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Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1983-1984
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1984
Report date:
1984

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
adopted May 12, 1981
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Thiodiethylene bis[3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate]
EC Number:
255-392-8
EC Name:
Thiodiethylene bis[3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate]
Cas Number:
41484-35-9
Molecular formula:
C38H58O6S
IUPAC Name:
2-[(2-{[3-(3,5-di-tert-butyl-4-hydroxyphenyl)propanoyl]oxy}ethyl)sulfanyl]ethyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propanoate
Details on test material:
- Physical state: solid
- Analytical purity: > 99.5% (CIBA-GEIGY Ltd, Analyse Number: 5659, 25.10.1984)

Test animals

Species:
rat
Strain:
other: Tif: RAIf
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Animal Production Stein, CIBA-GEIGY Ltd.,4332 Stein, Switzerland
- Age at study initiation: 4 weeks
- Weight at study initiation: males: 96 - 101 g, females: 95 - 102 g
- Housing: 5 per cage
- Diet: standard diet Nafag No. 890 Tox, ad libitum
- Water: tap water ad libitum
- Acclimation period: 9 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 10
- Air changes (per hr): 16 - 20
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: July 16, 1983 To: November 4, 1983

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
DIET PREPARATION
- Mixing appropriate amounts with (Type of food): The test substance was weighed on a calibrated Mettler balance. The pulverised food was then homogeneously mixed with the appropriate concentrations of the compound and 30% water was added before pelleting to ensure the necessary pellet quality. The pellets were subsequently airdried.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Prior to the initiation of the study, pretest feed samples were analysed for concentration of the test material. The same was undertaken periodically with the food batches applied during the test. These analysis were carried out in the Central Analytical Laboratories of CIBA-GEIGY LTD., Basle/Switzerland.
Duration of treatment / exposure:
93 - 103 days
Frequency of treatment:
daily
Doses / concentrationsopen allclose all
Dose / conc.:
60 ppm
Remarks:
Corresponding to 4.4 mg/kg bw in males and 4.5 mg/kg bw in females.
Dose / conc.:
200 ppm
Remarks:
Corresponding to 12.5 mg/kg bw in males and 13 mg/kg bw in females.
Dose / conc.:
600 ppm
Remarks:
Corresponding to 39 mg/kg bw in males and 40 mg/kg bw in females.
Dose / conc.:
2 000 ppm
Remarks:
Corresponding to 138 mg/kg bw in males and 140 mg/kg bw in females.
No. of animals per sex per dose:
20
Control animals:
yes, plain diet

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily (a.m. and p.m. on working days)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly (midweek)

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: weekly during the first month, monthly thereafter

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: before (day -4) and towards the end (day 87) of the treatment period
- Dose groups that were examined: control animals and in treated animals of the highest dose level

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the end of the treatment period
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: No
- How many animals: all animals
- Parameters checked: Erythrocytes (RBC), Mean Corpuscular Volume (MCV), Hematocrit (PCV), Hemoglobin (Hb), Mean Corpuscular Hemoglobin (MCH), Thrombocytes (PLT) (Platelet Count), Leucocytes (WBC) Total Count, Leucocytes (WBC) Differential Count, Prothrombin Time (PT)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the end of the treatment period
- Animals fasted: Yes
- How many animals: all animals
- Parameters checked: Glucose, Blood Urea Nitrogen (BUN), Total Bilirubin, Creatinine, Total Protein, Albumin, Globulins, A/G Ratio, Sodium (Na+), Potassium (K+), Chloride (Cl-), Calcium (Ca++), Phosphorus Inorganic, Asp. Aminotransferase ASAT (GOT), Ala. Aminotransferase ALAT (GPT), Alkaline Phosphatase (AP), Gamma-Glutamyl Transpeptidase (GGT), Cholesterol

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

OTHER: Hearing test:
- Time schedule for examinations: before (day -4) and towards the end (day 87) of the treatment period
- Dose groups that were examined: control animals and in treated animals of the highest dose level
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
- Besides the weight of the exsanguinated body the following organs were weighed: brain, heart, liver, kidneys, adrenals, thymus, gonads, spleen, thyroid
- The following organs and tissues were preserved in neutral 10% formalin:
skin, mammary area, spleen, mesenteric lymph node, axillary lymph node, sternum with bone marrow, femur with joint, skeletal muscle, trachea, lung, heart, aorta, submandibular salivary gland, liver, pancreas, oesophagus, stomach, small intestine, large intestine, kidney, urinary bladder, prostate, seminal vesicle, testis, epididymis, uterus, ovary, pituitary gland, adrenal gland, thyroid with parathyroid gland, thymus, peripheral nerve, brain, spinal cord, eye with optic nerve, orbital gland, extraorbital lacrimal gland, organs and tissues showing macroscopical changes

