pentasodium 4-hydroxy-7-[(sulfonatomethyl)amino]-3-[(4-{[2-(sulfonatooxy)ethyl]sulfonyl}phenyl)diazenyl]-8-[(2-sulfonato-4-{[2-(sulfonatooxy)ethyl]sulfonyl}phenyl)diazenyl]naphthalene-2-sulfonate

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

Study period:

From Jun. 24, 2002 to Aug. 14, 2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted according to OECD Guideline 471, EPA OPPTS 870.5100, EU Method B.13/14. and Japanese Substance Control Law (JSCL) Test Guideline III.1 in compliance with GLP

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

Japanese Substance Control Law (JSCL) Test Guideline III.1

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material (as cited in study report): Reaktiv-Scharlach F01-0467

Method

Target gene:

Not applicable

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

Plate incorporation test:
a: without metabolic activation:
50, 160, 500, 1600 and 5000 µg/plate
b: with metabolic activation:
50,160, 500, 1600 and 5000 µg /plate

Preincubation test:
a: without metabolic activation:
50, 160, 500, 1600 and 5000 µg/plate
b: with metabolic activation:
50, 160, 500, 1600 and 5000 µg /plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Soluble in DMSO

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(without metabolic activation)

Positive control substance:

sodium azide

Remarks:

None

Migrated to IUCLID6: (for strain TA 100 and TA 1535)

Controls 2

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(without metabolic activation)

Positive control substance:

9-aminoacridine

Remarks:

None

Migrated to IUCLID6: (for strain TA 1537)

Controls 3

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(without metabolic activation)

Positive control substance:

2-nitrofluorene

Remarks:

None

Migrated to IUCLID6: (for strain TA 98)

Controls 4

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(without metabolic activation)

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

None

Migrated to IUCLID6: (for strain WP2uvrA)

Controls 5

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(with metabolic activation-10 % rat liver)

Positive control substance:

other: 2-aminoanthracene for all strains

Remarks:

None

Controls 6

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(with metabolic activation-30 % Syrian golden hamster liver and preincubation)

Positive control substance:

other: 2-aminoanthracene for strain TA 100, TA 1535, TA 1537 and WP2uvrA

Remarks:

None

Controls 7

Untreated negative controls:

yes

Remarks:

(untreated control)

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

(with metabolic activation-30 % Syrian golden hamster liver and preincubation)

Positive control substance:

congo red

Remarks:

None

Migrated to IUCLID6: (for strain TA 98)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation) and preincubation

DURATION
- Incubation period: 48 h at approx. 37 °C
NUMBER OF REPLICATIONS: Three

DETERMINATION OF CYTOTOXICITY
- Method: A reduction in the number of spontaneously occurring colonies and visible thinning of the bacterial lawn were used as toxicity indicators.
Thinning of the bacterial lawn was evaluated microscopically.

Evaluation criteria:

The test substance was considered positive if
(a) at least a 2-fold increase in the mean number of revertants per plate of at least one of the tester strains over the mean number of revertants per plate of the appropriate vehicle control at complete bacterial background lawn
(b) a dose-related increase in the mean number of revertants per plate of at least one of the tester strains over the mean number of revertants per plate of the appropriate vehicle control in at least two to three concentrations of the test compound at complete bacterial background lawn.

If the test substance does not achieve either of the above criteria, it is considered to show no evidence of mutagenic activity in this system

Statistics:

Not reported (According to the OECD guideline 471, a statistical analysis of the data was not mandatory).

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: The test substance did not precipitate on the plates up to the highest investigated dose of 5000 µg/plate.


COMPARISON WITH HISTORICAL CONTROL DATA: Yes

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

Under the test conditions, the test substance is not mutagenic in the absence and presence of exogenous metabolic activation in the standard plate incorporation and the preincubation tests.

Executive summary:

A study was conducted to determine the mutagenic potential of test substance according to OECD Guideline 471, EPA OPPTS 870.5100, EU method B.13/14.and Japanese Substance Control Law (JSCL) Test Guideline III.1 in compliance with GLP.

 

Strains TA 100, TA 1535, TA 1537 and TA 98 of Salmonella typhimurium and Escherichia coli WP2uvrA were used in the mutagenicity assay.

Two independent mutagenicity studies were conducted, one as the standard plate test with the plate incorporation method and the other as a modified preincubation test (Prival test). The studies were performed in the absence and in the presence of a metabolizing system derived from a rat liver homogenate or a hamster liver homogenate. For both studies, the compound was dissolved in deionized water, and each bacterial strain was exposed to 5 dose levels.

Doses for both studies ranged from 50 to 5000µg/plate.

Control plates without mutagen showed that the number of spontaneous revertant colonies was within the laboratory's historical control range. All the positive control compounds gave the expected increase in the number of revertant colonies.

In the preincubation test the number of revertant colonies of the solvent controls in the presence of

S9-mix and the number of revertants colonies of the positive controls in the absence of S9-mix

with the strain WP2uvrA were out of the historical control data range, but the criteria for the

negative/positive responses were fulfilled.

Toxicity: In the mutagenicity experiments toxicity was not observed either with or without metabolic activation.

Plate incorporation test: Both in the absence and in the presence of rat liver(10 % (v/v)) metabolic activation system the test substance did not result in relevant increases in the number of revertants in any of the bacterial strains.

In the absence and in the presence of hamster liver S9-mix (30 % (v/v)) using the preincubation method according to Prival, test substance did not result in relevant increases in the number of revertant colonies with any of the tester strains.

Under the test conditions, the test substance is not mutagenic in the absence and presence of exogenous metabolic activation in the standard plate incorporation and the preincubation tests.