tetraethyl N,N'-(methylenedicyclohexane-4,1-diyl)bis-.sc.dl.sc.-aspartate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP guideline study, with acceptable restrictions: only 4 strains tested; E. coli WP2 strain or S. typhimurium TA 102 were not investigated.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

, only four strains tested (E. coli WP2 strain or S. typhimurium TA 102 were not investigated)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

136210-32-7

EC Number:

603-947-3

Cas Number:

136210-32-7

IUPAC Name:

136210-32-7

Constituent 2

Reference substance name:

Bis(4-(1,2-bis(ethoxycarbonyl)ethylamino)-3-methylcyclohexyl)methane

EC Number:

412-060-9

EC Name:

Bis(4-(1,2-bis(ethoxycarbonyl)ethylamino)-3-methylcyclohexyl)methane

Cas Number:

136210-32-7

Molecular formula:

C31H54N2O8

IUPAC Name:

bis(4-(1,2-bis(ethoxycarbonyl)ethylamino)-3-methylcyclohexyl)methane

Method

Target gene:

histidine locus

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9-Mix from the liver of Aroclor 1254 induced male rats

Test concentrations with justification for top dose:

0, 8, 40, 200, 1000, 5000 µg/plate (with and without S9-mix)

Vehicle / solvent:

The solvent employed for the test substance was ethanol and for the positive controls DMSO.

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: sodium azide, nitrofurantoin, 4-nitro-1,2-phenylene diamine, 2-aminoanthracene,

Remarks:

2-aminoanthracene (promutangen) only used with S9-mix, the other positive controls only used without S9-mix.

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation) for first trial and repeat

DETERMINATION OF CYTOTOXICITY
- gross appraisal of background growth
- a toxic effect was assumed when there was a marked and dose-dependent reduction in the mutant-count per plate, copared to the negative controls
- total bacterial counts

Evaluation criteria:

A reproducible and dose-related increase in mutant counts of at least one strain is considered to be a positive result. For TA 1535, TA 100 and TA 98 this increase should be about twice the amount of negative controls, whereas for TA 1537, at least a threefold increase should be reached. Otherwise, the result is evaluated as negative. However, these guidelines may be overruled by good scientific judgment. In case of questionable results, investigations should continue, possibly with modifications, until a final evaluation is possible.

Statistics:

Not specified.

Results and discussion

Test results

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:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: occured at the dose of 5000 µg/plate

ADDITIONAL INFORMATION ON CYTOTOXICITY:
At all tested doses, the substance had a weak, strain-specific bacteriotoxic effect, so that this range could nevertheless be used for assessment purposes.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Study was performed with Aspartic acid, N,N'-[methylenebis(2-methyl-4,1-cyclohexanediyl)]bis-, 1,1',4,4'-tetraethyl ester which is a structural analogue to Aspartic acid, N,N'-(methylenedi-4,1-cyclohexanediyl)bis-, 1,1',4,4'-tetraethyl ester. Both substances are diethyl esters of aspartic acid linked to a dicyclohexylmethyldiamine moiety. The difference between these two substances is merely the presence of two methyl groups connected to the cyclohexane rings. This structural analogy was confirmed by the Member State responsible for the notification of both substances under the NONS regulation. The Member State decided that test results obtained for one substance can be transferred to the other substance and that testing of both substances is usually not required. This decision is in accordance with the grouping of substances and read-across approach in Annex XI, 1.5 of the REACH Regulation.

Applicant's summary and conclusion

Executive summary:

Aspartic acid, N,N'-[methylenebis(2-methyl-4,1-cyclohexanediyl)]bis-,1,1',4,4'-tetraethyl ester showed no mutagenic activity in the Salmonella/microsome test (OECD TG 471) with the S. typhimurium strains TA 98, TA 100, TA 1535 and TA 1537 in the absence and in the presence of a metabolic activation system.