Reaction mass of Potassium aluminium tetrafluoride and Dipotassium aluminium pentafluoride monohydrate

Developmental toxicity / teratogenicity

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPP 83-3 (Prenatal Developmental Toxicity Study)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Test material form:

solid: crystalline

Details on test material:

Supplier: Atochem North America, Bryan, Texas
Test material: Kryocide
Common Name: Cryolite
Purity: 97.3%
Batch No.: 86-12
Description: Odorless white crystalline powder
Receipt date: September 12, 1990
Amount: 1 kg
Stability: The test material was considered stable when stored in a sealed container at room temperature

Test animals

Species:

mouse

Strain:

other: Crl:CD-1 (ICR) BR

Details on test animals or test system and environmental conditions:

One hundred fifty four sexually mature, virgin female mice, CrI:CD-1. (ICR) BR, were received from Charles River Breeding Laboratories, Inc., Montreal, Quebec, on June 11, 1991. The animals were approximately 10 weeks old upon receipt. Upon arrival and until pairing, all animals were individually housed in clean, wire-mesh cages suspended above cage-board. Body weights ranged from 18.3 to 32.8 g. All animals were housed for 16 days for acclimation purposes.

The basal diet used in this study was Purina Certified Rodent Chow #5002. Drinking water delivered by an automatic watering system and the feed were provided ad libitum throughout the acclimation period and during the study.

All animals were housed throughout the acclimation period and during the study in an environmentally-controlled room. Controls were set to maintain temperature at 72° + 3°F and relative humidity at 40-80%. Room temperature and relative humidity were recorded daily. Temperature ranged from 68°F to 72°F and relative humidity ranged from 46% to 70% during the study period. The single variation from the set temperature level did not apparently affect the outcome of the study. Light timers were calibrated to provide a 12-hour light/12-hour dark photoperiod. Air handling units were set to provide approximately 10 fresh air changes per hour.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 0.5% aqueous methylcellulose

Details on exposure:

The test mixtures were administered orally by gavage, via a stainless steel gavage cannula, once daily for 10 consecutive days initiating on gestation day 6 and continuing up to and including day 15 of gestation. A dosage volume of 5 ml/kg was used for all dosage levels. The control animals received 0.5% aqueous methylcellulose on a comparable regimen of 5 ml/kg. Individual dosages were based on the most recently recorded body weights to provide the correct mg/kg dose.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples of dosing preparations from each group, including the control group, were analyzed using a dissolution/ion-selective electrode method to determine homogeneity, 8-day stability and concentration. The suspensions were homogeneous, contained the proper amount of test material and were stable for eight days under refrigeration.

Details on mating procedure:

At the conclusion of the acclimation period, all available females were weighed and examined in detail for physical abnormalities. At the discretion of the study director, animals judged to be in good health and meeting acceptable body weight requirements (24.0 to 32.0 g) were placed in suspended wire-mesh cages with a resident male. The resident males were untreated, sexually mature mice utilized exclusively for breeding. These CrI:CD 1(ICR)BR male mice were also received from Charles River Breeding Laboratories, Inc., Montreal, Quebec, on June 11, 1991, and were the same age as the females (approximately 10 weeks old). The animals were paired on a 1:1 basis. The male mice were maintained under similar laboratory conditions as the females. The selected females were approximately 12 weeks old when paired for breeding.

Positive evidence of mating was confirmed by the presence of a copulatory plug in the vagina. Each mating pair was examined daily. The day on which evidence of mating was identified was termed day 0 of gestation and the animals were separated.

Duration of treatment / exposure:

from gestation days 6 through 15

Frequency of treatment:

once daily

Duration of test:

up to gestation day 18

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

30 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day

No. of animals per sex per dose:

25

Control animals:

yes, concurrent vehicle

Details on study design:

The experimental design for WlL-160005 consisted of three Kryocide treated groups and one control group. The bred females were consecutively assigned in a block design to groups containing 25 mice each by the following randomization procedure. The first mated female and the appropriate gestation day 0 designation were entered on the form and the female was assigned to group 1, the second mated female was assigned to group 2, and the third to group 3, etc. This process was continued daily until 25 females had been placed into each group. Surplus females were returned to individual wire-mesh cages for scheduled euthanization.
All mice were observed twice daily for moribundity and mortality. Individual detailed clinical findings, as appropriate, were recorded from days 0 through 18 of gestation (prior to compound administration during the dosing period). Animals were also observed for signs of toxicity approximately one hour following treatment during the dosing period. In addition, significant clinical findings were also observed and recorded for single animals on two days at the time of dosing. Animals aborted and animals moribund were euthanized at the study directors request, necropsied and the findings recorded. Females that delivered on gestation day 18 were necropsied. Because gestation day 18 is the normal length of gestation for this species, fetuses from dams that delivered on gestation day 18 were processed as described for those examined after Cesarean section if all implantation sites were accounted for.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: From days 0 through 18 of gestation

