Menthane, monohydroperoxy derivative

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vivo

Description of key information

In vitro assays The test substance is tested in an Ames test in Salmonella typhimuriun TA 100, TA1535, TA1537, TA97 and TA98 with and without metabolic activation. The test substance did not induce mutations in Salmonella typhimurium strains without metabolic activation, but with metabolic activation the results of the Salmonella typhimurium assay with test substance were ambiguous (Zeiger 1988). In an additional study (Dillon 1998) in Salmonella typhimurium TA100, TA102 and TA104, in the presence and absence of metabolic activation, cumene hydroperoxide was weakly mutagenic. A chromosome aberration test (TC 2012) with the test substance showed no clastogenic effects with and without metabolic activation in V79 cells The (slightly) positive results are confirmed in an Ames test with the structural analogue cumene hydroperoxide (Mortelmans 1988: positive with S9, negative without S9). Two studies by Hüls 1989, did not show positive responses in Salmonella TA 100, TA1535, TA1537, TA1538 and TA98 with cumene hydroperoxide and isopropyl hydroperoxide. In a recombination assay with Saccharomyces cerevisiae cumenehydroperoxide induced intrachromosomal recombinations (Brennan 1994). In vivo assay Cumene hydroperoxide is tested in an in vivo micronucleus assay in mice dosed topically at 0.75, 1.5, 3.0, 6.0 and 12 mg/kg bw for 93 days (NTP 2004). No increase of micronuclei was observed in erythrocytes (NCE) from collected peripheral blood.

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: DNA damage and/or repair

Type of information:

experimental study

Remarks:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study (standard NTP protocol)

Qualifier:

according to guideline

Guideline:

other: Standard NTP protocol

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

exposure period 13 weeks

Principles of method if other than guideline:

Micronucleus analysis in NTP toxicity studies (13 weeks).

GLP compliance:

not specified

Type of assay:

micronucleus assay

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

data not yet published

Route of administration:

dermal

Vehicle:

ethanol

Details on exposure:

data not yet published

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

once daily, 5 days/week

Post exposure period:

24 h

Remarks:

Doses / Concentrations:
0.75, 1.5, 3, 6, 12 mg/kg
Basis:

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

no

Tissues and cell types examined:

peripheral blood

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Mice are treated in a 13 week toxicity study as part of the NTP bioassay program

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): sampling collection time is 24 h in the case of single daily exposures


DETAILS OF SLIDE PREPARATION: a blood sample from male and female mice in each dose group is obtained; slides are prepared, fixed and stained


METHOD OF ANALYSIS: 1000 mature erythrocytes ( normochromatic erythrocytes, NCEs ) are scored per animal for the presence of micronuclei (MN); the percent PCE is determined in the blood as a measure of chemical induced toxicity to the bone marrow; all data are analysed seperately for male and female mice; the type of analysis is not given (data not yet published)

Evaluation criteria:

Mean MN-NCE/100 NCE

Statistics:

A formal statistical analysis of the data is performed that includes a trend test, to determine if there is an overall increase across all doses in the frequency of cells containing micronuclei, and a pairwise comparison of each dose group to the corresponding control, to see if any dose group is statistically different from the control group in the frequency of micronucleated cells.

Sex:

male/female

Genotoxicity:

negative

Vehicle controls validity:

valid

Additional information on results:

data not yet published

The trend test p-value for males was 0.264 and for females 0.777 ( a positive trend test is one in which the p-value is equal to or less than 0.025)

Males

Dose (mg/kg)	No. of animals scored	Mean MN-NCE/1000 NCE  (+- SEM)	Pairwise p-value
0*	5	2.80 (+- 0.70)
0.75	5	2.70 (+- 0.70)	0.554
1.5	5	2.90 (+- 0.29)	0.447
3	5	1.90 (+- 0.37)	0.906
6	5	2.80 (+- 0.34)	0.500
12	5	3.10 (+- 0.51)	0.348

*: vehicle control ethanol; SEM: standard error of mean

Females

Dose (mg/kg)	No. of animals scored	Mean MN-NCE/1000 NCE  (+- SEM)	Pairwise p-value
0*	5	1.90 (+- 037)
0.75	5	1.40 (+- 0.29)	0.808
1.5	5	2.10 (+- 0.29)	0.376
3	5	1.50 (+- 0.42)	0.754
6	5	1.30 (+- 0.25)	0.856
12	5	1.50 (+- 0.42)	0.754

*: vehicle control ethanol; SEM: standard error of mean

For the micronucleus frequency in any dose group to be considered significantly elevated over the control group, the p value must be equal to or less than 0.025 divided by the number of chemical treatment groups; if the number of treatment groups is 5, then the required pairwise p-value is 0.005; this adjustment in the pairwise value is a correction for multiple comparisons of the same data.

Conclusions:

Interpretation of results (migrated information): negative
Cumene hydroperoxide given dermally for 13 weeks to mice did not induce micronuclei in the peripheral blood of the test animals

Executive summary:

Cumene hydroperoxide was tested in a Micronucleus test (Standard NTP Protocol); the results are available in the Internet; the substance was applied dermally to male and female mice for 13 weeks and than erythrocytes were examined for the presence of micronuclei; under the conditions of this test cumene hydroperoxide was negative in the in vivo mouse micronucleus test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vivo:

The effects on the test substance in bacterial assays are weakly positive, which was confirmed by tests on structural analogues, o.a. cumenehydroperoxide, which showed positive responses. No effects were found in a chromosome aberration test in Chinese Hamster Lung cells with and without metabolic activation. The structural analogue cumene hydroperoxide was negative in an in vivo micronucleus test. No in vivo tests with the test substance are available, but the test with cumenehydroperoxide is considered to represent a worst case.

It is concluded that the test substance is not mutagenic.

Justification for selection of genetic toxicity endpoint
in vivo study with structural analogue according to standard protocol with sufficient detail to allow evaluation. The structural analogue is considered to be representative of a worst case situation (see attachment chapter 13 on read-across)

Justification for classification or non-classification

The test substance does not need to be classified for mutagenicity, as in a weight of evidence the combined results of in vitro and in vivo tests with either the test substance or the structural analogue cumene hydroperoxide were considered negative, i.e. non-mutagenic.