Disodium [[4-[bis[4-[(sulphonatophenyl)amino]phenyl]methylene]cyclohexa-2,5-dien-1-ylidene]amino]benzenesulphonate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Data is from publication.

Data source

Reference

Reference Type:

publication

Title:

Verapamil as a co-mutagen in the Salmonella/mammalian microsome mutagenicity test

Author:

Lynnette R. Ferguson and Bruce C. Baguley

Year:

1988

Bibliographic source:

Mutation Research, 209 (1988) 57-62

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:

To evaluate the mutagenic potential of Methyl Blue in Salmonella typhimurium TA1537, TA1598 and TA100 by AMES assay.

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Disodium [[4-[bis[4-[(sulphonatophenyl)amino]phenyl]methylene]cyclohexa-2,5-dien-1-ylidene]amino]benzenesulphonate

EC Number:

249-352-9

EC Name:

Disodium [[4-[bis[4-[(sulphonatophenyl)amino]phenyl]methylene]cyclohexa-2,5-dien-1-ylidene]amino]benzenesulphonate

Cas Number:

28983-56-4

Molecular formula:

C37H29N3O9S3.2Na

IUPAC Name:

Disodium [[4-[bis[4-[(sulphonatophenyl)amino]phenyl]methylene]cyclohexa-2,5-dien-1-ylidene]amino]benzenesulphonate

Details on test material:

- IUPAC Name: Disodium [[4-[bis[4-[(sulphonatophenyl)amino]phenyl]methylene]cyclohexa-2,5-dien-1-ylidene]amino]benzenesulphonate
- SMILES:O=S(=O)(c1ccccc1Nc1ccc(C(=C2C=CC(=N{+}c3ccccc3S(=O)(=O)O{-}.[Na]{+})C=C2)c2ccc(Nc3ccccc3S(=O)(=O)O{-})cc2)cc1)O{-}.[Na]{+}
- InChI:1S/C37H29N3O9S3.2Na/c41-50(42,43)34-19-13-31(14-20-34)38-28-7-1-25(2-8-28)37(26-3-9-29(10-4-26)39-32-15-21-35(22-16-32)51(44,45)46)27-5-11-30(12-6-27) 40-33-17-23-36(24-18-33)52(47,48)49;;/h1-24,38-39H,(H,41,42,43)(H,44,45,46)(H,47,48,49);;/q;2*+1/p-1
- Mol. formula: C37H26N3Na2O9S3
- Molecular Weight: 800.8182 g/mol

Specific details on test material used for the study:

- Name of test material :Methyl Blue
- Molecular formula : C37H26N3Na2O9S3
- Molecular weight : 800.8182 g/mol
- Smiles notation : c1cc(ccc1C(=C2C=CC(=[NH+]c3ccc (cc3)S(=O) (=O)[O-])C=C2)c4ccc(cc4)Nc5ccc(cc5)S (=O)(=O)[O-])Nc6ccc(cc6)S(=O)(=O)O.[Na+].[Na+]
- InChl : 1S/C37H29N3O9S3.2Na/c41-50(42,43)34-19-13-31(14-20-34)38-28-7-1-25(2-8-28)37(26-3-9-29(10-4-26)39-32-15-21-35(22-16-32)51(44,45 46)27-5-11-30(12-6-27)40-33-17-23-36(24-18-33)52(47,48)49;;/h1-24,38-39H,(H,41,42,43)(H,44,45,46)(H,47,48,49);;/q;2*+1/p-1
- Substance type: Organic
- Physical state: Solid

Method

Target gene:

Histidine

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA1537, TA1598 and TA100

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

not specified

Metabolic activation:

not specified

Test concentrations with justification for top dose:

100 µl

Vehicle / solvent:

50% of ethanol

Controls

Untreated negative controls:

yes

Remarks:

ethanol

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

not specified

Positive controls:

no

Details on test system and experimental conditions:

Details on test system and conditions
METHOD OF APPLICATION: in agar (plate incorporation)
DURATION
- Exposure duration: 3days

NUMBER OF REPLICATIONS: triplicate

OTHER: Reversion characteristics of each strain were tested in each experiment using the disc method of Zieger et al. (1982). The background number of revertants was 8 ± 3 for TA1537, 34 ±7 for TA98, and 171 ± 10 for TAI00.

Rationale for test conditions:

Not specified

Evaluation criteria:

Mutagenicity is expressed as revertants colonies/µg compound added to the plate, and is derived from the slope of the corresponding regression equation.

Statistics:

Mutagenicity data were analyzed according to the method of Moore and Felton (1983). A regression line was fitted for the number of mutants versus
drug concentration for the linear part of the curve. Mutagenicity was scored as negative if the regression coefficient was non-significant. To determine whether verapamil had a significant effect on mutagenicity, the slopes ( +__ SD) of the corresponding regression lines with and without verapamil were determined, and the significance of the difference was calculated using Student's t test.

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium, other: TA1537, TA1598 and TA100

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

not examined

Additional information on results:

All mutagenicity data, calculated as revertant colonies per /µg of added drug in the presence or absence of verapamil.

Remarks on result:

other: NO mutagenic effct were observerd

Any other information on results incl. tables

Sr no	Compound	Mutagenicity^bin bacterial strain^c
		TA1537	TA98	TA100
1	Methyl Blue	0	0	0

 

b Mutagenicity is expressed as revertant colonies/tzg compound added to the plate, and is derived from the slope of the corresponding

regression equation,.

c Bacterial strains are described in Maron and Ames (1983).

d Statistical significance of the differences: **p < 0.001; *p < 0.05.

e A value of 0 means that the correlation coefficient of the regression equation is not statistically significant.

Applicant's summary and conclusion

Conclusions:

Methyl Blue (28983-56-4) was evaluated for its mutagenic potential in Salmonella typhimurium TA1537, TA1598 and TA100. The test result was considered to be negative for the test.

Executive summary:

In Genetic toxicity in vitro study, Methyl Blue was assessed for its possible mutagenic potential. For this purpose AMES assay was performed in Salmonella typhimurium TA1537, TA1598 and TA100 by plate-incorporation assay. The test substance was exposed at the concentration of than 100µl of 50% ethanol, or 50% ethanol alone as negative control. Plates were allowed to harden, and then incubated for 3 days at 37°C before scoring colonies for reversion to Histidine independence. A dose range of each compound was tested, and each experimental point performed in triplicate on at leasttwo separate occasions. No mutagenic effect were observed at this dose .Therefore Methyl Blue was considered to be non mutagenic for AMES test. Hence the substance cannot be classified as gene mutant in vitro.