Registration Dossier
Registration Dossier
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EC number: 232-401-3 | CAS number: 8016-11-3
Toxicological information
Endpoint summary
Administrative data
Description of key information
A combined repeated dose toxicity with the reproduction/developmental toxicity screening test was carried out on rats according to OECD test guideline 422. A combined chronic toxicity/carcinogenicity study was carried out according to OECD test guideline 453.
Key value for chemical safety assessment
Repeated dose toxicity: via oral route - systemic effects
Link to relevant study records
- Endpoint:
- chronic toxicity: oral
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- weight of evidence
- Study period:
- 1986
- Reliability:
- other: The toxicological profile review for long term dietary exposure of rats to epoxidised soybean oil has been prepared by a BIBRA who are very reliable but the content is not necessarily drawn from a single scorable study
- Rationale for reliability incl. deficiencies:
- secondary literature
- Remarks:
- Since the data prepared by BIBRA may have been drawn from a variety of sources to construct the profile, it is not possible to allocate a score for reliability. However the reliability of the team preparing the review is considered to be 1 or 2. Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Justification for type of information:
- Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
- Principles of method if other than guideline:
- No specific guidelines are quoted. The study was completed to ensure compliance with a range of national and international requirements in relation to food contact. The study as completed was braodly in accordance with the requirements of OECD Guideline 453.
- GLP compliance:
- no
- Remarks:
- The study was undertaken before the effective date for GLP implementation in the USA
- Limit test:
- no
- Species:
- rat
- Strain:
- Wistar
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Scientific Agribusiness Consultants International
- Weight at study initiation: Batch 1 - between 20-50g; Batch 2 30-50g
- Fasting period before study: rats were not fasted
- Housing: Four to a cage in grid bottom aluminium cages measuring approx. 36x36x13 cm. These were suspended in racks carrying 20 cages above sheets of waterproof paper for collection of excreta. The paper was renewed daily.
- Diet : Laboratory Animal Diet No. 1 ad libitum
- Water : ad libitum
- Acclimation period: 14 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23 °C
- Humidity (%): 50 - 70 %
- Air changes (per hr): Partial air conditioning maintained a positive air pressure, with no recirculation of filtered 0.5um air to give 12-15 changes of air per hour. - Route of administration:
- oral: feed
- Vehicle:
- unchanged (no vehicle)
- Details on oral exposure:
- DIET PREPARATION
Spratt's Lab Diet No.1 was ground to a powder. ESBO (0.025, 0.25, 2.5 %) or 2.5 % SBO were added by weight and thoroughly mixed in a commercial food mixer. The ground diet served as the control. Fresh batches were prepared weekly. Administration was by admixture with the diet which was given ad lib for 104 weeks. The dietary levels were 0 % (controls), 0.025, 0.25% and 2.5% for epoxidised soybean oil and 2.5% for soybean oil - Analytical verification of doses or concentrations:
- no
- Details on analytical verification of doses or concentrations:
- No analyses were carried out to verify the concentration of ESBO or SBO.
- Duration of treatment / exposure:
- Administration was by admixture with the diet which was given ad lib for up to 104 weeks.
- Frequency of treatment:
- Test item was provided via diet so the diet was accessible all day
- Remarks:
- Doses / Concentrations:
0 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
0.025 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
0.25 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
2.5 %
Basis:
nominal in diet - No. of animals per sex per dose:
- The rats were distributed into five groups of 24 rats of each sex from each animal supply batch to give total of 48 animals of each sex in each group.
- Control animals:
- yes, plain diet
- Details on study design:
- No details supplied
- Positive control:
- No details supplied
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
Rats were observed at least once daily for abnormalities of appearance and behaviour or signs of ill-health. Those showing such signs were isolated and monitored more frequently. If the condition improved, the rat was returned to its original cage but if it deteriorated so that the welfare of the rat was impeded, or it was deemed unlikely to survive for another 24 hr the rat was killed for post-mortem examination and tissues kept for histological investigation. Any rat found dead in the cage was subjected to a post mortem examination and tissues preserved if not autolysed.
BODY WEIGHT: Yes
These were recorded on the first day of treatment (day 0), on days 2, 4, 7, 14, 21 and 28 and then at two-week intervals until week 104.
FOOD CONSUMPTION AND COMPOUND INTAKE : Yes
Both food and water were measured over the 24 hour period prior to bodyweight determinations.
HAEMATOLOGY: Yes
Blood was taken from a tail vein of selected rats from batch 1 from the groups given 0 (control), 0.025 and 0.25 % ESBO and from those given 2.5 % SBO. Ten males from each dose level were bled during week 13, 26, 53 and 79 and ten females were bled during week 15, 26, 53 and 79. The blood was examined for haemaglobin content, packed cell volume and erythrocyte and leucocyte counts. Appropriately stained microscope preparations were used to identify the different types of leucocytes and the proportion of erythrocytes present as reticulocytes. Examination of these slides was confined to those from the control group and the rats given 2.5 % ESBO or SBO. As part of the scheduled post-mortem examinations blood was taken from the aorta from each animal during the anaesthetic phase and examined as described above.
CLINICAL CHEMISTRY: Yes
Serum separated from the blood samples taken at the end of the study was used for biochemical analyses. This was analysed for the activities of glutamic-oxaloacetic and glutamic-pyruvic transaminases, lactic dehydrogenase and for content of glucose, urea, total protein and albumin.
URINALYSIS: Yes
During week 26, 52, 78-80 and 104 urine samples were collected from 10 rats of each sex from batch 1 of each dose level except those given 0.025 % ESBO. Collections made at week 13 were confined to the controls and 2.5 % ESBO group. The samples were obtained by placing the animals individually in metabolism cages without food or water. three samples were collected from each rat at each interval. The conditions of the collection and examinations were:
- Collected over a 6 hr period following the normal overnight feeding and drinking and examined for volume, concentration (by refractive index), pH and the presence of glucose, blood, bile salts, ketones and proteins
- Collected over a 2 hour period immediately following an oral water load of 25 ml/kg and examined for volume, concentration and content of cells.
- Collected over a 4 hour period commencing 16 hour after an oral water load of 25 ml/kg. Food but not water was available to the animals during the 16 hour period. These samples were examined for volume and concentration. - Sacrifice and pathology:
- After 104 weeks all surviving rats were killed, subject to post-mortem examination with recording of organ weights and smaples of tissues preserved. Feeding with the appropriate diet continued until all rats had been killed. The tissues from all rats were processed for microscopic examination.
GROSS PATHOLOGY: Yes
Rats surviving to the end of the study were weighed and the weights of brain, heart, liver, spleen, kidneys, stomach, small intestine, caecum (with and without contents), gonads, pituitary and thyroid recorded. When possible the following tissues, together with any other abnormal tissue, were preserved in 10 % buffered formalin: adrenal glands, aorta, bladder (urinary), brain, caecum, colon, eye, gonads, harderian gland, heart, kidneys, liver, lungs, lymph nodes, mammary gland, muscle (skeletal), nerve (sciatic), oesophagus, pancreas, pituitary gland, prostate, salivary glands, seminal besicles, skin, small intestine, spinal cord, spleen, stomach, thymus gland, thyroid gland, trachea, uterus, vagina.
HISTOPATHOLOGY: Yes
All tissues collected were preocessed for embedding in paraffin-wax, sectioned at approx. 5 um and stained with haematoxylin and eosin. Deviations from normal seen by the light microscope were noted. - Other examinations:
- No details supplied
- Statistics:
- Analyses of variance and least significance difference test for Body weights, Haematology, Urine volume, Refractive index, pH and cell count, Food intake, Water intake, Serum analyses, Organ weights and organ weights relative to bodyweight. Chi2 for heterogeneity using data from all groups and Fisher's exact test for comparing individual groups: Semi-quantitative urine analyses, histological findings, tumour incidence. In all cases treated groups were compared with the controls and a probability of less than 0.05 taken to indicate statistical significance.
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- There were a few deaths at Week 52 but these were not dose related. Please see below for more details.
- Mortality:
- mortality observed, treatment-related
- Description (incidence):
- There were a few deaths at Week 52 but these were not dose related. Please see below for more details.
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- Bodyweight changes at the high dose level were not indicative of treatment related effects
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- Slightly lower food consumption at higher doses, suggesting a compensatory nutritive value for the soybean oil, since bodyweights increased as diet intake went down.
- Food efficiency:
- not examined
- Water consumption and compound intake (if drinking water study):
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- No treatment related effects
- Clinical biochemistry findings:
- effects observed, treatment-related
- Description (incidence and severity):
- No treatment related effects
- Urinalysis findings:
- effects observed, treatment-related
- Description (incidence and severity):
- No adverse effects and possible beneficial effects on long term renal function from soybean oil administration
- Behaviour (functional findings):
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Description (incidence and severity):
- No treatment related effects.
- Gross pathological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- see results section below
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- see results section below
- Histopathological findings: neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- see results section below
- Details on results:
- CLINICAL SIGNS AND MORTALITY
No observations of condition or behaviour that could be related to treatment were reported. An eye infection at 11 and 12 weeks affected from 10-25 % of rats of each group indiscriminately and the animals recovered uneventfully. It is likely that this was as a result of dacryoadentis. In the first 52 weeks there were few deaths and they were not dose related. By week 78, 17-19 % of the controls had died. This increased to 38-42 % by week 96 and 48-52% at week 102. With the exception of the females given SBO and the females given the lowest dose (0.025 %) of ESBO there were less deaths in the treated groups than the controls. Please see table 1. Since this result did not demonstrate an adverse effect of treatment the data were not subject to detailed statistical analysis. However, in both sexes the number of deaths by 102 weeks in the high dose ESBO group was significantly (p<0.05 in males and p<0.01 in females by Fisher test) less than the control.
