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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information
Ames Methylenebis (dibutyldithiocarbamate) is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation) and is considered to be non-mutagenic to bacteria (Safepharm Limited, 2005). Chromosome aberration Methylenebis (dibutyldithiocarbamate) is negative in human lymphocytes and is considered to be non-clastogenic to human lymphocytes (Safepharm Limited, 2005). Mammilian gene mutation Methylenebis (dibutyldithiocarbamate) is negative using mouse lymphoma L5178Y cells and is not mutagenic in the mouse lymphoma L5178Y test system (WIL Research Europe B.V., 2012).
Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 02 July to 21 August 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted in accordance with relevant guideline and under GLP conditions.
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
yes
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Species / strain / cell type:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
0.03, 0.1, 0.3, 1, 3, 10, 33 and 100 μg/mL
Vehicle / solvent:
Dimethyl sulfoxide
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
In the absence of S9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
In the presence of S9 mix
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Preincubation period:
- Exposure duration: 3 hours (first experiment) and 24 hours (second experiment)
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells):

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

NUMBER OF REPLICATIONS:

NUMBER OF CELLS EVALUATED: 9.6E+05 cells/concentration

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

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

OTHER: Observations/measurements in the study were recorded electronically using the following programme: REES Centron Environmental Monitoring system version SQL 2.0: Temperature and humidity.
Evaluation criteria:
In addition to the criteria stated below, any increase of the mutation frequency should be evaluated for its biological relevance including a comparison of the results with the historical control data range.

The global evaluation factor (GEF) has been defined by the IWGT as the mean of the negative/solvent MF distribution plus one standard deviation. For the micro well version of the assay the GEF is 126.

A test substance is considered positive (mutagenic) in the mutation assay if it induces a MF of more than MF(controls) + 126 in a dose-dependent manner. An observed increase should be biologically relevant and will be compared with the historical control data range.

A test substance is considered equivocal (questionable) in the mutation assay if no clear conclusion for positive or negative result can be made after an additional confirmation study.

A test substance is considered negative (not mutagenic) in the mutation assay if:
a) None of the tested concentrations reaches a mutation frequency of MF(controls) + 126.
b) The results are confirmed in an independently repeated test.
Statistics:
None stated
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
up to 100 μg/mL
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
In the absence of S9-mix, the test substance did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the duration of treatment time.

In the presence of S9-mix, the test substance did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the concentration of the S9 for metabolic activation.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Conclusions:
Interpretation of results (migrated information):
negative

It is concluded that Methylenebis (dibutyldithiocarbamate) is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described in this report.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Ames

Key study:

According to an OECD 471 conducted to GLP, Methylenebis (dibutyldithiocarbamate) is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation). Methylenebis (dibutyldithiocarbamate) is considered to be non-mutagenic to bacteria under the conditions of this test (Safepharm Limited, 2005).

Supporting study:

According to an OECD 471 conducted to GLP, Methylenebis (dibutyldithiocarbamate) is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation). Methylenebis (dibutyldithiocarbamate) is considered to be non-mutagenic to bacteria under the conditions of this test (Safepharm Limited, 1997).

Chromosome aberration

According to an OECD 476, conducted to GLP, Methylenebis (dibutyldithiocarbamate) is negative in human lymphocytes. Methylenebis (dibutyldithiocarbamate) is considered to be non-clastogenic to human lymphocytes in vitro (Safepharm Limited, 2005).

Mammilian gene mutation

According to an OECD 476, conducted to GLP, Methylenebis (dibutyldithiocarbamate) is negative using mouse lymphoma L5178Y cells. Methylenebis (dibutyldithiocarbamate) is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions (WIL Research Europe B.V., 2012).


Justification for selection of genetic toxicity endpoint
The study was conducted according to appropriate OECD Guidelines, under GLP conditions at a reputable European Union laboratory.
The study in mammalian cells was chosen over that in bacterial cells, as the higher level study.

Justification for classification or non-classification

In accordance with Regulation No 1272/2008 Table 3.5.1, based on available data on Methylene bis (dibutyldithiocarbamate) is not classified for genotoxicity according to CLP.