THERMALLY OXIDIZED SOYA BEAN OIL AND THERMALLY OXIDIZED SOYA
BEAN OIL INTERACTED WITH MONO- AND DI-GLYCERIDES OF FATTY ACIDS
First draft prepared by
Dr B. Priestly
Department of Pharmacology
University of Adelaide
Thermally oxidized soya bean oil (TOSO) is made by oxidation of
refined soyabean oil in air at 190-250 °C. It is used as a
tin-greasing emulsion in baking. It is also interacted with food
fatty acids under vacuum at 130 °C to form thermally oxidized soya
bean oil interacted with mono- and diglycerides of fatty acids
(TOSOM) which has been used as an emulsifier in oleomargarine for
many years. TOSOM also imparts anti-spattering properties to
oleomargarine when used for frying. TOSO and TOSOM were considered
at the fifteenth, seventeenth, twentieth, and twenty-fourth meetings
of the FAO/WHO Joint Expert Committee on Food Additives. (Annex 1,
references 26, 32, 41, 53) under the name "esters of glycerol and
thermally oxidized soy bean fatty acids". Specifications were
tentatively assigned (Annex 1, reference 28) at the fifteenth
meeting, but withdrawn at the seventeenth meeting when it appeared
that the compound was a mixture and not a true ester. A
toxicological monograph was prepared at the seventeenth meeting
(Annex 1, reference 33) from data describing both the esterified and
unesterified oxidized soyabean oil, but an ADI was not established,
pending the evaluation of short-term and long-term studies on a
material of well-defined composition.
Since the previous evaluation, new data on metabolic
disposition and short-term and long-term studies in the rat have
become available and have been summarized and discussed in the
following monograph. Relevant material from the previously published
monograph has been included.
2. BIOLOGICAL DATA
2.1 Biochemical aspects
2.1.1 Absorption, distribution, and excretion
Groups of five male and five female rats received in their diet
either 20% soybean oil or thermally oxidized material. Highly
modified material reduced the absorption of dietary fat in
proportion to the degree of modification introduced. Delay in
absorption of modified oil material from the gastrointestinal tract
was measured by residuals found in the gut. Similar delaying effects
have been demonstrated on the presence of chylomicrons and fat in
the intestinal lymph fluid. The passage of intestinal contents is
delayed if modified oil is administered. Compared with normal oil
there is an early increase followed by only slight decrease in bile
flow following oral administration of modified oil. Intraperitoneal
administration of modified oil increases the diuretic effect of
intraperitoneal normal soybean oil. Study of the liver function
after eight weeks feeding of modified oil showed retention of the
bromo-sulphophthalein used as indicator compared with normal oil.
The in vitro effect of modified oil on the kinetics of various
cellular enzyme systems showed generally no inhibition of oxidative
metabolism (Kieckebusch et al., 1962).
Wistar rats, Tuck TO mice and Dunkin-Hartley guinea-pigs were
dosed orally with 20 mg/kg bw 14C-labelled refined soyabean oil,
200 mg/kg bw 14C-labelled Homodan O (a mixture of 2 parts soyabean
oil and 1 part TOSO), or 200 mg/kg bw 14C-labelled Emulsifier MO
(previously called Homodan MO and of equivalent specifications to
the emulsifier TOSOM). Urine, faeces and expired CO2 were
collected for up to three days.
After dosing with the control soyabean oil, the radiolabel
disappeared rapidly from the GI tract, there was a transient
accumulation in the tissues (mainly liver, kidney, lung, fat and
muscle) and most of the dose was recovered in the excreta within
24 h. In rats and mice, recovery of 14CO2 accounted for most of
the dose (50-70%), while in guinea-pigs, only 10-13% was recovered
as 14CO2. Urinary excretion accounted for 2% in rats and mice
and 4-8% in guinea-pigs. Faecal recovery accounted for the remainder
(5-15% rats and mice; 70-80% guinea-pigs). There were no sex
differences in disposition, and 7 days repeated administration of
unlabelled material prior to dosing did not alter the excretory
pattern. Only the liver, fat muscle and GI tract retained >1% of
the dose after three days.
The patterns of excretion of the 14C-labelled Emulsifiers O
and MO (TOSO and TOSOM) were comparable to that of soyabean oil in
the guinea-pig, but both rats and mice expired less 14CO2
(11-26%) and excreted more urinary metabolites (6-12%, 7-9%,
respectively). Faecal recoveries of radioactivity were 3-4-fold
greater for each emulsifier when compared with refined soyabean oil.
Tissue levels 1-4 h after dosing with TOSOM were approximately 3x
those observed after comparable dosing with unesterified TOSO. The
authors concluded that TOSOM is absorbed slightly better than the
unesterified TOSO, and that thermal oxidation of the soya bean fatty
acids results in retarded absorption in comparison with refined
soyabean oil (Phillips et al., 1978 a,b). The effect of thermal
oxidation on absorption is consistent with that seen with other food
oils. (Perkins et al., 1970).
