Benzenebutanoic acid, 4-[(4-methoxybenzoyl)amino]-.beta.-methyl-3-nitro-.gamma.-oxo-, methyl ester

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 February 1997 - 07 April 1997

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: compliant to GLP and testing guideline except for invalid positive control (MNNG) in the 1st experiment without S9; adequate coherence between data, comments and conclusions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

yes

Remarks:

In the 2nd experiment without metabolic activation, EMS and BrdU were used as positive controls because MNNG was probably too old and did not incude mutants. This change from the experimental protocol did not affect the results or conclusions of the study

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: 87/302/EWG, S.61

Deviations:

yes

Remarks:

idem above

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

426-290-2

EC Name:

-

Cas Number:

74149-72-7

Molecular formula:

C20H20N2O7

IUPAC Name:

methyl 4-[4-(4-methoxybenzamido)-3-nitrophenyl]-3-methyl-4-oxobutanoate

Details on test material:

- Name of test material (as cited in study report): UD-CG 115 nitrobenzamide (Gamma-NAP)
- Substance type: monosonstituent
- Analytical purity: 99.8%
- Lot/batch No.: 1/94
- Expiration date of the lot/batch: February 1999
- Storage conditions of test material: at room temperature (dark) and ambient humidity.

Method

Target gene:

HPRT

Species / strain

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (if applicable):

- Type and identity of media:
- Properly maintained: yes/no
- Periodically checked for Mycoplasma contamination: yes/no
- Periodically checked for karyotype stability: yes/no
- Periodically "cleansed" against high spontaneous background: yes/no

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induction (S9-mix)

Test concentrations with justification for top dose:

Between 5 and 50 µg/mL; higher doses elicit precipitation

Vehicle / solvent:

DMSO.
Rationale: DMF and acetone did not improve solubility.

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

Migrated to IUCLID6: MNNG, DMBA, BrdU

Evaluation criteria:

A positive response is defined if there is a reproducible and concentration-dependent increase in mutant frequency for a minimum of two
successive concentrations, irrespective of individual statistically significant. Comparisons were made against the negative control values in the
respective test. In addition, historical control frequencies obtained in similar gene mutation experiments were also taken into consideration.
Accordingly, the mutant frequency must exceed 30-35/10E6 in order to compensate for random fluctuations that are typical for this assay.

Results and discussion

Test results

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Remarks:

50 µg/mL

Vehicle controls validity:

other: yes except MNNG (1st exp. -S9)

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

Precipitates at 50 µg/mL.
-S9: Minor cytotoxicity (not dose-limiting) at 25-50 µg/mL: 81% and 76% survivals.

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

There was no dose-dependant increase of the number of the mutants and the rate of mutants over the total dose-range of 5 to 50 µg/mL
(limit of precipitation) in V79 mammalian cells.

Executive summary:

Not cytotoxic in V79 CHL cells.

Precipitates at 50 µg/mL.

Negative for mammalian gene mutation, with and without activation.