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Reaction mass of isomers disodium 8-(2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl)-5-(2-{2,5-dimethyl-4-[2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl)naphthalene-2-sulfonate, disodium 5-(2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl)-8-(2-{2,5-dimethyl-4-[2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl)naphthalene-2-sulfonate
EC number: 700-946-0 | CAS number: -
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
- Study period:
- 2013-03-19 to 2013-05-03
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 013
- Report date:
- 2013
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- tetrasodium 5-[(1E)-2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl]-8-[(1E)-2-{2,5-dimethyl-4-[(1E)-2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl]naphthalene-2-sulfonate 8-[(1E)-2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl]-5-[(1E)-2-{2,5-dimethyl-4-[(1E)-2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl]naphthalene-2-sulfonate
- EC Number:
- 700-946-0
- Molecular formula:
- C40H35N7Na2O6S2
- IUPAC Name:
- tetrasodium 5-[(1E)-2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl]-8-[(1E)-2-{2,5-dimethyl-4-[(1E)-2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl]naphthalene-2-sulfonate 8-[(1E)-2-{4-[benzyl(ethyl)amino]-2-methylphenyl}diazen-1-yl]-5-[(1E)-2-{2,5-dimethyl-4-[(1E)-2-(3-sulfonatophenyl)diazen-1-yl]phenyl}diazen-1-yl]naphthalene-2-sulfonate
- Test material form:
- solid: particulate/powder
1
Method
- Target gene:
- His
Species / strain
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
- Additional strain / cell type characteristics:
- not specified
- Metabolic activation:
- with and without
- Metabolic activation system:
- rat S9 liver microsomal fraction (Experiment I) and hamster S9 liver microsomal fraction Experiment II)
- Test concentrations with justification for top dose:
- Experiment I:
31.6, 100, 316, 1000, 2500 and 5000 µg/plate
Experiment II:
10.0, 31.6, 100, 316, 1000, 2500 and 5000 µg/plate - Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: DMSO
Controls
- Untreated negative controls:
- yes
- Remarks:
- A. dest., BSL Lot No. 130301, 130306, 130408
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO, Applichem Lot No. 2M005967 and 2K005471
- Positive controls:
- yes
- Positive control substance:
- other:
- Remarks:
- Positive control -S9: sodium azide for tester strains TA100, TA1535; 4-nitro-o-phenylene-diamine for TA98, TA1537; methylmethanesulfonate for TA102. Positive control +S9: 2-aminoanthracene for all strains except Congo Red TA 98, TA 100 +S9 (hamster)
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in agar (plate incorporation); preincubation
DURATION
- Preincubation period: 60 min; 100 µL of the test item preparation was pre-incubated with the tester strains (100 µL) and sterile buffer or the metabolic activation system (500 µL) for 60 min at 37 °C prior to adding the overlay agar (2000 µL) and pouring onto the surface of a minimal agar plate.
- Exposure duration: at least 48 h; After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.
NUMBER OF REPLICATIONS: For each strain and dose level, including the controls, three plates were used.
DETERMINATION OF CYTOTOXICITY
- Method: clearing or rather diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately ≤ 0.5 in relation to the solvent control - Evaluation criteria:
- The Mutation Factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).
A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs in at least one tester strain with or without metabolic activation.
A biologically relevant increase is described as follows:
- if in tester strains TA 98, TA 100 and E.coli the number of reversions is at least twice as high
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher than the reversion rate of the solvent control. - Statistics:
- According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.
Results and discussion
Test resultsopen allclose all
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- with
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- 2500 ug/plate and higher in experiment I 1000 ug/plate and higher in experiment II
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- 100 ug/plate and higher in experiment II
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- 2500 ug/plate and higher in experiment I 100 ug/plate and higher in experiment II
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- 2500 ug/plate and higher in experiment I
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).
Toxic effects of the test item were noted in two tester strains used in experiment I and in three tester strains in experiment II.
In experiment I toxic effects of the test item were observed in tester strain TA 100 at concentrations of 2500 µg/plate and higher (without metabolic activation). In tester strain TA 1537 toxic effects of the test item were noted at concentrations of 2500 µg/plate and higher (without and with metabolic activation).
In experiment II toxic effects of the test item were noted in tester strain TA 100 at concentrations of 1000 µg/plate and higher (without metabolic activation). In tester strains TA 1535 and TA 1537 toxic effects of the test item were noted at concentrations of 100 µg/plate and higher (without metabolic activation).
No biologically relevant increases in revertant colony numbers were noted in tester strains TA 98, TA 100, TA 1535 and TA 1537. Biologically relevant increases of revertant colony numbers were observed in tester strain E. coli WP2 uvrA at concentrations of 1000 µg/plate and 2500 µg/plate in experiment II (with metabolic activation). The threshold value of 2.0 was exceeded and a maximum mutation factor of 2.4 was reached at a concentration of 2500 µg/plate (with metabolic activation).
Applicant's summary and conclusion
- Conclusions:
- In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, test item caused gene mutations by base pair changes and others in the genome of the tester strain E. coli WP2 uvrA.
Therefore, test item is considered to be mutagenic in this bacterial reverse mutation assay. - Executive summary:
In a reverse gene mutation assay in bacteria, strains TA98, TA100, TA1535 TA1537 of S. typhimurium and E.coli WP2 uvrA were exposed to the test item at concentrations of 31.6, 100, 316, 1000, 2500 and 5000 µg/plate (experiment I) and 10.0, 31.6, 100, 316, 1000, 2500 and 5000 µg/plate (experiment II), in the presence and absence of mammalian metabolic activation according to the plate incorporation method (experiment I) and the pre-incubation method (experiment II).
Test item was tested up to the limit concentration of 5000 µg/plate in all tester strains used.
The positive controls induced the appropriate responses in the corresponding strains. There was evidence of induced mutant colonies over background.
This study is classified as acceptable. This study satisfies the requirement for Test Guideline OPPTS 870.5100; OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.
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