BODY WEIGHT AND WEIGHT GAIN
Neither 0.025 nor 0.25 % ESBO in the diet affected the body weights of males; no statistically significant difference from control weights were seen. Mean body weights of the males given 2.5 % ESBO were greater than the control and the differences were statistically significant compared with the controls, from week 58 to 92. For the remainder of the study the weights were greater than control but not always to a statistically significant degree. The mean bodyweights of the SBO-treated male rats were higher than those of the ESBO-treated animals and were statistically significantly different from the controls on most occasions from week 8 to the end of the experiment. The individual statistical comparisons were supported by an overall significant value in the analysis of variance from week 28. The pattern of bodyweights in the females was different. The weights of those given diet containing SBO were similar to the control up to week 74. After that time they tended to be heavier than the controls although the differences were seldom statistically significant. The difference appears to be mainly due to an irregularity in the growth curve of the controls at week 76 and 78. Although there was an early period (week 16-34) when the weights of the females given 2.5 % ESBO were higher than control by week 46-48 they were lower the difference first being statistically significant at week 54. Most of the values were significantly less than control up to week 90 although the differences were small, never being more than 8 % of the control weight. After week 90 the treated animals maintained their weight compared with a loss in the controls so that any differences were reduced. Statistically significantly lower body weights were seen in females given 0.25 % ESBO at some intervals between 64 and 74 weeks but, again the differences from control were small (not more than 6 % of the control).
FOOD CONSUMPTION AND COMPOUND INTAKE
The treated animals tended to eat slightly less than the controls although most of the values were within 10 % of the concurrent control. The analysis of variance of the data from all groups of one sex was statistically significant on very few occasions. The comparison of individual groups of one sex was statistically significant on very few occasions. The comparison of individual groups with the control revealed relatively few statistically significant differences. Many of these did not appear to be related to treatment as they were present only in the low dose groups or were similar at all dose levels. The intake by the male high dose ESBO group over the whole study was reduced by approximately 0.5 g/day and the females by 1 g/day. Similar reductions of food intake were apparent in the groups given SBO. The average intakes during the study by the males were approx. 10, 100 and 1000 g/kg for the rats given 0.025, 0.25 and 2.5 % ESBO respectively. The corresponding values for the females were approximately 14, 140 and 1400 mg/kg/day.
FOOD EFFICIENCY
The calculated intakes of ESBO and SBO in relation to bodyweight found that young rats consumed a higher dose level than the older rats. The intake by the females was greater than those of the males.
HAEMATOLOGY
The haematological measurements made on a proportion of the rats during the course of the study which related to the erythrocytes (red cell count, haemoglobin concentration, packed cell volume and reticulocyte count) showed little variation between the treated and control groups. There were isolated statistically significant differences, mainly increases, but these were not repeated at the later examinations. Similarly the total and differential white counts showed variations, sometimes statistically significant but these tended to be small and not to persist with prolonged treatment.
CLINICAL CHEMISTRY
There was a marked variation of results of the serum analyses in the old animals surviving to the end of the study. However, there were no apparent dose-related trends in the mean values. The only statistically significant differences from control were lower total protein concentrations in the male rats given SBO or the intermediate dose of ESBO and lower lactic dehydrogenase activity in those females given the highest dietary concentration of ESBO.
URINALYSIS
The majority of results of the renal concentration and dilution tests and of the urinary pH and cell excretion counts were similar in treated and control rats with no apparent dose - or time-related differences. In the males the only statistically significant difference was more concentrated urine collected in the 18-24 hour period when the rats were examined at 13 weeks. The females from the highest dose group of ESBO sampled at 78 and 104 weeks produced a lower volume of more concentrated urine than the controls in the 6-hour collection. In the same group the urinary cell excretion was higher than control from week 52 although, due to a marked variation, the differences were significant only at 1 year. The pH of the same urines was low although only by 10 % or less of the control. The only urine sample to give a reaction for bilirubin was from a control female at the end of the study. The number of samples with positive reactions for blood and the various concentrations of protein were similar in treated and control groups throughout the study. At the examination at 78-80 weeks, compared with controls, there were more males from the group given 2.5 % ESBO with glucose in the urine. After a further six months treatment this finding was not repeated even though the control incidence was still low. At the end of the study there were more positive reactions for ketones in the females given 2.5 % ESBO or 2.5 % SBO than in the controls.
ORGAN WEIGHTS
The organ weights and relative organ weights of both sexes given 0.025 or 0.25 % ESBO did not differ significantly from those of the controls. At the highest dose of ESBO (2.5 %) in males the liver, small intestine and thyroids were statistically significantly heavier than those of the control but these differences were not apparent when the values were expressed relative to body weight. The corresponding females had a significantly higher value for liver weight again not apparent when expressed relative to body weight. In the females the weights of the adrenal glands were less than the control and this persisted when the values were related to body weight. The male rats given SBO, with their higher body weights, had significant increases of most organ weights although again in relation to bodyweight they were comparable with the controls. In these rats the brain was one of the few organs not heavier than control and when expressed relative to the higher body weight the mean was significantly lowered. Both the organ weights and relative organ weights of the females given SBO were similar to the control values.
GROSS PATHOLOGY
Most of the findings in the treated male groups were of lower or similar incidence to the controls. None of the findings in the ESBO-treated males were present with a statistically significantly greater incidence than in the controls. All the male rats had some degree of glomerulonephrosis and within the control and ESBO-treated animals there was a trend for the higher dietary levels to have the less severe grades. An analysis for linear trend by Chi2 was significant (p<0.05) although there was no overall heterogeneity in the data. The distribution of the different grades of glomerulonephrosis was similar between the control and SBO-treated rats. The males given SBO had significantly higher incidences of Harderian glands showing secretion, congested lymph nodes and cystic seminal vesicles. In the females again most of the findings in the treated and the control groups were similar. As with the males there was a suggestion of a trend to less severe glomerulonephrosis but the differences or trend were not statistically significant. There were isolated non dose-related findings. The number of rats from the lowest dose with cystic spaces in the adrenal, with cardiac interstitial fibrosis and with congestion of the lymph nodes were higher than control. A similar incidence of lymph node congestion was seen in the rats given SBO. The SBO group also showed an increased incidence of pituitary haemorrhage compared with the control. There were a number of endometrial changes at the highest dose of ESBO. The incidence of cystic endometrium and of hyperplastic endometrium were significantly greater tan the control. Some animals had more than one of the three findings and the total numbers with one or more of the changes were 2, 4, 1, 10 and 4 for the control, three dose levels of ESBO and for SBO respectively. analysed by Chi2 these data showed significant heterogeneity and the incidence in the high dose ESBO group was significantly (p < 0.05) greater than the control. The difference between the 2.5 % ESBO and the SBO groups was not significant.
HISTOPATHOLOGY: NEOPLASTIC
in the males there were a total of 69 tumours, the most common being basophilic adenoma of the pituitary representing 35 % of the total with an even distribution through groups. The remainder were small incidences and only one, pancreatic-cell adenoma showed heterogeneity of distribution. This was due to the presence of four of the five tumours in the SBO group. Many of the tumours in males occurred in the controls alone with, the same incidence in treated and control groups or in the SBO group. Single cases of hepatocellular carcinoma, subcutaneous carcinoma, lymphoid leukaemia and thyroid adenoma were found in high dose (2.5 %) ESBO group with no corresponding control finding. A thyroid adenoma was found also in a male given 0.05 % ESBO. Some tumours; a Harderian-gland adenoma, a lymphoblastoma, two lymphosarcoma, a pancreatic-cell adenoma, a pituitary basophilic carcinoma, two subcutaneous fibrosarcoma, a parathyroid adenoma, and a fibrosarcoma in the abdomen were found in one of the lower-dose ESBO groups without a similar findings in the control or high dose group. There were more tumours in females with a total of 120 but of these 78 (65%) were pituitary adenoma with the highest incidence in the control but no significant heterogeneity between the groups. The remaining tumours were of low incidence in the ESBO-treated animals than in the controls. None of the individual tumours or the total incidences of rats affected showed significant heterogeneity among the groups or any significant increases in treated groups compared with the controls. Those present in the high-dose ESBO females without a corresponding control finding were single incidences of adrenal cortical carcinoma, pancreatic-cell adenoma and thymic lymphoblastomas as well as two thymic lymphosarcoma and uterine fibroleiomyoma. In the case of the uterine tumour there was also a single affected animal amoung the 0.25 % ESBO-treated rats. There were, in addition, single incidences of a number of tumours in the females given 0.025 or 0.25 % ESBO with no parallel finding in the controls or high dose. The lesions involved were a squamous carcinoma of the eye, a hepatic haemangioma, a mesenteric haemangioma, an oesophageal squamous carcinoma, an ovarian adenocarcinoma, a splenic reticulum-cell sarcoma, a splenic lymphosarcoma, a parathyroid adenoma and an uterine fibrolipoma. - Dose descriptor:
- NOEL
- Effect level:
- 1 other: g/kg
- Based on:
- test mat.
- Sex:
- male
- Basis for effect level:
- other: Please see results
- Dose descriptor:
- NOEL
- Effect level:
- 1.4 other: g/kg
- Based on:
- test mat.
- Sex:
- female
- Basis for effect level:
- other: Please see results
- Critical effects observed:
- not specified
- Conclusions:
- It was concluded that ESBO was not carcinogenic when fed to rats at up to 2.5 % of the diet. Further it was concluded that the NOEL was 2.5 % providing an average daily intake of approximately 1.0 g/kg in the males and 1.4 g/kg in the females.
- Executive summary:
Groups of 48 male and 48 female rats of a Wistar strain were fed diet containing 0.025, 0.25 or 2.5 % Epoxidised Soya Bean Oil (ESBO) for 2 years. Similar groups were given basal diet to serve as a control or 2.5 % Soya bean Oil (SBO).
There was no adverse effect on survival. The males given 2.5 % ESBO gained more weight than the controls whilst the females were slightly lighter. The same rats consumed slightly less food than controls, the difference being greater in females (approximately 1 g/rat/day) than males (approximately 0.5 g/rat/day). The water intake of the females given 2.5 % ESBO was lower than the control especially in the second year of the study.
Haematological examination and investigations of urine at 3, 6, 12, 18 and 24 months did not reveal any adverse effects. A lower volume of more concentrated urine was excreted by the females given 2.5 % ESBO compared with the controls with occasional increases in urinary cell excretion. The organ weights in females were similar to controls whilst in males given 2.5 % ESBO and more noticeably in those given 2.5 % SBO several organs were heavier than control. This was related to the growth changes since when expressed relative to bodyweight the values were normal.
