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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:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: liquid
Details on test material:
Name of test material (as cited in study report): 2-mercaptoethanol
- Physical state: liquid, colorless, clear
- Analytical purity: 99.6 corr. area-%
- Lot/batch No.: 63133756P0
- Expiration date of the lot/batch: 01.05.2014
- Stability under test conditions: Stability guaranteed by sponsor
- Storage condition of test material: Room temperature

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS- Source: Charles River Laboratories- Age at study initiation: 10-13 weeks- Weight at study initiation: - Housing: 1 rat per cage- Diet: Ground Kliba maintenance diet ad libitum- Water : filtered tap water ad libitum- Acclimation period: From GD 0 (day of supply) to the beginning of administration (GD6), the animals will be accustomed to the environmental conditions and to the dietENVIRONMENTAL CONDITIONS- Temperature (°C): 20-24°C- Humidity (%): 30-70%- Air changes (per hr): 15 airchanges per hour- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: isotonic saline solution
Details on exposure:
5 mL/kg body weight; for test substance preparation, the specific amount of test substance will be weighed, topped up with cooled isotonic saline solution (fresenius) in a graduated flask and intensely mixed by shaking; all constituents are cooled before and during preparation; preparations are prepared at intervals which guarantee that the test substance concentrations in the vehicle remain stable
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical investigations of the test substance preparations were carried out at the Analytical Chemistry Laboratory of Experimental Toxicology and Ecology of BASF SE, Ludwigshafen, Germany.Analytical verifications of the stability of the test substance in isotonic saline solution (Fresenius) over a period of 9 days in a refrigerator were conducted prior to the start of the study.Samples of the test substance preparations were sent to the analytical laboratory at the beginning of administration for verification of the concentrations.
Details on mating procedure:
Animals paired by the breeder (time-mated animals) were supplied at noon on the day of evidence of mating; this day is referred to as GD0
Duration of treatment / exposure:
Animals are treated once daily from GD6-GD19
Frequency of treatment:
Once daily
Duration of test:
Dams are sacrificed on day 20
Doses / concentrations
Remarks:
Doses / Concentrations:
5, 15, 25 mg/kg/d
Basis:
actual ingested
No. of animals per sex per dose:
25 females per dose
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: The high dose was selected based on signs of toxicity noted at a dose level of 50 mg/kg bw/d in a previously conducted OECD 422 study (test facility CIT, laboratory study number 24847 RSR, 2004) and in a maternal toxicity range-finding study (BASF project 10R0234/04R034) which preceded this definitive prenatal developmental toxicity study.In the OECD 422 combined repeated dose with the reproductive/developmental screening test, 2 out of 10 Sprague-Dawley dams in the 50 mg/kg bw/d group died immediately prior to or during parturition with another death occurring during early lactation. Although the OECD Guideline 414 study design does not continue the dose administration through the period immediately prior to and during parturition, the presence of this toxicity was pertinent in the selection of dose levels for the definitive prenatal developmental toxicity study.In the maternal toxicity range-finding study, 10 pregnant Wistar rats were administered the test substance by oral gavage from gestational day (GD) 6 through GD 19. One dam died on GD 18 and one dam was sacrificed in moribund condition on GD 20, showing signs of piloerection, semiclosed eyelids, apathy, and hypothermia. While it is possible that the mortality observed and the clinical signs were due to gavage errors in administering the test substance (as they were not observed in the OECD 422 study at the 50 mg/kg and higher dose levels), a conservative interpretation is that the toxicity observed was due to the test substance. For these reasons, the dose level of 50 mg/kg bw/d was considered to be potentially lethal to the dams in the OECD 414 study.The selected high dose for the present study represented half of this lethal dose. This approved procedure of decreasing a lethal dose by a factor of two to become the high dose in a subsequent regulatory study meets the principles of guidelines OECD 414 (adopted 2001) and OPPTS 870.3700 (US EPA), as well as ECHA practical guide 10 (“how to avoid unnecessary testing on animals”; chapter 4 “animal welfare”; ECHA-10-B-17-EN, 2010) which is in compliance with EU Directive 86/609/EEC on animal protection.The oral route was selected since this has proven to be suitable for the detection of a toxicological hazard.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes- Time schedule: daily, abnormalities and changes are documentedDETAILED CLINICAL OBSERVATIONS: Yes - Time schedule: dailyBODY WEIGHT: Yes- Bw is recorded on day 0, 1, 3, 6, 7, 8, 9, 10, 13, 15, 17, 19 and 20FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes - Food consuption is recoreded from GD0-1, 1-3, 3-6, 6-7, 7-8, 8-9, 9-10, 10-13, 13-15, 15-17, 17-19, 19-20POST-MORTEM EXAMINATIONS: Yes- Sacrifice on gestation day #20- Organs examined: Adrenal glands, Heart, Kidneys, Liver, Spleen
