Adiponitrile

Toxicity to reproduction

Administrative data

Endpoint:

fertility, other

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

September 21, 1981 through November 5, 1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study was not conducted according to guidelines but was conducted according to GLPs and the report contains sufficient data for interpretation of study results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

Female rats were exposed to adiponitrile by inhalation for 6 hours per day, 7 days per week, for 22 days. The females were then mated to untreated males. Exposures contined until copulation occured with a maximum of 33 days exposure time. Females were sacrificed on gestation day 13. Tissues and organs were examined for gross lesions, pregnancy status, nidation sites, and corpora lutea.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Adiponitrile

EC Number:

203-896-3

EC Name:

Adiponitrile

Cas Number:

111-69-3

Molecular formula:

C6H8N2

IUPAC Name:

hexanedinitrile

Details on test material:

As cited in the study report: The test material used in this study, adiponitrile (Lot No. 40, Environmental Health Laboratory sample T810038), was supplied by the sponsor and was indicated to have a purity of about 99%. Each batch of test material was analyzed before use in the exposure chambers. These analyses indicated no significant decomposition of the test material during the course of the study.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

As cited in the study report: Sprague-Dawley male and virgin female rats (Crl:CD(SD)BR) were purchased from Charles River Breeding Laboratories (Portage, Michigan) for this study. Animals were shipped by motor carrier and received on the day of shipment. Female rats were 60 days of age at receipt with body weights (measured on 10 females) ranging from 155 to 187 g. Male rats were 71 days of age at receipt with body weights (measured on 10 males) ranging from 257-323 g. Rats were quarantined for one week and examined for general health status and the presence of pinworms and ectoparasites. No significant health problems were noted during quarantine and the animals were released for study. Female rats were first exposed at the age of 73 days and male rats were mated at the age of 83 days. All animals were uniquely identified by a metal eartag and a bar-coded cage card. Males and females were individually caged (except during mating) in suspended stainless-steel wire cages. During exposure, females were caged in open-mesh inhalation style cages. Food (Purina Certified Rodent Chow No. 5002) and water (City of St. Louis drinking water treated with an ion-exchange water softener) were available ad libitum except food and water were not available for females during inhalation exposure. The animal room environment was routinely maintained at 72 +/- 2F, 40-60% relative humdiity and with a 12 hr/12 hr. dark light cycle. Although there were some excursions outside of the nominal temperature and humidity ranges, these excursions were minor in nature and were not considered to significantly affect the study.

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: conditioned air

Details on exposure:

As cited in the study report: Exposures were conducted in 10 m3 Rochester-style stainless steel and glass inhalation chambers for 6 hours per day, seven days per week. Atmospheres of adiponitrile were produced by metering the liquid through a Laskin-style nebulizer. Observations made during exposures included nominal concentration measurements, particle-size measurements, temperature and humidity measurements and observation of animals during exposure for signs of toxicity.

Details on mating procedure:

As cited in the study report: Two females were assigned, on a random basis, to an untreated male of the corresponding group. Females were caged with the assigned male consecutively (one male and female per cage) until confirmed copulation was observed or five nights occurred without confirmed copulation. Each morning the area under the cage and the female were examined for the presence of a copulatory plug. If the location of a copulatory plug was ambiguous, vaginal smears of appropriate females were taken and examined for the presence of sperm. The day on which confirmed copulation was observed was considered gestation day 0 and females with confirmed copulation were returned to individual caging conditions. Females which failed to mate with the initially assigned male were assigned, on a random basis, to another male which had copulated with another female in the same group. Such females were co-housed with the second assigned male until copulation was confirmed or 7 nights had elapsed without confirmed copulation. An exception to this procedure (and protocol deviation) occurred for one female (810066-1F006) which was co-housed with the second assigned male for only 6 nights because there were no remaining unmated females (including control group animals) to be exposed on the seventh day.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

As cited in the study report: Concentrations of adiponitrile in the exposure chambers were measured by gas chromatographic analysis of impinger-collected samples of chamber atmosphere. Data from studies conducted in the same exposure chambers (EHL Studies 810064 and 810065) had already established that one impinger was sufficient for quantitative recovery of adiponitrile. The frequency of sampling for analytical determination of adiponitrile was two samples per day for the low and mid exposure levels. Because of decreased precision, the number of analytical determinations was increased from two to four times per day on study day 5 (exposure day 5) through the end of the study for the high exposure level.