The incidence of most histological findings including tumours were similar in treated and control groups. There was a tendency for less severe glomerulonephrosis in the ESBO-treated rats. There was a marginally increased incidence of uterine changes in the females given 2.5 % ESBO. Since there were similar changes in the females given SBO these changes could not be clearly related to ESBO.
It was concluded that ESBO was not carcinogenic when fed to rats at up to 2.5 % of the diet. Further it was concluded that the NOEL was 2.5 % providing an average daily intake of approximately 1.0 g/kg in the males and 1.4 g/kg in the females.
- Endpoint:
- chronic toxicity: oral
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- weight of evidence
- Study period:
- 1960
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: see 'Remark'
- Remarks:
- The number of animals used at each dose level differ from the recomendations of current test guidelines, which indicate at least 20 animals per sex per group for rodent studies (rather than 15 as used in current study)should be used at each dose level, and for non-rodent studies a minimum of 4 animals per sex per group must be used (3 mongrel dogs used in current study). For this reason, the infomation published can only be supporting data rather than a key study. Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Justification for type of information:
- Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- no guideline available
- Principles of method if other than guideline:
- No guideline is referred to in this report.
Groups of 15 rats per sex were allocated to groups dosed at 0, 0.1, 0.5,1.0, 2.5 or 5.0% by dietary admixture. The groups were subdivided with 5 per sex per dose level scheduled for termination after one year and the remaining groups of ten scheduled for termination after two years of treatment.
Groups of 3 dogs were fed the same treated diet, at concentrations of 0, 1.0 or 5.0% but survivors were all terminated after one year.
A second study was run in conjunction with the Paraplex G-60 study.
For Paraplex G-62 the design was as described above except that the subgroups of 5 rats were terminated after six months rather one year. - GLP compliance:
- no
- Limit test:
- no
- Species:
- other: rats and dogs
- Strain:
- other: albino rats and mongrel dogs
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- No data provided on test animals or their environmental conditions prior to test
- Route of administration:
- oral: feed
- Vehicle:
- unchanged (no vehicle)
- Details on oral exposure:
- PARAPLEX G-60
ALBINO RATS
Fifteen young male and 15 female albino rats were placed on each of the following dietary levels of the test material: 0, 0.1, 0.5, 1.0, 2.5 and 5.0 %.
Finely ground Purina Dog Chow Kibbled Meal served as the basic diet, and into this was incorporated amounts of Paraplax G-60 calculated to achieve the above dietary concentrations.
MONGREL DOGS
Using the same basic diet, 3 mongrel dogs were placed on the following dietary concentrations of Paraplex G-60 mixed in finely ground Purina Dog Chow Kibbled Meal: 0.1, 1.0, and 5.0 %. They were fed once daily, the food being moistened with an equal weight of water.
PARAPLEX G-62
The diet used for the rats and dogs was the same as that described above for Paraplex G-60. The dose levels used were the same for rats and dogs as those detailed above. - Analytical verification of doses or concentrations:
- not specified
- Details on analytical verification of doses or concentrations:
- No data provided
- Duration of treatment / exposure:
- PARAPLEX G-60
Albino Rats - one or two years (sub-group of five rats per dose level scheduled for termination after one year)
Mongrel Dogs - one year
PARAPLEX G-62
Albino Rats - six months or two years (sub-group of five rats per dose level scheduled for termination after six months)
Mongrel Dogs - one year - Frequency of treatment:
- Once daily
- Remarks:
- Doses / Concentrations:
Rats: 0, 0.1, 0.5, 1.0, 2.5 and 5.0 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
Dogs: 0.1, 1.0, and 5.0 %
Basis:
nominal in diet - No. of animals per sex per dose:
- Albino Rats: 15 male and 15 female
Dogs: 3 dogs - Control animals:
- yes, concurrent no treatment
- Details on study design:
- PARAPLEX G-60
ALBINO RATS
Fifteen young male and 15 female albino rats were placed on each of the following dietary levels of the test material: 0, 0.1, 0.5, 1.0, 2.5 and 5.0 %.
Finely ground Purina Dog Chow Kibbled Meal served as the basic diet, and into this was incorporated amounts of Paraplax G-60 calculated to achieve the above dietary concentrations. The rats were individually caged and were weighed once weekly. Each group of 15 was subdivided into subgroups of 10 and 5 animals. At the end of one year, the survivors in the subgroups of 5 were sacrificed for histopathologic studies, organ to body weight measurements were made for liver, kidney and testes. The subgroups of 10 were continued on diet for a second year. Blood studies (haemoglobin, red blood cell and differential white cell counts) were made during the eleventh and twenty-fourth months. Histopathologic studies were made on two-year survivors from the 0, 2.5 and 5 % dietary groups.
MONGREL DOGS
Using the same basic diet, 3 mongrel dogs were placed on the following dietary concentrations of Paraplex G-60 mixed in finely ground Purina Dog Chow Kibbled Meal: 0.1, 1.0, and 5.0 %. They were fed once daily, the food being moistened with an equal weight of water. The dogs were weighed once weekly. Hematologic studies (haemoglobin, red and white cell and differential white cell counts) were made at the start of the study, at 6 months and at 12 months; after this the dogs were sacrificed for histopathologic studies.
PARAPLEX G-62
This study on rat and dogs was identical in all respects to that described above for Paraplex G-60 except that the rat subgroups of 5 were sacrificed at the end of 6 months and haematologic studies were made at 6 as well as at 12 and 24 months. - Positive control:
- No data
- Observations and examinations performed and frequency:
- PARAPLEX G-60
ALBINO RATS
At the end of one year, the survivors in the subgroups of 5 were sacrificed for histopathologic studies, organ to body weight measurements were made for liver, kidney and testes. The subgroups of 10 were continued on diet for a second year. Blood studies (haemoglobin, red blood cell and differential white cell counts) were made during the eleventh and twenty-fourth months. Histopathologic studies were made on two-year survivors from the 0, 2.5 and 5 % diets.
MONGREL DOGS
Haematologic studies (hemoglobin, red and white cell and differential white cell counts) were made at the start of the study, at 6 months and at 12 months; after this the dogs were sacrificed for histopathologic studies.
PARAPLEX G-62
This study on rat and dogs was identical in all respects to that described above for Paraplex G-60 except that the rat subgroups of 5 were sacrificed at the end of 6 months and hematologic studies were made at 6 as well as at 12 and 24 months. - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes - Other examinations:
- PARAPLEX G-60
ALBINO RATS
Blood studies (haemoglobin, red blood cell and differential white cell counts) were made during the eleventh and twenty-fourth months.
MONGREL DOGS
Haematologic studies (hemoglobin, red and white cell and differential white cell counts) were made at the start of the study, at 6 months and at 12 months
PARAPLEX G-62
This study on rat and dogs was identical in all respects to that described above for Paraplex G-60 except that the rat subgroups of 5 were sacrificed at the end of 6 months and haematologic studies were made at 6 as well as at 12 and 24 months. - Statistics:
- No definite effect on survival was apparent for rats. For rats of both sexes there was early depression of growth at the 5 % dietary level (P = < 0.05 at 2 and 4 weeks), this was followed by accelerated growth. Haematologic values were within normal limits at all dietary levels.
All dogs fed Paraplex G-60 survived to the end of the treatment year.
No details of statistical methods or tests applied to the data are included in the publication. - Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- Survival unaffected by treatment. The publication does not report any notable signs of clinical reaction to treatment
- Mortality:
- mortality observed, treatment-related
- Description (incidence):
- Survival unaffected by treatment. The publication does not report any notable signs of clinical reaction to treatment
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- Weight gain was reduced during early phase of treatment but recovered in rats following prolonged exposure but the dogs continued to show some weight gain depression to end of treatment period.
- Food consumption and compound intake (if feeding study):
- no effects observed
- Food efficiency:
- not examined
- Water consumption and compound intake (if drinking water study):
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- no effects observed
- Clinical biochemistry findings:
- not examined
- Urinalysis findings:
- not examined
- Behaviour (functional findings):
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Description (incidence and severity):
- Male rats had elevated relative liver weights at high dose levels for Paraplex G-60 and G-62. Females at high dose had show elevation of relatve kidney weight after dosing with G-62. No effects on dog organ weights.
- Gross pathological findings:
- effects observed, treatment-related
- Histopathological findings: non-neoplastic:
- no effects observed
- Details on results:
- PARAPLEX G-60
ALBINO RATS
No definite effect on survival was apparent. In both sexes there was early depression of growth at the 5 % dietary level. (P = < 0.005 at 2 and 4 weeks); this was followed by accelerated growth. Haematology values were within normal limits at all dietary levels. Organ to body weight measurements made on those animals that were sacrificed at the end of one year on the diet showed that male rats receiving 5 % diet had signifcantly elevated liver to body weight ratios. Histopathology of the heart, lung, liver, kidney, spleen, gastroenteric, thyroid, adrenal, pancreas, gonad, muscle and bone marrow tissues of the rats after one or two years on treated diet showed no lesions attributable to treatment.
MONGREL DOGS
All dogs dosed with diet containing Paraplex G-60 survived the one-year feeding period. Those in the high, 5 %, concentration group lost weight and from their consumption records, it appeared that they showed an aversion to their diet. The haematologic values remained within normal limits throughout. Histopathological studies, carried out on the same tissues as in the rats, showed no lesions attributable to treatment.
PARAPLEX G-62
ALBINO RATS
No adverse effect on survival is apparent. Among female rats, growth was significantly depressed during the early period on the diet. Analysis of regression of the data (linear at 2 and 8 weeks; curvilinear being required by the diet at 4 weeks) with 95 % confidence limits indicated the effective concentration to be about 3.75 % at 2 weeks and 1.0 % at 4 weeks, followed by no significant depression by 8 weeks. Among male rats, growth was also depressed during the early period. Analysis of linear regression of the data with 95 % confidence limits indicated the effective concentration to be about 2.1 % at 2 weeks, 1.6 % at 4 weeks and 3.7 % at 8 weeks, after which the effect gradually disappeared. Hematologic values were within normal limits at each concentration. Organ to body weight measurements were made on those animals that were sacrificed at the end of 6 months on the diet. Analysis of regression of the liver ratio data (curvilinear for females, linear for mlaes) with 95 % confidence limits indicate that significantly elevated rations result from the treatment and that the effective concentration should fall at 0.5 % for females and 2.3 % for males. Similar statistical treatment of the kidney ratio data indicated the occurence of significantly elevated ratios from the treatment for females only and that the effective concentration should fall at 1.6 %. Histopathologic studies made on the same tissues as listed for Paraplex G-60 showed no lesions attributable to treatment.
