Cuprate(4-), [2-[[[[2-hydroxy-3-sulfo-5-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]azo]phenylmethyl]azo]-4-sulfobenzoato(6-)]-, sodium

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:

April 12,1983 to April 22,1983

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliance with international guideline

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Cuprate(4-), [2-[[[[2-hydroxy-3-sulfo-5-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]azo]phenylmethyl]azo]-4-sulfobenzoato(6-)]-, sodium

EC Number:

291-103-1

EC Name:

Cuprate(4-), [2-[[[[2-hydroxy-3-sulfo-5-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]azo]phenylmethyl]azo]-4-sulfobenzoato(6-)]-, sodium

Cas Number:

90341-71-2

Molecular formula:

C22H14CuN4Na4O15S4

IUPAC Name:

Cuprate(4-), [2-[[[[2-hydroxy-3-sulfo-5-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]azo]phenylmethyl]azo]-4-sulfobenzoato(6-)]-, sodium

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material: Remazolbrillantblau 88

Method

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

Doses: 0, 4 , 20, 100, 500, 2500, 10000 µg/plate

Vehicle / solvent:

- Vehicle: water

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

other: N-Methyl-N'-nitro-N-nitrosoguanidine

Remarks:

without S9

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

other: 2-aminoantracene

Remarks:

with S9

Details on test system and experimental conditions:

METHOD OF APPLICATION: in AGAR
Top agar is prepared for the Salmonella strains by mixing 100 ml agar (0,6 % agar, 0,5 % NaCl) with 10 ml of a 0,5 mM histidine-biotin solution. With E. coli histidine is replaced by tryptophan (2,5 mL, 0,5 mM). The following ingredients are added (in order) to 2 ml of molten top ager at 45°C:
0,1 mL test compound solution
0,1 mL of an overnight nutrient broth culture of the bacterial tester strain
0,5 mL S-9 Mix (if required) or buffer
After mixing, the liquid is poured into a petridish with minimal agar (1,5 % agar, Vogel-Bonner E medium with 2 % glucose). After incubation for 48 to 72 hours at 37°C in the dark, colonies (his revertants) are counted.


- Preincubation period:48 to 72 hrs

NUMBER OF REPLICATIONS: 3

OTHER:
Bacteria are grown overnight in nutrient broth (25 g Oxoid Nutrient Broth 2 /Liter) at 37°C.

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 nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1538

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:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
In combination with the main experiment, toxicity testing was performed as follows: 0,1 ml of the different dilutions of the test compound were thoroughly mixed with 0,1 mL of 10^-6 dilution of the overnight culture of TA 100 and plated with histidine and biotin rich top agar (3 plates per dose). The solvent control is compared with the number of colonies per plate in the presence of the test compound. Results are given as a ratio of these values (= surviving fraction).

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Deviation:

This tes twas performed according to the methods described. No unforeseen circumstances were observed which have affected the quality and integrity of this report.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test compound did not cause a significant increase in the number of revertant colonies with any of the tester strains neither in the absence nor presence of S-9 Mix. No dose dependent effect was obtained . It is concluded that the test substance is not mutagenic in these bacterial test systems neither in the absence nor in the presence of an exogenous metabolizing system.

Executive summary:

Remazolbrillantblau BB was tested for mutagenicity with the strains TA 100, TA 1535, TA 1537, TA 1538, TA 98 of Salmonella typhimurium and Escherichia coli WP2 uvrA.The mutagenicity studies were conducted in the absence and in the presence of a metabolizing system derived from rat liver homogenate. A doserange of 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 positiv control compounds gave the expected increase in the number of revertant colonies.

Toxicity:

The test compound proved to be not toxic to the bacteria. 5000 µg/plate was chosen as top dose level for the mutagenicity study.

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 metabolic activation system, treatment of the cells with Remazolbrillantblau BB did not result in relevant increases in the number of revertant colonies.

Summarizing, it can be stated that Remazolbrillantblau BB is not mutagenic in these bacterial test systems neither with nor without exogenous metabolic activation at the dose levels investigated.