Trisodium 5-[[4-chloro-6-(methylphenylamino)-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[(2-sulphonatophenyl)azo]naphthalene-2,7-disulphonate

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:

1979

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

None

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: AMES, B.N., J. McCANN, and E. YAMASAKI (1975), Methods for Detecting Carcinogens and Mutagens with the Salmonella/ Mammalian-Microsome Mutagenicity Test. Mut. Res. 31, 347-364.

Deviations:

not specified

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Principles of method if other than guideline:

None

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Trisodium 5-[[4-chloro-6-(methylphenylamino)-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[(2-sulphonatophenyl)azo]naphthalene-2,7-disulphonate

EC Number:

274-417-3

EC Name:

Trisodium 5-[[4-chloro-6-(methylphenylamino)-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[(2-sulphonatophenyl)azo]naphthalene-2,7-disulphonate

Cas Number:

70210-20-7

Molecular formula:

C26H20ClN7O10S3.3Na

IUPAC Name:

trisodium 5-[[4-chloro-6-(methylphenylamino)-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[(2-sulphonatophenyl)azo]naphthalene-2,7-disulphonate

Test material form:

not specified

Specific details on test material used for the study:

None

Method

Target gene:

Histidine auxotrophic strains of Salmonella typhimurium

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium strains TA 98, TA 100, TA 1535, TA 1537 and TA 1538

Metabolic activation:

with and without

Metabolic activation system:

S9 fraction of liver from rats induced with Arochlor and cofactors

Test concentrations with justification for top dose:

None

Vehicle / solvent:

phosphate buffer

Controls

Untreated negative controls:

yes

Remarks:

Phosphate buffer

Positive controls:

yes

Positive control substance:

other: With metabolic activation: Strain TA 1535 with cyclophosphamide,

Details on test system and experimental conditions:

The tests were carried out in accordance with the method described by AMES et al. The bacteria on which the tests were performed were the histidineauxotrophic TA 98, TA 100, TA 1535, TA 15 37 and TA 1538, strains of Salmonella typhimurium.
The test was performed with the following concentrations of the trial substance without and with microsomal activation: 15, 45, 135, 405 and 1215 microg/0.1 ml. A repetition of the experiments was performed with the concentrations of 25, 75, 225, 6 75 and 2025 microg/ 0.1 ml. In these experiments tests on Strain TA 15 38 were included. The substance was dissolved in phosphate buffer. Phosphate buffer alone was used for the negative controls. In the experiments in which the substance was metabolically activated, activation mixture was added also. 1 ml activation mixture contains: 0.3 ml S9 fraction of liver from rats induced with Aroclor 1254 and 0.7 ml of a solution of co-factors.

Positive control experiments were carried out simultaneously with the following substances: 1) for Strain TA 98: daunorubicin-HCl (DAUNOBLASTINR), 5 and 10 microg/0.1 ml phosphate buffer; 2) for Strain TA 100: 4-nitroquinoline-N-oxide, 0.125 and 0.25 microg/0.1 ml phosphate buffer; 3) for Strain TA 1535: N-methyl-N1-nitro-N-nitrosoguanidine, 3 and 5 microg/0.1 ml phosphate buffer; 4) for Strain TA 15 37: 9(5) aminoacridine hydrochloride monohydrate, 50 and 100 microg/0.1 ml DMSO; 5) for Strain TA 15 38: 2-nitrofluorene, 5 and 10 microg/0.1 ml DMSO. The activation mixture was tested with Strain TA 15 35 and cyclophosphamide (ENDOXAN-ASTA ), 250 microg/0.1 ml phosphate buffer.
In the experiments with and without the addition of microsomal activation mixture three Petri dishes were prepared per strain and per group (i.e. per concentration or per control group).
The plates were incubated for about 48 hours at 37 deg C in darkness. When the colonies had been counted, the arithmetic mean was calculated.
A test substance is generally considered to be non-mutagenic if the colony count in relation to the negative control is not doubled at any concentration .

1.AMESf B.N., F.D. LEE, and W.E. DURSTON (1973), An Improved Bacterial Test System for the Detection and Classification of Mutagens and Carcinogens. Proc. Natl. Acad. Sei. USA 70, 782-786.
2.AMES, B.N., W.E. DURSTON, E. YAMASAKI, and F.D. LEE (1973), Carcinogens are Mutagens: A Simple Test System Combining Liver Homogenates for Activation and Bacteria for Detection.
Proc. Natl. Acad. Sei. USA 7_0, 2281-2285.
3.AMES, B.N., J. McCANN, and E. YAMASAKI (1975), Methods for Detecting Carcinogens and Mutagens with the Salmonella/ Mammalian-Microsome Mutagenicity Test. Mut. Res. 31, 347-364.

Rationale for test conditions:

None

Evaluation criteria:

None

Statistics:

None

Results and discussion

Test results

Key result

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

In the experiments performed with and without microsomal activation, comparison of the number of histidine-prototrophic mutants in the controls and after treatment with FAT 40034/B revealed no marked differences.

Remarks on result:

other: all strains/cell types tested

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:

FAT 40034/B was determined to be non-mutagenic in the bacterial reverse mutation assay.

Executive summary:

FAT 40034/B was tested for mutagenic effects on histidine-auxotrophic mutants of Salmonella typhimurium which included TA 98, TA 100, TA 1535, TA 15 37 and TA 1538. The investigations were performed with the following concentrations of the trial substance with and without microsomal activation: 15, 45, 135, 405 and 1215 microg/0.1 ml. A repetition of the experiments was performed with the concentrations of 25, 75, 225, 675 and 2025 microg/0.1 ml. In these experiments tests on Strain TA 1538 were included.

In the experiments performed with and without microsomal activation, comparison of the number of back-mutant colonies in the controls and the cultures treated with the various concentrations of FAT 40034/B revealed no marked deviations.

Based on the findings of the study, no evidence of the induction of point mutations by FAT 40034/B or by the metabolites of the substance formed as a result of microsomal activation was detectable in the strains of S. typhimurium used in these experiments. Hence, FAT 40034/B was determined to be non-mutagenic in the bacterial reverse mutation assay.