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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 December 1998 - 28 December 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD Guideline study under GLP conditions
Cross-reference
Reason / purpose for cross-reference:
reference to same study
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
10 December 1998 - 28 December 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD Guideline study under GLP conditions
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
- Physical state: colourless liquid with fragrant odor
- Purity: 100.0%
- Lot/batch No.: ACM1865, ACL2024, (Wako Pure Chemical Industries, Ltd.)
- Storage condition of test material: room-temperature, dark place
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Source: Charles River Laboratories Japan, Inc.
- Age at study initiation: 8 weeks old
- Weight at study initiation: male: 302 - 344 g, female: 207 - 248 g
- Acclimation period: 2 weeks
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatography
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
male: 42 days
female: 35-41 days
Frequency of treatment:
6 hours daily
Remarks:
Doses / Concentrations:
0, 150, 300, 600, 1200 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 150.4, 300.3, 599.9, 1199.7 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
10 rats/sex/dose
Control animals:
yes, concurrent vehicle
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly. Mated females: day 0, 7, 14 and 20 of pregnancy. Day 0 and 4 of postpartum. Scheduled sacrifice day.

FOOD CONSUMPTION: Yes

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Scheduled sacrifice day.
- Anaesthetic used for blood collection: Yes (identity)
- Animals fasted: Yes(male)
- Parameters: RBC, hemoglobin, hematocrit, MCV, MCH, MCHC, platelet, WBC, differential WBC

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Scheduled sacrifice day.
- Anaesthetic used for blood collection: Yes (identity)
- Animals fasted: Yes(male)
- Parameters: total protein, albumin, A/G ratio, T-bilirubin, glucose, T-cholesterol, triglyceride (male), phospholipid, GOT, GPT, LDH, ALP (male), γ-GTP, CPK, urea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus


URINALYSIS: Yes
- Time schedule for collection of urine: final week of dosing period
- Metabolism cages used for collection of urine: No data
- Parameters: pH, protein, glucose, ketone body, bilirubin, occult blood, urobilinogen
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
- Examined organs: skin, nasal cavity, nasopharynx, larynx, trachea, lung, bone marrow, lymph node, thymus, spleen, heart, tongue, salivary gland, esophagus, stomach, small intestine, large intestine, liver, pancreas, kidney, urinary bladder, pituitary gland, thyroid, parathyroid gland, adrenal gland, testis, epididymis, seminal vesicle, prostate, ovary, mammary gland, brain, spinal cord, peripheral nerve, eyeball, harderian gland, muscle, bone.
Other examinations:
Organ weight:
- Weighed organs: thymus, adrenal gland, testis, ovary, heart, lung, kidney, spleen, liver, brain, epididymis, seminal vesicle, prostate, uterus, pituitary gland.
Statistics:
chi-square test, Bartlett test, one-way analysis of variance, multiple analysis of Dunnett, Kruskal-Wallis rank test.
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY:
Five males (1 animal on week 1 of dosing, 3 animals on week 2, 1 animal on week 6 ) and 2 females (1 animal on week 2 of dosing, 1 animal on week 6 ) died in the 1200 ppm group.
Piloerection at one animal on week 1 of dosing, and piloerection, soiled perigenitalia due to urine and salivation at one animal on week 2 of dosing were observed in the 1200 ppm group.

ORGAN WEIGHTS:
The absolute and relative spleen weights and the relative heart weight increased statistically in surviving males received 1200 ppm.

GROSS PATHOLOGY:
In dead males received 1200 ppm, red zone of lung, atrophic thymus, and enlargement, pale and white zone of heart were observed.
In dead females received 1200 ppm, atrophic thymus, and enlarged heart were observed.
In surviving animals, no treatment-related changes were found in any treatment groups containing the 1200 ppm group.

HISTOPATHOLOGY:
In dead animals received 1200 ppm, congestion, edema and thrombus of lung, atrophy and karyorrhexis of thymus, congestion of bone marrow, atrophy and deposit of hemosiderin of spleen were found.
In surviving animals, no treatment-related changes were found in any treatment groups containing the 1200 ppm group.
Dose descriptor:
NOEC
Effect level:
600 ppm
Sex:
male/female
Basis for effect level:
other: Deaths occurred in both sexes of the 1200 ppm group.
Critical effects observed:
not specified
Executive summary:

JBRC of Japan (2000) reported a guideline (OECD 422) and GLP study of repeated acetonitrile exposure in rats. Animals were exposed to acetonitrile vapor by whole-body inhalation 6 hr /day for 42 days (males) or 35 -41 days (females). Evaluations included clinical signs, body weights, hematology, clinical chemistry, urinalysis, organ weights, and gross and microscopic pathology. The NOEL in this study was considered to be 600 ppm for both sexes based on mortality (5M, 2F) in the 1200 ppm exposure group.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report date:
2000