MONGREL DOGS
All dogs on Paraplex G-62 containing diets survived the one-year feeding period. Overall, it was observed that the dogs in the 0.1 and 1.0 % diet range increased in weight over the course of the year. Those dogs on the 5 % concentration lost weight and from their food consumption records, it appeared that they showed an aversion to their diet. Haematology values remained within normal limits throughout. Histopathology studies on ths smae tissue as in other animals descrived above revealed no lesions except for minimal fatty infiltration of the liver in one of the dogs receiving the 5% concentration. - Dose descriptor:
- NOAEL
- Remarks:
- highest concentration in diet with no adverse toxic effects on dogs or rats
- Effect level:
- 5 other: %
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: see 'Remark'
- Critical effects observed:
- not specified
- Conclusions:
- Two epoxidised soybean oils, Paraplex G-60 and Paraplex G-62, were added to the diets of rats for a two-year period and of dogs for one year in concentrations up to 5 %. No effects were observed on survival or on the blood picture and histologic examination of tissues revealed no lesions attributable to treatment. Early depression in growth was observed in rats from both materials (Paraplex G-62 being the most potent), but this effect disappeared on continued feeding. Dogs receiving the 5 % concentrations lost weight, an effect which persisted throughout the feeding period. Elevated liver to body weight ratios resulted at the 5 % concentration in male rats receiving Paraplex G-60 and at lower concentrations in both male and female rats receiving Paraplex G-62; kidney to body weight ratios also elevated with the latter material in female rats.
- Executive summary:
Two epoxidised soybean oils, Paraplex G-60 and Paraplex G-62, were added to the diets of rats for a two-year period and of dogs for one year in concentrations up to 5 %. No effects were observed on survival or on the blood picture and histologic examination of tissues revealed no lesions attributable to treatment. Early depression in growth was observed in rats from both materials (Paraplex G-62 being the most potent), but this effect disappeared on continued feeding. Dogs receiving the 5 % concentrations lost weight, an effect which persisted throughout the feeding period. Elevated liver to body weight ratios resulted at the 5 % concentration in male rats receiving Paraplex G-60 and at lower concentrations in both male and female rats receiving Paraplex G-62; kidney to body weight ratios also elevated with the latter material in female rats.
- Endpoint:
- short-term repeated dose toxicity: oral
- Remarks:
- combined repeated dose and reproduction / developmental screening
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- weight of evidence
- Study period:
- November 24, 2004 - June 13, 2005
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- GLP compliant study completed in accordance with current guidelines - OECD 422. Only minor deviations occurred with no notable effect on the outcome of the investigations - see 'Overall comments' for details. The study was conducted with ETP rather than ELSO (epoxidised linseed oil) but read-across between the two oils is considered appropriate., based on similarity of structure and function. Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Justification for type of information:
- Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
- Deviations:
- yes
- Remarks:
- see 'Overall Remarks'
- GLP compliance:
- yes
- Species:
- rat
- Strain:
- other: HanBrl:WIST (SPF)
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: RCC Ltd. Switzerland
- Age at study initiation: 10 weeks minimum
- Weight at study initiation: Males - 291-342 g Females - 180-223g
- Fasting period before study: Only fasted before collection of blood samples
- Housing: Animals were housed in Makrolon cages with wire mesh tops and standard granulated softwood bedding. During the prepairing period, males and females were housed individually. Cages of males were interspersed amongst those holding females to promote the development of regular estrous cycles. During the pairing period; rats were housed one male/one female in Makrolon pairing cages. After mating or at the end of the pairing period, the males and the females were housed individually again. During the lactation period (until day 4 of lactation), dams were housed together with their litters.
- Diet ( ad libitum): Pelleted standard Kliba-nafag 3433 rat/mouse maintenance diet (Batch no. 53/04)
- Water ( ad libitum): Fullinsdorf community potable tap water provided in cages bottles
- Acclimation period: Minimum of 6 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 5 °C
- Humidity (%): 30-70 %
- Air changes (per hr): 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours dark and at least 8 hours light - Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS:
The test item was weighed into a glass beaker on a tared precision balance and approximately 80 % of the vehicle added (w/v). Using an appropriate homogenizer a homogenous mixture was prepared. The remaining vehicle was added until the required final volume was achieved. Separate formulations were prepared for each concentration at weekly intervals. During the daily administration period homogeneity of the test item in the vehicle was maintained using a magnetic stirrer.
VEHICLE
- Justification for use and choice of vehicle (if other than water): ETP is stable in Corn Oil. Analytical evaluations were conducted by RCC. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Samples for determination of concentration, homogeneity and stability (7 days) of the dose formulations were taken immediately after the first dose preparation. Determination of concentration and homogeneity of these samples was performed prior to the first dosing occasion. Additionally, samples for determination of concentration and homogeneity were taken once during gestation.
In addition to these two sets of samples,.samples were also retained from each set of weekly dose formulations (three samples from top, middle and bottom). These samples were not analysed since no discrepancy (outside the specified range) was noted in those samples scheduled for analysis.
For determination of concentration and homogeneity three samples of approximately 2 g were taken from the top, middle and bottom of each formulation and transferred into flat bottomed flasks. Stability samples (approximately 2 g) were taken from the middle only. The samples were frozen (-25 °C to -15 °C) pending analysis. Analysis was performed using GC.
Actual sample concentrations had to be within the accepted limit of 80 % - 120 % of the nominal concentration. Results of homogeneity must not deviate more than 15 % from the respective mean value. Stability results were accepted if the deviation from mean homogeneity results was not higher than 10 %. After analysis and evaluation of the results, the dose formulation samples were discarded at the discretion of the principle investigator for the analytical phase. - Duration of treatment / exposure:
- After acclimatization, animals of both sexes received the test item for 14 days prior to pairing and during the pairing period. Daily dosing of the females was continued throughout pregnancy and up to Day 4 of lactation (4 or 5 days post-partum). Males were dosed for at least 28 days and until the day prior to scheduled necropsy. Males were terminated on day 29 and pups on day 4 post-partum. The dams were terminated on day 5 or 6 post-partum
- Frequency of treatment:
- ETP was administered once daily orally (by gavage). All animals received a dose volume of 2 mL/kg
- Remarks:
- Doses / Concentrations:
0 mg/kg body weight/day
Basis:
other: gavage - Remarks:
- Doses / Concentrations:
100 mg/kg body weight/day
Basis:
other: gavage - Remarks:
- Doses / Concentrations:
300 mg/kg body weight/day
Basis:
other: gavage - Remarks:
- Doses / Concentrations:
1000 mg/kg body weight/day
Basis:
other: gavage - No. of animals per sex per dose:
- 40 males, 10 per group
40 females, 10 per group
Please see Table 1 for more information - Control animals:
- yes, concurrent vehicle
- Details on study design:
- No details supplied
- Dose selection rationale: The purpose of this study was to generate preliminary information concerning the effects of Fatty acids, tall-oil, epoxidized,2-ethylhexylesters (ETP) on the possible health hazards likely to arise from repeated exposure over a relatively limited period of time. In addition, it
provided information on possible effects on male and female reproductive performance such as gonadal function, mating behavior, conception, development of the conceptus and parturition. ETP was administered once daily orally (by gavage) to males for at least 28 days and to female rats throughout the prepairing and pairing periods until day 3 of lactation. The choice of 100, 300 and 1000 mg/kg bw/day is a standard selection for a dose range-finding investigation.
- Rationale for animal assignment (if not random): P generation allocated randomly
- Rationale for selecting satellite groups: Not applicable
- Post-exposure recovery period in satellite groups: Not applicable - Positive control:
- No positive control included in the study.
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
All animals were checked at least twice daily for any mortalities. All animals were observed at least twice daily for signs of reaction to treatment and/or symptoms of ill health . Additionally, the females were observed for signs of difficult or prolonged parturition.
DETAILED CLINICAL OBSERVATIONS: Yes
Prior to the first administration and then at weekly intervals thereafter, a detailed clinical observation was performed outside the home cage. Animals were observed for the following: changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions and automatic activity (e.g. lacrimation, piloerection, pupil size, unusual respiratory pattern); changes in gait, posture and response to handling as well as the presence of clonic and tonic movements, stereotypies or bizarre behaviour.
At one time during the study (males: shortly before scheduled sacrifice; females: on day 3 or 4 post partum) relevant parameters from a modified Irwin screen test were performed for five P generation males and five P generation females randomly selected from each group. This FOB assessment was conducted following the daily dose administration. Animals were observed for the following: (a) Cage side observations: unusual body movements, abnormal behaviour, (b) Hand-held observations: palpebral closure, pinna reflex, lacrimation, pupil size, pupil reactivity, salivation (c) Open field observations: level of ambulatory activity including rearing, responsiveness to sharp noise (d) Categorical observations: hair coat, behaviour, respiration, muscle movements (e) Measurements/Counts: hindlimb/forelimb grip
BODY WEIGHT: Yes
The animals were weighed daily throughout the entire study.
FOOD CONSUMPTION AND COMPOUND INTAKE :
Males: Food consumption was recorded weekly during the pre-pairing and post-pairing period.
Females: Food consumption was recorded for the following periods: days 1-8 and 8-14 of the pre-pairing period; days 0-7, 7-14 and 14-21 post coitum and days 0-4 post partum. Food consumption was not recorded during the pairing period since these would have been mixed values of males and females.
HAEMATOLOGY: Yes
Blood samples were obtained on the day of scheduled necropsy from all P generation males after they had been fasted overnight. Blood samples of P generation females were obtained on day 5 or 6 post partum after the females had been fasted overnight. Blood samples were collected from the retro orbital sinus with the animals under light isoflurane anaesthesia.
