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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
November 6, 1992 through December 2, 1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline Study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995
Reference Type:
publication
Title:
Developmental toxicity of 2-butin-1,4-diol following oral administration to the rat
Author:
Hellwig J., M. Beth, and H.J. Klimisch
Year:
1997
Bibliographic source:
Toxicology letters, 92:221-230

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
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): 2-Butyne-1,4-diol (2-Butin-1,4-diol)
- Physical state: solid
- Analytical purity: 98.9% verified analytically before and after study
- Lot/batch No.: 27-0141
- Storage condition of test material: refrigerator under N2

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Additional details on type: sexually mature virgin females (Chbb:THOM (SPF)); disease free, randomized and tatooed for identification
- Source: Karl THOMAE, Biberach an der Riss, FRG
- Housing: singly in type DK III stainless steel wire mesh cages supplied by Becker & Co., Castrop-Rauxel
- Diet (e.g. ad libitum): ad libitum; ground Kliba 343 feed rat/mouse/ hamster supplied by KLINGENTALM(JHLE AG, Kaiseraugst, Switzerland
- Water (e.g. ad libitum): ad libitum, tap water from water bottles
- Age: 60 days old when recieved, 71 to 77 days old at begining of study (day 0 of study / day of pregnancy)
- Weight: 233.2g (mean) at day 0

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 24 °C
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12 (6:00 to 18:00 hrs)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Remarks:
double distilled
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Each day the test substance solutions were freshly prepared shortly before the test substance was administered. For the preparation of the solutions, an appropriate amount of test substance was weighed in a volumetric flask, subsequently topped up with doubly distilled water and intensively shaken. The volume administered each day was 10 ml/kg body weight. The calculation of the volume administered was based on the individual body weight determined at the beginning of the administration period (day 6 p.c.).
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability of the test substance solutions were determined by gas chromatography.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 2-4 untreated female rats were mated with one untreated male rat
- Length of cohabitation: 2 days
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
from day 6 - 15 post coitum
Frequency of treatment:
daily, in the morning
Duration of test:
Until 20 days post coitum
Doses / concentrations
Remarks:
Doses / Concentrations:
10; 40; 80 mg/kg bw/day
Basis:
other: nominal dose administered by oral gavage
No. of animals per sex per dose:
25 females per dose
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: Based on the results of a preceding study in which the stubstance was applied to 5 female Wistar rats (Chbb:THOM SPF) in doubly distilled water at doses of 20, 40 and 60 mg/kg body weight for 10 consecutive days (days 6 - 15 p.c.):

- 10 mg/kg body weight: as the expected no observed adverse effect level

- 40 mg/kg body weight: as a dose which might be a minimal toxic effect level for dams and/or fetuses or another no adverse effect level

- 80 mg/kg body weight: as the dose level at which some overt signs of maternal toxicity (e.g. body weight loss, reduced food consumption) were expected

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily, more often if clinical signs of toxicity were elicited

BODY WEIGHT: Yes
- Time schedule for examinations: on days 0, 1, 3, 6, 8, 10, 13, 15, 17, and 20 post coitum
- Corrected body weight body gain was calculated after terminal sacrifice (terminal body weight on day 20 p.c. minus weight of the uterus before it was opened minus body weight on day 6 p.c.)

FOOD AND WATER CONSUMPTION: on days 1, 3, 6, 8, 10, 13, 15, 17, and 20 post coitum

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: uterus, ovaries
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: number of dead fetuses
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No data
Statistics:
The Dunnet-Test was used for a simultaneous comparison of several dose groups with the control. The hypothesis of equal means was tested. This test was performed two-sided and was used for the statistical evaluation of the following parameters: food consumption, body weight, body weight change, corrected body weight gain, weight of uterus before it was opened, number of corpora lutea, number of implantations, number of resorptions and number of live fetuses, proportion of preimplantation loss, postimplantation loss, resorptions and live fetuses in each litter, litter mean fetal body weight and litter mean placental weight.

Fischer's Exact test was used for a pairwise comparison of each dose group with the control for the hypothesis of equal proportions. This test was performed one-sided and was used for female mortality, females pregnant at terminal sacrifice and the number of litters with fetal findings.

The Wilcoxon-Test was used for a comparison of each dose group with the control for the hypothesis of equal medians. This test was performed one-sided and was used for the proportion of fetuses with malformations, variations, retardations and/or unclassified observations in each litter.
Indices:
conception rate, preimplantation loss, postimplantation loss
Historical control data:
Yes. Body weight, reproduction data, placental weights, mean fetal weights, fetal external malformations, feetal external variations and unclassified findings, fetal soft tissue malformations and variations and unclassified findings, fetal skelatal malformations and variations and retardations, and other data.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: High dose group only in 2 animals

Details on maternal toxic effects:
One dam (No. 94) of test group 3 (80 mg/kg body weight/day) died intercurrently on day `8 p.c. after showing some adverse clinical findings (apathy, poor general state, vaginal hemorrhage and piloerection). At necropsy mottled liver and marginal emphysema of the lungs were recorded for this dam (see 4.2.2.1.). Another dam of this test group (No. 100) showed piloerection twice during the treatment period (days 8 and 9 p.c.). These clinical findings are possibly related to the test substance administration. There were no abnormal clinical findings for any other dam in the study.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day
Basis for effect level:
other: maternal toxicity

Results (fetuses)

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

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day
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

The following findings were obtained and assessed as substance-related:

Test group 3 (80 mg/kg body weight/day): A statistically significant reduction in food consumption at the beginning of the treatment period (days 6 - 8 p.c.) [about 21% less than the controls] and a statistically significant body weight loss between days 6 - 8 p.c. In addition, one intercurrent death (dam No. 94) on day 8 p.c. and some adverse clinical symptoms (apathy, poor general state, vaginal hemorrhage and pilo-erection) in this dam occurred shortly before death and, for another dam (animal No. 100), piloerection was observed on days 8 and 9 p.c. There was also an increased occurrence of one skeletal variation (3.9% affected fetuses/litter with accessory 14th rib(s)).

