Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate

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)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from publication

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Gene mutation toxicity study of the test chemical

Author:

Errol Zeiger et. al.

Year:

2000

Bibliographic source:

Mutation Research, 2000

Reference 2

Reference Type:

other: Secondary source

Title:

Gene mutation toxicity study of the test chemical

Author:

NTRL

Year:

2010

Bibliographic source:

NTP (National Toxicological Program) A81251, 2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: As mentioned below

Principles of method if other than guideline:

Gene mutation toxicity study of the test chemical

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate

EC Number:

247-368-0

EC Name:

Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate

Cas Number:

25956-17-6

Molecular formula:

C18H16N2O8S2.2Na

IUPAC Name:

disodium 6-hydroxy-5-[(2-methoxy-3-methyl-4-sulfonatophenyl)diazenyl]naphthalene-2-sulfonate

Test material form:

solid

Details on test material:

IUPAC name: Disodium 6-hydroxy-5-[(2-methoxy-4-sulphonato-m-tolyl)azo]naphthalene-2-sulphonate
Mol. formula: C18H14N2Na2O8S2
Molecular Weight: 496.4266 gm/mol
Smiles: c12c(cc(cc2)S(=O)(=O)[O-])ccc(c1/N=N/c1c(cc(c(c1)C)S(=O)(=O)[O-])OC)O.[Na+].[Na+]
InChI: 1S/C18H16N2O8S2.2Na/c1-10-7-14(16(28-2)9-17(10)30(25,26)27)19-20-18-13-5-4-12(29(22,23)24)8-11(13)3-6-15(18)21;;/h3-9,21H,1-2H3,(H,22,23,24)(H,25,26,27);;/q;2*+1/p-2/b20-19+;;

Method

Target gene:

Histidine

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 100

Additional strain / cell type characteristics:

not specified

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1535

Additional strain / cell type characteristics:

not specified

Species / strain 3

Species / strain / cell type:

S. typhimurium TA 97

Additional strain / cell type characteristics:

not specified

Species / strain 4

Species / strain / cell type:

S. typhimurium TA 98

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

with 10 % & 30 % HLI = induced male Syrian hamster liver S9 and 10% & 30 % RLI = induced male Sprague Dawley rat liver S9

Test concentrations with justification for top dose:

100-10000 µg/Plate

Vehicle / solvent:

Water

Controls

Controls 1

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

Water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

For strains tested with S9

Controls 2

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

Water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

For strains TA100 and TA1535 tested in the absence of S9

Controls 3

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

Water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

For strain TA97 tested in the absence of S9

Controls 4

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

Water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

For strain TA98 tested in the absence of S9

Details on test system and experimental conditions:

PREINCUBATION : In the standard protocol (preincubation) for conducting the Ames assay, a test tube containing a suspension of one strain of Salmonella typhimurium plus S9 mix or plain buffer without S9, is incubated for 20 minutes at 37º C with the test chemical.
Control cultures, with all the same ingredients except the test chemical, are also incubated. In addition, positive control cultures are prepared; these contain the particular bacterial tester strain under investigation, the various culture ingredients, and a known potent mutagen.
After 20 minutes, agar is added to the cultures and the contents of the tubes are thoroughly mixed and poured onto the surface of Petri dishes containing standard bacterial culture medium. The plates are incubated, and bacterial colonies that do not require an excess of supplemental histidine appear and grow.
These colonies are comprised of bacteria that have undergone reverse mutation to restore function of the histidine-manufacturing gene. The number of colonies is usually counted after 2 days.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Test results 2

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Test results 3

Species / strain:

S. typhimurium TA 97

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Remarks on result:

other: No mutagenic potential observed

Any other information on results incl. tables

 Strain: TA100

Dose	No Activation  (Negative)		No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)		10% RLI  (Negative)		10% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	111	9.5	123	2.7	126	2.3	120	5	138	4.7	116	6.4
100	118	16.9	134	6.3	138	7.3	119	9.1	126	3.7	128	1.5
333	123	7.8	120	5	136	3.3	120	5.8	134	5.3	134	11.3
1000	105	4.6	123	6.1	132	5.7	131	2.9	122	10.5	136	4.9
3333	127	3.8	133	4.2	126	3.2	137	4.7	125	3.5	119	5.5
10000	122	5.3	110	5.5	116	8	116	6	136	10.2	126	8.5
Positive Control	845	23.7	861	20.1	483	10.7	544	17.6	548	16.5	592	7.9

 

Strain: TA1535

Dose	No Activation  (Negative)		No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)		10% RLI  (Negative)		10% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	10	1	12	1.5	11	1.2	12	1.2	14	1.9	13	1.5
100	10	.3	10	.9	10	0	11	.7	10	.9	9	.7
333	10	1.2	9	.6	10	.7	11	.9	13	3	8	.7
1000	10	.3	10	2.1	11	2.3	9	.3	13	1.3	11	1.3
3333	9	.3	9	.6	9	.9	12	1.2	12	2	10	1.2
10000	9	.3	10	.6	10	1	11	2.5	10	1	11	2
Positive Control	890	19.6	857	11.9	90	6.7	94	5.4	117	7.5	142	4.9

Strain: TA97

Dose	No Activation  (Negative)		No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)		10% RLI  (Negative)		10% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	140	6.5	163	9.7	165	8.6	167	15.1	189	11.1	178	3.2
100	148	11	161	8.8	164	3.2	175	7.2	194	3.5	191	3.2
333	140	2.5	158	14.7	170	10.8	164	3	184	8.5	163	3.7
1000	154	3.8	167	5.5	164	17.7	176	6.4	178	5.2	165	8.5
3333	131	4.2	138	6.1	165	12.3	164	8.7	199	.3	180	16
10000	120	3.8	163	21.8	171	2.1	156	15.6	192	9.5	183	20.7
Positive Control	426	16.4	458	5.6	510	7	631	24.1	541	13.5	565	22.6

Applicant's summary and conclusion

Conclusions:

In bacterial gene mutation study, the test chemical in water at doses 100-10000 µg/Plate were observed to be not mutagenic in Salmonella typhimurium strains TA100,TA1535,TA97 and TA98 with and without addition of S9 liver fractions from Aroclor induced hamsters. The same has been observed for rats as well.

Executive summary:

Genetic toxicity in vitro study was assessed for test chemical. For this purpose AMES test was performed.The test material was exposed to Salmonella typhimurium TA100, TA1535, TA98 and TA97 in the presence and absence of metabolic activation S9. The concentration of test material used in the presence and absence of metabolic activation were 0, 10, 30,100, 333, 1000 and 3333 µg/plate. Mutagenic effects were observed in neither strains, in the presence and absence of metabolic activation. Therefore test chemical was considered not to be mutagenic in Salmonella typhimurium TA100, TA1535, TA98 and TA97 by AMES test. Hence the substance cannot be classified as gene mutant in vitro.