Octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline without GLP, acceptable with restrictions

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1981

Report date:

1981

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1982

Report date:

1982

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 409 (Repeated Dose 90-Day Oral Toxicity Study in Non-Rodents)

Deviations:

yes

Remarks:

6 animals per dose group instead of 8; homogeneity of the test substance formulation was not determined, organ weights were limitedly determined. 28-day recovery period included

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate

EC Number:

218-216-0

EC Name:

Octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate

Cas Number:

2082-79-3

Molecular formula:

C35H62O3

IUPAC Name:

octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate

Details on test material:

- Physical state: white powder

Test animals

Species:

dog

Strain:

Beagle

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: WIGA, Sulzfeld/Germany
- Age at study initiation: 26-35 weeks
- Weight at study initiation: males 8.6-12.6 Kgs; females 7.6-12.2 kgs
- Housing: 2 per each kennel
- Diet : Pelleted standard diet (Nafag, No. 941) 350 g/day/animal
- Water (e.g. ad libitum): tap water

ENVIRONMENTAL CONDITIONS
- Temperature (°C): floor temperature: approx. 23; room temperature: 20±3
- Photoperiod (hrs dark / hrs light): 10/14

Administration / exposure

Route of administration:

other: oral diet (pellets)

Vehicle:

polyethylene glycol

Details on oral exposure:

DIET PREPARATION
- The test substance was weighed and diluted with PAG 400 into an Erlenmeyer flask on a Mettler balance (Type PC 4400). The pulverized dog food was then homogenously mixed with the appropriate concentrations of the compound and about 10 % of water was added before pelleting to ensure the necessary pellet quality. The pellets were subsequently air-dried.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- Method: liquid chromatography with UV-detection at 280 nm
- Sampling times: pre-test, week 4 and 9

Duration of treatment / exposure:

91 days

Frequency of treatment:

once (350 g)/day, 7 days a week

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
1000, 3000 and 10000 ppm
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
31.5, 92.2 and 295.4 mg/kg bw/day in males and 34.5, 97.2 and 335.7 mg/kg bw/day in females
Basis:
actual ingested

No. of animals per sex per dose:

6 (five plus 1 for recovery group)

Control animals:

yes, concurrent no treatment

Details on study design:

- Post-exposure recovery period in satellite groups: 28 days (1 animal per sex and dose group)

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: mortality, signs of local and systemic toxicity

BODY WEIGHT: Yes
- Time schedule for examinations: daily

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 : Yes

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: Yes

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: pretest, at week 13, and 17 (recovery period, test week 4)
- Dose groups that were examined: all

HAEMATOLOGY: Yes
- Time schedule for collection of blood: week 1, 4, 8 and 12, and at recovery period test week 1
- Anaesthetic used for blood collection: No
- Animals fasted: No
- How many animals: all
- Parameters checked: Haemoglobin (Hb), Erythrocytes (RBC), Haematocrit (PCV), Mean Corpuscular Volume (MCV), Mean Corpuscular Haemoglobin (MCH), Reticulocytes, Thrombocytes, Prothrombin Time, Partial Thromboplastin Time, Thrombin Time, Leucocytes (WBC), Differential Count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: week 1, 4, 8 and 12, and at recovery period test week 1
- Animals fasted: No
- How many animals: all
- Parameters checked: Glucose, Urea (urea-N), Total Bilirubin, Total protein, Protein Electrophoresis, Asp. Aminotransferase (GOT), Ala. Aminotransferase (GPT), Lactate Dehydrogenase (LDH), Alkaline Phosphatase (AP), G-Glutamyl transpeptidose (G-GTP), Cholesterol, Creatine Kinase (CK), Creatinine, Elektrolytes (Na, K, Cl, Ca, P)

URINALYSIS: Yes
- Time schedule for collection of urine: week 1, 4, 8 and 12, and at recovery period test week 1
- Metabolism cages used for collection of urine: No, obtained by catheterization
- Animals fasted: No
- Parameters checked: color, specific gravity, pH, protein, glucose, ketone, blood, bilirubin, urobilinogen, urine sediment

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: monthly
- Dose groups that were examined: all
- Battery of functions tested: sensory activity (simple noise test: calling the dog)

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

For both, the following organs were examined: Brain, spinal cord, eye, pituitary, salivary gland, heart, thymus, thyroid, lungs, trachea, spleen, bone (with marrow), lymph nodes, aorta, urinary bladder, oesophagus, stomach, small and large intestine, adrenal glands, pancreas, liver, kidneys, ovaries/testes, prostate/uterus, skin, skeletal muscle, sciatic nerve, gall bladder and mammary gland.