Test group2 (40 mg/kgbody weight/day): No substance-related effects on dams, gestational parameters or fetuses

Test group 1(10 mg/kg body weight/day): No substance-related effects on dams, gestational parameters or fetuses.

The following findings were NOT considered substance-related:

Conception rate: The conception rate varied between 72% (test group 1 - 10 mg/kg body weight/day) and 88% (control group and test groups 2 and 3 - 40 and 80 mg/kg body weight/day). There were no substance-related and/or biologically relevant differences between the groups 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. The differences evident (i.e. the statistically significant increase in preimplantation loss and the statistically significantly lower number of live female fetuses/dam in test group 1 - 10 mg/kg body weight/day) are considered to be incidental and within the normal range of deviations for animals of this strain and age given the historical control data. Moreover, these findings do not show any relation to dosing.

Placental weights: Mean placental weights of the high dose group were slightly, but statistically significantly increased. This is considered random, because this value is fully in the historical control range.

External malformations: External malformations were recorded for two control and one low dose fetuses. All of these fetuses showed anasarca and for one of the two control fetuses a cleft palate was additiionally noted. These malformations, which show no relationship to dosing and are also present in the historical control data at a low frequency, are considered to be spontaneous in nature.

External unclassified observations: One unclassified observation (placentae fused) was recorded for two fetuses of the control group, for one fetus each of test groups 1 and 2 (10 and 40 mg/kg body weight/day) and 3 fetuses of test group 3 (80 mg/kg body weight/day). This finding is not associated with the treatment and is regarded as incidental.

Soft tissue malformations: The examination of the organs of the fetuses revealed one kind of malformation in the low and the intermediate dose group. Dilatation of one or both heart ventricles occurred in one fetus of test group 1 (10 mg/kg body weight/day) and in 3 fetuses of test group 2 (40 mg/kg body weight/day). Due to the missing dose-response relationship and due to the fact that both soft tissue malformations are also present in the historical control data at a low incidence, these malformations are considered to be spontaneous in nature and not related to the test substance administration.

Soft tissue variations: Soft tissue variations (dilated renal pelvis and/or hydroureter) were detected in all test groups. Both variations are commonly found in the rat strain used and all respective values of test groups 1 - 3 (10, 40 and 80 mg/kg body weight/day) are fully in the range of the historical control. Due to an unexpected low occurrence of these findings in the actual control group, the total rate of affected fetuses/litter with soft tissue variations is slightly, but statistically significantly increased in test group 3 (80 mg/kg body weight/day); this is, however, without any biological relevance and not regarded as induced by the test substance.

Skelatal malformations: Various malformations of the sternum [sternebrae bipartite, ossification centers dislocated, sternebrae severely fused (bony plate)], the ribs [rib(s) absent], and/or the vertebral column [thoracic/lumbar vertebral body/bodies dumbbell-shaped (asymmetrical), bipartite asymmetrical and/or absent] were seen. All skeletal malformations appeared without a dose-response relationship and are considered to be spontaneous in nature. With the exception of severely fused sternebrae, which occurred only in the control group, all skeletal malformations can be found at a comparable fetal/litter incidence in the historical controls.

Skelatal variations: The variations elicited were related to the ribs [shortened 13th, accessory 14th or rudimentary cervical rib(s)], the sternum [sternebrae of irregular shape or bipartite (symmetrical)] and the vertebral column [accessory thoracic vertebra]. Skeletal variations appeared without a clear dose-response relationship, can be found in a similar frequency in the historical control data, and/or the differences between the groups are without biological relevance. For both statistically significantly increased skeletal variations [i.e. sternebrae of irregular shape, test group 2 (40 mg/kg body weight/ day) and accessory 14th rib(s), test groups 2 and 3 (40 and 80 mg/kg body weight/day)], this effect is interpreted as an embryotoxic effect representing a manifestation of a non-specific stress on the dams rather than as being a teratogenic effect of the test substance at this dose level.

 

Skelatal retardations: In all groups signs of skeletal retardations occurred substantiated by incomplete or missing ossification of skull (incl. hyoid) bones, vertebral column, sternebrae, hindlimbs, metacarpal and/or metatarsal bones. All differences between the groups in respect to skeletal retardations are without any biological relevance and do not show any relation to treatment.

Applicant's summary and conclusion

Conclusions:
For this prenatal toxicity study in rats, the no observed adverse effect level (NOAEL) for dams and fetuses is 40 mg/kg body weight/day.
Executive summary:

Thus, under the conditions of this full-scale prenatal toxicity study, 2-Butyne-l,4-diol caused overt signs of maternal toxicity at 80 mg/kg bodyweight/day substantiated by reduced food consumption, impaired body weight gains, the intercurrent death of one dam and some adverse clinical signs (all these findings occurred at the beginning of the treatment period). Forty (40) and 10 mg/kg body weight/day were tolerated by the dams without any substance-induced findings. Marginal signs of developmental toxicity were observed at the highest dose level (80 mg/kg body weight/day), substantiated by an increased number of affected fetuses/litter with accessory 14th rib(s), a skeletal variation. No substance-induced teratogenic effects, however, were observed up to and including the dose of 80 mg/kg body weight/day and there were no signs of embryo-/fetotoxicity at 40 mg/kg and 10 mg/kg bodyweight/day. For this prenatal toxicity study in rats, the no observed adverse effect level (NOAEL) for dams and fetuses is 40 mg/kg body weight/day.