Trisodium 2-[[4,5-dihydro-3-methyl-5-oxo-1-[4-[[2-(sulphonatooxy)ethyl]sulphonyl]phenyl]-1H-pyrazol-4-yl]azo]naphthalene-1,5-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:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed publication

Data source

Reference

Reference Type:

publication

Title:

Genotoxicity Testing of the Food Colours Amaranth and Tartrazine

Author:

Aparajita Das and Anita Mukherjee

Year:

2004

Bibliographic source:

Int J Hum Genet, 4(4): 277-280 (2004)

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Ames mutagenicity assay was performed to evaluate the mutagenic nature of the test compound tartrazine.

GLP compliance:

not specified

Type of assay:

bacterial gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo)pyrazole-3-carboxylate

EC Number:

217-699-5

EC Name:

Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo)pyrazole-3-carboxylate

Cas Number:

1934-21-0

Molecular formula:

C16-H12-N4-O9-S2.3Na

IUPAC Name:

trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo)pyrazole-3-carboxylate

Constituent 2

Reference substance name:

Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo) pyrazole-3-carboxylate

IUPAC Name:

Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo) pyrazole-3-carboxylate

Details on test material:

Details on test material
- Name of test material (as cited in study report): Tartrazine
- Molecular formula (if other than submission substance): C16-H12-N4-O9-S2.3Na.
C16-H9-N4-O9-S2.3Na
- Molecular weight (if other than submission substance): 534.3681 g/mol
- Substance type: Organic
- Physical state: No data available
Purity No data available
- Impurities (identity and concentrations): No data available

Specific details on test material used for the study:

- Name of test material: Tartrazine
- IUPAC name: Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo) pyrazole-3-carboxylate
- Molecular formula: C16H12N4O9S2.3Na
C16H9N4O9S23Na
- Molecular weight : 534.3681 g/mol
- Substance type: Organic
- Physical state: No data available
- Purity No data available
- Impurities (identity and concentrations): No data

Method

Target gene:

Histidine

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA97a, TA98 and TA100

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

not specified

Metabolic activation system:

No data

Test concentrations with justification for top dose:

0, 100, 250, 500 or 1000 μg /plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Sterile double distilled water
- Justification for choice of solvent/vehicle: No data available

Controls

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

Sterile double distilled water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

sodium azide

other: nitro phenylene diamine for TA97a and TA98

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: No data available
- Exposure duration: 48 hrs
- Expression time (cells in growth medium): 48 hrs
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): No data available
SPINDLE INHIBITOR (cytogenetic assays): No data available
STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: Three plates were used for each set.

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data available

OTHER EXAMINATIONS:
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other: No data available

OTHER: No data available

Rationale for test conditions:

No data

Evaluation criteria:

Increase in the number of mutagenic revertants

Statistics:

ANOVA test was performed at 0.05 level

Results and discussion

Test results

Species / strain:

S. typhimurium, other: TA97a, TA98 and TA100

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No data

Remarks on result:

other: No mutagenic potential

Any other information on results incl. tables

Table: Mutagenicity of food colours in tester strains of Salmonella typhimurium

Dose μG/plate	Mean of the No. Revertant Colonies ± S.D.
	TA97a	TA98	TA100
100	126.70 ± 19.55	64.7 ± 4.04	110.7 ± 12.5
250	131.30 ± 18.50	60.0 ± 3.46	105.0 ± 15.13
500	118.00 ± 9.54	57.7 ± 21.2	125.0 ± 5.00
1000	113.30 ± 25.50	26.7 ± 3.21	115.0 ± 13.53
Solvent control	122.30 ± 9.61	21.3 ± 9.02	121.0 ± 3.61
NPD	827.70 ± 106.53	522.0 ± 50.48	-
SA	-	-	1624.67 ± 89.76

 

NPD= Nitrophynylenediamine

SA= Sodium azide

S.D.= Standard Deviation

Anova Value of TA97a (Tartrazine —1.256, ns)

TA 98 (Tartrazine –7.96*)

TA100 (Tartrazine—2.74, ns)

P< 0.01 (5.06)

Applicant's summary and conclusion

Conclusions:

Tartrazine did not induce gene mutation in Salmonella typhimurium strains TA97a, TA98 and TA100 and hence it is not likely to classify as a gene mutant in vitro.

Executive summary:

Ames mutagenicity assay was performed to evaluate the mutagenic nature of the test compound tartrazine using the plate incorporation assay.

 

Tartrazine was dissolved in sterile double distilled water and was tested at a concentration of 0, 100, 250, 500 and 1000 μg /plate. The plates were inverted within an hour and placed in a dark vented incubator at 37⁰C for 48 hours. Positive controls (for TA97a and TA98, 20 μg/plate nitro phenylene diamine and for TA100, 1.5 μg/plate sodium azide) and negative controls were maintained concurrently for all the experiments. Three plates were used for each set. After 48 hours of incubation, the revertant colonies were counted.

Tartrazine did not induce gene mutation in Salmonella typhimurium strains TA97a, TA98 and TA100 and hence it is not likely to classify as a gene mutant in vitro.