Blood samples were collected early in the working day to reduce biological variation caused by circadian rhythms. The following haematology parameters were determined: Erythrocyte count, Haemoglobin, Haematocrit, Mean corpuscular volume, red cell volume distribution width, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, haemoglobin concentration distribution width, platelet count, total leukocyte count, differential leukocyte count, coagulation, thromboplastin time, activated partial thromboplastin time.
CLINICAL CHEMISTRY: Yes
The following clinical biochemistry parameters were determined: glucose, urea, creatinine, bilirubin (total), cholesterol (total), Aspartate aminotransferase, Alanine aminotransferase, bile acids, alkaline phosphatase, Gamma-glutamyl-transferase, Sodium, Potassium, Chloride, Calcium, Phosphorus inorganic, protein (total), albumin, globulin, albumin/globulin ratio.
LACTATION AND LITTER DATA:
Day 0 of lactation was the day on which a female had delivered all her pups. The litters were examined for litter size, live birth, stillbirth, and any gross anomalies. The sex ratio of the pups was recorded. The dams were caged together with their litters until day 4 of lactation. Pups were weighed individually (without identification) on days 0 (if possible), and with identification on days 1 and 4 post partum. The dams and pups were observed daily for survival and behavioural abnormalities in nesting and nursing. - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
Males were sacrificed on day 29 after treatment for 28 days. Pups were sacrificed on day 4 post partum. Females were sacrificed on day 5 or 6 post partum. The animals were examined macroscopically for any structural abnormalities or pathological changes, with special attention paid to the organs of the reproductive system. The number of implantation sites and corpora lutea was recorded for all dams with litters. Dead pups (except where excessively cannibalized) and pups killed at day 4 of lactation were examined macroscopically.
Of all parental animals the following tissues were preserved in neutral phosphate buffered 4 % formaldehyde solution: gross lesions, prostate, testes (in Bouin’s fixative), seminal vesicles with coagulation gland, ovaries, epididymides (in Bouin’s fixative), brain, spinal cord, small and large intestines (incl. Peyer’s patches), stomach, liver, kidneys, adrenals, spleen, heart, uterus with vagina, thymus, trachea and lungs (preserved by inflation with fixative and then immersion), thyroid, lymph nodes (mesenteric and mandibular), urinary bladder, peripheral nerve, and bone marrow.
HISTOPATHOLOGY: Yes
Full histopathology was carried out on the preserved organs and tissues of the animals in the vehicle control and high dose group (with special emphasis on stages of spermatogenesis and histopathology of interstitial testicular cell structure). For liver and thyroid examinations were extended to the animals of the other dosage groups, since treatment-related changes were seen in the highest dose group. All gross lesions were examined.
ORGAN WEIGHTS: Yes
For all adult female and male animals in each group the following organs were trimmed from any adherent tissue, as appropriate, and their wet weight taken: liver, brain, adrenals, thymus, kidneys, spleen, heart, testes and epididymides. - Other examinations:
- Deailed above
- Statistics:
- Dunnett t-test, Steel (rank) test, Fischer's Exact test
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- One female in group 4 died during the blood sampling procedure. This was considered an accident. No further deaths occured.
- Mortality:
- mortality observed, treatment-related
- Description (incidence):
- One female in group 4 died during the blood sampling procedure. This was considered an accident. No further deaths occured.
- Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- No effects
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- Food consumption was slightly higher in the male group 4. This was incidental and of no toxicological relevance.
- Food efficiency:
- not examined
- Description (incidence and severity):
- Test substance was administered by oral gavage
- Water consumption and compound intake (if drinking water study):
- not examined
- Description (incidence and severity):
- Test substance was administered by oral gavage
- Ophthalmological findings:
- not examined
- Description (incidence and severity):
- Not required in range-finding examination
- Haematological findings:
- no effects observed
- Description (incidence and severity):
- No effects
- Clinical biochemistry findings:
- no effects observed
- Description (incidence and severity):
- No effects
- Urinalysis findings:
- not examined
- Description (incidence and severity):
- Not required
- Behaviour (functional findings):
- effects observed, treatment-related
- Description (incidence and severity):
- Locomotor activity was statistically higher in group 3 and 4 males than in the respective control. This was attributable to normal biological variation.
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Description (incidence and severity):
- During necropsy of parent animals no treatment-related findings were noted. For males treated at 300 or 1000 mg/kg/day, mean absolute and relative liver weights were dose-dependently increased.
- Gross pathological findings:
- no effects observed
- Description (incidence and severity):
- No effects
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Please see below
- Histopathological findings: neoplastic:
- not examined
- Description (incidence and severity):
- Not required
- Details on results:
- CLINICAL SIGNS AND MORTALITY
No mortalities were observed. No treatment-related effects were seen. Neither food consumption nor body weight development were affected at any dose level.
BODY WEIGHT AND WEIGHT GAIN
No effects seen in males or females at any dose level
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Food consumption was slightly higher in the male group 4. This was incidental and of no toxicological relevance.
HAEMATOLOGY
No effects seen
CLINICAL CHEMISTRY
No effects seen
NEUROBEHAVIOUR
Locomotor activity was statistically higher in group 3 and 4 males than in the respective control. This was attributable to normal biological variation.
ORGAN WEIGHTS
During necropsy of parent animals no test item-treated findings were noted. For males treated at 300 or 1000 mg/kg/day, mean absolute and relative liver weights were dose-dependently increased. See table 3 for further information.
GROSS PATHOLOGY
The intergroup incidence of various spontaneous changes gave no indication of an effect of treatment. No treatment related macroscopic abnormalities were noted.
HISTOPATHOLOGY:
Liver- Minimal hepatocellular hypertrophy in animals treated at 1000 mg/kg bw/day. This change was considered to represent an adaptive reaction most likely induced by an increased biotransformation of the test item. Therefore, it was not assessed as an adverse effect.
Thyroid- Increased incidence of minimal follicular cell hypertrophy in animals treated at 1000 mg/kg bw/day. No test item- related histopathological findings were noted in the reproductive organs of either sex. In particular, the assessment of the integrity of the spermatogenetic cycle did not reveal any evidence of impaired spermatogenesis.
REPRODUCTION DATA
Fertility rate was high resulting in at least 9 litters per group for evaluation of reproduction data. There were no treatment-related effects on precoital time, fertility indices, mean duration of gestation, number of implantations, post-implantation loss, pup survival or litter size from birth through to scheduled pup sacrifice on Day 4 post partum at any dosage.
Litter Data:
No test item-related abnormal findings were noted for pups at first litter check or during the first 4 days post partum. Sex ratios at first litter check and on day 4 post partum were unaffected by treatment with the test item.
Mean pup weights on day 0 and day 1 post partum were unaffected by treatment with the test item. Mean pup weight development during the first 4 days post partum was unaffected by treatment with the test item.
F1 Pups Necropsy findings:
No test item-related macroscopic findings were noted during necropsy of F1 pups.
Please see table 2 for F0 animal breeding statistics. - Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Sex:
- male/female
- Remarks on result:
- not determinable due to adverse toxic effects at highest dose / concentration tested
- Dose descriptor:
- NOEL
- Effect level:
- 100 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: Higher doses resulted in increased liver weight
- Key result
- Critical effects observed:
- no
- Conclusions:
- In order to generate preliminary information on the possible health hazards likely to arise from repeated exposure over a relatively limited period of time, Fatty acids, tall-oil, epoxidized,2-ethylhexylesters (ETP) was administered orally by gavage at dosages of 0 (corn oil), 100, 300 or 1000 mg/kg body weight/day to male rats for 28 days and to female rats throughout the pre-pairing, pairing and gestation periods until the individual day 4 post partum.
Administration of 1000 mg/kg/day resulted in increased liver weights (males and females) and minimal hepatocellular hypertrophy (males and females), that were considered to represent an adaptive change most likely induced by an increased biotransformation of the test item. Therefore, it was considered not to be adverse.
Administration of 300 mg/kg/day resulted in increased liver weights (males and females). In the absence of any histopathological correlate, these increased weights were considered not to be adverse.
Based in these data, the NOAEL was considered to be 1000 mg/kg bodyweight/day.
In the absence of any adverse effects on reproductive parameters, the NOAEL and NOEL for reproduction/developmental toxicity were considered to be 1000 mg/kg bodyweight/day. - Executive summary:
The purpose of this study was to generate preliminary information concerning the effects of Fatty acids, tall-oil, epoxidized,2-ethylhexylesters (ETP) on the possible health hazards likely to arise from repeated exposure over a relatively limited period of time. In addition, it provided information on possible effects on male and female reproductive performance such as gonadal function, mating behaviour, conception, development of the conceptus and parturition.
ETP was administered once daily orally (by gavage) to males for at least 28 days and to female rats throughout the prepairing and pairing periods until day 3 of lactation.
The following dose levels were applied:
Group 1: 0 mg/kg body weight/day (vehicle control)
Group 2: 100 mg/kg body weight/day
Group 3: 300 mg/kg body weight/day
Group 4: 1000 mg/kg body weight/day
A standard dose volume of 2 mL/kg body weight with a daily adjustment to the actual body weight was used. Control animals were dosed with the vehicle alone (corn oil).
The following results were obtained:
Parent animals:
General Tolerability
No mortalities were observed. No test-item related effects were seen. Neither food consumption nor body weight development were affected at any dosage.
Reproduction Data:
Fertility rate was high resulting in at least 9 litters per group for evaluation of reproduction data. There were no treatment-related effects on precoital time, fertility indices, mean duration of gestation, number of implantations, post-implantation loss, pup survival or litter size from birth through to scheduled pup sacrifice on Day 4 post partum at any dosage.
Clinical Laboratory Investigations:
The assessment of clinical chemistry and haematology parameters indicated no differences between animals treated with the test item and vehicle controls.
Terminal Examinations:
During necropsy of parent animals no test item-treated findings were noted. For males treated at 300 and 1000 mg/kg/day, mean absolute and relative liver weights were dose-dependently increased.
