α-methylcyclohexylmethyl acetate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Veilex2_Ames test (OECD TG 471)_Read across CP acetate: negative

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2007-12-14

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

E. coli WP2 uvr A

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and 5,6-benzoflavone

Test concentrations with justification for top dose:

Dose range finding test 1:
- 4.88, 19.5, 78.1, 313, 1250, and 5000 µg/plate
Dose range finding test 2:
- 2.44, 4.88, 9.77, 19.5, 39.1, and 78.1 µg/plate without S9
- 9.77, 19.5, 39.1, 78.1, 156, and 313 µg/plate with S9
Main test
- 2.44, 4.88, 9.77, 19.5, 39.1, and 78.1 µg/plate without S9
- 9.77, 19.5, 39.1, 78.1, 156, and 313 µg/plate with S9

Vehicle / solvent:

- Solvent used: DMSO
- Justification for choice of solvent: the test substance was found to be soluble in DMSO up to 50 mg/mL

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: see section "Any other information on materials and methods incl. tables"

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation method

DURATION
- Preincubation period: 20 minutes
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in duplicate.

DETERMINATION OF CYTOTOXICITY
- Method: on the basis of a decline in the number of spontaneous revertants.

Evaluation criteria:

The test substance was judged to be positive when the number of revertant colonies increased to twice or more that in the negative control in a concentration-dependent manner and also the reproducibility of the test results was obtained. In all other cases, it was judged to be negative.

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
- Dose range finding test 1: The number of revertant colonies in the test substance treatment groups was less than twice that in the solvent control in the groups of treatment with and without S9 mix. The bacterial growth inhibition was observed at 78.1 µg/plate or more in all test strains without S9 mix and at 313 µg/plate or more in all test strains with S9 mix. The precipitation of the test substance was not observed at any doses in the groups of treatment with and without S9 mix.
- Dose range finding test 2: The number of revertant colonies in the test substance treatment groups was less than twice that in the solvent control groups of treatment with and without S9 mix. The bacterial growth inhibition was observed at 39.1 µg/plate or more in TA100, TA1535, TA98, and TA1537 and at 78.1 µg/plate in WP2uvrA in the groups of treatment without S9 mix and at 156 µg/plate or more in TA100, TA1535, TA98, and TA1537 and at 313 µg/plate in WP2uvrA in the groups of treatment with S9 mix. The precipitation of the test substance was not observed at any doses in the groups of treatment with and without S9 mix.

MAIN STUDY:
The number of revertant colonies in the test substance treatment groups was less than twice that in the solvent control groups of treatment with and without S9 mix. The bacterial growth inhibition was observed at 39.1 µg/plate or more in TA100, TA1535, TA98, and TA1537 and at 78.1 µg/plate in WP2uvrA in the groups of treatment without S9 mix and at 156 µg/plate or more in TA100, TA1535, TA98, and TA1537 and at 313 µg/plate in WP2uvrA in the groups of treatment with S9 mix. The precipitation of the test substance was not observed at any doses in the groups of treatment with and without S9 mix.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

The substance is not mutagenic in the Salmonella typhimurium reverse mutation assay performed according to OECD 471.

Executive summary:

The mutagenic activity of the substance was evaluated in accordance with OECD 471 and according to GLP principles. A preincubation assay was performed with S. typhimurium strains TA100, TA1535, TA98, TA1537, and E. coli WP2uvrA with and without metabolic activation (Rat liver S9-mix induced by a combination of phenobarbital and 5,6-benzoflavone). Adequate solvent (DMSO) and positive controls were included. Two dose range finding tests were performed and one main test. In the main test all stains were dosed with 2.44, 4.88, 9.77, 19.5, 39.1, and 78.1 µg/plate without metabolic activation and 9.77, 19.5, 39.1, 78.1, 156, and 313 µg/plate with metabolic activation. The experiment was performed in duplicate. The number of revertant colonies in the test substance treatment groups was less than twice that in the solvent control groups of treatment with and without S9 mix. Under the conditions of the test the substance is not mutagenic.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

For Veilex 2 no information is available on Ames. For covering the endpoint CP Acetate is used. The summary of this information is presented first and thereafter the read across justification.

Read across CP Acetate

The mutagenic activity of the substance was evaluated in accordance with OECD 471 and according to GLP principles. A preincubation assay was performed with S. typhimurium strains TA100, TA1535, TA98, TA1537, and E. coli WP2uvrA with and without metabolic activation (Rat liver S9-mix induced by a combination of phenobarbital and 5,6-benzoflavone). Adequate solvent (DMSO) and positive controls were included. Two dose range finding tests were performed and one main test. In the main test all stains were dosed with 2.44, 4.88, 9.77, 19.5, 39.1, and 78.1 µg/plate without metabolic activation and 9.77, 19.5, 39.1, 78.1, 156, and 313 µg/plate with metabolic activation. The experiment was performed in duplicate. The number of revertant colonies in the test substance treatment groups was less than twice that in the solvent control groups of treatment with and without S9 mix. Under the conditions of the test the substance is not mutagenic.

