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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
4th October to 20th November 1985
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted to GLP.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1986
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
not specified
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): DINOSEB TECHNICAL GRADE
- Physical state: Broum-yellowish, resinous
- Analytical purity: 96.1%
- Lot/batch No.: DBS 071085
- Expiration date of the lot/batch: Stable for 3 years when stored under conditions of -20 degrees centigrade (reported by the sponsor, telefax of May 9/ 1986)
- Stability in the vehicle: Stable for 90 minutes at least (Results of analysis performed for RCC Project 045303)
- Storage condition of test material: In a freezer at -20 degrees centigrade

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: KFM, Kleintierform, Madoerin AG, CH 4414 Fuellinsdorf, Switzerland
- Age at study initiation: at least 11 weeks old
- Weight at study initiation: 189 grams to 236 grams
- Housing: The animals were housed individually in Makrolon type-3 cages with standard softwood bedding (Lignocel, Schill AG, CH 4132 Muttenz,
Switzerland).
- Diet (e.g. ad libitum): Pelleted standard Kliba 343 rat maintenance diet (Klingentalmuehle AG, CH 4304 Kaiseraugst, Switzerland) ad libitum.
- Water (e.g. ad libitum): Tap water was available, ad libitum.
- Acclimation period: 10 days (minimum) under test conditions, after veterinary examination.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2 degrees centigrade
- Humidity (%): 55 % ± 10 %
- Air changes (per hr): 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light/12 hours dark, at least 8 hours music/light period.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test material was weighed into a glass beaker on a tarred precision balance (Mettler PK 300) and the vehicle added (w/v). The mixtures were prepared using a homogenizer. The low and middle dose mixtures were prepared from the high dose Stock mixture. The homogeneity of the test article in the vehicle was maintained using a magnetic stirrer during the daily administration period. Caused by the dilution factor 1:3, the exact dose level of the low and middle dose was 1.1 and 3.3 mg/kg body weight, respectively.

The test article/vehicle mixtures were prepared daily prior to administration.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration and homogeneity of the test article/vehicle mixtures were determined at the initiation and termination of the treatment period. Analyses were performed in the RCC Analytical Chemistry Laboratory, according to a method supplied by HOECHST AG. The concentrations found were in the range of 95.5% to 123.3% of nominal values. The homogeneity varied in the range of +4.9% to -2.7% of the mean concentration. The stability of the test item in corn oil over a period of 90 minutes was determined during the RF study.
Details on mating procedure:
- Impregnation procedure: co-housed
- If co-housed: the females were paired overnight with sexual mature males
- M/F ratio per cage: 1:1
- Length of cohabitation: overnight
- Proof of pregnancy: The day on which spermatozoa mere found in the vaginal smear or a vaginal plug were observed, was designated as day 0 post coitum.
Duration of treatment / exposure:
The test article was administered orally by intubation, once daily in the morning from day 6 through day 15 post coitum. All groups received a volume of 5 ml/kg body weight with a daily adjustment of individual volume to the actual body weight.
Frequency of treatment:
once daily in the morning
Duration of test:
On day 21 post coitum, all females were sacrificed and the foetuses removed by ceasarean section.
Doses / concentrations
Remarks:
Doses / Concentrations:
1, 3 & 10 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
25 mated female rats
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dosages were based upon the results of a preliminary study (RCC Project 045270, reported on July 8th 1985) and information available to the individual members of the Task Force. This data indicated maternal lethality at doses above 10 mg/kg. In the RCC study, the doses were 0, 3, 10 and 30 mg/kg. All females in the 30 mg/kg group died after two doses. In the 10 mg/kg group body weight gain was inhibited until gestation day 9. Food consumption was increased from days 11 to 21. There were no effects at 3 mg/kg except for increased food consumption from days 11 to 21.
Based on the results, dose levels of 1, 3 and 10 mg/kg body weight were chosen for the main study.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily, minimum.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Twice daily, minimum.

BODY WEIGHT: Yes
- Time schedule for examinations: Daily, from day 0 until day 21 post coitum.

FOOD CONSUMPTION: Yes
The data were recorded on day 6, 11, 16 and 21 post coitum

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21, all females were killed by CO2 asphyxiation and the foetuses removed by caesarean section.
- Organs examined: Postmortem examinations, including gross macroscopic examination of all internal organs, with emphasis upon the uterus, uterine contents, position of fetuses in the uterus and number of corpora lutea were performed and the results recorded. The foetuses were removed from the uterus, sexed, weighed, examined for gross external abnormalities and submitted to one of the following procedures individually:

1) Slicing technique of Wilson for examination of the viscera and brain. One half of the number of live foetuses from each litter was fixed in a mixture of ethyl alcohol, formol and acetic acid. After evaluation the individual sections were preserved in a solution of ethyl alcohol and glycerine (one fetus per container). Descriptions of any abnormalities were recorded.

