Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-compliant guideline study, available as unpublished report, no restrictions, fully adequate for assessment.
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): potassium tetrafluoroborate
- Physical state: white powder
- Analytical purity: 98.9%
- Lot/batch No.: BWF41213
- Expiration date of the lot/batch: 13 December 2009
- Storage condition of test material: ambient temperature

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany, SPF-bred colony
- Age at study initiation: (P) 8-9 wks
- Weight at study initiation: (P) within 20% of the mean weight for each sex
- Housing: in macrolon cages with wood shavings (Lignocel, Type 3/4) as bedding material and strips of paper (Enviro-dri) as environmental enrichment. During the premating period, the animals were housed in groups of 4/sex. For mating, 1 male and 1 female were housed together. Mated female swere housed individually in macrolon cages, which were then placed in another cage rack. After delivery, the cage containing the dam with litter were transferred to another cage rack
- Diet: cereal-based (closed formula) roden diet (Rat & Mouse No. 3 Breeding Diet, RM3) from a commercial supplier (SDS Special Diets Services, ad libitum
- Water: tap water in polypropylene bottles, cleaned weekly and filled as needed, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
1% solution in water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test substance was suspended in 1% CMC. The dosing solutions were prepared weekly and stored at 2-10 ºC. The miscibility of the test substance in vehicle were checked by visual inspection before the start of the study.

VEHICLE
- Concentration in vehicle: 0, 20, 58.2, 175 and 500 mg/mL
- Amount of vehicle: 2 mL/kg bw
- Lot/batch no.: 101K0185
- Purity: 1% solution in water
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analyses to determine the stability, homogeneity and content of the test substance in the vehicle was conducted, by quantitative determination of the level of potassium tetrafluoroborate by determination of boron using ICP-AES and determination of fluoride using GC-FID with headspace.
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: until pregnancy occurred
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged individually for the birth and rearing of the pups until day 4 of lactation
Duration of treatment / exposure:
During a 2-week premating period, during the mating, gestation and lactation period until sacrifice. Animals were not dosed on the day of necropsy.
Frequency of treatment:
daily
No. of animals per sex per dose:
12/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Rationale for animal assignment: computer randomization proportionally to body weight.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily in the morning hours by cage-side observations. On working days, all cages were checked again in the afternoon for dead or moribund animals to minimise loss of animals from the study. On Saturdays and Sundays only one check per day was carried out.

BODY WEIGHT: Yes
- Time schedule for examinations: shortly before the time of dosing (randomization) and on the first day of dosing and weekly thereafter during the premating period. In addition, body weights of the male animals were measured on day 3 and 5 of the study. Males were weighed approximately weekly during the mating period until sacrifice. Females were weighed approximately weekly during mating and mated females were weighed on days 0, 7, 14 and 21 during presumed gestation and on day 1 and 4 of lactation. All animals were weighed on the day of sacrifice.

FOOD CONSUMPTION: Yes
Food consumption of male rats was measured approx. weekly, except during the mating period. In addition, food consumption of the male animals was measured on day 3 and 5 of the study, except for male animals of lowest dose group.
Food consumption of female rats was measured weekly during the premating period and during the gestation period from gestation days 0-7, 7-14 and 14-21, and once during the lacation period from day 1 to 4.

WATER CONSUMPTION: Yes
During daily observations, until day 5 of gestation.

GROSS NECROPSY
Animals were examined grossely for macroscopic changes.

HISTOPATHOLOGY / ORGAN WEIGHTS
Samples of the following organs were preserved: ovaries (after counting of the corpora lutea), uterus (after counting of the implantation sites), organs and tissues showing macroscopic abnormalities.
Fetal examinations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, number of litters lost entirely, presence of gross anomalies

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities
Statistics:
Clinical findings were evaluated by Fisher's probability test. Body weight, body weight gain, organ weights and food consumption data were subjected to one-way analysis of variance (ANOVA) followed by Dunnett's multiple comparison tests. Fisher's exact probability test were used to evaluate the number of mated and pregnant females and females with live pups. Number of implantation sites, live and dead pups were evaluated by the Mann Whitney U-test. Histopathological changes were evaluated by Fisher's exact probability test.
Indices:
REPRODUCTIVE INDICES
The following data were collected:
- number of succesful copulations (number of females mated)
- number of males that became sire
- number of pregnant females as demonstrated by the presence of implantation istes observed at autopsy.
- number of females surviving delivery
- number of females with liveborn and (all) stillborn pups

Pre-coital time = time between the start of mating and successful copulation
Duration of gestation = time between gestation day 0 and day of delivery
Mating index = (number of females mated/number of females placed with males) x 100
Female fertility index = (number of pregnant females/number of females placed with males) x 100
Female fecundity index = (number of pregnant females/number of females mated) x 100
Gestation index = (number of females with live pups/number of females pregnant) x 100

OFFSPRING VIABILITY INDICES
The following data were collected:
- number of pups delivered (live and stillborn)
- number of live pups at day n
- number of pups lost
- number of litters lost entirely
- number of corpora lutea
- number of male pups at day n
- number of implantation sites
- number of lost implantations
- litter size

Live birth index = (number of pups born alive/number of pups born) 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 n = (number of live male pups on day n/number of live pups on day n) x 100
Number of lost implantations = number of implantation sites-number of pups born alive
Pre-implantation loss = [(number of corpora lutea-number of implantation sites/number of corpora lutea] x 100
Post-implantation loss = [(number of implantation sites - number of pups born alive)/number of implantation sites] x 100

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
CLINICAL SIGNS AND MORTALITY
Two female animals of the 350 mg/kg bw/day died during lactation period. Daily clinical observations during the study did not reveal any remarkable findings in animal appearance, general condition or behaviour among the dosing and control groups.

