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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

AMES test

In the bacterial reverse mutation test using S.typhimurium strains TA98, TA100, TA1535 and TA1538, test substance did not induce any significant increase of His revertants. Thus, it is concluded that test substance is not genotoxic.

In vitro gene mutation assay in Mammalian cells

In the in vitro chromosome aberration test using Chinese Hamster Lung (CHL) cells, no abberation effects were seen.Therefore, the  test substance is considered to be non mutagenic in Chinese Hamster Lung (CHL) cells.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from peer-reviewed journal
Qualifier:
no guideline available
Principles of method if other than guideline:
The in vitro chromosome aberration test was carried out using Chinese Hamster Lung (CHL) cells to evaluate genetic potential of substance dimethyl sulphone (Methylsulfonylmethane).
GLP compliance:
no
Type of assay:
in vitro mammalian chromosome aberration test
Target gene:
Chinese Hamster Lung (CHL) cells
Species / strain / cell type:
other: Chinese Hamster Lung (CHL) cells
Details on mammalian cell type (if applicable):
Details on mammalian cell line
- Type and identity of media: The cell were cultured in Eagles minimum essential medium (EMEM) supplemented with 10% fetal bovine serum. The cell were incubated in a 95% air and 5% CO₂ atmosphere at 37°C. The cell were plated at a density of 2.5ẋ10⁵ cells on 6cm plates and grown for 22h, and then colcemid (0.25g/ml) was added 2hr before harvest.
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: No data available
- Periodically checked for karyotype stability: No data available
- Periodically "cleansed" against high spontaneous background: No data available
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
rat liver S9 fraction
Test concentrations with justification for top dose:
0, 5, 2.5, 1.25 mg/ml
Vehicle / solvent:
Vehicle
- Vehicle(s)/solvent(s) used: No data available
- Justification for choice of solvent/vehicle: No data available
Untreated negative controls:
yes
Negative solvent / vehicle controls:
not specified
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
methylmethanesulfonate
Details on test system and experimental conditions:
METHOD OF APPLICATION: plate incorporation

DURATION
- Preincubation period: No data available
- Exposure duration: 24hr
- Expression time (cells in growth medium): 22 hr
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): 24 hr

SELECTION AGENT (mutation assays): No data available

SPINDLE INHIBITOR (cytogenetic assays): No data available

STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: No data available

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: No data available

OTHER EXAMINATIONS
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
Rationale for test conditions:
No data
Evaluation criteria:
No data
Statistics:
No data
Key result
Species / strain:
other: Chinese Hamster Lung (CHL) cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
valid
Positive controls validity:
valid
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: No data available

RANGE-FINDING/SCREENING STUDIES: No data available

COMPARISON WITH HISTORICAL CONTROL DATA: No data available

ADDITIONAL INFORMATION ON CYTOTOXICITY: No data available
Remarks on result:
other: no mutagenic potential
Conclusions:
In the in vitro chromosome aberration test using Chinese Hamster Lung (CHL) cells, no abberation effects were seen.Therefore, the test substance is considered to be non mutagenic in Chinese Hamster Lung (CHL) cells.
Executive summary:

The in vitro chromosome aberration test was carried out using Chinese Hamster Lung (CHL) cells to evaluate the potential of test substance damaging chromosome. The cell were exposed to test substance at concentration 0, 5, 2.5, 1.25 mg/ml for 24 hr. Methylmethane sulfon ate (MMS)(0.02 mg/ml) and Benzo[a]pyrene (B[a]P) (0.02mg/ml) used as positive control substances. After treatment with test substance , no significant increase in the number of aberrant cells was observed. In presence or absence of S9 mix, the chromosome aberration produced by treatment with Methylsulfon ylmethane was less than 2 %. These results indicate that methylsulfonylmethane did not increase chromosome aberration as compared with the negative control. Therefore, the test substance is considered to be non genotoxic.

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from peer-reviewed journal
Qualifier:
no guideline available
Principles of method if other than guideline:
The potential genotoxicity of test substance was evaluated in S. typhimurium strains TA98, TA100, TA1535 and TA1538 by Ames test.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
No data available
Target gene:
Salmonella typhimurium TA98, TA100, TA1535 and TA1538
Species / strain / cell type:
other: TA98, TA100, TA1535 and TA1538
Details on mammalian cell type (if applicable):
No data available
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
2500, 5000, 1000 µg/plate
Vehicle / solvent:
Vehicle
- Vehicle(s)/solvent(s) used: No data available
- Justification for choice of solvent/vehicle: No data available
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
sodium azide
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
other: 9-aminoacridine hydrochloride (9-AA)
Remarks:
For Strain TA1538 (50 µg/plate)
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 hr
- Expression time (cells in growth medium): No data available
- 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: 3 replicate plates per dose

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: No data available

OTHER EXAMINATIONS: No data available
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
Rationale for test conditions:
No data available
Evaluation criteria:
No data available
Statistics:
No data available
Key result
Species / strain:
other: TA98, TA100, TA1535 and TA1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
Positive controls validity:
valid
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: No data available