2) The remaining foetuses (the other half of the number of live foetuses) were placed in a solution of potassium hydroxide for clearing and stained with alizarin red (modified technique). The skeletons were examined and all abnormalities were recorded. The specimens were preserved individually in plastic bags.

The uteri (and contents) of all pregnant females were weighed on the scheduled day of neoropsy and used to determine the corrected body weight gain.
The uteri of all females which were found at necropsy to be not pregnant were placed in an aqueous solution of ammonium sulfide to accentuate possible hemorrhagic areas of implantation sites.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: half per litter
Statistics:
The following statistical methods mere used to analyse body weights, food consumption and reproduction data:

Univariete one-way analysis of variance was used to assess the significance of intergroup differences if the variables could be assumed to follow a normal distribution.
The Dunnett many-one t-test, based on a pooled variance estimate, was used for intergroup comparisons (i.e. single treatment groups against the control group).

A one-way univariate analysis of variance based on Wilcoxon ranks together with the Kruskall-Wallis test was applied to the reproduction data parameters.

Fisher's exact test for 2x2 tables was applied if the variables could be dichotomized without loss of information.

Individual values, means, standard deviations and t-statistics were rounded off before printing.
Indices:
No data
Historical control data:
Yes

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Oral administration of the test material to mated female rats caused maternal toxicity at the highest dose level of 10 mg/kg bw/day, indicated by reduction of food consumption, nearly unchanged body weights during the first three days of the treatment period and reduced body weight gain during the entire treatment period.

Effect levels (maternal animals)

Dose descriptor:
NOEL
Effect level:
3 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
The test material did not reveal any teratogenic potency up to the highest dose level of 10 mg/kg bw/day.

Effect levels (fetuses)

Dose descriptor:
NOEL
Effect level:
10 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: teratogenicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

No death occurred and no sign or symptom was observed in any female of any group. At necropsy, no abnormal finding was noted.

The oral administration of the test material caused reduced food consumption of the dams treated at 10 mg/kg bw/day, between days six to eleven post coitum (first recording period during the treatment period) and nearly unchanged body weights until day eight post coitum (third test article administration). The calculations of the mean body weight gain during the entire treatment period resulted in a reduction in this group, when compared to that of the vehicle control group. No test article-related differences between the vehicle control group and any dose group were noted in any of the reproduction data recorded.

The evaluation of the data of foetuses yielded test article-related significantly reduced mean body weights in foetuses of dams treated at 10 mg/kg bw/day when compared to that of the vehicle control group.

During the external and visceral investigations of foetuses, no malformation or anomaly was noted in any group. During the skeletal investigations of foetuses for malformations and/ or anomalies, similar findings of nonspecific variations were noted in the control, mid-dose and high-dose groups. No finding was noted in any foetus of the low-dose group. The frequency of these common findings was within the normal range for animals of this rat strain.

The comparison of the stage of skeletal development yielded a slightly increased quota of incompletely ossified crania, of still absent ossifications of phalangeal nuclei, calcanea and cervical vertebrae in the foetuses of group treated at 10 mg/kg bw/day. This result corresponded to the significantly reduced body weights of foetuses and was considered to be an effect of the maternal toxicity.

Applicant's summary and conclusion

Conclusions:
The no-observable-effect level (NOEL) in the maternal and foetal organism was 3 mg/kg bw/day. The test material did not reveal any teratogenic potency up to the highest dose level of 10 mg/kg bw/day when administered to mated female Wistar rats under the described conditions of this study.
Executive summary:

The purpose of this embryotoxicity study was to assess the effects of the test material on embryonic and foetal development when administered daily to mated female rats by oral gavage from day 6 through day 15 post coitum. The study was conducted in accordance with OECD 414 using pregnant female Wistar rats.

On day 21 post coitum, all females were sacrificed and the foetuses removed by caesarean section. The investigations of females/dams and foetuses were performed in accordance with international recommendations. All parameters recorded were evaluated and reported.

The results from this study showed that the oral administration of the test material to mated female rats caused maternal toxicity at the highest dose level of 10 mg/kg bw/day, indicated by reduction of food consumption, nearly unchanged body weights during the first three days of the treatment period and reduced body weight gain during the entire treatment period. The effects on body weights and skeletal development of foetuses were also considered to be caused by the maternal toxicity.

The no-observable-effect level in the maternal and foetal organism was 3 mg/kg bw/day.

The test material did not reveal any teratogenic potency up to the highest dose level of 10 mg/kg bw/day when administered to mated female Wistar rats under the described conditions of this study.