Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate

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

Reliability:

2 (reliable with restrictions)

Data source

Reference

Reference Type:

publication

Title:

Methodology for the testing of food dyes for genotoxic activity: experiments with RED 2G (C.I. 18050)

Author:

R.B. Haveland-Smith, R.D. Combes, B.A. Bridges

Year:

1979

Bibliographic source:

Mutation Research, 64 , 241—248

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: as mentioned below

Principles of method if other than guideline:

The mutagenicity of Red 2G was evaluated in Salmonella typhimurium strain TA1538 by Fluctuation tests.

GLP compliance:

not specified

Type of assay:

bacterial gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Red 2G

IUPAC Name:

Red 2G

Constituent 2

Reference substance name:

Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate

EC Number:

223-098-9

EC Name:

Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate

Cas Number:

3734-67-6

Molecular formula:

C18H15N3O8S2.2Na

IUPAC Name:

disodium 5-acetamido-4-hydroxy-3-(phenyldiazenyl)naphthalene-2,7-disulfonate

Test material form:

not specified

Details on test material:

- Name of test material (as cited in study report): Red 2G- Molecular formula (if other than submission substance): C18-H15-N3-O8-S2.2Na- Molecular weight (if other than submission substance): 509.426 g/mol - Substance type: Organic - Physical state: Aqueous solutions or suspensionsPurity- 80%- Impurities (identity and concentrations): As, Pb ,Sb, BaSO4, Cr, Cu, Zn,Free aromatic amines, Synthetic intermediates (excluding free aromatic amines),Subsidiary colures were present.

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

Calcium precipitated microsome from rat liver (Male, 5-week old Sprague-Dawley rats were supplied with 0.1% w/v sodium pentobarbitone and 2% w/v D-glucose in their drinking water for one week.)

Test concentrations with justification for top dose:

Fluctuation tests without microsomal activation: 10 mg/ ml Fluctuation tests with microsomal activation: 10 mg /ml

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Deionised water

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: Rimmel Mahogany Silk 2-acetylaminofluorene

Details on test system and experimental conditions:

Fluctuation test without microsomal activationThe fluctuation test was performed according to the method of Green and Muriel with the following modifications. A single colony from a plate of each of the bacterial strains (S. typhimurium TA1538) was inoculated into 10 ml of the appropriate growth medium and incubated overnight at 37°C with shaking. The mutation media were as follows: (a) TA1538: solution SA, 1.20.5 ml; solution SB, 4.5 ml; solution SC, 25 pl; solution SD, 25 pl. Sub-lethal concentrations of each test agent were prepared in the mutation media. 30 µl of inoculum culture were pipetted into this mixture, which was then dispensed in 2-ml aliquots into 50 test tubes (75 × 12 mm, rimless). The number of turbid tubes was scored after 72 h incubation at 37 ° C.Fluctuation test with microsomal activation0.1 ml of overnight TA1538 culture was pipetted into 0.9 ml solution SA (plus 9 mg /m1 D-glucose, 17 µg /ml L-histidine and 0.9 µg /ml D-biotin). The mutation media were as follows: (a) WP2uvrA: solution F (plus D-glucose, L-tryptophan and bacteria), 1 ml; calcium-precipitated microsome, 1 ml; 8 mg/ ml G-6-P, 0.25 ml; 5 units /ml G-6-PDH, 0.1 ml; water or water soluble test agent, 0.5 ml; (b) TA1538: as Green et al. (DMSO-soluble agents were added to the mutation media in 5-pl volumes).The mixture was dispensed in 100-µl aliquots into each of 50 test tubes and incubated for 24 h at 37°C. 2 ml solution F (plus 0.4% w/v D-glucose) was dispensed into each of the TA1538 tubes. These were then incubated for 72 hour at 37°C before scoring for turbidity.

Evaluation criteria:

The tubes were scored for turbidity.

Statistics:

No data available.

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

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):negativeRed 2G was determined to give negative results for mutagenicity in Salmonella typhimurium strain TA1538 in Fluctuation tests conducted with and without metabolic activation at the concentration of 10mg /ml.

Executive summary:

Genotoxicity study for Red 2G was conducted in Salmonella typhimurium strain TA1538 at the concentration of 10mg /ml in presence and absence of metabolic activation by Fluctuation tests.

The tubes were scored for turbidity. When dyes such as Red 2G was used in this system, it may be impossible to detect the turbidity in the tubes by eye or to use a growth indicator such as bromothymol blue, due to masking by the colour. In this case, the presence of viable prototrophic revertants was verified by streaking loopfuls from each tube onto unsupplemented agar.

No significant change was observed in the tubes. Therefore Red 2G was determined to give negative results for mutagenicity in Salmonella typhimurium strain TA1538 in Fluctuation tests conducted with and without metabolic activation at the concentration of 10mg /ml.