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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Follows relevant OECD guidlines, also EPA, EEC-Directive 2000/32/EC L136 Annex 4E, B17 and GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
mammalian cell gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Reaktionsprodukt aus borsaure mit mea and tea
- Molecular formula (if other than submission substance):C8H25BN2O7
- Molecular weight (if other than submission substance):272.1043
- Smiles notation (if other than submission substance):OCCN(CCO)CCO.OCCN.OB(O)O
- InChl (if other than submission substance):Boric acid (H3BO3), reaction products with ethanolamine and triethanolamine
- Structural formula attached as image file (if other than submission substance): see Fig. ATTACHED BELOW
- Substance type:reaction product
- Physical state:liquid
- Analytical purity:100%
- Impurities (identity and concentrations):none known
- Composition of test material, percentage of components:19.3% boric acid, 17.7 monethanolamine, 63.0% triethanolamine
- Isomers composition:
- Purity test date:2001-11-13
- Lot/batch No.:ESDB01441
- Expiration date of the lot/batch:2003-07-12
- Radiochemical purity (if radiolabelling):
- Specific activity (if radiolabelling):
- Locations of the label (if radiolabelling):
- Expiration date of radiochemical substance (if radiolabelling):
- Stability under test conditions:
- Storage condition of test material:
- Other:

Method

Target gene:
HPRT
Species / strain
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial supernatent fraction from liver of Aroclor 1254-treated Sprague-Dawley rats and an NADP generating system (S9-mix)
Test concentrations with justification for top dose:
The test compound was dissolved in cell culture medium and tested at the following concentrations:

without S9 -mix: 10,50,1,00,250,1000,2500 and 5000 µg/ml

with S9 -mix:       10,50,1,00,250,1000,2500 and 5000µg/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: [none]
- Justification for choice of solvent/vehicle:none necessary as test substance was dissolved directly into the test system (MEM)
Controls
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium;
DURATION
- Preincubation period:24 hours
- Exposure duration:4 hours
- Expression time (cells in growth medium):
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells):4 or 5 days

SELECTION AGENT (mutation assays):
SPINDLE INHIBITOR (cytogenetic assays):
STAIN (for cytogenetic assays):mehtylene blue

NUMBER OF REPLICATIONS:6 for each concentration

NUMBER OF CELLS EVALUATED:1000 cells

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other:

OTHER EXAMINATIONS:
- Determination of polyploidy:
- Determination of endoreplication:
- Other:

OTHER:
Evaluation criteria:
The test compound is classified as mutagenic if:
it reproducably induces with one of the test compound concentrations a mutation frequency that is three times higher than the spontaneous mutant frequency in this experiment.
there is a reproducible concentration related increase in the mutation frequency. Such an evaluation may be considered independantly from the enhancement factor for induced mutants.
survival of the corresponding group is at least 30%
Statistics:
Statistical analysis: The biometry of the results of the test compound was performed with the Mann-Whitney U-test (Considered appropriate by reviewer)

Results and discussion

Test results
Species / strain:
Chinese hamster lung fibroblasts (V79)
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:
not applicable
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: monitored, no effects
- Effects of osmolality:monitored, no effects
- Water solubility:>1g/l
- Precipitation: no precipitation
Remarks on result:
other: strain/cell type: lung
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
CAS No. 68512-53-8 did not induce gene mutations, i.e. is not mutagenic, in this HPRT-test with V79 Chinese hamster cells, either in the presence or in the absense of metabolic activation under the conditions described in this report
Executive summary:

In this study the potential of CAS No. :68512 -53 -8 (Batch ESDB01441) to induce gene mutations at the HPRT locus in V 79 cells of the Chinese hamster lung in vitro was investigated.

The test compound was dissolved in cell culture medium and tested at the following concentrations:

without S9 -mix: 10,50,1,00,250,1000,2500 and 5000 µg/ml

with S9 -mix:       10,50,1,00,250,1000,2500 and 5000µg/ml

The concentration ranges were based on the results of preliminary testing for solubility and toxicity. Slight toxicity was observed with and without metabolic activation at the higest concentration.

This GLP study was initiated on 14th Jan. 2002.

Up to the highest investigated dose the test compound induced no relevant and reproducable increase in mutant colony numbers in two independant experimants.

Appropriate reference mutagens used as positive controls showed a significant increase in mutant colonies, thus indicating the sensitivity of the assay, and the efficacy of the S9-mix.

In conclusion, CAS No.:68512 -53 -8 did not induce gene mutations in the HPRT-test with V79 Chinese hamster cells, both in the presence as well as in the absence of a metabolic activation system, under the experimental conditions described.

CAS No.:68512 -53 -8 is not mutagenic in this in vitro HPRT assay with V79 Chinese hamster lung cells