Alkali salt of sulfonated aryl azo amino sulfonyl aryl azo sulfonyl aryl azo aryl sulfonate

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:

07 July 2014 to 20 Oct 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Compliant with GLP and testing guideline, coherence among data, results and conclusions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial gene mutation assay

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): Reactive Brown DYHY 0331/0334

Method

Target gene:

The test item Reactive Brown DYHY0331/0334 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) or uninduced Syrian hamster (Prival modification)

Test concentrations with justification for top dose:

In Main Assay I and II: 5000, 2500, 1250, 625 and 313 µg/plate.

Vehicle / solvent:

sterile water for injection

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, Trypan Blue

Details on test system and experimental conditions:

Toxicity and Main Assay I were performed using the plate incorporatin method; Main assay II using the pre-incubaton method

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 results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

No relevant increase in the number of revertant colonies was observed in the plate incorporation or pre-incubation assay, at any dose level, with any tester strain, in the absence or presence of any S9 metabolic activation system.

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

It is concluded that the test item, Reactive Brown DYHY0331/0334, does not induce reverse mutation in Salmonella typhimurium or Escherichia coli in
the absence or presence of S9 metabolism, under the reported experimental conditions.

Executive summary:

The test item Reactive Brown DYHY0331/0334 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. In the preliminary toxicity test and in Main Assay I, treatments were performed using the plate incorporation method in the absence and presence of a cofactor-supplemented S9 metabolic fraction prepared from the livers of rats treated with phenobarbitone and b-naphthoflavone. Based on the chemical structure of the test item (azodyes), Main Assay II was performed using the preincubation method and a reductive metabolic activation system (Prival modification).

The test item was used as a solution in sterile water for injection.

The test item Reactive Brown DYHY0331/0334 was assayed in the toxicity test at a maximum concentration of 5000 μg/plate and at four lower concentrations spaced at approximately half-log intervals: 1580, 500, 158 and 50.0 μg/plate. No toxicity was observed with any tester strain at any dose level, in the absence or presence of S9 metabolism. Plates treated with the test item presented a dose dependent black colour of the agar, which did not interfere with the scoring of colonies.

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: 5000, 2500, 1250, 625 and 313 μg/plate.

No toxicity was observed with any tester strain at any dose level, in the absence or presence of S9 metabolism. Plates treated with the test item presented a dose dependent black colour of the agar, which did not interfere with the scoring of colonies.

As no relevant increase in revertant numbers was observed at any concentration tested, a Main Assy II was performed using the pre-incubation method in the presence of flavin mononucleotide and uninduced hamster liver S9.

The dose-range used was the same as in Main Assay I.

No toxicity was observed with any tester strain at any dose level, in the absence or presence of S9 metabolism. Plates treated with the test item presented a dose dependent black colour of the agar, which did not interfere with the scoring of colonies.

The test item did not induce two-fold increases in the number of revertant colonies in the plate incorporation or pre-incubation assay, at any dose level, in any tester strain, in the absence or presence of any metabolic activation system metabolism.

It is concluded that, under the reported experimental conditions, the test item Reactive Brown DYHY0331/0334 does not induce reverse mutation in Salmonella typhimurium or Escherichia coli in the absence or presence of the standard S9 metabolic activation or using a reductive metabolic activation, as described by Prival and Mitchell for azo-dyes.