Histopathology:
The following findings distinguished test item-treated animals from vehicle controls:
Liver- Minimal hepatocellular hypertrophy in animals treated at 1000 mg/kg/day. This change was considered to represent an adaptive reaction most likely induced by an increased biotransformation of the test item. Therefore, it was not assessed as an adverse effect.
Thyroid- Increased incidence of minimal follicular cell hypertrophy in animals treated at 1000 mg/kg/day. No test item- related histopathological findings were noted in the reproductive organs of either sex. In particular, the assessment of the integrity of the spermatogenetic cycle did not reveal any evidence of impaired spermatogenesis.
Litter Data:
No test item-related abnormal findings were noted for pups at first litter check or during the first 4 days post partum. Sex ratios at first litter check and on day 4 post partum were unaffected by treatment with the test item.
Mean pup weights on day 0 and day 1 post partum were unaffected by treatment with the test item. Mean pup weight development during the first 4 days post partum was unaffected by treatment with the test item.
F1 Pups Necropsy findings:
No test item-related macroscopic findings were noted during necropsy of F1 pups.
- Endpoint:
- short-term repeated dose toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 28 June 1973 to 6 November 1973
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: The study was conducted and reported by BIBRA - while the in life phase pre-dates study guideline adoption and GLP, the report prepared in 1991 has been QA reviewed and is considered reliable. .
- Justification for type of information:
- Read across to a study result from an investigation using a similar material is justified for members of the Epoxidised Oils and Derivatives group. Four epoxidised oils (linseed, soybean, 2-ethylhexyl tallate and fatty acids, C14-C22, 2-ethylhexylesters) have been identified as sharing common structural and functional similarities, recognised in an OECD SIDS review as a single category, and therefore justifying read-across between data for different members of the group. Consequently data sharing between ESBO epoxidised soybean oil and epoxidised linseed oil is commonly utilised in the preparation of this dossier and other read-across bridges are used for other members of the EOD group where appropriate.
- Reason / purpose for cross-reference:
- reference to other study
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
- Deviations:
- yes
- Remarks:
- Conducted prior to guideline adoption. 19-20 week dosing period with two interim terminations after 2 or 6 weeks
- Principles of method if other than guideline:
- No specific guidelines are quoted. The study was completed to provide some toxcity data for a material used as a plasticiser and also with food contact applications. The study as completed was broadly in accordance with the requirements of OECD Guideline 408.
Groups of 15 males and 15 female rats were given dietary concentrations of 0, 0.05, 0.5 or 5.0% ELO. A negative control roup was administered linseed oil at 5.0% (non-epoxidised)
Interim groups were terminated after 2 or 6 weeks treatment at 0, 0.05, 0.5 or 5.0% ELO. Each interim group consisted of five males and five females. - GLP compliance:
- no
- Remarks:
- The study was undertaken before GLP implementation. However the report prepared by BIBRA was audited.
- Limit test:
- no
- Species:
- rat
- Strain:
- Wistar
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: A Tuck & Son Ltd, Rayleigh, Essex, UK
- Weight at study initiation: 40-60g
- Fasting period before study: rats were not fasted for dietary administration
- Housing: Five to a cage in grid bottom aluminium cages measuring approx. 45x36x16 cm. These were suspended in racks carrying 24 cages above sheets of paper for collection of excreta. The paper was renewed daily.
- Diet : Spratt's Laboratory Animal Diet No. 2 ad libitum
- Water : ad libitum domestic potable water in cage bottles
- Acclimation period: 6 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): circa 20 °C
- Humidity (%): 45 - 70 %
- Air changes (per hr): no recirculation of filtered 0.5um air to give 15-20 changes of air per hour.
- Light dark cycle: 14 hours light/10 hours dark - Route of administration:
- oral: feed
- Vehicle:
- unchanged (no vehicle)
- Details on oral exposure:
- DIET PREPARATION
Spratt's Lab Diet No.1 was ground to a powder. ELO (0.05, 0.5, 5 %) or 0.5 or 5% LO were added by weight and thoroughly mixed in a commercial food mixer. The ground diet served as the control. Fresh batches were prepared weekly. Administration was by admixture with the diet which was given ad lib for up to 20 weeks.
No diet analyses were conducted.
Exposure levels were calculated to be:
Linseed oil
% mg/kg bw/day (males) mg/kg bw/day (females)
5.0 3879.7 4200.0
Epoxidised Linseed Oil
0.05 39.3 41.8
0.5 402.0 448.9
5.0 4292.2 4609.0 - Analytical verification of doses or concentrations:
- no
- Details on analytical verification of doses or concentrations:
- No analyses were carried out to verify the concentration of ELO or LO.
- Duration of treatment / exposure:
- Administration was by admixture with the diet which was given ad libitum for up to 20 weeks. Interim sacrifice groups were dosed for 2 or 6 weeks, the main study males for 19 weeks and main study females for 20 weeks.
- Frequency of treatment:
- Test item was provided via diet so the diet was accessible all day. Fresh diet prepared at weekly intervals and new diet suppled daily to each cage.
- Remarks:
- Doses / Concentrations:
0 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
0.05 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
0.5 %
Basis:
nominal in diet - Remarks:
- Doses / Concentrations:
5 %
Basis:
nominal in diet - No. of animals per sex per dose:
- 15 per sex per group in main study. Five per sex/group in each of two interim termination sub groups
- Control animals:
- yes, plain diet
- Details on study design:
- - Dose selection rationale: The results of a structural analogue ESBO (see long term repeatdose study with ESBO also prepared by BIBRA)
- Rationale for animal assignment (if not random): random
- Rationale for selecting satellite groups: 2-week and 6 week interim sacrifice groups equate to sub acute exposure scenarios to obtain additional information from the study.
Very few investigations were available at the time the study was conducted. In vivo data were in short supply. In vitro results indicated ELO was of low toxicity to HeLa cells, there was no evidence of mutagenicity from an Ames test with several bacterial strains with or without metabolic activation. The remaining information came from the structuarla analogue ESBO. Consequently most ofthe remaining data in this dossier is obtained by read-across to ESBO. - Positive control:
- No details supplied
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
Rats were observed regularly but no records of conditional or behavioural changes were maintained.
BODY WEIGHT: Yes
These were recorded on the first day of treatment (day 0), twice per week for the first two weeks and then at weekly intervals to termination at week 6, 19 or 20.
FOOD CONSUMPTION AND COMPOUND INTAKE : Yes
Both food and water were measured over the 24 hour period prior to each bodyweight determination.
HAEMATOLOGY: Yes
Blood was taken from the dorsal aorta at termination (both interim sacrifices and final termination). The blood was examined for haemaglobin content, packed cell volume and erythrocyte and total and differential leucocyte counts plus reticulocyte counts.
URINALYSIS: Yes
Urine samples were collected from 12 rats of each sex from the main study during week 12-13 (males) or week 14-15 (females). Urine samples were collected from all rats in the interim sacrifice groups - in week 2 and 5.
The samples were obtained by placing the animals individually in metabolism cages without food or water. The conditions of the collection and examinations were:
- Collected over a 6 hr period following the normal overnight feeding and drinking and examined for volume, concentration (by refractive index), pH and the presence of glucose, blood, bile salts, ketones and proteins
- Collected over a 2 hour period immediately following an oral water load of 25 ml/kg and examined for volume, concentration and content of cells.
- Collected over a 4 hour period commencing 16 hour after an oral water load of 25 ml/kg. Food but not water was available to the animals during the 16 hour period. These samples were examined for volume and concentration. - Sacrifice and pathology:
- After 2, 6, 19 or 20 weeks all surviving rats were killed, subject to post-mortem examination with recording of organ weights and samples of tissues preserved. The tissues from all rats were processed for microscopic examination.
ORGAN WEIGHTS: Yes
adrenals, brain, caecum, gonads, heart, kidneys, liver, pituitary gland, small intestine, spleen, stomach, thyroid gland
GROSS PATHOLOGY: Yes
Rats surviving to the end of the study were weighed and the weights of brain, heart, liver, spleen, kidneys, stomach, small intestine, caecum (with and without contents), gonads, pituitary and thyroid recorded. When possible the following tissues, together with any other abnormal tissue, were preserved in 10 % buffered formalin: adrenal glands, aorta, bladder (urinary), brain, caecum, colon, eye, gonads, harderian gland, heart, kidneys, liver, lungs, lymph nodes, mammary gland, muscle (skeletal), nerve (sciatic), oesophagus, pancreas, pituitary gland, prostate, salivary glands, seminal besicles, skin, small intestine, spinal cord, spleen, stomach, thymus gland, thyroid gland, trachea, uterus.
HISTOPATHOLOGY: Yes
All tissues collected were processed for embedding in paraffin-wax, sectioned at approx. 5 um and stained with haematoxylin and eosin.
Additional liver sections were stained with Oil Red O as a marker of intracellular lipid deposition. - Other examinations:
- No details supplied
- Statistics:
- The results of the ELO and LO groups were initially compared statistically with the control group. Then the treated ELO groups were analysed independently with the control.
Comparisons were made with two sided least signifiant, t-test or sample t-test with Satterthwaite's correction or by Mann-Whitney U test. The tests were selected on degree of normality or homoscedasticity as assessed by Kolmogorov-Smirnov tests and Bartlett's test.
Statistical tests were applied to bodyweights, food and water consumption, haematology, urinalysis, organ weights and relative organ weights and histopathological findings. - Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- No unscheduled mortalities. No records maintained for other observations
- Mortality:
- mortality observed, treatment-related
- Description (incidence):
- No unscheduled mortalities. No records maintained for other observations
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- Bodyweight changes at the high dose level were not indicative of treatment related effects
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- On several occasion during treatment period food consumption by all ELO groups ws reduced in comparison with basal diet controls. Generally consumption was similar for ELO and LO groups.
- Food efficiency:
- not examined
- Water consumption and compound intake (if drinking water study):
- effects observed, treatment-related
- Description (incidence and severity):
- ELO treated rats tended to drink less than base diet controls but water consumption was similar for ELO and LO treated groups
- Ophthalmological findings:
- not examined
- Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- No toxicologically significant changes noted
- Clinical biochemistry findings:
- not examined
- Urinalysis findings:
- effects observed, treatment-related
- Description (incidence and severity):
- Minor changes oserved but with no adverse toxicological effects.