Other examinations:

The following organs were weighed: heart, liver, kidneys, adrenals, thyroid, gonads, brain and pituitary.

Statistics:

For each time point and parameter a uni-variate statistical analysis was conducted. Each treated group was compared to the control group in respect of dispersion and displacement (Y. Lepage, Biometrika, 58: pp. 213-217, 1971). In addition a trend test was applied considering all groups (H.R. Jonckheere, Biometrika, 41: pp. 133-145, 1954).

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY:
No animal died during the experiment.
A slight to moderate diarrhea was observed practically during the whole administration period in the control and all treated groups. Towards the end of the administration period this finding was less frequent and during the recovery period with very few exceptions no diarrhea was observed.
While sampling urine during week 4 a modulated vagina was observed in dogs 38, 42 and 45. This finding was no more observed at later stages.

BODY WEIGHT AND WEIGHT GAIN:
The body-weight gain was similar in all animals.

FOOD CONSUMPTION:
The food consumption of all animals was comparable.

FOOD EFFICIENCY:
The mean food conversion was similar in all animals.

OPHTHALMOSCOPIC EXAMINATION:
No findings related to the compound administration were recorded.

HAEMATOLOGY:
The values were generally unremarkable and comparable to those of the controls.
Practically in all counts the values of the eosinophil granulocytes were at the upper normal limit or slightly higher. A subclinical parasitic infection (Strongyloides spp.) in the population was suspected, but a coprologic survey during week 16 gave only negative results. During week 15 two dogs were given 12.5 mg/dog SYNACTHEN R , a synthetic ACTH preparation. 7 hours later the number of eosinophils was below of 50 % of the pretest level, indicating that the function of the adrenal cortex was intact.

CLINICAL CHEMISTRY:
The values were generally unremarkable and comparable to those of the controls.
An elevation of bilirubin values with a positive trend from the control to the highest dosage group was seen in all treated groups at weeks 4, 9 and 13. The range of individual values was however wide, particularly in the females of the 10000 ppm group. Elevated bilirubin levels as observed in the treatment groups are usually correlated with:
- jaundice
- increase in the activity of alkaline phosphatase
- increase of urinary excretion of bile pigments
- histopathologic changes in the liver
Since none of these conditions was observed, the elevated bilirubin concentration in the treatment groups is considered as artifactions and without toxicological relevance.

URINALYSIS:
The values were generally unremarkable and comparable to those of the controls.

NEUROBEHAVIOUR:
No impairment of the auditory perception was found.

ORGAN WEIGHTS:
Organ weights and ratios for the compound treated dogs were comparable to those of the control animals with the exception of a slightly increased incidence of higher liver weights and ratios in the dogs of the 3000 and 10000 ppm groups. There was +20% and +60% increase in the ratios (liver/body) in males, and +19% and +21% increase in the ratios in females of the 3000 and 10000 ppm groups at the end of treatment period, respectively (p<0.05). After the end of recovery period, there was slight decrease in the 3000 ppm male and 45% increase in the ratios of 10000 ppm male. In females, there was +26% and 19% increase in the ratios of 3000 and 1000 ppm groups, respectively. Only one animal per dose group was used in the recovery group.

GROSS PATHOLOGY:
No compound related gross anatomical changes were noted.

HISTOPATHOLOGY: NON-NEOPLASTIC:
In two treated dogs (32 f from the 1000 ppm group and 13 m from the 3000 ppm group) and in one control animal (28 f) minute green brown pigmented droplets were found in very occasional biliary canaliculi of the liver. However no correlation between elevated or transitionally elevated concentration of serum bilirubin levels noted in a few treated dogs at biochemical examination, and histopathological findings in the liver could be established. All other changes seen in some control and treated dogs described within the microscopical findings in individual dogs were only incidental in nature mostly due to parasitic infestation. They were not related to the treatment.

Effect levels

Dose descriptor:

NOAEL

Effect level:

ca. 34 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: non-reversible increase in liver weight at higher dose level

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The test substance was administered to dogs in a daily diet over a period of 3 month. The strong increase in liver weight and liver / body weight ratio at the dogs of the 3000 and 10000 ppm group after treatment period and non-reversible effects after recovery period can be considered as an adverse effect. Thus, the NOAEL is considered to be 1000 ppm (34 mg/kg bw).

Executive summary:

The test article was administered to dogs in a daily diet over a period of 3 month. None of the animals died, gross pathology and histopathology were without findings at any dose levels. In-life observation revealed no signs of systemic effects but a significant increase in liver weight was observed. The effect was reduced during post-observation. Since the increase in liver weight at the 3000 ppm group (female) was at the end of the recovery period 26 %, the NOAEL for the test compound is considered to be 1000 ppm (34 mg/kg bw).