Acetic acid, 2-cyano-2-[3-[(3-methoxypropyl)amino]-2-cyclohexen-1-ylidene]-, 2-ethoxyethyl ester, (2Z)-

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

BASF SE, Ludwigshafen, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2-ethoxyethyl 2-cyano-2-[(1Z)-3-[(3-methoxypropyl)amino]cyclohex-2-en-1-ylidene]acetate

EC Number:

700-860-3

Cas Number:

1419401-88-9

Molecular formula:

C17 H26 N2 O4

IUPAC Name:

2-ethoxyethyl 2-cyano-2-[(1Z)-3-[(3-methoxypropyl)amino]cyclohex-2-en-1-ylidene]acetate

Test material form:

other: solid

Details on test material:

Please refer to confidential details on test material.

Method

Target gene:

Salmonella typhimurium
histidine auxotrophy (his-) to histidine prototrophy (his+)
Escherichia coli
tryptophan auxotrophy (trp-) to tryptophan independence (trp+)

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9 fraction from Phenobarbital (i.p.) and β-Naphthoflavone (orally) induced rat liver

Test concentrations with justification for top dose:

First and second experiment with and without S9 mix:
0; 33; 100; 333; 1 000; 2 625 and 5 250 μg/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

With S9 mix: 2-aminoanthracene (2-AA); without S9 mix: N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylenediamine (NOPD), 9-aminoacridine (AAC), 4-nitroquinoline-N-oxide (4-NQO)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 20 min
- Exposure duration: 48-72 hours


NUMBER OF REPLICATIONS: 3 test plates per dose or per control


DETERMINATION OF CYTOTOXICITY
• decrease in the number of revertants
• clearing or diminution of the background lawn (= reduced his- or trp- background growth)
• reduction in the titer

Evaluation criteria:

Acceptance criteria
Generally, the experiment is considered valid if the following criteria are met:
• The number of revertant colonies in the negative controls was within the range of the historical negative control data for each tester strain.
• The sterility controls revealed no indication of bacterial contamination.
• The positive control substances both with and without S9 mix induced a distinct increase in the number of revertant colonies within the range of the historical positive control data or above.
• Fresh bacterial culture containing approximately 10E9 cells per mL were used. For approval the titer of viable bacteria was ≥ 10E8 colonies per mL.

Assessment criteria
The test substance is considered positive in this assay if the following criteria are met:
• A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system was observed.
A test substance is generally considered non-mutagenic in this test if:
• The number of revertants for all tester strains were within the historical negative control range under all experimental conditions in at least two experiments carried out independently of each other.

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

Remarks:

standard plate test: decrease in his+ revertants, slight reduction in the titer from about 2625 µg/plate onward; In the preincubation assay: slight decrease in his + or trp+ revertants, slight reduction in the titer at a concentration of 2625 µg/plate.

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

SOLUBILITY
No test substance precipitation was found with and without S9 mix.

MUTAGENICITY:
A biologically relevant increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either without S9 mix or after the addition of a metabolizing system.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.