- Behaviour (functional findings):
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Description (incidence and severity):
- Absolute and relative liver and kidney weights tended to increase in high dose group
- Gross pathological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- No treatment related macroscopic observations
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Increased hepatocyte vacuolation, attributed to high lipid content of diet containg 5% ELO
- Histopathological findings: neoplastic:
- not examined
- Details on results:
- CLINICAL SIGNS AND MORTALITY
There were no mortalities for any of the ELO, LO or control groups in the main study. The interim sacrifice groups, killed after 2 or 6 weeks of treatment also had no unscheduled deaths.
No record of clinical observations was retained in the study data and the report gives no details of any clinical signs of reaction to treatment.
BODY WEIGHT AND WEIGHT GAIN
Minor fluctuations in bodyweight were noted for males and females in each of the treatment groups in comparison with basal diet controls but no notable differences were apparent when weights were compared with linseed oil treated controls. Over the course of the 20 weeks of the main study, bodyweights were significantly lower during the fist half of the study for males and females dosed at 5% ELO . In the second half of the study the weights were slightly increased such that by termination there were no significant differences between ELO or LO treated rats and the diet controls.
ELO had no notable adverse effect on bodyweight or weight gain
FOOD CONSUMPTION AND COMPOUND INTAKE
Food and water consumption showed minor variations at various points of recording. Generallya slight reduction in water consumption was commonly recorded forthe ELO treated groups but no clear dose related effects were apparent. Food and water consumption were compared, for ELO groups, with both LO and basal diet controls and the slight significant differences showed no consistent trends to indicate any adverse effects.
HAEMATOLOGY
There were few statistically significant differences between ELO treated and control groups for any of the haematology parameters.
Blood samples were collected after 2, 6 or 19/20 weeks.
After two weeks, reticulocyte counts were reduced for the high dose males and females and intermediate group males. No differences between ELO and LO groups. (There was only one male control value for reticulocytes).
After 6 weeks, the high dose males had a significantly increased lymphocyte count, but a similar increase occurred for LO treated rats.
At study conclusion there were no significant differences for any parametrs for the treated females. The high dose males had a significantly lower eosinophil count.
None of the haematology parameters showed a consistent pattern indicative of a treatment or dose related effect.
URINALYSIS
Renal dilution tests were conducted after 2, 6 or 19 weeks of treatment. The significant results in the high doe males were not replicated among the females and my ahave been a consequence of the larger urine volumes poduced by the males (despite generally lower water consumption values).
Semi-quantitative analyses of the 6-h urine samples showed no toxicologicallysignificant changes for ELO or LO treated rats.
ORGAN WEIGHTS
2-week interim sacrifice
The absolute and relative liver weights were significantly increased for males dosed at 5% ELO (and for the 5% LO treated rats).
The absolute liver weights for the 5% ELO treated females were slightly but not significantly increased, while the relative weight increase was significant.
The relative kidney weights for the high dose group males were increased in comparison with controls and 5% LO treated rats.
Absolute spleen weights were reduced for the high dose males and females. Pituitary weights in the high dose group were also slightly reduced.
Empty caecum weights relative to bodyweight were increased for the high dose females.
Apart from the liver and kidney changes in the high dose group, none of these changes were considered toxicologically notable.
6 week interim sacrifice
Absolute stomach weights were slightly lower for the high dose males. The absolute and relative stomach weights were also significantly lower for males treated at 0.5% ELO. No effects were noted for females.
Relative liver weights were significantly increased for males and females dosed at 5% ELO.
Absolute pituitary gland weights were significantly lower for the high dose males. but relative weights were unaffected.
Relative kidney weights for male and female rats treated at 5% ELO were significantly increased in comparison with controls.
Minor changes were noted for caecum, stomach, adrenal weights in some groups but no dose-relationship.
Terminal sacrifice
The absolute and relative liver and kidney weights for males and females treated at 5% were statistically significantly greater than the controls and the 5% LO treated rats. The percentage increase was 16-30% for both sexes.
Other changes were noted - similar to the 2 and 6 week termination groups - affecting stomach, caecum, pituitary gland, showing slight or significant differences from controls but showing no consistent trends in relation to dose or treatment.
GROSS PATHOLOGY
No treatment-related changes were observed during necropsy.
HISTOPATHOLOGY: NEOPLASTIC
Thickening of the alveolar wall was evident in ELO-treated male and female rats dosed at 5%. No macroscopic changes in the lungs were noted.
Vacuolation of hepatocytes was commonly observed in both control and treated rats. The degree of vacuolation was assigned from examination of Oil Red O stained sections. The degree of vacuolation was greater for the high dose males and females and associated with the presence of lipid.
A higher incidence of tracheal mucosa inflammatory cell infiltrate was apparent for the low dose females - 0.05% - Dose descriptor:
- NOEL
- Effect level:
- 0.5 other: %
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: Please see results
- Dose descriptor:
- NOAEL
- Effect level:
- 5 other: %
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: Minor changes were noted at the highest dose level - 5% ELO in diet, equivalent to more than 4200 mg/kg bw/day. None of the effects were considered toxicologically significant
- Critical effects observed:
- not specified
- Conclusions:
- 5% ELO in diet administered to rats for up to 20 weeks, resulted in no adverse toxicologically significant effects. A number of slight variaions occurred involving liver and kidney weights, reduced water consumption and hepatocyte vacuolation which were attributed to treatment with ELO. Consequently the NOAEL is considered to be 5% or greater than 4200 mg/kg bw/day but the absolute n effect level was set at the next lower dose level - 0.5% ELO or 402 mg/kg bw/day.
Comparison of ELO with the structual analogue epoxidised soybean oil suggests that the NOAEL of 5% is a good representation of the low toxicity and innocuous nature of the epoxidised oil group. - Executive summary:
Groups of 15 male and 15 female rats of a Wister strain were fed diet containing 0.05, 0.5 or 5 % Epoxidised Linseed Oil (ELO) for 19 -20 weeks. Similar groups were given basal diet to serve as a control or 0.5 or 5.0% Linseed Oil (LO). In addition two interim sacrifice points were scheduled after 2 or 6 weeks of treatment (five male and five female rats were dosed at 0.05, 0.5 or 5% ELO for each sacrifice point).
Bodyweights were recorded at regular ntervals through the study. Food and water intakes were also measured regularly. Terminal blood samples were collected from the dorsal aorta, after 2, 6 or 19/20 weeks. Standard haematology and urinalysis parameters were measured. Various organs were weighed at termination and preserved tissues were examined microscopically.
Initially the bodyweights for the high dose ELO group were reduced in comparison with linseed oil and untreated diet controls, but weight gains were similar for all groups by the end of the treatment period.
Food and water consumption were similar for all treated and control groups.
There were no changes in any of the haematology or urinalysis parameters indicative of treatment-related toxicity.
Increased relative liver weights were evident in the males and females dosed at 5% ELO, in the two interim sacrifices and main study sacrifice. Relative kidney weights in this dose group were also increased for both sexes.
There were no histopathological changes to account for the renal weight increase. The high dose livers showed a greater degree of vacuolated lipid filled hepatocytes. These fatty changes were considered to account for the liver weight increase noted macroscopically but were considered attributable to a higher lipid content of the diet (5% ELO) rather than to any toxic effect of Epoxidised Linseed Oil.
None of the effects observed at 5% ELO were considered to be toxicologically significant. The no observed effect level or NOEL was set at the next lower dose (0.5% ELO) equivalent to 402 mg/kg bw/day. The NOAEL was 5% ELO or 4292 mg/kg bw/day.
For comparison, it was concluded that ESBO (epoxidised soybean oil) following a two year dietary administration, was not carcinogenic when fed to rats at up to 2.5 % of the diet. Further it was concluded that the NOEL was 2.5 % providing an average daily intake of approximately 1000 mg/kg in the males and 1400 mg/kg in the females.
Much of the data generated for ESBO is used via read-across for ELO. The results of these two sub-chronic or chronic exposure studies confirm that ELO is unlikely to prove more toxic than ESBO and so the data is conservative for read-across for epoxidised linseed oil.
