3-(diethylamino)-7-oxo-7H-[1]benzopyrano[3',2':3,4]pyrido[1,2-a]benzimidazole-6-carbonitrile

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:

weight of evidence

Study period:

1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Test guideline 3

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

3-(diethylamino)-7-oxo-7H-[1]benzopyrano[3',2':3,4]pyrido[1,2-a]benzimidazole-6-carbonitrile

EC Number:

257-884-8

EC Name:

3-(diethylamino)-7-oxo-7H-[1]benzopyrano[3',2':3,4]pyrido[1,2-a]benzimidazole-6-carbonitrile

Cas Number:

52372-36-8

Molecular formula:

C23H18N4O2

IUPAC Name:

17-(diethylamino)-11-oxo-14-oxa-3,10-diazapentacyclo[11.8.0.0²,¹⁰.0⁴,⁹.0¹⁵,²⁰]henicosa-1(21),2,4,6,8,12,15(20),16,18-nonaene-12-carbonitrile

Test material form:

solid: particulate/powder

Details on test material:

Disperse Red 362

Method

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

rat liver homogenate

Test concentrations with justification for top dose:

First experiment 25, 125, 625, 3125 and 6250 µg/plate - all strains
Second experiment: 4, 20, 100, 500 and 1000 µg/plate - TA 100

Vehicle / solvent:

Acetonitril

Controls

Controls 1

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 S9

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: N-methyl-N-nitro-N-nitrosoguanidine

Remarks:

TA1535, TA100 w/o S9

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylendiamine

Remarks:

TA98 w/o S9

Controls 4

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 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

E. coli WP2 uvrA w/o S9

Details on test system and experimental conditions:

METHOD OF APPLICATION:
1. Plate incorporation
- Incubation: 37°C 48 to 72 hours

- Triplicate plating


DETERMINATION OF CYTOTOXICITY
- decrease in number of revertants
- clearing or diminuation of background lawn
- reduction of titer

Evaluation criteria:

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

Negative: number of revertants are within historical negative control range in two independent experiments

Results and discussion

Test results

Test results 1

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 100

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 6

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with

Genotoxicity:

positive

Remarks:

at precipitating concentrations

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 7

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with

Genotoxicity:

positive

Remarks:

precipitation from 625 µg/plate onwards

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 8

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with

Genotoxicity:

positive

Remarks:

at precipitating concentrations

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 9

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with

Genotoxicity:

positive

Remarks:

precipitation from 500 µg/plate onwards

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: from 500 µg/plate onwards

Applicant's summary and conclusion

Conclusions:

The test substance is mutagenic in the Salmonella typhimurium reverse mutation assay under the experimental conditions chosen here.

Executive summary:

The test substance was tested for its mutagenic potential based on the ability to induce mutations in selected loci of several bacterial strains, i.e. Salmonella typhimurium and Escherichia coli, in a reverse mutation assay.The tester strains Salmonella typhimurium TA 1535, TA 100, TA 1537, TA 98 and E.coli WP2 uvrA were used in the standard plate test.

Dose ranges from 25 to 6250 µg/plate were used in all strains in the first experiment and 4 to 1000 µg/plate in TA 100 in the second experiment with and without metabolic activation.

Solubility: Precipitation of the test substance was found from about 500 µg/plate onwards

Toxicity: A slight decrease in the number of revertants was observed from about 3125 µg/plate onwards

Mutagenicity: An increase in the number of his revertants was observed after the addition of a metabolizing system using the strains TA 1535, TA 100, TA 1537 and TA 98.

Conclusion: According to the results of the present study, the test substance is mutagenic in the Salmonella typhimurium reverse mutation assay under the experimental conditions chosen here.