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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not specified
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
other: SIDS Dossier data
Title:
Unnamed
Year:
2001
Report Date:
2001

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
not specified
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Details on test material:
Trade Name: WINGSTAY 29
CAS No: 68442-68-2
EINECS No: 270-485-3
EINECS Name: Benzenamine, N-phenyl-, styrenated
CAS Name: Benzenamine, N-phenyl-, styrenated
Substance type: organic
Physical status: liquid
Purity: > 98 % w/w
Result: Molecular weight: 320

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male
Details on test animals and environmental conditions:
Not specified

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
Corn oil
Details on exposure:
Not applicable.
Duration of treatment / exposure:
single dose
Frequency of treatment:
Once
Post exposure period:
24 and 48 hours
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 500, 1000 and 2000 mg/kg
Basis:
actual ingested
No. of animals per sex per dose:
Five male mice per dose.
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide

Examinations

Tissues and cell types examined:
polychromaticerythrocytes (PCE) and nonchromatic erythrocytes (NCE)
Details of tissue and slide preparation:
Bone marrow cells were harvested 24 and 48 hours after dosing. All dose levels, the vehicle control and a positive control (Cyclophosphamide) were evaluated at the 24 hours. At 48 hours, only the vehicle control and high dose were evaluated.
Bone marrow was taken from the hind limbs. Slides were prepared from the bone marrow extracts, fixed with methanol and stained in May Grunwald Solution and Giemsa. Two thousand micronucleated polychromatic erythrocytes were evaluated for micronuclei.
Evaluation criteria:
The ratio of polychromatic erythrocytes (PCE) to nonchromatic erythrocytes (NCE) cells was determined from the first 500 erythrocytes on each slide.
Statistics:
Statistical analyses were performed using Analysis of Variance and Dunnett’s t-test.

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
not specified
Negative controls validity:
not specified
Positive controls validity:
not specified
Additional information on results:
The test substance did not produce any signs of clinical toxicity. Statistically lower PCE:NCE ratios, while not dose related, did strongly indicate that the test substance was cytotoxic to the bone marrow. The test substance did not produce any statistically significant increase in micronucleated PCEs relative to the vehicle control at the 24-hour and 48-hour harvest interval.
The positive control induced a statistically significant increase in micronucleated PCEs compared to the vehicle control.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
The test substance was tested up to the limit dose (2000 mg/kg) and did not cause chromosome damage in the mouse bone marrow micronucleus assay under the conditions of this test.
Executive summary:

Study conducted to OECD test guidelines in compliance with GLP. Data included for OECD SIDS dossier. The test substance did not produce any signs of clinical toxicity. Statistically lower PCE:NCE ratios, while not dose related, did strongly indicate that the test substance was cytotoxic to the bone marrow. The test substance did not produce any statistically significant increase in micronucleated PCEs relative to the vehicle control at the 24-hour and 48-hour harvest interval. The positive control induced a statistically significant increase in micronucleated PCEs compared to the vehical control. Result: The test substance was tested up to the limit dose (2000 mg/kg) and did not cause chromosome damage in the mouse bone marrow micronucleus assay under the conditions of this test.

Read across to supporting substance, CAS No. 68442 -68 -2, by structural analogue. This substance has been supported under Environmental Protection Agency’s (EPA’s) High Production Volume (HPV) Challenge Program. The American Chemical Councils RAPA Panel, has derived a “Substituted Diphenylamines” category of chemicals for this substance, please refer to EPA reference 201-14700A located at

 

http://www.epa.gov/hpv/pubs/summaries/subdipha/c13378rt.pdf

 

Relying on several factors specified in EPA’s guidance document on “Development of Chemical Categories in the HPV Challenge Program,” in which use of chemical categories is encouraged, the following closely related chemicals constitute a chemical category:

 

Structural Similarity. A key factor supporting the classification of these chemicals as a category is their structural similarity (see Figure 1). All share a common starting material; Diphenylamine (Benzenamine, N-phenyl-, CAS# 122-39-4), a common synthetic pathway, and all compounds in this category are diamines with various substitutions.

 

Similarity of Physicochemical Properties. The similarity of the physicochemical properties of these materials parallels their structural similarity. All are off-white to light brown solids or viscous liquids intended for use as antioxidants in finished rubber articles or as antidegradant additives that extend the useful life of heavy-duty industrial functional fluids used in high-speed, high-temperature and/or high-load applications. As a class, these amine-based antidegradant compounds are less migratory (more polymer-bound) and less staining than the Substituted p-Phenylenediamine antidegradants. The use of these materials requires that they be stable under high temperatures. Their low volatility is due to their low vapor pressure, highly viscous or solid form. The existing information for these materials indicates that they have low water solubility and high flash points.

 

Toxicological Similarity. Review of existing published and unpublished test data for Substituted Diphenylamines shows the aquatic and mammalian toxicity among the materials within this category are similar.

 

Mammalian Toxicology - Mutagenicity. Data from bacterial reverse mutation assays, in vitro and in vivo chromosome aberration studies, as well as additional supporting in vitro and in vivo genetic toxicity studies were reviewed, and the findings indicate a low concern for mutagenicity either for aryl or alkyl substituted materials. Similarly, the data for a mixed aryl/alkyl substituted molecule also indicates a lack of mutagenicity. Data are available for several members of the category or close structural analogs, and these data can be bridged to the other members of the category. Therefore, for the purposes of the HPV Program, the category has been adequately tested for mutagenicity, and no additional mutagenicity testing is proposed.

 

Conclusion. Based upon the data reviewed in “Substituted Diphenylamines” category of chemicals, the physicochemical and toxicological properties of the Substituted Diphenylamine category members are similar and follow a regular pattern as a result of that structural similarity. Therefore, the definition of a chemical category has been met, and read across is considered appropriate for the category of chemical.