[No public or meaningful name is available]

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:

01-Mar-2006 to 14-Apr-2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

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

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

No. 338 Red

IUPAC Name:

No. 338 Red

Details on test material:

- Substance type: Red powder
- Lot/batch No.: 50670

Method

Target gene:

- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene

Species / strain

Species / strain / cell type:

bacteria, other: Salmonella typhimurium : TA1535, TA1537, TA98 et TA100. Escherichia coli : WP2uvrA-

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Concentration range in the range finding test (without and with metabolic activation): 15, 50, 150, 500, 1500 and 5000 µg/plate
Concentration range in the main test (without and with metabolic activation): 15, 50, 150, 500, 1500 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Test compound was soluble in DMSO and DMSO has been accepted and approved by authorities and international guidelines

Controls

Controls 1

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

without S9

Migrated to IUCLID6: for WP2uvrA, TA100 and TA1535

Controls 2

Positive control substance:

9-aminoacridine

Remarks:

without S9

Migrated to IUCLID6: for TA1537

Controls 3

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

without S9

Migrated to IUCLID6: for TA98

Controls 4

Positive control substance:

other: 2-aminoanthracene for WP2uvrA, TA100, TA1535 and TA1537

Remarks:

with S9

Controls 5

Positive control substance:

benzo(a)pyrene

Remarks:

with S9

Migrated to IUCLID6: for TA98

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hour

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in triplicate in each strain. Two independent experiments were conducted.

NUMBER OF CELLS EVALUATED: 10E8 per plate

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies.

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was determined.

Evaluation criteria:

There are several criteria for determining a positive result, such as a dose-related increase in the revertant frequency over the dose range tested and/or a reproduceble increase at one or more concentrations in at least one bacterial strain with or without S9.

Results and discussion

Test results

Test results 1

Species / strain:

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

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

( 5000 µg/plate)

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

( 5000 µg/plate)

Test results 3

Species / strain:

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

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

( 5000 µg/plate)

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

( 5000 µg/plate)

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation was observed at the dose level of 5000 µg/plate.
- A pink colour, becoming darker with increasing concentrations, was noted from 150 µg/plate and above

RANGE-FINDING/SCREENING STUDIES:
- Toxicity was observed at the dose level of 5000 µg/plate in the absence and presence of S9-mix in all tester strains.

COMPARISON WITH HISTORICAL CONTROL DATA:
- The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

MUTAGENICITY
- In the absence of S9, no increases in the number of revertant colonies were observed

- In the presence of S9 the following dose related increases in the number of revertant colonies were observed
Range finding test
TA1535: 6.6-fold
TA1537: 4.7-fold
TA98: 78-fold
TA100: 9.6-fold
WP2uvrA: 9.5-fold

Main test
TA1535: 4.0-fold
TA1537: 15-fold
TA98: 126-fold
TA100: 11-fold
WP2uvrA: 8.8-fold

Applicant's summary and conclusion

Conclusions:

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

The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

The test material was considered to be mutagenic.