No data available.
2.1.3 Effects on enzymes and other biochemical
No data available.
2.2 Toxicological studies
2.2.1 Acute toxicity
No data available
2.2.2 Short-term studies
For 12 to 16 weeks groups of 20 male and 20 female rats
received in their diets either 20% untreated soybean oil or 5%
variously modified soybean oil plus 15% olive oil or 2.5% modified
oil plus 17.5% olive oil. Only highly modified oils caused
significant reduction in growth and food intake, and increased
mortality. No definite effect was noted at the 2.5% level. Motor
activity of animals receiving high doses of highly modified oil was
increased compared with controls. The weights of major organs were
similarly increased for the groups on the more highly modified oils.
Gross and histo-pathology showed some pathological changes in the
thyroid and kidney of the group receiving the most highly modified
material (Kieckebusch et al., 1962).
Three groups each of six rats were treated for 17 weeks with an
esterified product at 0, 0.084 and 0.84% of their basic diet. There
was no significant effect on weight gain nor macroscopic appearance.
Liver and stomach were examined histologically and found to be
normal (Dam, 1952).
Two groups of nine male rats received for 36 weeks feeds
containing oleomargarine made with an esterified product at 0.3 and
3% levels. No controls were used. No weight differences nor gross
pathological changes were noted. The histology of kidneys, liver and
small intestine was normal (Aaes-Jorgensen et al., 1954).
Groups of 20 Wistar rats of each sex were fed diets containing
1, 5 or 10% of a commercial tin-greasing emulsion, for 91 days. The
emulsion was specified only with regard to its content (1%) of TOSO.
Intake of the emulsion was determined to be 0.88-0.97, 4.6-4.88, or
9.38-10.16 g/kg bw/day in the three dosed groups, making the upper
level of TOSO intake in this study approximately 100 mg/kg bw/day.
There were no effects noted on growth, food intake, urinalysis,
haematology or blood biochemistry. There were no effects on organ
weight or histopathology. The Committee considered this study to be
of minimal usefulness. The dose levels were too low to produce any
toxicity, and only 10/20 rats from each group were subjected to
haematological analysis, blood biochemistry or autopsy (Kemper
Four groups of 4 female pigs received an esterified product for
98 days at dietary levels of 0, 0.4, 2 and 10%. No significant
effects were noted on growth rate, food consumption, blood
parameters, liver and kidney function, serum chemistry, organ
weights, nor gross and histopathology due to administration of the
test compound (Gyrd-Hansen & Rasmussen, 1968).
2.2.3 Long-term/carcinogenicity studies
A three-generation study was performed using an average level
of 15% esterified product in the diet. The parent generation (57
females, 15 males) was observed for at least 24 months. Growth, body
weight gain and appearance were similar to controls (seven males,
seven females) receiving 15% soybean oil. Five animals in the test
group developed tumours but none in the control group. There were 16
survivors in the test group and two in the control group.
The F1 generation (37 males, 37 females) was also observed
for 24 months. Thirty-seven males and females received soybean oil
as controls. Twenty-four test and 10 control animals survived two
years. Two test animals and three controls developed tumours. The
F2 generation (57 females, 27 males) was observed for over two
years. Seventy males and females received soybean oil as controls.
Four test and 19 control animals survived 24 months. No animals in
the test group but three in the control group developed tumours.
Neither the F1 nor the F2 generation showed significant
differences from controls as regards growth, body weight gain, nor
gross and histopathology (Harmsen, 1959, 1960).
In another experiment an esterified product was fed orally at
the rate of 3 g/day and injected s.c. to 29 rats at 1 ml weekly for
five months and 2 ml bi-weekly for a further three months. Animals
were observed for 27 months. No tumours developed. A control series
of 30 rats treated similarly with oral esterified product and
injected s.c. with refined soybean oil showed four tumours after 24
months observation, none at the site of injection. The F1 test
generation (nine males, nine females) received 3 g esterified
product orally and 1 ml i.p. for eight weeks, followed by a further
1 ml i.p. for four months. After 11 months six survived without any
tumours being noted. The control group (14 males, 4 females)
received 1 ml soybean oil s.c. for five months and 2 ml s.c. for
three months. During the 29-month observation period, one rat
developed a tumour at the site of injection (Harmsen, 1961).
Rats fed with 20% of thermally oxidized soybean oil in the diet
had a significantly longer life-span (807 ± 32 days) than those fed
with 20% of fresh soybean oil (673 ± 42 days) (mean ± S.E.): a
similar difference was observed between other oxidized and fresh
fats and oils (Kaunitz et al., 1966).
