5-ethylidene-8,9,10-trinorborn-2-ene

Genetic toxicity: in vivo

Administrative data

Endpoint:

genetic toxicity in vivo

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Remarks:

Study conducted in an established laboratory according to the Rodent Dominant Lethal Assay Guideleines of the U.S. EPA (1982) and in full compliance with GLP.

Data source

Reference

Reference Type:

publication

Title:

Absence of dominant lethal effects in male CD® rats exposed to 5-ethylidene-2-norbornene vapor.

Author:

Neeper-Bradley, T. and Ballantyne, B.

Year:

1996

Bibliographic source:

Toxic Substance Mechanisms 15, 389-404.

Materials and methods

Principles of method if other than guideline:

This study design was based on guidelines supplied by the United States EPA for the "Rodent Dominant Lethal Assay" (1982). Other sources examined for development of the study design included Green et al (1976; 1985) and Ehling et a1. (1978).

GLP compliance:

yes

Type of assay:

rodent dominant lethal assay

Test material

Test material information

Constituent 1

Reference substance name:

5-ethylidene-8,9,10-trinorborn-2-ene

EC Number:

240-347-7

EC Name:

5-ethylidene-8,9,10-trinorborn-2-ene

Cas Number:

16219-75-3

Molecular formula:

C9H12

IUPAC Name:

5-ethylidenebicyclo[2.2.1]hept-2-ene

Details on test material:

Test substance: 5-Ethylidene-2-norbornene (CAS No. 16219-75-3)
Reference no 1-SCD-113.
Purity: 99%

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

All animals were assigned a unique number and identified by cage tags.

TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Portage, MI)
- Age at study initiation: 70 days of age
- Housing: During the acclimation period and prior to mating, animals were housed separated by sex, 1 male or 2 females to a cage, in stainless steel wire-mesh cages. After the exposure period, the males were housed in stainless steel wire-mesh cages with 2 females weekly, for mating. After successful mating, the females were housed individually.
- Diet: Ground, certified Rodent Diet (RMH 3200, Agway, Inc.) was available ad libitum, except during exposures.
- Water: Tap water was available ad libitum, except during exposures.
- Acclimation period: The acclimation period for each shipment of animals was approximately 2 weeks. During this period, the animals were weighed at least 2 times at scheduled intervals.

ENVIRONMENTAL CONDITIONS
- Temperature: 66-75°F.
- Humidity: 40-70%
- Photoperiod: 12-hour photoperiod (approximately 0500 to 1700 hours for the light phase).

Administration / exposure

Route of administration:

inhalation: vapour

Vehicle:

No

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Exposure cages were stainless steel with glass windows for observation and were approximately 1330 liters
- System of generating particulates/aerosols: Liquid ENB was metered from a piston or syringe pump into a heated glass evaporator (55-66 Celsius) similar in design to that described by Snellings and Dodd (1990). The resulting ENB vapor was carried into the exposure chamber by a countercurrent air stream that entered the bottom of the evaporator and flowed into the chamber from the top of the evaporator.
- Temperature: 21.8-22.6 Celsius.
- Humidity: 55.1-61.4%,
- Air flow rate: approximately 300 liters/minute
- Air change rate: 13 air changes each hour

Duration of treatment / exposure:

6 hours/day for 5 days

Frequency of treatment:

5 consecutive days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

5 ppm (analytical)

Doses / concentrations 2

Dose / conc.:

52 ppm (analytical)

Doses / concentrations 3

Dose / conc.:

254 ppm (analytical)

No. of animals per sex per dose:

20 exposed males per group
40 untreated females per group were used for each mating week

Control animals:

other: Yes, exposed to filtered air. Following the final exposure to filtered air, positive control males received a single intraperitoneal injection of 0.5 mg TEM/kg body weight.

Positive control(s):

Triethylenemelamine

Examinations

Evaluation criteria:

Reproductive/gestational parameters examined included: No. impregnated females/no. females paired (mating index, females):
No. impregnating males/no. males paired (mating index, males):
No. pregnant females/no. females impregnated (fertility index, females):
No. males producing pregnant females/no. males impregnating (fertility index, males):
No. corpora lutea/pregnant female:
No. total implantations/pregnant female:
Preimplantation loss/pregnant female (%):
No. nonviable implantations (early and late resorptions and
dead fetuses)/pregnant female:
Postimplantation loss/pregnant female (%):
No. females with 1 or more dead implantations (%):
No. females with 2 or more dead implantations (%):
Ratio of nonviable implantations/total implantations:
FL% (the dominant lethal factor) =[ 1 - live implantations/female of test group ] X 100 / live implantations/female of control group

Statistics:

The unit of comparison was the male or the pregnant female (Weil, 1970). Results of the quantitative continuous variables (for example, male body weights) were intercompared for the 3 ENB-treated groups and vehicle control group by use of Levene's test for equal variances (Levene, 1960), analysis of variance (ANOVA), and t-tests. The t-tests were used when the F value from the ANOVA was significant. When Levene's test indicated homogeneous variances, and the ANOVA was significant, the pooled t-test was used. When Levene's test indicated heterogeneous variances, all groups were compared by an ANOVA for unequal variances (Brown and Forsythe, 1970) followed, when necessary, by the separate variance t-test. The positive control group was compared to the vehicle control group using the previously described set of statistics.
Nonparametric data were examined statistically using the Kruskal-Wallis test (Sokal and Rohlf, 1969) followed by the Mann-Whitney U test (Sokal and Rohlf, 1969) when appropriate. Frequency data were compared using the Fisher's Exact Test (Sokal and Rohlf, 1969). For all statistical tests, the probability value of < 0.05 (two-tailed) was used as the critical level of significance.

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

yes

Remarks:

slight transient body weight effects at high dose

Negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

ENB did not produce evidence of dominant lethality in rats using a standard protocol (Neeper-Bradley and Ballantyne, 1996). Male rats exposed to analytically measured ENB vapor concentrations of 0, 5, 52, or 254 ppm for 6 hr/day for 5 consecutive days were mated with unexposed females.

Reproductive factors, including the number of fertile males and the number of gravid females with viable implants, were unaffected by exposure to ENB vapor. No significant preimplantation loss or dominant lethal effects were observed in females mated to ENB-exposed males during the 10-week breeding period. Males exposed to 254 ppm had slightly reduced body weight gains after the 5-day treatment period, but body weights and body weight gains were comparable to the controls thereafter. Triethylenemelamine (0.5 mg/kg, single dose on day 5 as a positive control) produced clear dominant lethal effects.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The ENB-exposed males showed no clinical signs of toxicity. The only objective finding was slight decrease in body weight gain at 254 ppm over the exposure period, but which resolved by the end of the first postexposure week. No reproductive or gestational effects, including dominant lethality, were noted in the ENB-treated groups during the 10 wk of postexposure mating. This finding confirms the absence of testicular toxicity in the rat from repeated exposure to high concentrations of ENB vapor.

Executive summary:

Neeper-Bradley and Ballantyne (1996) reported that ENB did not induce dominant lethal mutations in rats exposed to concentrations up to 254 ppm for 5 consecutive days (6 hr/day). The NOEC for toxicity (body weight gain) was 52 ppm. The NOEC for dominant lethality was greater than 254 ppm.