Isoprene

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:

other: GLP compliant, guideline study, published in peer reviewed literature, no restrictions, fully adequate for assessment.

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1989

Reference 2

Reference Type:

publication

Title:

Inhalation developmental toxicity of isoprene in mice and rats.

Author:

Mast TJ et al

Year:

1990

Bibliographic source:

Toxicologist 10(1):42 (abstract)

Reference 3

Reference Type:

publication

Title:

Unnamed

Year:

1995

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Isoprene

EC Number:

201-143-3

EC Name:

Isoprene

Cas Number:

78-79-5

Molecular formula:

C5H8

IUPAC Name:

2-methylbuta-1,3-diene

Details on test material:

- Name of test material (as cited in study report): isoprene
- Analytical purity: >99.7%

Test animals

Species:

mouse

Strain:

CD-1

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Portage, MI, USA
- Age at study initiation: 12 week
- Housing: Virgin females individually caged 8 days prior to exposure, mated females individually caged on 0 dg
- Diet: pelleted NIH-07 diet (Ziegler Bros ., Inc ., Gardner, PA, USA) ad libitum (except during exposure)
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature: 75±3°F
- Humidity: 55±15%
- Air changes (per hr): no data
- Photoperiod: 12 hrs dark /12 hrs light

IN-LIFE DATES: no data

Administration / exposure

Route of administration:

inhalation: vapour

Vehicle:

other: air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 2 .3 m3 stainless-steel chamber (1 .7 m3 active mixing volume) contained 3 levels of caging, each of which was split into 2 offset tiers. The cage units accommodated individual animal cages, feed troughs and automatic water dispensers. Stainless-steel catch pans designed to aid in maintaining a uniform concentration of vapour throughout the chamber, as well as for the collection of urine and faeces, were suspended below each cage unit.
- Rate of air: 15 ft3/min. The uniform mixture was diverted along the inner surfaces of the chamber and the catch pans created mixing eddies
- Method of conditioning air: HEPA- and charcoal-filtered.
- Isoprene vapour was generated by a rotary evaporation system which assured uniformity of chemical-laden atmosphere within the chamber.
- Temperature and humidity in air chamber: 75±3°F, 55±15%
- Air flow rate: 12 to 18 CFM

TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatography

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentrations of isoprene vapour in the chambers were monitored at 20-30 min intervals by gas chromatography (minimum detectable limit = 0.04 ppm isoprene). The grand means of chamber concentrations for all exposure levels were 100% of the target with relative SDs of 2 to 6%. At least 99% of all individual concentration measurements were within ±10% of the specified operating limits for the exposure target level. There was no measurable concentration in the control and holding chambers. The maximum concentration observed in the room was 0 .8 ppm.

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 2-3 females/male
- Length of cohabitation: overnight
- Further matings: none
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug referred to as day 0 of gestation (0dg)

Duration of treatment / exposure:

12 consecutive days. gestation days 6-17

Frequency of treatment:

6 hours/day

Duration of test:

days 0-18 of gestation

No. of animals per sex per dose:

10 virgin females/group and 33 positively mated females /group.

Control animals:

yes, sham-exposed

Details on study design:

Virgin mice were exposed for 12 consecutive days concurrently with mated animals and were killed 1 day after their last exposure day.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Mated mice 0, 6, 9, 12, 15 and 18 dg. Virgin mice 3 days prior to exposure and on exposure days 1, 5, 10 and at termination.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 18 mated mice, 1 day after final exposure virgin mice
- Organs weighed: liver and kidneys
- Organs examined: ovaries, uterus

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No data (ovaries examined at another laboratory and results not included in this publication)
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- Live foetuses were weighed examined for gross defects, and their sex was determined by internal examination of the gonads
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter plus any with gross external abnormalities]
- Skeletal examinations: Yes: [all per litter]
- Head examinations: Yes: [half per litter]

Statistics:

All means and SDs for animal data were calculated with SAS statistical software. Mean body weights (as a mean of litter means for foetal data) were analyzed with an analysis of variance (ANOVA) model for unbalanced data. Response variables, either body weight or the arcsin transformations of proportional incidence data, were analyzed against the class variable, "treatment", in a one-way ANOVA model. A Tukey's t-test (two-tailed) was used to assess statistically significant differences between control and exposed groups. If appropriate, the dose-response relationship was determined by means of an orthogonal trend test. In the case of proportional data the t-tests and trend analyses were performed on transformed variables. The litter was used as the basis for analysis of foetal variables.

Historical control data:

reported

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
There were statistically significant reductions in maternal body weight and body weight gain and, in uterine weight at the highest dose. Liver to body weight ratios for pregnant dams were significantly increased in the 1400 and 7000 ppm groups compared to the control group, and kidney to body weight ratios were significantly increased in the 7000 ppm group.

Effect levels (maternal animals)

Dose descriptor:

NOAEC

Effect level:

3 900 mg/m³ air (nominal)

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
There was an exposure-correlated reduction in foetal body weight (statistically significant for females only at 780 mg/m3, for males only at 3900 mg/m3 and for males and females at 19,503 mg/m3). There was no increase in the incidence of foetal malformations although two foetuses with cleft palate were found, one in each of the two highest exposure groups (1400 and 7000 ppm). An increased incidence of supernumerary ribs was observed in the exposed groups. This skeletal variation is generally considered to be a secondary effect of maternal toxicity or stress and its significance is unclear. The incidence of supernumerary ribs (percent of foetuses examined) was 20.1, 23.8, 33.6, and 40.3% at 0, 780, 3900, and 19,503 mg/m3.

Effect levels (fetuses)

1

Dose descriptor:

NOAEC

Effect level:

19 503 mg/m³ air (nominal)

Basis for effect level:

other: teratogenicity

2

Key result

Dose descriptor:

LOAEC

Effect level:

780 mg/m³ air (nominal)

Basis for effect level:

other: Developmental Toxicity. statistically significant differences in foetal weight in the absence of maternal toxicity; no NOAEC

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The NOAEC for maternal toxicity was 3900 mg/m3 (1400ppm). The NOAEC for teratogenicity was 19,503 mg/m3 (7000 ppm). The NOAEC for developmental toxicity based on foetal weight effect was not established and is therefore <780 mg/m3 (<280 ppm).

Executive summary:

Exposure of Swiss (CD-1) mice to 0, 780, 3900 or 19,503 mg/m³ (0, 280, 1400 or 7000 ppm) isoprene resulted in significant reductions in maternal body weight and uterine weight at the highest exposure. There was no NOAEC for developmental toxicity, which was evident as an exposure-correlated reduction in foetal body weight (statistically significant for females only at 780 mg/m³ , for males only at 3900 mg/m³ and for males and females at 19,503 mg/m³). The incidence of foetuses with supernumerary ribs was statistically significant at 3900 mg/m³. There was no significant increase in the incidence of malformations. Two foetuses with cleft palate were observed, one in each of the two highest exposure groups. This low incidence of a relatively common finding in mice, is considered not to be indicative of teratogenicity / developmental toxicity.