[7-hydroxy-8-[(2-methoxyphenyl)azo]-2-naphthyl]trimethylammonium chloride

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:

weight of evidence

Reliability:

4 (not assignable)

Data source

Reference

Reference Type:

secondary source

Title:

OPINION ON Basic Red 76

Author:

Scientific Committee on Consumer Safety SCCS

Year:

2011

Bibliographic source:

Scientific Committee on Consumer Safety SCCS - adopted at its 10th plenary meeting of 22 March 2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

not specified

Type of assay:

bacterial gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

[7-hydroxy-8-[(2-methoxyphenyl)azo]-2-naphthyl]trimethylammonium chloride

EC Number:

269-941-4

EC Name:

[7-hydroxy-8-[(2-methoxyphenyl)azo]-2-naphthyl]trimethylammonium chloride

Cas Number:

68391-30-0

Molecular formula:

C20H22ClN3O2

IUPAC Name:

7-hydroxy-8-[(2-methoxyphenyl)diazenyl]-N,N,N-trimethylnaphthalen-2-aminium chloride

Constituent 2

Reference substance name:

Basic Red 76

IUPAC Name:

Basic Red 76

Details on test material:

Details on test materialName of test material (as cited in study report): Basic Red 76Molecular formula (if other than submission substance): C20H22N3O2 ClMolecular weight (if other than submission substance): 371.86g/molSubstance type: Organic Physical state: SolidPurity: 98.6% HPLC Impurities (identity and concentrations): water content = 4.1%(w/w); Monomethyl sulphate 15.9% (w/w); o-anisidine = 19ppm; Chloromethane = 0.3% (w/w); Methyl acetate = 0.1% (w/w); Methyl formate = 0.4% (w/w); 7-Hydroxy-N,N,N-trimethyl-napthalen-2-aminium chloride = <500 ppm (detection limit); Sulphated ash 0.3% (w/w); Chloride 2.7% (w/w); Sodium = 190 ppm

Method

Species / strain

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Liver S9-fraction from phenobarbital/β-naphthoflavone-induced rats was used as exogenous metabolic activation system.

Test concentrations with justification for top dose:

experiment I: 10, 33, 100, 333, 1000, 2500 and 5000 μg/plate without and with S9-mixexperiment II: 33, 100, 333, 1000, 2500 and 5000 μg/plate without and with S9-mix

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: in accordance with the OECD guideline

Details on test system and experimental conditions:

Details on test system and conditionsMETHOD OF APPLICATION: in medium; in agar (plate incorporation); preincubation; in suspension; as impregnation on paper diskExperiment I was performed according to the direct plate incorporation test, experiment II with the pre-incubation method.DURATIONPreincubation period: 60 minutes Exposure duration: 48 hours incubation with and without S9 mixNUMBER OF REPLICATIONS: triplicates in 2 individual experimentsNUMBER OF CELLS EVALUATED: No dataDETERMINATION OF CYTOTOXICITYMethod: mitotic index; cloning efficiency; relative total growth; other: No dataOTHER EXAMINATIONS:Determination of polyploidy: No dataDetermination of endoreplication: No dataOther:experiment I: direct plate incorporation with 48 h incubation without and with S9-mixexperiment II: pre-incubation method with 60 minutes pre-incubation and at least 48 h incubation without and with S9-mixTest concentrations were based on the results of a pre-experiment with strains TA98 and TA100 for toxicity and mutation induction both without and with S9-mix. Toxicity was evaluated for 8 concentrations up to the prescribed maximum concentration of 5000 μg/plate on the basis of a reduction in the number of revertant colonies and/or clearing of the bacterial background lawn. Becauserelevant toxic effects were not observed in any of the strains at the maximal concentration, 5000 μg/plate was used as the top concentration. Since in this pre-experiment evaluable plates were obtained for five concentrations or more in the strains used, the pre-experiment is reported in experiment I.

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:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:Test concentrations were based on the results of a pre-experiment with strains TA98 and TA100 for toxicity and mutation induction both without and with S9-mix. Toxicity was evaluated for 8 concentrations up to the prescribed maximum concentration of 5000 μg/plate on the basis of a reduction in the number of revertant colonies and/or clearing of the bacterial background lawn. Because relevant toxic effects were not observed in any of the strains at the maximal concentration, 5000 μg/plate was used as the top concentration.

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):negativeBasic Red 76 was investigated for the induction of gene mutations in strains of Salmonella typhimurium (Ames test).In the main tests toxic effects evident as clearing of the bacterial background lawn were observed in experiment I in TA98 and TA100 and in experiment II in all strains predominantly at higher concentrations. Toxic effects evident as a reduction in the number of revertants were observed at higher concentrations without and with metabolic activationin nearly all strains tested. A biologically relevant increase in revertant colonies was not observed in any of the strains tested at any dose level in the absence or presence of S9-mix in both experiments.Under the experimental conditions used, Basic Red 76 was not mutagenic in this gene mutation tests in bacteria both in the absence and the presence of S9 metabolic activation.

Executive summary:

Basic Red 76 was investigated for the induction of gene mutations in strains ofSalmonella typhimurium(Ames test).

Salmonella typhimuriumTA98, TA100, TA102, TA1535 and TA1537 were used for the AMES assay. DMSO was used as a vehicle.

 

Concentration: experiment I: 10, 33, 100, 333, 1000, 2500 and 5000 μg/plate without and with S9-mix

experiment II: 33, 100, 333, 1000, 2500 and 5000 μg/plate without

and with S9-mix

Treatment: experiment I: direct plate incorporation with 48 h incubation without and with S9-mix

experiment II: pre-incubation method with 60 minutes pre-incubation

and at least 48 h incubation without and with S9-mix

 

Liver S9-fraction from phenobarbital/β-naphthoflavone-induced

rats was used as exogenous metabolic activation system. Test concentrations were based on the results of a pre-experiment with strains TA98 and TA100 for toxicity and mutation induction both without and with S9-mix. Toxicity was evaluated for 8 concentrations up to the prescribed maximum concentration of 5000 μg/plate on the basis of a reduction in the number of revertant colonies and/or clearing of the bacterial background lawn. Because relevant toxic effects were not observed in any of the strains at the maximal concentration, 5000 μg/plate was used as the top concentration. Since in this pre-experiment evaluable plates were obtained for five concentrations or more in the strains used, the pre-experiment is reported in experiment I. Experiment I was performed according to the direct plate incorporation test, experiment II with the pre-incubation method. Negative and positive controls were in accordance with the OECD guideline.

 

In the main tests toxic effects evident as clearing of the bacterial background lawn were observed in experiment I in TA98 and TA100 and in experiment II in all strains predominantly at higher concentrations. Toxic effects evident as a reduction in the number of revertants were observed at higher concentrations without and with metabolic activation

in nearly all strains tested. A biologically relevant increase in revertant colonies was not observed in any of the strains tested at any dose level in the absence or presence of S9-mix in both experiments.

 

Under the experimental conditions used, Basic Red 76 was not mutagenic in this gene mutation tests in bacteria both in the absence and the presence of S9 metabolic activation