Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1997
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report Date:
1997

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
GLP compliance:
yes (incl. certificate)
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Name of the test substance: RA 3643
Appearance: white powder

Batch no. : 51/95
Date of receipt: 1995-12-21
Storage: at room temperature, dark
Used for: validation of the analytical method for the determination of RA 3643 in the diet

Batch no. : 74
Date of receipt: 1996-02-01
Storage: at room temperature, dark
Used for: 14 day range-finding study and determination of the stability of RA 3643 in the diet

Batch no. : 87/96
Date of receipt: 1996-04-15
Storage: initially at room temperature in the dark, from 1996-04-23 in a refirgerator (2-10°C)
Used for: sub chronic toxicity study

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
Male (46) and female (46) Wistar rats were obtained from a colony maintained under SPF-conditions at Charles River Wiga GmbH, Sulzfeld Germany. They were about 4 weeks old.
The body weights on the experimental start date ranged from 157.4 g to 189.1 g for males, and from 118.2 g to 145.2 g for females.
Housing conditions were conventional. The number of air changes was about 10 per hour, and lighting was artificial by fluorescent tubes, time switch controlled at a sequence of 12 h light (7.30-19.30), 12 h dark. The temperature and relative humidity in the experimental room, monitored continuously by means of a thermohygrograph, were 22-24°C and 50-70%, respectively.
Occasionally, the values were out of these ranges. The temperature exceeded the upper value twice.
The humidity was lower than 50% for two periods of a few hours and higher than 70% for three such short periods. In addition, the humidity exceeded 70% for about ten longer periods.
From the start of the treatment period, the animals were housed in groups of five, separated by sex, in suspended stainless steel cages with wire mesh floor and front. During the acclimatization period they were housed in similar cages in groups of five or six.
Feed (in powder) and water (tap water) were provided ad libitum from the arrival of the rats until the end of the study.

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
The oral route was used because this route is recommended in the guidelines for testing of cosmetic ingredients for their safety evaluation. The TNO rodent diet was used as the carrier. The test substance was incorporated into this diet by mixing in a mechanical blender. Because the test substance contained particles of different sizes, the substance was ground in an electric coffee mill prior to diet mixing. Fresh batches of experimental diets were prepared at 2-3 week intervals and stored at c.-20°C in a freezer until use.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
The test substance was administered in the diet, at constant concentrations, for 13 consecutive weeks.
Frequency of treatment:
in the diet
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0.15 % w/w
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
0.5 % w/w
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
1.5 % w/w
Basis:
nominal in water
No. of animals per sex per dose:
Four groups of 10 males and 10 females each: a vehicle control group and three test groups.
Control animals:
yes, concurrent vehicle

Examinations

Observations and examinations performed and frequency:
Each animal was observed:
- clinical signs
- body weight
- food consumption and food efficiency
- intake of the test substance
- water consumption
- ophthalmoscopic examination
- haematology
- clinical chemistry
- renal concentration test and urinalysis
- gross necropsy
- histopathological examination

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):
no effects observed
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:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
The concentrations of the test substance found in diet samples analyzed after storage in the animal room for 7 days or in a freezer for five weeks showed that the substance was stable under simulated experimental conditions. Analysis of 5 samples per diet, taken at different locations, showed that the test substance was homogeneously distributed in the diet at all dose levels.The content of the test substance was close to the intended level in all diets analyzed.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
831 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Dose descriptor:
NOAEL
Effect level:
963 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

The objective of this study was to examine the possible sub chronic oral toxicity of the test substance RA 3643 in rats. This substance was administered in the diet, at concentrations of 0, 0.15, 0.5 or 1.5%, to groups of 10 males and 10 female Wistar rats for 13 consecutive weeks.

The ingestion of RA 3643 was well tolerated, as evidenced by the absence of treatment-related changes in the appearance, general condition and behaviour of the animals, their growth, food and water consumption, red blood cell and cloting potential values, total white blood cell counts, clinical chemistry values, renal concentrating ability, composition of the urine, organ weights, gross necropsy findings and histopathological findings.

In comparison with the controls, the mean percentage of neutrophils was higher and that of lymphocytes lower in all groups of female rats given RA 3643.

The differences showed no dose-response relationship and probably resulted from the relatively low neutrophil and high lymphocyte count in the control group.

Most importantly, the differences were not reflected in dose-related or significant changes in the absolute numbers of these cell types and are, therefore, regarded as chance findings, unrelated to treatment.

Applicant's summary and conclusion

Conclusions:
Since the ingestion of RA 3643 at dietary levels up to 1.5% for 13 consecutive weeks was tolerated without signs of toxicity, the dietary concentration of 1.5% was a no-observed-adverse-effect level of RA 3643 under the conditions of this study. This dietary level provided a mean intake of 831 and 963 mg of RA 3643/kg body weight/day in male and female rats, respectively.