BODY WEIGHT
Mean body weight and/or body weight change of the female animals of 350 mg/kg bw/day group were statistically significantly decreased during several periods of the study. During the premating period, no effect on body weight or body weight change was observed in female animals treated with KBF4. Mean body weight of the female animals of the 350 mg/kg bw/day group was statistically significantly decreased from gestation day 7-21. Mean body weight change of the female animals of this group was statistically significantly decreased from gestation days 7-14. Mean body weight of the female animals of this group was statisitcally significantly decreased on lactation day 1. During the lactation period no effect was observed on mean body weight change of the KBF4-treated females.

FOOD CONSUMPTION
Food consumption of the female animals of the 350 mg KBF4/kg body weight group was statistically significantly decreased from day 14-21 of the premating period, during GD 0-14 (g/animal/day) and GD 0-7 (g/kg/day) and day 1-4 of lactation (g/animal/day). One animal of this group did not eat any food from lactation day 1-4. Furthermore, no remarkable differences were observed in the KBF4-treated groups.

REPRODUCTIVE PERFORMANCE
In each group 12 females were placed with males. All females of the KBF4-treated groups and 11 females of the control group were mated within 4 days. The number of pregnant females, the number of females with live born pups and the number of males that became sires amounted to 8, 9, 9, and 5 for the control, 40, 116.5 and 350 mg/kg bw/day groups, respectively. The mating index was 100% in all KBF4-treated groups and 92% in the control group. The female fecundity index and female fertility index were comparable between the control, 40 and 116.5 mg/kg bw/day groups and ranged from 67-75%. In the 350 mg/kg bw/day group, the fecundity index, female fertility index was 42%. The gestation index was 100% in all groups. The duration of gestation was comparable between the groups. The number of corpora lutea and implantation sites was statistically significantly decreased in the 350 mg/kg bw/day group; no effect was observed in other treated groups when compared to the control group. Pre- and post-implantation loss was comparable in all groups.

GROSS PATHOLOGY
At scheduled necropsy no treatment-related gross changes were observed.

HISTOPATHOLOGY
Microscopic examination did not reveal treatment related histopathological changes in any of the sampled organs and tissues.

OTHER FINDINGS
Effects on water consumption were observed (not exactly measured) in the female animals of the 350 mg/kg bw/day group from week 3 onwards. In consultation with the study director it was decided to stop registration of water consumption after day 5 of gestation.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
116.5 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

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

Details on embryotoxic / teratogenic effects:
VIABILITY (OFFSPRING)
The number of litters with live born pups was 8, 9, 9 and 5 for the control, 40, 116.5 and 350 mg/kg bw/day groups, respectively. The number of live pups per litter on postnatal day 1 in the control, 40, 116.5 and 350 mg/kg bw/day groups was 11.9, 12.1, 11.8 and 9.4, respectively. Pup mortality on postnatal day 4 amounted to 3, 13, 3 and 29 (incidences 3.2, 12, 2.8 and 62%) in the control, 40, 116.5 and 350 mg/kg bw/day groups, respectively; in the 350 mg/kg bw/day group the mortality was statistically significantly increased. The death of the pups of two dams (10 and 11 pups, respectively) could be secondary to the death of the dams on postnatal days 2 and 3, respectively. In the control, 40, 116.5 and 350 mg/kg bw/day groups 0(8), 1(8), 0(9) and 3(2) litters, respectively, were lost entirely between postnatal days 1-4 (between brackets the number of litters with live pups on postnatal day 4). The number of live pups per litter on postnatal day 1 was 11.9, 12.1, 11.8 and 9.4 in the control, 40, 116.5 and 350 mg/kg bw/day groups, respectively, and on postnatal day 4 the number of pups was 11.5, 12.0, 11.4 and 9.0 in the control, 40, 116.5 and 350 mg/kg bw/day groups, respectively. The number of live pups per litter of the 350 mg/kg bw/day weight group was statistically significantly decreased on postnatal day 1. On postnatal day 4 only pups of 2 litters in the high-dose group were alive. The viability index on postnatal days 1-4 was 97, 88, 97 and 38% for the 0, 40, 116.5 and 350 mg/kg bw/day groups, respectively.

BODY WEIGHT (OFFSPRING)
No differences were observed for pup weight and pup weight changes on postnatal days 1 and 4. On postnatal day 1 the number of runts (pup weight less than mean pup weight of the control group minus 2 standard deviations) was statistically significantly increased in the 116.5 and 350 mg/kg bw/day groups. In addition, the number of runts was statistically significantly increased in the 116.5 mg/kg bw/day group on postnatal day 4.

