Registration Dossier

Administrative data

Endpoint:
extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
Remarks:
based on generations indicated in Effect levels (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Sept - Dec 1976
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well performed and reported non-guideline study with scientific sound design.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1999
Report Date:
1999

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 443 (Extended One-Generation Reproductive Toxicity Study)
Deviations:
not applicable
Principles of method if other than guideline:
Rats were fed diets containing the test item at different levels and mated. The litters were reared and observations were made on fertility of females, number of young born per litter, sex ratio, grossly visible abnormalities, mortality, body weights and resorption percentage.
GLP compliance:
no
Remarks:
performed before GLP guidelines
Limit test:
no
Justification for study design:
Study was performed before the OECD 443 study was effective. However, the study design followed is comparable to the OECD 443 basic design.

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: The Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands
- Age at study initiation:8 weeks
- Weight at study initiation: (P) Males: 218 -219 g; Females: 148 -149 g
- Housing: in screen-bottomed, stainless steel cages
- Diet (e.g. ad libitum): stock diet (containing 29.7% yellow maize, 36% whole wheat, 11% defatted soy-bean mael, 4% meat scraps, 7% fish meal, dried whey, brewer's yeat, grass meal, soy-bean oil, vitamin preparations, trace mineralized salt, steamed bone meal), ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24-26°C
- Humidity (%): 50%



IN-LIFE DATES: From: Sept. 1976 To: Dec. 1976 (end of subsequent chronic toxicity study)

Administration / exposure

Route of administration:
oral: feed
Vehicle:
other: stock diet
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency):
The test diets were normally prepared in batches of 40 kg once every 4 to 6 weeks, but occasionally batches of 20 kg were prepared.
The test material was thoroughly mixed into stock diet by means of a mechanical blender at levels of 0 (control), 0.6, 1.2 or 2.4%. The diets were freshly prepared every two to three weeks and stored in an unheated room at ambient temperature.

Details on mating procedure:
After the pre-mating period (30 days) the rats were mated within their diet group. Each male was housed with two females in a cage for one week. At week 2 and 3 of the mating period each male rat was transferred to another "mating" cage within the same diet group. So, three different males were available for each dam. After a mating period of three weeks, the females were caged individually, until their litters had been weaned.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples were taken at intervals and sent to Hoechst AG, Frankfurt, Germany, for determination of DOS2.

Results of determination of Dioctadecylsulphide (DOS2) in test diets (diet analyses from retrieved from TNO Combined chronic toxicity and carcinogenicity study with dioctadecylsulfide (DOS2) performed in parallel)


Intended % DOS2
in the diet % DOS2 in feed samples in week
45 60 85

0 0.04 0.04 < 0.01
0.6 0.56 0.60 0.57
1.2 1.17 1.24 1.13
2.4 2.28 2.39 2.37

Duration of treatment / exposure:
Males: 7 weeks (4 weeks pre-mating, 3 weeks mating)
Females: 13 weeks (4 weeks pre-mating, 3 weeks mating, 3 weeks gestation, 3 weeks lactation)
Frequency of treatment:
continuously
Details on study schedule:
- Age at mating of the mated animals in the study: 15 weeks
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0.6 %
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
1.2%
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
2.4%
Basis:
nominal in diet
No. of animals per sex per dose:
15 males/dose
30 females/dose
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: Administration of DOS2 at dose levels of 0.1 and 1% to rats for 90 days revealed no treatment-related abnormalities (Hoechst 1967)
Positive control:
No

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
In the pre-mating period of 30 days food intake was recorded at weekly interval. Food efficiency ratios were calculated from the gain in body weight and the total food consumption over 4 weeks.
Oestrous cyclicity (parental animals):
no data
Sperm parameters (parental animals):
no data
Litter observations:
Records were made of the number of pups in each litter, sex ratio at birth and the weight of the litter at day 1, 10 and 20 of lactation. Pups were inspected grossly for club-feet, cleft palate and hydrocephalus. Litters containing more than ten siblings were randomly reduced to 10 on day 1 of lactation, in order to equalize the stress of lactation among the dams.
Postmortem examinations (parental animals):
After weaning their young, the mothers were sacrificed and the implantation sites in the uterus were counted after staining with ammonium sulfide solution. The males were discarded.
Postmortem examinations (offspring):
n.a., since the offspring was used a subsequent combined chronic toxcity/carcinogenicity study (please refer to study report no. R 6693; IUCLID Chapter 7.5.1; WoE_130 week oral toxicity (diet)_rat_TNO_1980)

Post mortem examiniations on the offspring were performed at the termination of the this chronic study.
Statistics:
Statistical procedures used in the evaluation of data were as follows:
- for pup body weights: one-way analysis of variance (ANOVA) followed bby Dunnett's multiple comparison tests
- for number of females pregnant, females with liveborn, females with (all) stillborn pups, number of live- and stillborn pups, number of pups/litters lost, number of male pups and number of implantation sites: Fisher's exact probability test
- for mean number of pups delivered, mean no. of pups alive, mean number of implantations and post-implantation loss: Kruskal-Wallis followed by Mann-Whitney U-tests.