The mutagenicity of Veilex #2 (CAS#13487-27-9) using read across from substance CP Acetate (CAS#25225-10-9)

Introduction and hypothesis for the analogue approach

Veilex#2 has acyclohexylethyl-backbonewith anacetate estergroup attached. For this substance nomutagenicitydata are available. In accordance with Article 13 of REACH, lacking information may .be generated by other means, i. e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across. For assessing themutagenicityin bacterial cells of Veilex#2 the analogue approach is selected because for a closely related analogue, CP Acetatemutagenicityinformation is available which can be used for read across.

Hypothesis: Veilex#2 has similar genotoxic potential compared to CP Acetate as thetwo methyl groups attached to the meta-position of the cyclohexylethyl backbone of CP Acetate are not expected to influence mutagenicity.

Available information: For the target chemical Veilex#2 no information is available regarding mutagenicity. A well conducted Ames test with CP Acetate according to OECD TG 471 and in accordance with GLP is available (K1).

Target chemical and source chemical(s)

Chemical structures of the target chemical and the source chemicals are shown in the data matrix, including physico-chemical properties and toxicological information, thought relevant for sensitization, of all substances.

Purity / Impurities

Itis not expected that the impurities of the target and source chemical affect the read-across justification. Veilex#2 and CP acetate are both mono-constituent presenting a purity of close to 100%

Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group.

In accordance with ECHA guidance (2015, RAAF) the analogue was selected from a group of related cyclohexyl-esters all negative in the Ames test and CP Acetate being the closest structural analogue. For support also information from CP Formate is presented (see Data matrix below)

Structural similarities and differences:Veilex #2 as well as the structural analogue have a common cyclohexylethyl backbone. CP Acetate, has two methyl groups attached to the meta-position of this backbone, while Veilex#2 does not have these methyl substituents. An acetate ester is attached to the cyclohexylethyl backbone of both Veilex#2 and CP Acetate.These two methyl group difference between the target and source chemicals will not influence the mutagenicity of thesechemicals.

Toxico-kinetic similarities and differencesare expressed because mutagenicity needs to be considered for both local and systemic endpoints:

Absorption:Veilex#2 and CP Acetate have similar molecular weight and physico-chemical properties indicating similar absorption characteristics. Both substances are liquids, the molecular weight (170.3 and 198.3 g/mol, respectively) and log Kow (Ca. 3.2 and 4.42, respectively) indicate that Veilex#2 and its source chemical are bioavailable via all routes of exposure:

Metabolism:It can be anticipated that Veilex#2 will metabolise to acetic acid (CAS#64-19-7) and 1-cyclohexylethanol (CAS#1193-81-3) while CP Acetate will metabolise acetic acid and to 1-(3,3-dimethylcyclohexyl)ethanol (CAS#25225-09-6) as presented in Figure 1.

Uncertainty of the prediction:There is no remaining uncertainty, in view of similarities in structure, toxico-kinetic profile (absorption and metabolism). The reactivity profile between the substances is also considered similar because both have an acetic ester group attached to a cyclohexyl ring. Therefore, read across can be applied. There are no structural alerts regarding mutagenicity for the target chemical, source chemical and their metabolites (Toxtree V2.6.13 and OECD toolbox V3.3.0.132). Therefore the mutagenicity data of CP Acetate can be used for read across to Veilex#2. According to ECHA guidance (2015, RAAF) the quality code 5 can be applied because of the close relation between the two substances.

Fig. 1 The metabolisation pathway of Veilex #2 and CP Acetate.

Data matrix

The relevant information on physico-chemical properties and toxicological characteristics are presented in the Data matrix below.

Conclusions on the result of the Ames test

Hazard: For Veilex#2 no information regarding mutagenicity is available. An Ames test according to OECD TG 471 is available for the closely related analogue CP Acetate. CP Acetate was not mutagenic in the Ames test (OECD TG 471 and GLP). Based on this result of CP Acetate it can be concluded that Veilex#2 is not mutagenic in the Ames test either.

Data matrix for the read across to Veilex#2 from CP Acetate

Common names	Veilex#2	CP Acetate	CP Formate
Chemical structures
	Target	Key source	Supporting
CAS no	13487-27-9	25225-10-9	25225-08-5
EC no	REACH (2018)	Not registered yet	939-618-9
Empirical formula	C10H18O2	C12H22O2	C11H20O2
Physico-chemical data	Measured by IFF	Predicted with EpiSuite	Measured by IFF
Molecular weight	170.25	198.304	184
Physical state	liquid	liquid	liquid
Melting point,^oC	< -20	13.46	-20oC
Boiling point,^oC	215.7	>204	219.9
Vapour pressure, Pa	38.5	6.7	13.4
Water solubility, mg/L	43	7.462	26
Log Kow	3.2	4.42	3.8
Human health endpoints
Genotoxicity – Ames test	Read across	Not mutagenic, OECD TG 471, K1)	Not mutagenic (OECD TG 471, K1)

 

Justification for classification or non-classification

Based on the results of the Ames test performed with the read across substance CP Acetate, the test substance does not have to be classified for mutagenicity in accordance with Regulation (EC) No. 1272/2008.