Cyclohexanamine, 4,4'-methylenebis[2,6-dimethyl-

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study (OECD 471).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

4-[(4-amino-3,5-dimethylcyclohexyl)methyl]-2,6-dimethylcyclohexan-1-amine

EC Number:

634-657-5

Cas Number:

65962-45-0

Molecular formula:

C17H34N2

IUPAC Name:

4-[(4-amino-3,5-dimethylcyclohexyl)methyl]-2,6-dimethylcyclohexan-1-amine

Details on test material:

- Name of test material (as cited in study report): 4,4´-Diamino-3,3´,5,5´-tetramethyl-dicylohexylmethane
- Physical state: liquid
- Analytical purity: 91.3 %

Method

Target gene:

histidine gene
tryptophan

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

S9-mix from rat livers

Test concentrations with justification for top dose:

1st experiment: 0, 33, 100, 333, 1000, 2500, 5000 µg/plate (+/- S-9)
2nd experiment: 0, 10, 100, 333,1000, 2500 µg/plate (+/- S-9)
3rd experiment: 0, 10, 33, 100, 333, 1000, 2500 µg/plate (+/- S-9)
4th experiment: 0, 3.3, 10, 33, 100, 333, 1000µg/plate (- S9)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Complete solubility of the test substance in DMSO.

Controls

Untreated negative controls:

yes

Remarks:

sterility control

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: with S-9 mix: 2-aminoanthracene; without S-9 mix: N-methyl-N-nitro-N-nitrosoguanidine for the strains TA 100 and TA 1535, 4-nitro-o-phenylendiamine for the strain TA 98, 9-aminoacridine for strain TA 1537 ; 4-nitroquinoline-N-oxide for strain E.coli

Details on test system and experimental conditions:

Standard Plate Test:
Test tubes containing 2 ml soft agar kept in a water bath at about 42-45°C and the remaining components are added in the following order:
0.1 ml test solution or vehicle
0,1 ml fresh bacterial culture
0.5 ml S9 mix (with metabolic activation) or 0.5 ml phosphate buffer (without metabolic activation)
After mixing, the samples are poured onto Vogel-Bonner agar plates.

Preincubation Test:
0.1 ml test solution or vehicle, 0.1 ml bacterial suspension and 0.5 ml S9 mix (with metabolic activation) or phosphate buffer (without metabolic activation) are incubated at 37°C for the duration of about 20 minutes. Subsequently, 2ml of soft agar is added, and after mixing, the samples are poured onto the agar plates.

Both Tests:
In each experiment 3 test plates per dose or per control used, after incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies ar counted.

Evaluation criteria:

Positive results:
doubling of the spontaneous mutation rate,
dose-response relationship,
reproducibility of the results

Statistics:

Mean and standard deviation calculated in result tables.

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

4,4´-Diamino-3,3´,5,5´-tetramethyl-dicylohexylmethane is not a mutagenic in the the bacterial reverse mutation test in the absence and presence of metabolic activation.

Executive summary:

4,4´-Diamino-3,3´,5,5´-tetramethyl-dicylohexylmethane was tested for mutagenicity in the Salmonelle typhimurium /Escherichia coli reverse mutation assay both in the standard plate test and in the preincubation test with and without the addition of a metabolizing (S9-mix) obtainded from rat liver using the Salmonelle strains TA1535, TA100, TA1537, TA98 and Escherichia coli WP2 uvrA (AMES test). No relevant increase in the number of revertants was detected in any strain with and without metabolic activation.