All tests were two sided. As a level of significance p<0.05 was considered.
Reproductive indices:
- female fertility index = (number of pregnant females/number of females placed with males) x 100
- gestation index = (number of females with live pups/number of females pregnant) x 100
- number of lost implantations = number of implantation sites - number of pups born alive
- post-implantation loss = [(number of implantation sites - number of pups born alive)/number of implantation sites] x 100
Offspring viability indices:
- live birth index = (index of pups born alive/number of pups born) x 100
- viability index (days 1-20) = (number of live weanlings/number of pups alive on day 1 post partum) x 100
- pup mortality day n = (number of dead pups on day n/total number of pups on day n) x 100
- sex ratio day 1 = (number of live male pups on day n/number of live pups on day 1) x 100

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed

Details on results (P0)

No abnormalites of conditions or behaviour were observed in any of the groups during the pre-mating, mating or lactation period. The mean body weights of the various groups were approximately the same. Neither food consumption, nor food efficiency figures showed significant differences amongst the groups.

From the control, 0.6, 1.2 and 2.4% Hostanox SE 10 groups, 30, 30, 28 and 28 females were pregnant. The female fertility index was 100, 100, 97 and 93% for the control, 0.6, 1.2 and 2.4 % Hostanox SE 10 groups, respectively.
The number of females with liveborn pups was 30, 29, 27 and 25 for the control, 0.6, 1.2 and 2.4% groups, respectively. One female of the 1.2 and 2.4% Hostanox SE 10 groups delivered only dead pups. Stillborn pups were observed in 2, 2, 3, and 2 the control, 0.6, 1.2 and 2.4% group, respectively. The gestation index was 100, 97, 96 and 89% for the control, 0.6, 1.2 and 2.4% Hostanox SE 10 groups, respectively.
Post-implantation loss was 13.4, 15.2, 14.7 and 18.4% for the control, 0.6, 1.2 and 2.4% dose group, respectively.
In the different test groups all males could fertilize one or more female. In the control group two males out of 15 appeared to be not fertile.

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
> 2.4 other: % in diet
Sex:
male/female
Basis for effect level:
other: none of the parameters examined was adversely affected; corresponding to 1200 mg/kg bw/d (considering a diet conversion factor (ppm to mg/kg bw/d) of 20 for older rats)

Results: F1 generation

General toxicity (F1)

Clinical signs:
not specified
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
slightly decreased
Body weight and weight changes:
no effects observed
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not examined
Description (incidence and severity):
offspring was used for subsequent chronic study
Gross pathological findings:
not examined
Description (incidence and severity):
spring was used for subsequent chronic study
Histopathological findings:
not examined
Description (incidence and severity):
spring was used for subsequent chronic study

Details on results (F1)

The mean litter number of liveborn pups was comparable amongst the groups. The number of stillborn pups was low in all groups. Pup mortality calculated on pup basis was slightly increased from lactation days 1-10; 2, 6, 10 and 8 pups died in the control, 0.6, 1.2 and 2.4% Hostanox SE groups, respectively. Pup mortality in the control group was relatively low. No malformations were observed in any of the groups. Mean pup weights and pup weight changes (growth) of all groups were comparable on lactation days 1, 10 and 20.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
> 2.4 other: % in diet
Sex:
male/female
Basis for effect level:
other: none of the parameters examined was adversely affected; corresponding to 1200 mg/kg bw/d (considering a diet conversion factor (ppm to mg/kg bw/d) of 20 for older rats)

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Hostanox SE 10 was studied for its reprodcutive toxic/teratogenic properties in a one-generation combined chronic toxicity study in male and female Wistar rats. The NOAEL was considered to be higher than 2.4% (highest dose tested) in diet. None of the fertility/viability parameters examined was adversely affected by the feeding of the test substance.
Executive summary:

Hostanox SE 10 was examined in a reproduction study in rats by feeding the test substance at dietary levels of 0 (control), 0.6, 1.2 and 2.4% (corresponding to 0, 300, 600 and 1200 mg/kg bw/d). Only one litter was reared. Observations were made on the fertility of females, the number of live and dead pups born per litter, sex ratio of the pups on lactation day 1, grossly visible abnormalities, pup mortality and pup body weights on lactation days 1, 10 and 20 and post-implantation loss.

None of the parameters examined was adversely affected by the feeding of the test substance.

The F1 -generation was used for a subsequent chronic toxicity study (130 weeks duration, please refer to IUCLID Chapter 7.5.1). No visceral malformations could be found at the termination of this chronic study and therefore, no teratogenic effects were recorded.