BODY WEIGHT: Yes
- Time schedule for examinations: Gestation days 0, 6-16 and 18

FOOD CONSUMPTION: Yes
- Time schedule: From days 0 through 18 of gestation

WATER CONSUMPTION: No data

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 18

Ovaries and uterine content:

On day 18 of gestation, all surviving females were euthanized for a scheduled Cesarean section. The uteri and ovaries were examined and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Mean gravid uterine weights and net body weight changes were calculated for each group.

Fetal examinations:

On day 18 of gestation, atl surviving females were euthanized for a scheduled Cesarean section. Fetuses were weighed, sexed and examined for external, skeletal and soft tissue malformations and developmental variations.

Statistics:

All analyses were conducted using two-tailed tests for a minimum significance level of 5% comparing each treated group to the vehicle control group. Each mean was presented with the standard deviation (S.D.) and the number animals (N) used to calculate the mean. All statistical tests were performed by a Digital Microvax 3400 computer with appropriate programming.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Maternal toxicity was expressed by two mortalities in the 300 mg/kg bw/day group. Another animal from this group was euthanized at the study directorts request. One mortality in the 100 mg/kg bw/day group was the only other potential maternal effect in the study. Clinical signs were observed in the animal euthanized in the 300 mg/kg bw/day group and in the animal found dead in the 100 mg/kg bw/day group. Necropsy findings in these animals were limited to reddened mucosa of the glandular portion of the stomach, dark red areas of the glandular portion of the stomach, pale liver and dark red contents in the stomach. Clinical findings in animals surviving to the scheduled necropsy could not be attributed to test article administration. No adverse effects were apparent on body weight gain and food consumption at any dose level.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

30 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Although there was no indication that embryonic survivability or fetal growth were affected at this dose level, anatomical development was apparently affected; s notably increased (although not statistically significant) incidence of the fetal variant bent ribs was expressed at this dose level. In conjunction, two fetuses in two separate litters had bent limb bones, a limited and non statistically significant incidence of a fetal malformation. Neither bent ribs nor bent limb bones have occurred in control mice in this laboratory. However, abnormal morphology of the limb bones was not reproduced in a previous developmental toxicity study with the substance. A single maternal mortality in the 100 mg/kg bw/day group was the only indication of potential maternal toxicity at this dose level. However, because fetal anatomical development, survival and fetal growth were not affected at this dose level, 100 mg/kg/day was considered to be the NOAEL (no observable adverse effect level) for developmental toxicity. The 30 mg/kg bw/day dose level was considered to be the NOAEL for maternal toxicity.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day

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

Executive summary:

Trisodium hexafluoroaluminate (cryolite) was investigated for prenatal developmental toxicity in mice in a GLP compliant study according to EPA Guideline 83 -3. Cryolite was administered by oral gavage to female CD-1 mice (25/group) at dose levels of 0, 30, 100 or 300 mg/kg bw/day once daily from gestation days 6 to 15 (WIL Research Laboratories, 1991). There was increased mortality at 300 mg/kg bw/day. The glandular portion of the stomach was red beginning at 100 mg/kg bw/day. In addition, females in the 300 mg/kg bw/day group exhibited dark red contents of the stomach. Fetuses at 300 mg/kg bw/day exhibited bent ribs and bent limb bones. The study reveals severe maternal toxicity in terms of mortality and signs of toxicity in the gastrointestinal tract induced at dosages of >100 mg cryolite/kg bw/day leading to derivation of a NOAEL for maternal toxicity of 30 mg cryolite/kg bw/day.

Based on the observation of skeletal anomalies at the dose level of 300 mg cryolite/kg bw/day also a NOAEL for developmental toxicity of 100 mg cryolite/kg bw/day can be derived from the study. As these anomalies were only reported at dose levels showing severe maternal toxicity, the effects are not considered to be indicative for a substance specific teratogenic potential of cryolite.