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Acetonitrile
EC Number:
200-835-2
EC Name:
Acetonitrile
Cas Number:
75-05-8
Molecular formula:
C2H3N
IUPAC Name:
acetonitrile
Details on test material:
- Physical state: colourless liquid with fragrant odor
- Purity: 100.0%
- Lot/batch No.: ACM1865, ACL2024, (Wako Pure Chemical Industries, Ltd.)
- Storage condition of test material: room-temperature, dark place
Specific details on test material used for the study:
- Physical state: colourless liquid with fragrant odor
- Purity: 100.0%
- Lot/batch No.: ACM1865, ACL2024, (Wako Pure Chemical Industries, Ltd.)
- Storage condition of test material: room-temperature, dark place

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc.
- Age at study initiation: (P) x 8 weeks
- Weight at study initiation: (P) Males: 302 - 344 g; Females: 207 - 248 g
- Acclimation period: 2 weeks


Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatography
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: up to 9 days
- Proof of pregnancy: vaginal plug or sperm in vaginal smear (day 0 of pregnancy)
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
male: 42 days
female: 35-41 days
Frequency of treatment:
6 hours daily
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 150, 300, 600, 1200 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 150.4, 300.3, 599.9, 1199.7 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
10 rats/sex/dose
Control animals:
yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly. Mated females: day 0, 7, 14 and 20 of pregnancy. Day 0 and 4 of postpartum. Scheduled sacrifice day.

FOOD CONSUMPTION: Yes
Oestrous cyclicity (parental animals):
From the start day of acclimation period until the day of successful copulation.
Sperm parameters (parental animals):
Parameters examined in male parental generations:
sperm count, sperm motility, sperm morphology in epididymis.
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight, physical or behavioural abnormalities


GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals at the scheduled sacrifice day in the combined general toxicity study.
- Maternal animals: All surviving animals on day 4 of postpartum.


GROSS NECROPSY
- Gross necropsy performed in the combined general toxicity study.


HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues examination performed in the combined general toxicity study.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring on day 4 of postpartum was subjected to postmortem macroscopic examination.


GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
Statistics:
chi-square test, Bartlett test, one-way analysis of variance, multiple analysis of Dunnett, Kruskal-Wallis rank test
Reproductive indices:
Gestational period, gestation rate, implantation index, delivery index, birth index, live birth index, sex ratio.
Offspring viability indices:
viability index=(number of pups alive on day 4/number of pups alive on day 0) × 100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
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:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
effects observed, treatment-related
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
See the part of repeated dose general toxicology. (7.5.3 Repeated dose toxicity: inhalation)

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
Prolonged estrus stages were found in 3 females of the 1200 ppm group.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Fertility rates decreased slightly in the 1200 ppm group without statistically significance.

ORGAN WEIGHTS (PARENTAL ANIMALS)
See the part of repeated dose general toxicology. (7.5.3 Repeated dose toxicity: inhalation)

GROSS PATHOLOGY (PARENTAL ANIMALS)
See the part of repeated dose general toxicology. (7.5.3 Repeated dose toxicity: inhalation)

HISTOPATHOLOGY (PARENTAL ANIMALS)
See the part of repeated dose general toxicology. (7.5.3 Repeated dose toxicity: inhalation)

Effect levels (P0)

Dose descriptor:
NOEC
Effect level:
600 ppm
Sex:
male/female
Basis for effect level:
other: In the 1200 ppm groups, fertility rate was slightly low , and estrous cycles changed in some animals. Mortality also occurred.

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 200 ppm
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Target system / organ toxicity (F1)

Key result
Critical effects observed:
no

Overall reproductive toxicity

Key result
Reproductive effects observed:
yes
Lowest effective dose / conc.:
600 ppm
Treatment related:
yes
Relation to other toxic effects:
reproductive effects as a secondary non-specific consequence of other toxic effects
Dose response relationship:
yes
Relevant for humans:
not specified

Applicant's summary and conclusion

Executive summary:

The IPH of Japan (1980) has reported results of a guideline (OECD 422) and GLP reproduction /developmental toxicity screening test of acetonitrile in rats. Groups of 10 males and 10 females were exposed to acetonitrile vapor in whole-body inhalation chambers for 6 hours daily for 6 weeks (42 days) for males (from 2 weeks before mating until 2 weeks after the end of the mating period), and for 35 -41 days for females (from 2 weeks before mating until day 19 of pregnancy) at concentrations of 0, 150, 300, 600 or 1200 ppm. In the 1200 ppm group, the fertility rate was slightly lower than controls, and the changes of estrous cycles were found in some animals in the study. Mortality also occurred at the 1200 ppm exposure level. Based on these results the NOECs for systemic toxicity and reproductive toxicity were considered to be 600 ppm in both sexes.