RANGE-FINDING/SCREENING STUDIES: No data available

COMPARISON WITH HISTORICAL CONTROL DATA: No data available

ADDITIONAL INFORMATION ON CYTOTOXICITY: No data available
Remarks on result:
other: no mutagenic potential
Conclusions:
In the bacterial reverse mutation test using S.typhimurium strains TA98, TA100, TA1535 and TA1538, test substance did not induce any significant increase of His revertants. Thus, it is concluded that test substance is not genotoxic.
Executive summary:

The reverse mutation tests were carried out using S. typhimurium strainsTA98, TA100, TA1535 and TA1538. A 0.1 ml volume of the test substance solutions (2500, 5000, 1000 µg/plate) was mixed with 0.1ml of the 10 h cultured bacteria and then bacterial culture was mixed with 0.5ml of the S9 mix or a 0.1 M phosphate buffer (pH 7.4) and incubated for 20min at 37 °C. After incubation a top agar (2ml) was added and the mixture overlaid on a minimal glucose agar plate. After 48 hrs of incubation, the revertant colonies were counted.The treatments were performed in the presence and absence of metabolic activation. There was no increase in the revertant numbers as a result of test substance treatment for any of the S.typhimurium strain with and without the S9 mix. Thus, it was concluded that test substance was non mutagenic in S.typhimurium strains TA98, TA100, TA1535 and TA1538.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

The micronucleus assay results confirmed that dimethyl sulphone (methylsulfonylmethane) at doses up to 5000 mg/kg did not exhibit any genotoxicity in mouse bone marrow.

Link to relevant study records
Reference
Endpoint:
genetic toxicity in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from peer-reviewed journal
Qualifier:
no guideline available
Principles of method if other than guideline:
The in vivo micronucleus test was carried out in mouse bone marrow, to find out genetic effect of dimethyl sulphone (Methylsulfonylmethane).
GLP compliance:
no
Type of assay:
other: micronucleus assay
Species:
mouse
Strain:
not specified
Sex:
not specified
Details on test animals and environmental conditions:
No data available
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: Carboxymethylcellulose
- Justification for choice of solvent/vehicle: No data available
- Concentration of test material in vehicle: No data available
- Amount of vehicle (if gavage or dermal): 0.5% w/v (vehicle control group)
- Type and concentration of dispersant aid (if powder): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Details on exposure:
No data available
Duration of treatment / exposure:
48 hr
Frequency of treatment:
Once
Post exposure period:
No data available
Dose / conc.:
1 250 other: mg/kg
Dose / conc.:
2 500 other: mg/kg
Dose / conc.:
5 000 other: mg/kg
No. of animals per sex per dose:
No data available
Control animals:
not specified
Positive control(s):
Positive controls: mitomycin C
- Justification for choice of positive control(s): No data available
- Route of administration: intra peritoneal (i.p)
- Doses / concentrations: 4 mg/kg
Tissues and cell types examined:
bone marrow
Details of tissue and slide preparation:
No data available
Evaluation criteria:
No data available
Statistics:
No data available
Sex:
not specified
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls validity:
valid
Negative controls validity:
not specified
Positive controls validity:
valid
Remarks on result:
other: non mutagenic
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: No data available
- Solubility: No data available
- Clinical signs of toxicity in test animals: No data available
- Evidence of cytotoxicity in tissue analyzed: No data available
- Rationale for exposure: No data available
- Harvest times: No data available
- High dose with and without activation: No data available
- Other:

RESULTS OF DEFINITIVE STUDY
- Types of structural aberrations for significant dose levels (for Cytogenetic or SCE assay): No data available
- Induction of micronuclei (for Micronucleus assay): No data available
- Ratio of PCE/NCE (for Micronucleus assay): No data available
- Appropriateness of dose levels and route: No data available
- Statistical evaluation: No data available
Conclusions:
The micronucleus assay results confirmed that test substance at doses up to 5000 mg/kg did not exhibit any genotoxicity in mouse bone marrow.
Executive summary:

The in vivo micronucleus test was carried out in mouse bone marrow, to find out genetic effect of test substance . Methylsulfonylmethane was administered to group of mice using an oral gavage at doses of 1250, 2500, 5000mg/kg. The vehicle control groups received carboxymethyl cellulose at an equivalent oral volume of 0.5% w/v, whereas the positive control group received an i.p. dose of mitomycin C at 4mg/kg. No significant difference in body weight was noted between test substance treated mice and the solvent control. The number of micronucleated polychromatic erythrocytes, where the frequency in the solvent control was 0.4±0.5, the frequency in the positive control was 12.2±1.2, which was significantly higher and the frequency after test substance treatment with 1250, 2500, 5000mg/kg was 0.1±0.5, 0.5±0.5 and 0.4±0.5 respectively. Therefore, the test substance treatment did not cause any change in the frequency of micronucleated polychromatic erythrocytes when compared with the solvent control. Consequently, the micronucleus assay results confirmed that test substance at doses up to 5000 mg/kg did not exhibit any genotoxicity in mouse bone marrow.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Gene mutation in vitro:

Data for the various test chemicals was reviewed to determine the mutagenic nature of dimethyl sulfone, methanesulfonylmethane (67-71-0). The studies are as mentioned below:

AMES test

The reverse mutation tests were carried out using S. typhimurium strainsTA98, TA100, TA1535 and TA1538. A 0.1 ml volume of the test substance solutions (2500, 5000, 1000 µg/plate) was mixed with 0.1ml of the 10 h cultured bacteria and then bacterial culture was mixed with 0.5ml of the S9 mix or a 0.1 M phosphate buffer (pH 7.4) and incubated for 20min at 37 °C. After incubation a top agar (2ml) was added and the mixture overlaid on a minimal glucose agar plate. After 48 hrs of incubation, the revertant colonies were counted.The treatments were performed in the presence and absence of metabolic activation. There was no increase in the revertant numbers as a result of test substance treatment for any of the S.typhimurium strain with and without the S9 mix. Thus, it was concluded that test substance was non mutagenic in S.typhimurium strains TA98, TA100, TA1535 and TA1538.

Supported by other studies.

The mutagenic potential of test substance was insvestigated in a reverse mutation assay in Salmonella typhimurtum strains TA98, TA100, TA102, TA1535 and TA1537 in the presence and absence of S9 liver homogenate fraction. Two independent experiments were conducted. The first trial was conducted using the plate incorporation assay with and without metabolic activation, while the second one was conducted without metabolic activation and the preincubation method with metabolic activation. In both experiments, five concentrations of MSM ranging from 50 to 5000 ug/plate were tested. No significant cytotoxic effects of MSM were noted. test substance did not induce any significant increase in the number of reversions, either in the presence or absence of metabolic activation.Therefore,test substance  did not meet the criteria for a potential mutagen, at concentrations up to 5000 µg/plate. Thus, it was concluded that test substance was non mutagenic at concentrations up to 5000 µg/plate in S.typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 with and without the metabolic activation.

Pre-incubation Ames test was carried out according to the method of Maron and Ames (1983), to observe genetic effect of test substance in S.typhimurium strains TA98, TA100 and TA102. All strains were tested at dose 0.003-300 µmol/plate both with and without metabolic activation (RAT, LIVER, S-9, AROCLOR 1254). Four plates were used per dose and incubated for 3 days and the colonies counted with an automated Fisher Count-All colony counter. 2-nitrofluorene, sodium azide, mitomycin C, 2-aminofluorene used as a positive control substances. Water is used as solvent as well as negative control. In results,test substance dose not induced any mutagenic activity in any of the tester strains. Therefore, the test substance is considered to be non mutagenic in Salmonella typhimurium strains TA98, TA100 and TA102 with and without metabolic activation.

In vitro gene mutation assay in Mammalian cells

The in vitro chromosome aberration test was carried out using Chinese Hamster Lung (CHL) cells to evaluate the potential of test substance damaging chromosome. The cell were exposed to test substance at concentration 0, 5, 2.5, 1.25 mg/ml for 24 hr. Methylmethane sulfon ate (MMS)(0.02 mg/ml) and Benzo[a]pyrene (B[a]P) (0.02mg/ml) used as positive control substances. After treatment with test substance , no significant increase in the number of aberrant cells was observed. In presence or absence of S9 mix, the chromosome aberration produced by treatment with Methylsulfon ylmethane was less than 2 %. These results indicate that methylsulfonylmethane did not increase chromosome aberration as compared with the negative control. Therefore, the test substance is considered to be non genotoxic.

Genetic toxicity in vivo

The in vivo micronucleus test was carried out in mouse bone marrow, to find out genetic effect of test substance . Methylsulfonylmethane was administered to group of mice using an oral gavage at doses of 1250, 2500, 5000mg/kg. The vehicle control groups received carboxymethyl cellulose at an equivalent oral volume of 0.5% w/v, whereas the positive control group received an i.p. dose of mitomycin C at 4mg/kg. No significant difference in body weight was noted between test substance treated mice and the solvent control. The number of micronucleated polychromatic erythrocytes, where the frequency in the solvent control was 0.4±0.5, the frequency in the positive control was 12.2±1.2, which was significantly higher and the frequency after test substance treatment with 1250, 2500, 5000mg/kg was 0.1±0.5, 0.5±0.5 and 0.4±0.5 respectively. Therefore, the test substance treatment did not cause any change in the frequency of micronucleated polychromatic erythrocytes when compared with the solvent control. Consequently, the micronucleus assay results confirmed that test substance at doses up to 5000 mg/kg did not exhibit any genotoxicity in mouse bone marrow.

Based on the data summarized, dimethyl sulfone, methanesulfonylmethane (67-71-0) did not induce gene mutation .Hence it is not likely to be mutagenic in vitro.

Justification for classification or non-classification

Thus based on the above annotation and CLP criteria the test chemical dimethyl sulfone, methanesulfonylmethane (67-71-0)did not induce gene mutation .Hence it is not likely to be mutagenic in vitro.