Methyl formylphenylacetate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Sept 9-26, 2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Principles of method if other than guideline:

The chemical intermediate SCH 1000 Formylester NV of SCH 1000 BR-synthesis was investigated in a modified bacterial mutagenicity test as described by Ames et al. (1975). This high throughput microtitre-based version, called Ames II, is based on the same genetic principle (base-pair substitution and frameshift mutations in the his operon of S. typhimurium) as the traditional Ames assay (Gee et al, 1998).

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Methyl formylphenylacetate

EC Number:

227-577-3

EC Name:

Methyl formylphenylacetate

Cas Number:

5894-79-1

Molecular formula:

C10H10O3

IUPAC Name:

methyl 3-oxo-2-phenylpropanoate

Method

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 98

Species / strain 2

Species / strain / cell type:

S. typhimurium, other: TA mix (TA 7001, 7002, 7003, 7004, 7005, 7006)

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

the test article was tested over a concentration range of 10 to 1000 µg/ml medium with and without microsomal rat liver enzymes concurrently with positive and negative controls

Vehicle / solvent:

DMSO

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

2-nitrofluorene

other: 2-aminoanthracene

Details on test system and experimental conditions:

The Ames II test was conducted with and without addition of microsomal liver enzymes from rats (Aroclor 1254-induced). Following base-pair and frameshift-specific tester strains were used: S. typhimurium TA Mix (TA 7001, TA 7002, TA 7003, TA 7004, TA 7005 and TA 7006) and TA 98, respectively. The Salmonella strains and method are described by Gee et al. (1998).
The assay was performed according to the instruction manual for the Ames II (Xenometrix, Boulder/USA). 0.01 ml of the vehicle, test article or positive control were incubated with
0.24 ml bacterial ovemight culture (ca 10^7/ml)/exposure medium in 24-well plates for 90 min at 37°C, 250 rpm. With metabolic activation 0.2 ml strain mixture and 0.04 ml S9-mix (30%) were used. After 90 min the exposed cultures were diluted with pH indicator medium lacking histidine and aliquoted into 48 wells of a 384-well plate (3 replicates) using a 8-channel pipettor. The plates were incubated for 48 hrs at 37°C.

Rationale for test conditions:

the experiment is regarded valid, if the vehicle control shows the normal spontaneous revertant and the diagnostic mutagens cause the expected increase in the mutation rate.

Evaluation criteria:

The individual test chemicals were classified according to the following criteria: Negative: ≤8/48 wells Equivocal: 9-12/48 wells Positive: ≥13/48 wells

The historical control range experienced in this laboratory covering more than 100 experi¬ments is 0-7/48 wells.

A concentration-dependent increase of revertant wells (mean of triplicate) over the vehicle control range is indicative for a genotoxic activity of the test article

Statistics:

The purple pH indicator bromocresol tums yellow as the pH drops due to the accumulation of catabolites from the metabolic activity of revertant cells. The number of positive wells (yellow) out of a total of 48 wells is indicative of the reversion frequency per replicate/ concentration and was compared to the number of spontaneous revertant wells of the solvent control. Each test point contains 48 wells of a 384-well plate. In each 48-well section, the wells were scored for the number of revertant wells (yellow) and the mean value of the triplicates was calculated.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Species / strain:

other: TA mix (TA 7001, 7002, 7003, 7004, 7005, 7006)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

based on the described result it is concluded, that SCH 1000 Formylester NV, when tested up to toxic concentrations, caused neither base-pair substitution nor frameshift mutations in bacteria. No evidence of genotoxic activity was observed in as series of S. typhimurium tester strains (TA mix and TA 98) in the absence and presence of metabolic activation. The test compound is, therefore, classified as "Ames II negative"