GROSS PATHOLOGY (OFFSPRING)
Macroscopic observations in stillborn pups and pups that died between postnatal days 1-4 did not reveal any treatment related abnormalities.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
ca. 116.5 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
not specified
Basis for effect level:
changes in litter size and weights

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The NOAEL in rats is 116.5 and 40 mg/kg bw for maternal toxicity and developmental toxicity, respectively.
Executive summary:

In a GLP compliant reproduction/developmental toxicity screening test, performed according to OECD Guideline 421, Wistar rats were orally exposed to potassium tetrafluoroborate. 1% carboxymethylcellulose solution in water (CMC) was used as vehicle. Groups of 12 rats were dosed daily with 0, 40, 116,5 and 350 mg KBF4/kg body weight during 4 weeks premating, mating, gestation and up to day 4 of lactation. Two female animals of the 350 mg KBF4/kg body weight died during the lactation period. Daily clinical observations during the study did not reveal any remarkable findings in animals' appearance, general condition or behavior among the dosing and control groups. Effects on water consumption were observed (not exactly measured) in the 350 mg KBF4/kg body weight group from week 3 onwards. Mean body weight and or body weight change of the animals of the 350 mg KBF4/kg body weight were statistically significantly decreased during several periods of the study. Food consumption of the 350 mg KBF4/kg body weight group was statistically significantly decreased during several periods of the study. In each group 12 females were placed with males. All females of the KBF4-treated groups and 11 females of the control group were mated within 4 days. The number of pregnant females, the number of females with live born pups and the number of males that became sires amounted to 8, 9, 9 and 5 for the control, 40, 116.5 and 350 mg KBF4/kg body weight groups, respectively. The mating index was 100% in all KBF4-treated groups and 92% in the control group. The female fecundity index and female fertility index were comparable between the control, 40 and 116.5 mg KBF4/kg body weight groups and ranged from 67-75%. In the 350 mg KBF4/kg body weight group fecundity index and female fertility index was 42%.The gestation index was 100% in all groups. The duration of gestation was comparable between the groups. The number of corpora lutea and implantation sites was statistically significantly decreased in the 350 mg KBF4/kg body weight group; no effect was observed in the other KBF4-treated groups when compared to the control group. Pre- and post-implantation loss was comparable in all groups. The number of litters with live born pups was 8, 9, 9 and 5 for the control, 40, 116.5 and 350 mg KBF4/kg body weight groups, respectively. The number of live pups per litter on post natal day (PN) 1 in the control, 40, 116.5 and 350 mg KBF4/kg body weight groups was 11.9, 12.1, 11.8 and 9.4, respectively. Pup mortality on PN 4 was comparable in all groups except for the mid-dose group and amounted to 3, 13, 3, 29 (incidences 3.2, 12, 2.8 and 62%) in the control, 40, 116.5 and 350 mg KBF4/kg body weight groups, respectively; in the 350 mg KBF4/kg body weight group the mortality was statistically significantly increased. The death of the pups of dam C63 (10 pups) and C67 (11 pups) may be secondary to the death of the dams on PN 2 and 3, respectively. In the control, 40, 116,5 and 350 mg KBF4/kg body weight groups 0(8), 1(8), 0(9) and 3(2) litters, respectively were lost entirely between PN 1-4 (between brackets the number of litters with live pups on PN 4). The number of live pups per litter on PN 1 was 11.9, 12.1, 11.8 and 9.4 in the control, 40, 116.5 and 350 mg KBF4/kg body weight groups, respectively and on PN 4 the number of pups was 11.5, 12.0, 11.4 and 9.0 in the control, 40, 116.5 and 350 mg KBF4/kg body weight groups, respectively. The number of live pups per litter of the 350 mg KBF4/kg body weight group was statistically significantly decreased on PN 1. On PN 4 only pups of 2 litters of the mid-dose group were alive. No difference was observed in the sex ratio between the groups. No differences were observed on pup weight and pup weight changes on PN 1 and 4. On PN 1 the number of runts (pup weight less than mean pup weight of the control group minus 2 standard deviations) was statistically significantly increased in the 116.5 and 350 mg KBF4/kg body weight groups. In addition, the number of runts was statistically significantly increased in the 116.5 KBF4/kg body weight group on PN 4. Macroscopic observations in stillborn pups and pups that died between PN 1 -4 are did not reveal any treatment related abnormalities. At scheduled necropsy no treatment related gross changes were observed. Microscopic examination did not reveal treatment related histopathological changes in any of the sampled organs and tissues. Based on the effects on mortality, body weight and food consumption in the 350 mg KBF4 group/ kg body weight group, the NOAEL for maternal toxicity is 116.5 mg KBF4/kg body weight/day. The NOAEL for developmental toxicity is 40 mg KBF4/kg body weight based on the increased pup mortality (PN 1-4) in the 350 mg KBF4/kg body weight group and the increased number of runts in the 116.5 and 350 mg KBF4/kg body weight groups.