O-isopropyl ethylthiocarbamate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

O-isopropyl ethylthiocarbamate

EC Number:

205-517-7

EC Name:

O-isopropyl ethylthiocarbamate

Cas Number:

141-98-0

Molecular formula:

C6H13NOS

IUPAC Name:

N-ethyl(propan-2-yloxy)carbothioamide

Constituent 2

Reference substance name:

Carbamothioic acid, ethyl, O-(1-methyl-ethyl) ester

IUPAC Name:

Carbamothioic acid, ethyl, O-(1-methyl-ethyl) ester

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): IPETC
- Physical state: Liquid
- Stability under test conditions: Stable
- Storage condition of test material: ambient

Method

Species / strain

Species / strain / cell type:

mammalian cell line, other: human peripheral lymphocytes

Details on mammalian cell type (if applicable):

Human peripheral blood was obtained by venipuncture from healthy donors known to be without any medication and collected in heparinised vessels. Small innocula of whole blood (0.5 mL) were added to tubes containing 5 mL of complete culture medium. The tubes were sealed and incubated at 37°C with occasional shaking to prevent clumping.

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Post-mitochondrial fraction (S9 fraction) from rats treated with Aroclor 1254 was prepared according to MARON and AMES (1983). S9 was collected from 20 - 30 rats. The pooled fraction was tested for:

Test concentrations with justification for top dose:

10, 26, 103, 257, 1029 and 2571 µg IPETC per mL medium

Vehicle / solvent:

dimethyl sulfoxide (DMSO)

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

DMSO

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

was used as the positive control for the study in the absence of metabolic activation.

Controls 2

Untreated negative controls:

yes

Remarks:

DMSO

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

was used as the positive control for the study in the presence of metabolic activation.

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION

- Exposure duration: 4 hours in experiment 1 and 24 hours in experiment 2

SPINDLE INHIBITOR (cytogenetic assays): colcemid® to accumulate cells in a metaphase-like stage of mitosis (c-metaphase).

Evaluation criteria:

The test item is judged to have mutagenic properties with respect to chromosomal or chromatid change, if the following criteria are fulfilled:
o the number of chromosomal aberrations is significantly increased compared with the solvent control
o the increase observed is concentration-dependent
o both duplicate cultures lead to similar results
o the increase should not occur in the severely cytotoxic range (mitotic index <0.25), as it is known that high cytotoxicity causes artefacts in the form of aberrations in in vitro chromosomal aberration tests
o a reproducible increase in the number of cells with chromosomal aberrations.

Statistics:

The assessment was carried out by a comparison of the number of chromosome aberrations of the samples with those of the solvent control, using the exact test of R. A. FISHER as recommended by the UKEMS guidelines (The United Kingdom Branch of the European Environmental Mutagen Society: Report of the UKEMS subcommittee on guidelines for mutagenicity testing, part III, Statistical evaluation of mutagenicity test data, 1989).

Results and discussion

Test results

Key result

Species / strain:

mammalian cell line, other: human peripheral lymphocytes

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

cytotoxicity was noted in the experiment without and with metabolic activation (24-h or 4-h exposure, respectively) starting at 1029 µg IPETC /mL. Haemolysis was noted at concentrations of 2571 and 5143 µg/mL in both experiments.

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: strain/cell type: human peripheral lymphocytes

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Under the present test conditions, IPETC, tested up to cytotoxic concentrations, in the absence and in the presence of metabolic activation employing two exposure times (without S9) and one exposure time (with S9) revealed no indications of mutagenic properties with respect to chromosomal or chromatid damage.
In the same test, Mitomycin C and cyclophosphamide induced significant damages, which confirmed the validity of this assay.

Executive summary:

The registered substance was tested for its potential to induce chromosome aberrations in an Chromosome Aberration study in mammalian cells (human peripheral lymphocytes) according to OECD Guideline 473 with and without metabolic activation.
Under the present test conditions, IPETC, up to cytotoxic concentrations, in the absence and in the presence of metabolic activation employing two exposure times, revealed no indications of mutagenic properties with respect to chromosomal or chromatid damage. In the test, Mitomycin C and cyclophosphamide induced significant damages, which confirmed the validity of this assay.