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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro DNA damage and/or repair study
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Data is from peer reviewed publication
Justification for type of information:
Data is from peer reviewed publication

Data source

Reference
Reference Type:
publication
Title:
The genotoxicity of a variety of aniline derivatives in a DNA repair test with primary cultured rat hepatocytes
Author:
Naoki Yoshimi, Shigeyuki Sugie, Hitoshi Iwata, Kenji Niwa, Hideki Mori, Chise Hashida and Hidesuke Shimizu
Year:
1988
Bibliographic source:
Mutation Research, 206 (1988) 183-191

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: As mention below
Principles of method if other than guideline:
The genotoxicity of N-methylanthranilic acid methyl ester, was examined by a DNA repair test with rat hepatocytes.
GLP compliance:
not specified
Type of assay:
DNA damage and repair assay, unscheduled DNA synthesis in mammalian cells in vitro

Test material

Constituent 1
Chemical structure
Reference substance name:
Methyl N-methylanthranilate
EC Number:
201-642-6
EC Name:
Methyl N-methylanthranilate
Cas Number:
85-91-6
Molecular formula:
C9H11NO2
IUPAC Name:
methyl 2-(methylamino)benzoate

Method

Target gene:
No data
Species / strain
Species / strain / cell type:
hepatocytes: ACI rat hepatocytes
Details on mammalian cell type (if applicable):
- Type and identity of media: Williams' Medium E
- Properly maintained: No data
- Periodically checked for Mycoplasma contamination: No data
- Periodically checked for karyotype stability: No data
- Periodically “cleansed” against high spontaneous background: No data
Additional strain / cell type characteristics:
not specified
Metabolic activation:
not specified
Metabolic activation system:
No data
Test concentrations with justification for top dose:
10-3, 10-4, 10-5, 10-6 M
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
other: N-2-fluorenylacetamide
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: No data
- Exposure duration: 20 hrs
- Expression time (cells in growth medium): 20 hrs
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data
SPINDLE INHIBITOR (cytogenetic assays): No data
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: Duplicate

NUMBER OF CELLS EVALUATED: 50 cells/coverslip

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Other: No data

OTHER: The test was performed in accordance with the method of Williams et al. Hepatocytes were isolated from the livers of male ACI rats weighing 200-250 g. The isolated hepatocytes were allowed to attach for 2 h on plastic coverslips in primary culture using Williams' Medium E. The cultures were then washed and exposed to the test chemical and [Me- 3H]thymidine (10 µCi/ml; 49 Ci/mmole) for 20 h. It was logarithmically diluted before addition to the cultures. At the end of incubation, the cultures were washed, and the coverslips were mounted on glass slides. The slides were dipped in Sakura NR-M2 photographic emulsion and exposed for 14 days. Autoradiographic grains were counted on a television screen (Olympus, type S) with a microscopic attachment.

The data were expressed as the average net counts/nucleus for 3 coverslips + the standard deviation (50 cells/coverslip). Two experiments were performed for the test compound for the assays with rat hepatocytes.
Evaluation criteria:
The test chemical was considered positive when the mean net nuclear grain count was more than 5 grains above background and statistically greater than that of controls (unpaired t-test; P < 0.01).
Statistics:
Mean ± standard deviation

Results and discussion

Test results
Species / strain:
hepatocytes: ACI rat hepatocytes
Metabolic activation:
not specified
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
Positive controls validity:
not specified
Additional information on results:
No data available
Remarks on result:
other: strain/cell type:
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Chemical

Dose M

UDS grains/nucleus

% od UDS positive cells

DNA repair

N-methylanthranilic acid methyl ester

10-3

0.7 ± 1.4

0

-

10-4

-0.1 ± 1.5

0

10-5

-0.5 ± 1.2

0

10-6

-0.5 ± 1.0

0

Applicant's summary and conclusion

Conclusions:
The given test material test substance is negative in the rat hepatocyte/DNA repair test.
Executive summary:

The hepatocyte/DNA repair test which measures unscheduled DNA synthesis (UDS) is known to be sensitive to various classes of DNA-reactive carcinogens and is regarded as a reliable short-term test for the detection of chemical carcinogens.The genotoxicity of test substance, was examined by a DNA repair test with rat hepatocytes.

 

The test was performed basically in accordance with the method of Williams et al. The test material was dissolved in DMSO and the positive control used was N-2-fluorenylacetamide. The hepatocytes were exposed to the test chemical for 20 hrs. At the end of incubation, the cultures were washed, and the coverslips were mounted on glass slides. The slides were dipped in Sakura NR-M2 photographic emulsion and exposed for 14 days. Autoradiographic grains were counted on a television screen (Olympus, type S) with a microscopic attachment.

 

The test chemical was evaluated to be positive only when the mean net nuclear grain count was more than 5 grains above background and statistically greater than that of controls.

 

The results of the hepatocyte/DNA repair test suggests that N-methylanthranilic acid methyl ester is negative for genotoxicity in vitro.