Groups of 60 Wistar rats of each sex were fed diets containing
1.2% TOSO for 132-137 weeks. The test substance for these groups,
designated TOSO(G), was provided by Grindstedvaerket A/S, Denmark
and met official Danish specifications. Further groups of 60 rats
were fed a diet containing 0.3 or 1.2% of TOSO(N), a material of
similar specification, but obtained from Nexus ApS, Holland. Average
intake of TOSO was 0.125 and 0.5 g/kg bw/day. Other groups were fed
diets containing 3, 6 or 12% TOSOM for 132-137 weeks. The test
substance was provided by Grindstedvaerket A/S, Denmark and met
official Danish specifications. Average intake of TOSOM was 1.3, 2.7
and 5.4 g/kg bw/day in males and 1.8, 3.6 and 7.4 g/kg bw/day in
females. Wistar rats (120 of each sex) served as controls. All diets
were adjusted to a total caloric value of 12% edible fatty acids by
supplementation with mono- and diglycerides. There were minor
variations in growth, food consumption and blood bio-chemistry
throughout the study, but there was no discernible treatment-related
effect. Survival was >50% in all groups at 2 years. A decrease
(p <0.05) in total white cell count and percentage of lymphocytes
and an increase in percentage of monocytes was observed at the
highest TOSOM dose rate and in all groups receiving TOSO, but only
in some of the blood samplings. However, there was no consistent
dose or time pattern and, with the exception of an increased
percentage of monocytes at weeks 10 and 38 in rats treated with
TOSO, the parameters were within the normal range for Wistar rats.
In rats treated with TOSO, the number of rats bearing malignant
tumours was increased slightly (p <0.05) at the 1.2% dose level in
male rats given TOSO(G) and in female rats given TOSO(N) 16/60 vs
24/119; 23/60 vs 24/118 respectively), although there were no
effects on the overall number of tumour-bearing rats, or in the
number of rats with multiple tumours. The incidence of tumours at
selected sites achieved statistical significance in the case of
cortical adenomas in males which had received 1.2% TOSO(G) (5/60 vs
1/119); adenocarcinoma of the uterus in females which had received
1.2% TOSO(N) (7/60 vs 3/118) and subcutis fibrosarcoma in males
which had received 1.2% TOSO(N) (5/60 vs 3/119).
In rats treated with TOSOM, there were no treatment-related
effects on the overall number of tumour-bearing rats, or on the
number of rats with malignancies or multiple tumours. The incidence
of mammary adenoma (7/60 vs 4/118) in females and adrenal medullary
adenoma (4/59 vs 1/119) in males were the only sites at which a
significant (p <0.05) increase compared to the controls occurred.
There was no consistency across treatments, no evidence of
reduced tumour latency, and the observed incidences were within the
range of historical controls. The authors concluded that the effects
were not related to dosing with either TOSO or TOSOM. There were no
compound-related changes in the incidence of non-neoplastic lesions.
In summarizing both the neoplastic and non-neoplastic data, the
authors excluded the data from the oral cavity, then summarized it
separately. It was suggested that a relatively high incidence of
chronic inflammation and carcinomas in situ or squamous-cell
carcinomas at these sites in control and treated rats (14 of 119 in
controls and 4 to 10 of 60 in treated groups) was not associated
with the emulsifiers, but was apparently caused by penetrative
lodgement of dietary oat and barley chaff particles in the nasal and
pharyngeal cavities. The Committee was not convinced by this
explanation but could see no reason to cast doubt on the authors'
conclusion that neither TOSO nor TOSOM was responsible for the
effect (Gry et al., 1987).
2.2.4 Reproduction studies
No data available.
2.3 Observations in humans
No data available.
The Committee considered studies showing that after oral
administration of radiolabelled material to rats and mice, tissue
levels of TOSOM were slightly greater than those of TOSO. The
absorption of both thermally oxidized substances was lower than that
of refined soya bean oil.
A 2.5 year study in rats used dietary levels of 3, 6, or 12%
TOSOM (equal to 1.3, 2.7 and 5.4 g/kg bw/day in males; 1.8, 3.6 and
7.4 g/kg bw/day in females), and 0.3 or 1.2% thermally oxidized soya
bean oil (from two sources) (equal to 130 and 540 mg/kg bw/day in
males; 180 and 740 mg/kg bw/day in females). The only effects noted
were transient minor variations in total leukocyte counts and
percentages of lymphocytes and monocytes, which were not considered
to be of toxicological significance. For some sites, the incidence
of tumours in treated animals was slightly higher (P<0.05) than in
controls. However, the incidences were within the range of those of
historical controls for Wistar rats, there was no dose-response
relationship, and there was no consistency of effect for the three
compounds tested. The Committee concluded that neither TOSO nor
TOSOM is carcinogenic in rats.
Using a safety factor of 200, the Committee allocated an ADI of
0-3 mg/kg bw for TOSO and 0-30 mg/kg bw for TOSOM, based on the
observation that the highest doses in the 2.5 year rat study
(approximately 600 and 6000 mg/kg bw/day respectively) produced no
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