the tissues indicated
a possible edematous condition.
- 25 -
-------
RESULTS AND DISCUSSION
Gain in weight was greater for the 10% organic molasses
group than for the other groups tested. Although the ad lib
control group gained the next largest amount, it was on an
unrestricted food intake and thus had the lowest EFU of any
group tested. In this respect, the EFU of the 10% organic
molasses group was significantly greater than that of the ad
lib control group, the 5% organic molasses and the 10% cane
groups. The pair-fed control groups whose diets were exactly
the same as the ad lib group had EFU values significantly
higher than the ad lib group. This may be due to the fact
that scatter played a large part in recording intake of the
unrestricted group. The inorganic molasses diets were less
likely to scatter because of their more solid consistency.
Nevertheless, the data indicate that the organic molasses
supplied energy of at least equal to cane molasses or sucrose.
Although the 5% organic molasses group had significantly
lighter adrenals than the 5% cane molasses group and the 5%
pair fed control, the adrenals were heavier than the ad lib
control, and the actual difference was not believed meaningful.
In support of this belief, the 10% organic molasses group had
adrenal weight of the same or greater magnitude than all other
groups except the 5% cane molasses group. The 10% organic
molasses group had a significantly heavier heart weight than
its pair-fed control, but not significantly heavier than the
ad lib control.
The greater amino acid content of the organic molasses
diets (hydrolyzed bacterial protein) may account in part for
the greater growth and EFU. Both organic molasses groups
showed significantly heavier livers and kidneys than their
comparable groups due to the increased utilization of the
higher amino acid content of their diets. There were no sig-
nificant differences among the groups as to lung weight ratios,
indicating that edema was not.responsible for the increased
weight gains of organic-molasses-fed animals. All evidence
indicates that organic molasses at the 10% level is an ade-
quate energy substitute for cane molasses in the diet.
- 26 -
-------
Table 7 Diet Preparation
to
Groups
90% Casein
Cottonseed Oil
Sucrose
Vitamin Mix
Mineral Mix
Non-Nutritive Fiber
Molasses
(40% Solids) Organic Molasses
(40% Solids) Cane
H20
Control
1+6-1-7
190
44
504
10
35
87
0
0
130
5% Organic
Molasses
2
190
44
460
10
35
87
109
0
65
10% Organic
Molasses
3
190
44
416
10
35
87
218
0
0
5% Cane
4
190
44
460
10
35
87
0
109
65
10% Cane
5
190
44
416
10
35
87
0
218
0
Total
1000
1000
1000
1000
1000
-------
Table 8 Efficiency of food utilization for weanling rats fed for 21 days *
Group
K)
00
Body Weight
0 Day 21 Day
21 Day
Weight Gain
21 Day
Food Consumption
with 5% Pair Fed
Groups 49.2
7. Sucrose Control
with 10% Pair Fed
Groups 49.4
172.0+13.0
178.8+14.7
122.8
129.4
323.2
320.4
E.F.U. **
1.
2.
3.
4.
5.
6.
Sucrose
Control
Organic
Molasses 5%
Organic
Molasses 10%
Cane 5%
Cane 10%
Sucrose Control
49
49
47
49
53
\jj_
.5
.5
.3
.3
.3
181
171
191
175
166
.5+33.
.3+35.
.4+21.
.6+15.
.0+20.
4
4
5
8
4
Lirai
132
121
144
126
112
ms
.0
.8
.1
.3
.7
urai
401
342
343
323
317
ms
.5
.7
.8
.1
.6
32.7
35.2
42.0
39.1
35.4
38.0
40.4
*10 rats/group
** gram gain/100 gram eaten E.F.U. equals Efficiency of Food Utilization
*** standard deviations
-------
Table 9 Mean Organ Weight Ratios of Weanling Rats Fed for 21 Days
fO
VO
Group^
1. Sucrose Control
2. Organic
Molasses 5%
3. Organic
Molasses 10%
4. Cane 5%
5. Cane 10%
6. Sucrose Control
Pair Fed with
5% Organic
Molasses
7. Sucrose Control
Pair Fed with
10% Organic
Molasses
Adrenals
0.0291.0067
0.0271.0077
Heart
Liver
Kidneys
0.381.032
5.571.65
0.991.084
0.411.020
5.821.79
1.021.093
Lunc
0.0171
0.0221
0.0271
0.0341
0.0271
.0069**
.0050
.0047
.0109
.0022
0
0
0
0
0
.411.
.391.
.451.
.431.
.431.
075**
033
042
051
077
6
7
6
5
5
.451.
.061.
.501.
.661.
.511.
56**
96
70
30
43
1.071
1.231
1.241
11.11
1.091
.084**
.134
.117
.144
.093
0
0
0
0
0
"
.621.086
.651.062
.621.044
.671.109
.671.092
0.621.070
0.631.066
* 10 Rats per Group
** Standard Deviations
-------
BIBLIOGRAPHY
1. Porteus, Ian King, to Norstel and Templewood Hawksley
Ltd., U. S. Patent No. 3,155,611, 1964.
2. Zimmerman, F. J. and Diddams, D. G.: The Zimmerman
Process and its Application in the Pulp and Paper
Industry^ TAPPI, 43_, 710 (1960) .
3. Teletzke, G. H.: Wet Air Oxidation. Chem. Eng. Prog.,
6_0 1,33 (1964).
4. Hackler, Lonnie R., Neuman, A. L., and Johnson, B. Conner:
Feed from Sewage, III. Dried Activated Sewage Sludge
as a Nitrogen Source for Sheep. J. Animal Sci., 16,
125-9 (1957).
5. Hurwitz, E. : The Use of Activated Sludge as an Adjuvant
to Animal Feeds. Eng. Bull., Ext. Ser. No. 94, 395-
414 (1957).
6. Ant-Wuorinun, Olli: New Method of Wood Saccharification
with Sulfurous Acid. Svensk Papperstidn, 45, 149-57
(1942) .
7. McKinney, L. L. and Sollars, W. F.: Extraction of Soy-
bean Protein with Sulfurous Acid. Ind. Eng. Chem.,
4_1, 1058 (1949).
8. Parsons, T. R., and Baker, B. E.: Preparation and
Analysis of Sulfurous Acid Hydrolyzates of Casein.
J. Sci., Food and Agriculture, 1_> 261-5, 1956.
9. Johnsen, Vernon L., La Grange, Matern, Eugene V. and
Bennett, Raymond S., to Wilson & Co., Inc. U. S.
Patent No. 3,308,113, March 7, 1967.
10. Mulbarger, M. C. Unpublished Communication. Wilson &
Co., May 11, 1967.
- 30 -
------- |