Reaction products of 4-amino-5-hydroxynaphthalene-2,7-disulfonic acid, coupled twice with diazotized 2-[(4-aminophenyl)sulfonyl]ethyl hydrogen sulfate, sodium salts

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:

28. Aug. to 25. Oct. 1985

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Report date:

1985

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Method

Target gene:

his operon

Species / strain

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S-9 mix from rat (Ames) and hamster (Prival)

Test concentrations with justification for top dose:

Preliminary Test: 4, 20, 100, 500, 2500, 10000 µg/plate
Main Test: 4, 20, 100, 500, 2500, 5000 µg/plate

Vehicle / solvent:

aqua bidest

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

all strains with rat S9; TA 1535, TA 1537, and TA 100 with hamster S9

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

all strains with rat S9

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: benzidine

Remarks:

TA98 with hamster S9

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA100, TA1535 w/o S9

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

TA1537 w/o S9

Controls 6

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

TA98 w/o S9

Details on test system and experimental conditions:

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

DURATION
- Preincubation period: 30 min (Prival)
- Expression time (cells in growth medium): 48 to 72 hours

DETERMINATION OF CYTOTOXICITY
- Method: thinning of bacterial lawn; reduced rate of spontaneously occurring colonies (surviving fraction)

Evaluation criteria:

Cytotoxicity: reduced growth rate or thinning of bacterial lawn
Positive:
- dose-related and reproducible increase in number of revertant colonies
- doubling of spontaneous mutation rate in at least one tester strains either with or without S9

Statistics:

-

Results and discussion

Test results

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 nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results:
negative

The test substance is not mutagenic in the standard plate test (Ames Test) and in the preincubation method according to Prival.

Executive summary:

The test item Reactive Black 5 was tested for mutagenicity with the strains TA100, TA1535, TA1537, TA98 of Salmonella typhimurium.

The mutagenicity studies were conducted in the standard plate test (Ames Test) and in a modified preincubation test (Prival Test). The studies were performed in the absence and in the presence of a metabolizing system derived from rat or hamster liver homogenate. A dose range of six different doses from 4 µg/plate to 5000 µg/plate was used.

Control plates without mutagen showed that the number of spontaneous revertant colonies was similar to that described in the literature. All the positive control compounds gave the expected increase in the number of revertant colonies.

Toxicity: The test compound proved to be not toxic to the bacterial strains.

5000 µg/plate was chosen as top dose level for the mutagenicity study.

a) Ames test:

Mutagenicity: In the absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Also in the presence of a metabolic activation system, treatment of the cells with the the test item

b) Prival Test:

In the presence of hamster Liver S9 using the preincubation method according to Prival the test item 'fl. getrocknet' did not induce a significant increase in the number of revertant colonies, with any of the tester strains.

Summarizing, it can be stated that the test item is not mutagenic in the standard plate test (Ames Test) and in the preincubation method according to Prival.