diethyl pyridine-2,4-dicarboxylate

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

Study period:

9.9.2002 - 2.4.2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test has been performed under GLP and according to OECD guideline.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

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:

other: ICH S2A and S2B

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Mexoryl SBU

Cas Number:

41438-38-4

Molecular formula:

C11H13N04

IUPAC Name:

Mexoryl SBU

Constituent 2

Reference substance name:

diethyl pyridine-2,4-dicarboxylate

EC Number:

680-341-5

Cas Number:

41438-38-4

Molecular formula:

C11H13N04

IUPAC Name:

diethyl pyridine-2,4-dicarboxylate

Details on test material:

- Name of test material: MEXORYL SBU- The test material is the same than the one mentioned in section 7.2.1

Method

Target gene:

Histidine

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Metabolic activation system:

S9 Mix

Test concentrations with justification for top dose:

Experiment 1: 52, 164, 512, 1600 and 5000 µg/plateExperiment 2: 492, 878, 1568, 2800 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO- Justification for choice of solvent/vehicle: not reported

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

other: Without S9: 2-Nitrofluorene (TA98), Sodium azide (TA100, TA1535), 9-Aminoacridine (TA1537), t-Butyl hydroperoxide (TA102). With S9: 2-Aminoanthracene (all strains)

Details on test system and experimental conditions:

METHOD OF APPLICATION: Experiment 1: direct plate incorporation methodExperiment 2: preincubation with S9, plate incorporation without S9DURATION- Preincubation period: 60 min (Experiment 2 with S9)- Exposure duration: at least 48 h NUMBER OF REPLICATIONS: 3DETERMINATION OF CYTOTOXICITY- Method: qualitatively estimated based on the reduction in bacterial lawn

Evaluation criteria:

Biological relevance of the results should be considered first. Statistical methods (Dunnett's test at p <= 0.05 and linear regression analysis) are used as an aid in evaluating the test results and statistical significance should not be the only determinant of a positive response.Therefore mutagenicity evaluation of the test item is based on the validity of the study and statistically significant, dose related or reproducible increase(s) in the number of revertant cells with evidence of a biological effect.

Statistics:

Dunnett's test at p <= 0.05 and linear regression analysis

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS- Precipitation: No precipitate was noted at any dose level. RANGE-FINDING/SCREENING STUDIES: Since the plates from at least 3 dose levels of the preliminary study could be scored, these results were included and discussed as the actual mutagenicity data for the strain TA100 in the experiment carried out using the plate incorporation method (Experiment 1).COMPARISON WITH HISTORICAL CONTROL DATA: The mean negative control counts are within or close to the range of historical data. ADDITIONAL INFORMATION ON CYTOTOXICITY: No signs of cytotoxicity were noted at any dose level, either with or without metabolic activation but the test item was tested up to limit concentrations.

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):negativeWhen tested up to the maximum recommended dose level of 5000 µg/plate, and using both the plate incorporation and the preincubation methods, the test item MEXORYL SBU did not induce biologically significant increases in the number of revertants in the five Salmonella typhimurium strains used (TA98, TA100, TA1535, TA1537 and TA102), either with or without metabolic activation.

Executive summary:

Five histidine-dependent strains of Salmonella typhimurium (TA98, TA100, TA1535, TA1537 and TA102) were used to evaluate the mutagenic potential of the test item Mexoryl SBU in the presence and absence of metabolic activation (with and without S9). The study was carried out using both the plate incorporation method and the preincubation method (37 ± 2°C for approximately 60 minutes under stirring). The test item was tested as a clear colourless solution in dimethylsulfoxide (DMSO).

The first experiment was performed using the plate incorporation method at the dose levels of 52, 164, 512, 1600 and 5000 µg/plate (half-log progression), both in the presence and absence of metabolic activation. No precipitate and no signs of cytotoxicity were noted, either with or without metabolic activation. No statistically or biologically significant increases in the number of revertants were noted in

any of the five strains used, either with or without metabolic activation.

The second experiment was performed using the plate incorporation method without metabolic activation and using the preincubation method with metabolic activation, at the dose levels of 492, 878, 1568, 2800 and 5000 µg/plate (quarter-log progression). No precipitate and no signs of cytotoxicity were noted, either with or without metabolic activation. A statistically significant (p <= 0.01) increases in the number of revertants was only noted at the dose level of 2800 µg/plate in the strain TAl00 in the absence of metabolic activation. This increase was slight and not dose-related. This finding was therefore not considered to be representative of any biologically relevant increase in the number of mutants.

Under the experimental conditions and according to the criteria of the test study plan, it is concluded that the test item Mexoryl SBU did not induce any mutagenic effect in the Ames test, either with or without metabolic activation.