Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

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:
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:
Mutagenicity of Azo Dyes Following Metabolism by Different Reductive/Oxidative Systems
Author:
Thomas M. Reid, Kenneth C. Morton, Ching Y. Wang, and Charles M. King
Year:
1984
Bibliographic source:
Environmental Mutagenesis 6: 705-717 (1984)

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
Gene mutation toxicity study was performed for Direct blue 218 to evaluate its mutagenic nature
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: amorphous powder
Details on test material:
Name of the test chemical: Tetrasodium [μ-[[3,3'-[(3,3'-dihydroxy[1,1'-biphenyl]-4,4'-diyl)bis(azo)]bis[5-amino-4-hydroxynaphthalene-2,7-disulphonato]](8-)]]dicuprate(4-)
Common Name: C.I Direct Blue 218
IUPAC name: tetrasodium (3E)-5-amino-3-{2-[4-(4-{2-[(2E)-8-amino-1-oxo-3,6-disulfonato-1,2-dihydronaphthalen-2-ylidene]hydrazin-1-yl}-3- hydroxyphenyl)-2-hydroxyphenyl]hydrazin-1-ylidene}-4-oxo-3,4-dihydronaphthalene-2,7-disulfonate dicopper
Molecular Formula: C32H20Cu2N6Na4O16S4
Molecular Weight: 1087.84 g/mol
SMILES Notation: c12c3c(c(S(=O)(=O)[O-])cc1cc(S(=O)(=O)[O])cc2N)N=Nc1ccc(cc1O[Cu]O3)c1cc2c(N=Nc3c(cc4c(c3O[Cu]O2)c(cc(c4)S(=O) (=O)[O-])N)S(=O)(=O)[O-])cc1.[Na+].[Na+].[Na+].[Na+]
InChI: 1S/C32H24N6O16S4.2Cu.4Na/c33-19-11-17(55(43,44)45)5-15-9-25(57(49,50)51)29(31(41)27(15)19)37-35-21-3-1-13(7-23(21)39) 14-2-4-22(24(40)8-14)36-38-30-26(58(52,53)54)10-16-6-18(56(46,47)48)12-20(34)28(16)32(30)42;;;;;;/h1-12,39-42H,33-34H2,(H,43,44,45)(H,46,47,48)(H,49,50,51)(H,52,53,54);;;;;;/q;2*+2;4*+1/p-8/b37-35-,38-36-;;;;;;
Substance Type: Organic
Physical State: Solid Deep purple to dark blue amorphous powder
Specific details on test material used for the study:
- Name of test material: Direct blue 218
- IUPAC name: Copper,[tetrahydrogen-3,3'-[(3,3'-dihydroxy-4,4'-biphenylylene)bis(azo)]bis[5-amino-4-hydroxy-2,7-naphthalenedisulfonato](4-)]di-,tetrasodium salt (7CI)
- Molecular formula: C32H16Cu2N6O16S4.4Na
- Molecular weight: 1087.82 g/mol
- Substance type: Organic
- Physical state: No data
- Purity: >98%
- Impurities (identity and concentrations): <2%

Method

Target gene:
Histidine
Species / strain
Species / strain:
S. typhimurium TA 1538
Details on mammalian cell lines (if applicable):
Not applicable
Additional strain characteristics:
not specified
Cytokinesis block (if used):
No data
Metabolic activation:
with and without
Metabolic activation system:
Rat S9 was prepared from the livers of Aroclor-induced male Fisher rats and hamster S9 was prepared from uninduced female hamsters
Test concentrations with justification for top dose:
0, 0.25 or 0.50 µmole/plate
Vehicle:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The test chemical was soluble in DMSO
Controls
Negative controls:
not specified
Solvent controls:
yes
Remarks:
DMSO
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
other: Benzidine congeners (Dimethoxybenzidine)
Details on test system and conditions:
METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 mins
- Exposure duration: 72 hr
- Expression time (cells in growth medium): 72 hr
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

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

NUMBER OF REPLICATIONS: Triplicate

NUMBER OF CELLS EVALUATED: No data

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

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

OTHER: No data
Rationale for test conditions:
No data
Evaluation criteria:
The plates were observed for revertants
Statistics:
No data

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
not specified
Positive controls valid:
yes
Remarks on result:
other: No mutagenic potential
Additional information on results:
No data

Any other information on results incl. tables

Table: Mutation data for the test chemical Direct blue 218 and the positive control chemical

Dose (µmole/plate)

Reduction system (activation system)

Bacterial (rat)

None (rat)

FMN (hamster)

None(hamster)

Bacterial (hamster)

Direct blue 218

 

 

 

 

 

0

43

35

33

-

-

0.25

40 (63)

14

27

-

-

0.50

47 (47)

16

21

-

-

Dimethoxybenzidine

 

 

 

 

 

0

43

35

33

27

26

0.25

790

843

235

169

108

0.50

1073

1203

316

125

167

1.0

1491

1287

366

170

124

Applicant's summary and conclusion

Conclusions:
Direct blue 218 did not induce gene mutation in the Salmonella typhimurium strain TA1538 both in the presence and absence of rat and hamster liver S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.
Executive summary:

Gene mutation toxicity study was performed for Direct blue 218 to evaluate its mutagenic nature. The study was performed as per the preincubation protocol and reduction of the test chemical using Salmonella typhimurium strain RA1538 both in the presence and absence of rat and hamster liver S9 metabolic activation system at doses of 0, 0.25 or 0.50 µmole/plate. DMSO was used at the vehicle. The plates were incubated for 48 hrs after 20 mins preincubation before the evaluation of the revertant colonies could be made. Concurrent solvent and negative control chemicals were also included in the study. Direct blue 218 did notinduce gene mutation in theSalmonella typhimurium strain TA1538 both in the presence and absence of rat and hamster liver S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.