Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

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:
23 December 2014 to 27 April 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
Test item: Disperse Yellow DYLA 1306

Method

Target gene:
The test item Disperse Yellow DYLA 1306 was examined for the ability to induce gene mutations in tester strains of Salmonella typhimurium and Escherichia coli, as measured by reversion of auxotrophic strains to prototrophy.
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Metabolic activation system:
S9 liver homogenate from induced rat (rat mixed induction).
Test concentrations with justification for top dose:
According to solubility, in Main Assay I: 248, 124, 62.0, 31.0 and 15.5 µg/plate.
Main Assay II: 248, 165, 110, 73.3, 48.9 and 32.6 μg/plate (TA100 +S9)
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: 2-aminoanthracene
Details on test system and experimental conditions:
Toxicity and Main Assay I were performed using the plate incorporation method.
A second separate experiment were performed on the TA100 tester strain in order to confirm the positive results obtained in the first experiment.
Evaluation criteria:
For the test item to be considered mutagenic, two-fold (or more) increases in mean revertant numbers must be observed at two consecutive dose levels or at the highest practicable dose level only. In addition, there must be evidence of a dose-response relationship showing increasing numbers of mutant colonies with increasing dose levels.
Statistics:
Doubling rate (Chu et al. 1981).
Regression line.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium, other: S. typhimurium TA 1535, TA 1537, TA 98 and E. coli WP2
Metabolic activation:
with
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with
Genotoxicity:
positive
Remarks:
in the Main Assay 1, test result was questionable, in Main Assay II, there was a weak positive result
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
The test item induced increases in the number of revertant colonies, which were at least two-fold greater than the control value at the highest dose levels with TA100 tester strain in the presence of S9 metabolic activation.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
It is concluded that the test item Disperse Yellow DYLA 1306 induces reverse mutation in Salmonella thyphimurium TA100 tester strain in the presence of S9 metabolism, under the reported experimental conditions.
Executive summary:

The test item Disperse Yellow DYLA 1306 was examined for the ability to induce gene mutations in tester strains of Salmonella typhimurium and Escherichia coli, as measured by reversion of auxotrophic strains to prototrophy. The five tester strains TA1535, TA1537, TA98, TA100 and WP2 uvrA were used. Experiments were performed both in the absence and presence of metabolic activation, using liver S9 fraction from rats pre-treated with phenobarbitone and betanaphthoflavone. The test item was used as a solution in dimethylsulfoxide (DMSO).

Toxicity test: Based on results obtained in a preliminary solubility trial, the test item Disperse Yellow DYLA 1306 was assayed in the toxicity test at a maximum concentration of 248 µg/plate and at four lower concentrations spaced at approximately half-log intervals: 78.4, 24.8, 7.84 and 2.48 µg/plate. Precipitation of the test item was observed at the end of the incubation period at the two highest concentrations. No toxicity was observed with any tester strain in the absence or presence of S9 metabolism. Increases in revertant numbers were observed in the presence of S9 metabolism with TA100 tester strain.

Main Assay: On the basis of toxicity test results, in Main Assay I, using the plate incorporation method, the test item was assayed at the following dose levels: 248, 124, 62.0, 31.0 and 15.5 μg/plate.

Precipitation of the test item was observed at the end of the incubation period at the two highest concentrations both in the absence and presence of S9 metabolic activation.

No toxicity was observed with any tester strain at any dose level in the absence or presence of S9 metabolism. The test item induced increases in the number of revertant colonies in the presence of S9 metabolism with TA100 tester strain reaching a two-fold increase at the highest concentration tested compared to the vehicle control, but not compared to the untreated control (MF 1.8 -fold).

Following the approval of a Protocol Amendment, an additional confirmatory experiment (Main Assay II) was performed in which TA100 tester strain was treated in the presence of S9 metabolism at the dose levels of 248, 165, 110, 73.3, 48.9 and 32.6 μg/plate.

Dose related increases in revertant numbers were observed. These increases were greater than twice the control values at the two highest dose levels, thus confiming the positive response with TA100 tester strain in the presence of S9 metabolism. No further experiment was undertaken.

Conclusion

It is concluded that the test item Disperse Yellow DYLA 1306 induces reverse mutation in Salmonella typhymurium TA100 tester strain in the presence of S9 metabolism, under the reported experimental conditions.