HISTOPATHOLOGY: Yes
- After the fixation organ samples from each control and test rat were taken, embedded in paraplast, sectioned at 3-5 micron, stained with haematoxylin and eosin and subjected to microscopic examination.
Statistics:
For each time point and parameter a uni-variate statistical analysis was conducted. Due to the routine manner of the analysis system, parameter free methods were applied. Each treated group was compared to the control group in respect of dispersion and displacement. In addition a trend test was applied considering all groups.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
No clinical symptoms and no signs of local and/or systemic toxicity were observed.
Mortality:
no mortality observed
Description (incidence):
No death occurred during the 3 months test period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
The mean body weight gain of all treated male and female groups was similar to that of the respective control groups.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
The mean food consumption of all treated male and female groups was similar to that of the respective control groups. Specific food consumption in relation to body weight - usually referred to as food conversion ratio - in treated animals was similar to that of the respective control groups.
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
The mean water consumption of all treated male and female groups was similar to that of the respective control groups.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Ophthalmic inspections performed before (day -4) and towards the end (day 87) of the application period revealed no evidence of a reaction to the treatment.
Haematological findings:
no effects observed
Description (incidence and severity):
The findings in the hematological investigation were unremarkable and considered as incidental in nature and not related to the treatment with the test substance.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
The values of treated rats were unremarkable and comparable to those of the control group. Parameters achieving a level of statistical significance in their difference from control values were considered to have arisen fortuitously.
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Mean organ weights and ratios are presented in the attached summary tables. Both absolute and relative liver weight showed a dose dependent increase reaching the level of statistical significance in treated male groups 3, 4 and 5 (200, 600 and 2000 ppm) and in treated female group 5 (2000 ppm). Additional statistically significant differences in organ weights, as indicated by asterisks in the attached mean tables, between treated and control groups were noted. Since no systematic pattern emerged, except a trend to higher kidney and testis weights in males and thyroid weight in males and females at higher dosages - which corresponds to the higher weight of exsanguinated body in these groups - these differences were attributed to spontaneous variation rather than to the treatment.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No abnormal substance related findings were observed.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Minimal hypertrophy of hepatocytes was present in the centrilobular region of the liver in all treated males of group 5 (2000 ppm), in 6 males of group 4 (600 ppm) and in 3 females of group 5 (2000 ppm). Only 19 liver samples were examined in female group 5 (liver of animal no 197 was missing). All other findings in control and treated animals are considered to be incidental in nature.
Histopathological findings: neoplastic:
not examined
Description (incidence and severity):
Hearing test: Hearing tests performed before (day -4) and towards the end (day 87) of the application period revealed no treatment related effects on the auditory perception.

Effect levels

open allclose all
Dose descriptor:
NOEL
Effect level:
60 ppm
Sex:
male/female
Basis for effect level:
organ weights and organ / body weight ratios
Dose descriptor:
NOAEL
Effect level:
>= 140 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
female
Basis for effect level:
other: highest dose tested
Dose descriptor:
NOAEL
Effect level:
>= 138 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
other: highest dose tested

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Executive summary:

In a 90 day repeated dose toxicity study by the oral route following OECD guideline 408, a total of 40 RAIf rats (20 each sex) were fed with the test material at dosages of 60, 200, 600 and 2000 ppm for three months. According to analytical results the calculated mean daily intake was 4.4-138 mg/kg bw (male) and 4.5-140 mg/kg bw (female). No death occurred. The body weight gain, the food and water consumption of males and females were similar to the control. No clinical symptoms and no signs of local and/or systemic toxicity were observed and no effects in eye and hearing tests were noted. Haematology and blood chemistry showed no effect associated to the test substance. The organ weight analysis showed a dose-dependent increase of the liver weight at and above 200 ppm in males and at 2000 ppm in females. The only histopathological finding was a minimal hypertrophy in the centrilobular region of the liver in males at and above 600 ppm and females at 2000 ppm. It can be inferred from the observations made during the above study that a "no observable effect level" (NOEL) for the test material when offered to rats continuously in their feed over a period of 3 months is 60 ppm, corresponding to a mean daily intake of 4.4 mg/kg body weight for males and 4.5 mg/kg bw for females. Because the observed changes were considered adaptive in nature, the NOAEL was set 138 mg/kg body weight (highest dose tested)