Duration of treatment / exposure:

Females (P): 6 hours / day before mating

Frequency of treatment:

7 days / week for 22 days

Details on study schedule:

As cited in the study report: Twenty-four females were assigned on a random basis (with equivalent body weight distribution) to each treatment group or the control group. The target exposure levels were 13 mg/m3 (low exposure level group), 30 mg/m3 (mid exposure level group) and 100 mg/m3 (high exposure level group). The control group females were handled by the same inhalation procedures as the treatment groups except that these animals were only exposed to house supply air containing no adiponitrile. Fifteen males were nominally assigned, on a random basis to give equivalent body weight distribution, to each treatment or control group. Males were not exposed to adiponitrile in inhalation chambers. After sufficient time on study to cover three to four estrus cycles of the rat (22 exposure days, 22 days on study), females were mated to males. Exposure of females continued until copulation was confirmed (maximum exposure of 33 days for some females).

Doses / concentrations

Remarks:

Doses / Concentrations:
13.0, 31.8, and 104 mg/m3
Basis:
analytical conc.

No. of animals per sex per dose:

24 female animals per dose

Control animals:

yes, concurrent vehicle

Positive control:

no

Examinations

Parental animals: Observations and examinations:

Males and females (after assignment to the study) were weighed once per week. Mated females were weighed on gestation days 0 and 13. Females were given a thorough physical examination once per week and were observed for clinical signs before and after exposures. All animals were checked twice daily for mortality and gross abnormalities. Vaginal smears were taken on 5 consecutive days for unmated females.

Oestrous cyclicity (parental animals):

Determined by vaginal smears.

Sperm parameters (parental animals):

not determined

Litter observations:

not determined

Postmortem examinations (parental animals):

As cited in study report: Females of each treatment or control group were sacrificed on gestation day 13 or the nearest normal working day after gestation day 13 (up to gestation day 15). Females without confirmed copulation were sacrificed in the second week after the last day of co-housing. Males were sacrificed after mating and no necropsies were performed on males. Each female was given an external examination and sacrificed by exposure to chloroform. The tissues and organs of the thoracic and abdominal cavities were examined for gross lesions, pregnancy status was determined and, for pregnant females, nidation sites were classified and counted and corpora lutea were counted.

Postmortem examinations (offspring):

not determined

Statistics:

As cited in study report: Data for body weights were analyzed using Dunnett's test. Counted data was analyzed, when appropriate, by the Mann-Whitney U test, and data which measured a response or lack of response (mating efficiency, pregnancy rate) were analyzed, when appropriate, by Fisher's exact test, and the uncorrected chi-square test. A critical value of p=0.05 was used and the Bonferroni inequality was assumed for comparison of multiple treatments with control values for all tests except Dunnett's test.

Reproductive indices:

Mating efficiciency, pregnancy rate, number of live implants, pre- and postimplantation loss rates

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): The only finding that appeared to be treatment-related was reddish-brown fur discoloration of the head, neck and shoulders region, which was observed in several females in the high exposure level group in the third week of exposure. Although this finding could not be explained, it did not appear to indicate toxic effects.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS): Mean body weights of females in the high exposure level group were statistically different from the control values in the second and third exposure weeks. However, these effects were very small in magnitude, with mean weights being 3 to 4% lower than mean control group weights.

Effect levels (P0)

Dose descriptor:

NOAEC

Effect level:

104 mg/m³ air (analytical)

Sex:

female

Basis for effect level:

other: Highest dose tested. Based on pregnancy rate, mating efficiency, number of live implants, pre- and postimplantation loss rates

Results: F1 generation

General toxicity (F1)

Clinical signs:

not examined

Mortality / viability:

not examined

Body weight and weight changes:

not examined

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

Overall reproductive toxicity

Reproductive effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

There were no significant detectable female fertility effects at any of the exposure levels tested as judged by mating efficiency, pregnancy rates, number of live implants or pre- or postimplantation loss. The fertility NOAEC was the highest concentration tested, 104 mg/m3 air. There was a slightly statistically significant reduction in mean body weights of the females during exposure weeks 2 and 3. Because this effect was observed prior to mating, the exposure concentration of 104 mg/m3 could be considered a maternal systemic LOAEC with no relation to fertility effects.

Executive summary:

Female Sprague-Dawley rats were exposed by the inhalation route (6 hrs/day, 7 days/week) to adiponitrile at target exposure levels of 13 mg/m3, 30 mg/m3 and 100 mg/m3 for 22 exposure days (22 days on study) and mated to untreated males to assess female fertility. Exposure of the females was continued until the day of mating and females were sacrificed at mid-gestation (gestation days 13-15). Mean daily analytical exposure levels (13.0 mg/m3, 31.8 mg/m3 and 104 mg/m3) were within 6% of the target levels. Under the conditions of this study no toxic effects were observed in the females of any exposure group other than slightly lower body weights of high exposure level females in the second and third weeks of exposure. There were no treatment related adverse clinical signs or lesions detected at gross necropsy. There were also no significant, detectable effects on female fertility at any of the exposure levels tested. Treatment group females were comparable to control group females in mating efficiency, pregnancy rate, number of live implants and pre- and post implantation loss rates.