Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Data is from peer reviewed journal

Data source

Reference
Reference Type:
publication
Title:
Evaluation of Laser Dye Mutagenicity Using the Ames/Salmonella Microsome Test
Author:
Barbara J.Y. Wuebbles and James S. Felton
Year:
1985
Bibliographic source:
Environmental Mutagenesis 7511-522 (1985)

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: as per mentioned below
Principles of method if other than guideline:
Method as described by Ames et al 1975
GLP compliance:
not specified
Type of assay:
bacterial gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
Details on test material
- Name of test material (as cited in study report): Coumarin 7
- Molecular formula (if other than submission substance): C20-H19-N3-O2
- Molecular weight (if other than submission substance): 333.389
- Substance type: Organic
- Physical state: Solid/powder
- Purity: No data available
- Impurities (identity and concentrations): contaminants present

Method

Target gene:
Histidine
Species / strain
Species / strain:
other: Salmonella typhimurium strains TA1538, TA98 and TA100
Details on mammalian cell lines (if applicable):
Not applicable
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
no data
Test concentrations with justification for top dose:
0.1 mg/plate and 1 mg/plate
Vehicle:
DMSO
Controls
Negative controls:
not specified
Solvent controls:
not specified
True negative controls:
not specified
Positive controls:
not specified
Positive control substance:
not specified
Remarks:
no data
Details on test system and conditions:
Details on test system and conditions
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period:
- Exposure duration: 2 days
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): No data available
SPINDLE INHIBITOR (cytogenetic assays): No data available
STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: duplicate

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data available

OTHER EXAMINATIONS: No data available
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other: No data available

OTHER: No data available
Evaluation criteria:
A sample was judged mutagenic if it produced greater than twice the spontaneous background colonies at more than one dose or at the highest dose tested.
Statistics:
No data available

Results and discussion

Test results
Species / strain:
other: Salmonella typhimurium strains TA1538, TA98 and TA100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
Additional information on results
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data available
- Effects of osmolality: No data available
- Evaporation from medium: No data available
- Water solubility: No data available
- Precipitation: No data available
- Other confounding effects: Less soluble in DMSO

RANGE-FINDING/SCREENING STUDIES: Spot test was performed with and without activation using the Salmonella typhimurium strains TA1538, TA98 and TA100. The spot test results were inconclusive. The dyes were than screened at 0.1 mg/plate and 1 mg/plate.

COMPARISON WITH HISTORICAL CONTROL DATA: No data available

ADDITIONAL INFORMATION ON CYTOTOXICITY: No data available

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative with and without

The given test material does not induce gene toxicity in the Salmonella typhimurium strains TA1538, TA98 and TA100. Hence it is negative for gene mutation.
Executive summary:

Liquid-dye lasers are being used in a variety of applications because of their versatility. To date there is little information available on many such dyes since there has been minimal testing of their toxicity and mutagenicity. Because laser technology is an important part of a growing number of research and development procedures and the dyes are potentially genotoxic, special precautions and handling procedures may be required.

One of the families of dyes that are involved in the study is coumarins. Amongst the 12 coumarins studied, the test material Coumarin 7 is also studied for mutagenicity. Bacteria were grown overnight in Oxoid nutrient broth, then refrigerated at 4-5OC for a few hours before use. 0.1 ml of bacterial culture was added to 2 ml of 45°C molten top agar containing 0.01 mg histidine HCI and 0.012 mg biotin/ml, followed by the test sample in ≤0.2 ml DMSO. Finally, 0.5 ml of sodium phosphate buffer, pH 7.4 (no activation), or 0.5 ml of Aroclor-induced rat S9 mixture was added, and the mixture was poured on minimal glucose agar plates. Histidine revertant colonies were counted on a Biotran II automated colony counter after a 2-day incubation at 37°C. A sample was judged mutagenic if it produced greater than twice the spontaneous background colonies at more than one dose or at the highest dose tested. To estimate mutagenic potency (revertant/µg) from linear dose-response curves, we used the method of Moore and Felton. All compounds were tested to a level at which they were toxic or to the limits of solubility. Duplicate plates were run on all tests except high-performance liquid chromatography (HPLC) fractions. All results were verified in repeat experiments. In the above mentioned study, coumarin 7, the test material failed to induce gene mutation in theSalmonella typhimuriumstrains TA1538, TA98 and TA100 with and without metabolic activation.

Thus the given test material, Coumarin 7, is negative for gene mutation in vitro.