Ovaries and uterine content:
The ovaries and uterine content was examined after termination:Examinations included:- Weight of the unopened uterus- Number of corpora lutea- Number of implantations- Number of early resorptions- Site of implantations in the uterus
Fetal examinations:
After removal of fetuses from the uterus, the following examinations have been made:- weight of each fetus- sex- weight of placentas- gross pathological examination of fetuses after dissection from uterus - half of the fetuses of each dam is skines, fixed in ethyl alcohol and the skeloton and cartilage stained (method by Kimmel and Trammell)- the other half of the fetuses is fixed in Harrisons fluid and soft tissues examined
Statistics:
Dunnett's test, Fishers exact test and Wilcoxon test

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effectsDetails on maternal toxic effects:There were no substance-related or spontaneous mortalities in any females of all test groupsNo clinical signs or changes of general behavior, which may be attributed to the test substance, were detected in any female.The mean food consumption and the mean body weight of the dams in test groups 1-3 (5, 15 or 25 mg/kg bw/d) were generally comparable to the concurrent control group throughout the whole study period.No treatment-related changes among hematological and clinical chemistry parameters were observed.The mean gravid uterus weights of the animals of test groups 1-3 (5, 15 and 25 mg/kg bw/d) were not influenced by the test substance.The conception rate reached 100% in all test groups (0, 5, 15 and 25 mg/kg bw/d).There were no test substance-related and/or biologically relevant differences between test groups 0, 1, 2 and 3 (0, 5, 15 and 25 mg/kg bw/d) in conception rate, in the mean number of corpora lutea and implantation sites or in the values calculated for the pre- and the postimplantation losses, the number of resorptions and viable fetuses.There were no testsubstance-related differences between the test groups 0, 1, 2 and 3 (0, 5, 15 and 25 mg/kg bw/d) in absolute and relative organ weights.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
25 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 effectsDetails on embryotoxic / teratogenic effects:The sex distribution of the fetuses in test groups 1-3 (5, 15 and 25 mg/kg bw/d) was comparable to the control fetuses.The mean placental weights of the low-, mid- and high-dose groups were comparable to the corresponding control group.The mean fetal weights of test groups 1, 2 and 3 (5, 15 and 25 mg/kg bw/d) were not influenced by the test substance.External malformations were recorded for one control fetus and one mid-dose fetus (15 mg/kg bw/d). One male control fetus had more than one malformation affecting the head (i.e. mandibular micrognathia, aglossia and microphthalmia). The total incidence of external malformations in treated animals did not differ significantly from the control group and was comparable to the historical control data.No soft tissue malformations were recorded; Two soft tissue variations, i.e. dilated renal pelvis and dilated ureter, were detected in all test groups including the control (test groups 0-3; 0, 5, 15 and 25 mg/kg bw/d). The incidences of these variations were neither statistically significantly different from control nor dose-dependent and therefore, not considered biologically relevant. Skeletal malformations were recorded for two control fetuses and two mid-dose fetuses (15 mg/kg bw/d). One male control fetus had associated external malformations. Furthermore, one male mid-dose fetus (15 mg/kg bw/d) had more than one malformation affecting the skeleton in different areas. The total incidence of skeletal malformations in treated animals did not differ significantly from the control group.For all test groups, skeletal variations of different bone structures were observed, with or without effects on corresponding cartilages. The observed skeletal variations were related to several parts of fetal skeletons and appeared in the majority of cases without a relation to dosing. The overall incidences of skeletal variations were comparable to the historical control data.Additionally, some isolated cartilage findings without impact on the respective bony structures, which were designated as unclassified cartilage observations, occurred in all test groups. The observed unclassified cartilage findings were related to the skull, the cervical vertebrae, the ribs and the sternum and did not show any relation to dosing.Soft tissue malformations did not occur in any fetuses in this study. There were noted external and skeletal malformations in two control and three mid-dose fetuses. The distribution of total malformations about the groups was not related to dose.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: developmental toxicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The NOAEL for maternal and developmental toxicity was determined to be 25 mg/kg bw.