Referenceopen allclose all
No further information
Table 1 Averages of Ratios of Organ to Body Weight of Rats Receiving Paraplex G-60 in their Diet for One Year
Sex |
Dietary Conc. (%) |
Number of Rats |
Organ to Body Weight Ratios x 10-3 |
||
Liver |
Kidney |
Testes |
|||
Female |
0 |
5 |
36.8± 6.4 |
7.01± 1.44 |
- |
|
0.1 |
5 |
35.9± 25.5 |
6.99± 3.36 |
- |
|
0.5 |
4 |
34.3± 9.9 |
7.03± 2.13 |
- |
|
1.0 |
4 |
38.8± 6.0 |
7.14± 3.09 |
- |
|
2.5 |
5 |
36.6± 5.0 |
7.99± 1.92 |
- |
|
5.0 |
4 |
34.4± 10.8 |
7.06± 3.21 |
- |
Male |
0 |
5 |
36.1± 10.3 |
6.58± 1.67 |
6.35± 2.14 |
|
0.1 |
5 |
33.3± 10.0 |
6.05± 0.36 |
6.06± 1.53 |
|
0.5 |
5 |
36.0± 13.0 |
6.94± 2.11 |
6.56± 2.61 |
|
1.0 |
5 |
39.1± 16.4 |
6.62± 0.61 |
6.17± 1.03 |
|
2.5 |
5 |
39.6± 7 .2 |
7.00± 2.03 |
6.37± 0.83 |
|
5.0 |
4 |
42.4± 11.5 |
7.66± 3.82 |
6.21± 1.46 |
Table 2 Histopathologic Findings in Rats Receiving Paraplex G-60 in Their Diet for Two Years
Sex |
Dietary Conc. (%) |
Number of Rats |
Histopathologic Findings |
Female |
0 |
6 |
Lymphosarcoma and leucsarcoma in 1 rat. Severe bronchiectasis in 1 rat |
|
2.5 |
8 |
Tubular vacuolic nephrosis in 1 rat. Sclerosing adenosis or sweat gland hyperplasia in the breast of 1 rat. Cavernous hemangioma of the adrenal of 1 rat. |
|
5.0 |
9 |
Myositis chronic of striated muscle in 1 rat. Pneumonia in 1 rat. Reticulum cell sarcoma of the lungs and atropy of adrenal zona glomerulosa in 1 rat. |
Male |
0 |
5 |
Pycnosis of nuclei in loops of Henle and pulmonary edema and congestion in 1 rat. Granulocytic hyperplasia of bone marrow in 1 rat. Lung abscesses and protein casts in tubules of 1 rat. |
|
2.5 |
6 |
Hepatocarcinoma of liver, chronic glomerulonephiritis and bronchiolar adenomatosis of lungs in 1 rat. Moderate fatty infiltration in liver of 1 rat. |
|
5.0 |
3 |
Atrophy of adrenal zona reticularis in 1 rat. Protein casts in kidney tubules in 1 rat. |
Table 3 Body Weight Changes in Dogs Receiving Paraplex G-60 in Their Diet for One Year
Dietary Conc. (%) |
Dog No. |
Body Weight (kg) |
|||||
Start |
2 Wk |
5 Wk |
10 Wk |
26 Wk |
52 Wk |
||
0.1 |
1 |
11.5 |
12.9 |
13.3 |
13.6 |
14.5 |
13.5 |
|
2 |
12.4 |
13.6 |
14.6 |
15.5 |
15.7 |
15.5 |
|
3 |
16.8 |
19.8 |
19.9 |
21.0 |
21.5 |
22.8 |
1.0 |
4 |
9.2 |
9.4 |
12.7 |
13.9 |
14.4 |
14.5 |
|
5 |
11.1 |
11.6 |
18.0 |
18.7 |
17.5 |
17.9 |
|
6 |
15.3 |
17.4 |
9.5 |
9.7 |
10.4 |
11.0 |
5.0 |
7 |
14.2 |
13.3 |
9.0 |
8.9 |
8.8 |
8.5 |
|
8 |
9.7 |
9.5 |
16.0 |
16.0 |
15.7 |
14.8 |
|
9 |
18.0 |
17.0 |
12.5 |
11.3 |
11.4 |
10.6 |
Table 4 Averages of Ratios of Organ to Body Weight of Rats Receiving Paraplex G-62 in Their Diet for Six Months
Sex |
Dietary Conc. (%) |
Number of Rats |
Organ to Body Weight Ratios x 10-3 |
||
Liver |
Kidney |
Testes |
|||
Female |
0 |
5 |
36.5± 9.2 |
7.59± 1.17 |
- |
|
0.1 |
5 |
35.7± 11.9 |
7.77± 1.53 |
- |
|
0.5 |
5 |
36.2± 8.6 |
8.10± 1.14 |
- |
|
1.0 |
5 |
44.6± 11.4 |
8.87± 1.39 |
- |
|
2.5 |
4 |
43.2± 5.0 |
9.09± 0.45 |
- |
|
5.0 |
5 |
42.5± 13.9 |
8.59± 2.44 |
- |
Male |
0 |
4 |
39.0± 15.9 |
8.25± 4.07 |
7.79± 2.48 |
|
0.1 |
5 |
43.1± 12.5 |
8.74± 4.69 |
7.45± 2.80 |
|
0.5 |
5 |
37.4± 6.9 |
7.22± 1.11 |
7.31± 1.33 |
|
1.0 |
4 |
43.4± 14.6 |
8.73± 3.05 |
6.98± 1.46 |
|
2.5 |
4 |
43.3± 24.5 |
8.73± 3.44 |
8.27± 1.15 |
|
5.0 |
4 |
50.8±15.0 |
9.64± 3.12 |
7.91± 3.66 |
Table 5 Histopathologic Findings in Rats Receiving Paraplex G-62 in Their Diet for Two-Years
Sex |
Dietary Conc. (%) |
Number of Rats |
Histopathologic Findings |
Female |
0 |
7 |
Bronchiectasis in 3 rats. Angiectasis in adrenals in 1 rat. Granulocytic hyperplasia of bone marrow angiectasis in adrenals and bronchiectasis in 1 rat. Tubular nephresis in kidneys, focal necrosis in liver, ovarian cyst, chronic myocarditis and angiectais and focal necrosis in adrenal gland in 1 rat. |
|
2.5 |
8 |
Giant fibroadenoma of the breast in 1 rat. Fatty infiltration of the liver and adrenal congestions in 1 rat. |
|
5.0 |
5 |
Papillary cystadenoma of the ovary and loss of lipoid in the adrenal cortex in 1 rat. Angiectasis in adrenals in 1 rat. Follicular cyst of ovary and congestion and angiectasis of adrenals in 1 rat. Angiectasis and focal degeneration of adrenals in 1 rat. |
Male |
0 |
3 |
Bronchiectasis in 2 rats. |
|
2.5 |
4 |
Mild tubular nephrosis of kidney and vacuolation of liver nuclei in 1 rat. Mild tubular nephrosis of kidney, atrophic testes and marked congestion of all organs in one rat. Acute focal bronchopneumonia in 1 rat. Focal fatty vacuolation of adrenal in 1 rat. |
|
5.0 |
6 |
Tubular and bile nephrosis and acute cystitis in 1 rat. Vascular hamartoma of liver, mesothelioma of epididymus and bronchiectasis and acute granulomas in 1 rat. Mild tubular nephrosis of kidney, generalised venous congestion and prominent pancreatic islets in 1 rat. Tubular nephrosis in 1 rat. |
Table 6 Body Weight Changes in Dogs Receiving Paraplex G-62 in Their Diet for One Year
Dietary Conc. (%) |
Dog No. |
Body Weight (kg) |
|||||
Start |
2 Wk |
5 Wk |
10 Wk |
26 Wk |
52 Wk |
||
0.1 |
1 |
10.7 |
11.5 |
11.4 |
12.8 |
15.3 |
15.5 |
|
2 |
9.8 |
10.2 |
10.6 |
11.6 |
13.8 |
15.4 |
|
3 |
20.3 |
21.6 |
21.4 |
20.6 |
22.2 |
23.6 |
1.0 |
4 |
14.9 |
16.2 |
16.2 |
16.5 |
18.4 |
17.1 |
|
5 |
7.4 |
8.4 |
8.2 |
8.5 |
8.7 |
7.4 |
|
6 |
19.1 |
20.8 |
20.0 |
20.5 |
20.4 |
21.3 |
5.0 |
7 |
8.6 |
8.0 |
7.9 |
7.1 |
7.6 |
7.7 |
|
8 |
10.9 |
9.7 |
9.3 |
8.7 |
8.5 |
8.7 |
|
9 |
12.9 |
11.9 |
11.3 |
9.6 |
9.3 |
9.1 |
Table 2: F0 animals breeding for F1 litters
Group (mg/kg/day) |
1 (0) |
2 (100) |
3 (300) |
4 (1000) |
# of females paired |
10 |
10 |
10 |
10 |
# of females mated |
10 |
10 |
10 |
10 |
# of pregnant females |
10 |
10 |
10 |
10 |
# of females delivering pups |
10 |
10 |
10 |
10 |
# of females with live pups on day 4 post partum |
10 |
9 (A) |
10 |
10 |
A = For female 52 only two dead pups were noted at first litter check
Table 3: Organ Weights (Gram)
|
Group 1 0 mg/kg |
Group 2 100 mg/kg |
Group 3 300 mg/kg |
Group 4 1000 mg/kg |
|
Liver Males |
Mean |
8.33 |
8.73 |
9.38* |
9.71* |
St. Dev. |
0.78 |
1.00 |
0.81 |
0.56 |
|
N |
10 |
10 |
10 |
10 |
|
|
|||||
Liver Females |
Mean |
8.02 |
9.03 |
8.40 |
9.43* |
St. Dev. |
0.81 |
1.42 |
1.46 |
0.98 |
|
N |
10 |
10 |
10 |
9 |
|
calculated intakes:
Exposure levels were calculated to be:
Linseed oil
% mg/kg bw/day (males) mg/kg bw/day (females)
5.0 3879.7 4200.0
Epoxidised Linseed Oil
0.05 39.3 41.8
0.5 402.0 448.9
5.0 4292.2 4609.0
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed
- Dose descriptor:
- NOAEL
- 1 000 mg/kg bw/day
- Study duration:
- subchronic
- Species:
- rat
Repeated dose toxicity: inhalation - systemic effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: inhalation - local effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: dermal - systemic effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Repeated dose toxicity: dermal - local effects
Endpoint conclusion
- Endpoint conclusion:
- no study available
Additional information
In parent animals:
No mortalities were observed. No test-item related effects were seen. Neither food consumption nor body weight development were affected at any dosage. There were no treatment-related effects on reproductive endpoints. The assessment of clinical chemistry and haematology parameters indicated no differences between animals treated with the test item and vehicle controls. During necropsy no test item-treated findings were noted. For males treated at 300 and 1000 mg/kg/day, mean absolute and relative liver weights were dose-dependently increased. At necropsy liver- minimal hepatocellular hypertrophy in animals treated at 1000 mg/kg/day. This change was considered to represent an adaptive reaction most likely induced by an increased biotransformation of the test item. Therefore, it was not assessed as an adverse effect. Thyroid- Increased incidence of minimal follicular cell hypertrophy in animals treated at 1000 mg/kg/day. No test item- related histopathological findings were noted in the reproductive organs of either sex. In particular, the assessment of the integrity of the spermatogenetic cycle did not reveal any evidence of impaired spermatogenesis.
Litter Data:
No test item-related abnormal findings were noted for pups at first litter check or during the first 4 days post partum. Sex ratios at first litter check and on day 4 post partum were unaffected by treatment with the test item.
Mean pup weights on day 0 and day 1 post partum were unaffected by treatment with the test item. Mean pup weight development during the first 4 days post partum was unaffected by treatment with the test item. No test item-related macroscopic findings were noted during necropsy of F1 pups.
Justification for classification or non-classification
The data indicate low toxicity for the substance following repeated oral dosing. According to Regulation (EC) No. 1272/2008, no classification is warranted for STOT-RE.
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