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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study according to OECD Guideline 414, adopted 1981 (exposure not through the entire period of gestation to the day before caesarean section as recommended in the current guideline).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
Principles of method if other than guideline:
adopted 1981
Also in accordance with Commission Directive 87/302/EEC of Nov . 18, 1987 Part B
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Purity >= 98.4%
BASF No. 89/811
test substance (TS), stability of the test substance, and solution of the test substance (stability, homogeneity) were analysed

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
- sexually mature, virgin Wistar rats
- supplied by Karl Thomae, Biberach, Germany
- acclimatization period at least 2 weeks
HOUSING AND DIET
- rats singly housed
- temperature 20-24°C, relative humidity 30-70%, day/night rythm 12h/12h
- room desinfected before use
- food (Kliba 343 feed) and tap water ad libitum (analysed: no contaminations)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: bidistilled water
Details on exposure:
- Concentration in vehicle: 0 (control), 1, 4, or 10 g/100 ml
- Total volume applied: 10 ml/kg bw in both groups
- TS solution prepared in intervals of 10 days (stable solution)
- Number of animals per group: 25
- due to technical reasons, study carried out in 2 sections
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
- test substance (TS), stability of the TS, and solution of the test substance (stability) were analysed
- stability of the TS and TS solutions was confirmed
Details on mating procedure:
- virgin Wistar rats (mean weight 242 g; randomization) mated with untreated fertile males of the same breed
- if sperm was detected in the vaginal smear in the morning, this day was considered day0 (rats were 88-90 days old)
Duration of treatment / exposure:
gestation day (GD) 6 to 15
Frequency of treatment:
once daily
Duration of test:
GD 20
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 400 or 1000 mg/kg bw d
Basis:
actual ingested
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
This study followed a limit test (0 or 1000 mg/kg bw/day) in the same species (showing increased incidences in skeletal variations).

Examinations

Maternal examinations:
- Body weight gain: recorded on gestation day 0, 1, 3, 6, 8, 10, 13, 15, 17, 20 (corrected bw determined)
- Food consumption: determined on the same days than bw
- Clinical observations: clinical symptoms recorded once daily
Ovaries and uterine content:
Measured parameters at termination are weight of uterus, no. of corpora lutea, live fetuses and dead implantations, early and late resorptions, dead fetuses; calculation of conception rate, preimplantation loss and postimplantation loss.
Fetal examinations:
Measured parameters are bw, sex, external findings, viability, placental weight; one half of the fetuses per dam prepared for soft tissue examination (method according to Barrow and Taylor, J Morph 127, 291-306, 1969), the other half for skeletal examination (method according to Dawson, Stain Tech 1, 123, 1926); detected changes differentiated in malformation, variation, retardation and unclassified observations.
Statistics:
- two-sided DUNNETT-Test for comparison of one dose group with control
- one-sided FISHER's EXACT Test for a pairwise comparison of each dose group with control for hypothesis of equal proportions
- one-sided WILCOXON-Test for comparison of the dose group with control for the hypothesis of equal medians
- significance level p<0.05
Indices:
see above
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:
- Number of nonpregnant rats: 1 rat in control group, 0 in low dose group, 4 rats in mid dose group and 0 in high dose group
- None of the determined parameters (see freetext Material and Methods) revealed statistically significant or toxicologically relevant results with exception of the bw of pregnant rats in the high dose group on gestation day 8 (32% lower than control value; transient, significant effect; authors comment: marginal effect but possibly treatment related)

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
400 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
1 000 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
400 mg/kg bw/day
Basis for effect level:
other: developmental toxicity
Dose descriptor:
LOAEL
Effect level:
1 000 mg/kg bw/day
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
- No statistical significant effects observed (compare with measured parameters in Material & Methods) except the effects described below.
- Sex distribution: in the mid dose group statistically significant more females (no dose dependency), authors comment: no biological relevance.
- Placental weight: in the mid dose group significantly decreased placental weights of male fetuses, but value within laboratory historical control range.
- External malformations: 1 anophthalmia in low dose group and in high dose group 1 fetus with cleft palate and 1 with microphthalmia; considered to be spontaneous in nature by the authors (also low incidence in laboratory historical controls).
- Soft tissue malformations: significantly increased incidence of affected fetuses/litter (1.8%) in the mid dose group, but this value is within the laboratory historical control data (0-1.9%); considered by the authors to be spontaneous in nature and not treatment related.

- Skeletal abnormalities
1) Nonsignificant increase in dumbbell ossification of thoracic vertebral bodies (most often observed in the high dose group; 6.3% versus 3.6% in control), but this variation is outside the laboratory historical control concerning litter incidence (40% versus 19.5% in control); compare also with the LIMIT Test (see also this Section).
2) Significant increase in number of affected fetuses/litter with accessory 14th rib in the high dose group (variation); this variation is regarded to be incidental in nature because litter and fetal incidences are within laboratory historical control range and this variation is not observed in the LIMIT Test (see also this Section).

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
1 000 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

AUTHORS CONCLUSION
No overt signs of teratogenicity observed up to 1000 mg/kg bw/day; however, a slight increase of dumbbell ossification of thoracic vertebral bodies was seen; this variation was significantly increased in a previously conducted LIMIT Test at the same dose level; a borderline effect due to TS treatment cannot be ruled out; at the same dose level marginal maternal toxicity (decreased body weight) occurred. The NOAEL for dams and fetuses is 400 mg/kg bw/day.

Applicant's summary and conclusion

Conclusions:
In Wistar rats the NOAEL for maternal toxicity and developmental toxicity is 400 mg/kg bw/day, the LOAEL is 1000 mg/kg bw/day (reduced body weight in dams and increased skeletal variations in offspring). No teratogenic effects were detected.
Executive summary:

GLP guideline study according to OECD Guideline 414, adopted 1981 (exposure not through the entire period of gestation to the day before caesarean section as recommended in the current guideline).

In a developmental toxicity study pregnant Wistar rats (n = 25 per group) received 0, 100, 400, or 1000 mg/kg bw/day by gavage on gestation days (GD) 6 to 15. The animals were sacrificed on GD 20. No maternal toxicity was detected concerning food consumption, clinical symptoms and pathological alterations at necropsy; however, a decrease in body weight at GD 8 in the high dose group was found. No substance-related differences between the treatment and control group were noted regarding uterus weight, mean number of corpora lutea, live fetuses and dead implantations, early and late resorptions, dead fetuses or in the values calculated for conception rate, pre- and post-implantation losses. Examination of fetuses did not reveal any obvious substance-related effects except a slight increase in dumbbell ossifications of thoracic vertebral bodies which were also outside the laboratory historical control range concerning litter incidence.This variation was significantly increased in a previously conducted LIMIT Test at the same dose level; a borderline effect due to TS treatment cannot be ruled out.

Conclusion: In Wistar rats the NOAEL for maternal toxicity and developmental toxicity is 400 mg/kg bw/day, the LOAEL is 1000 mg/kg bw/day (reduced body weight in dams and increased skeletal variations in offspring). No teratogenic effects were detected.ed.