Dimethyl disulphide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1998

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Version / remarks:

1998

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Dimethyl disulphide

EC Number:

210-871-0

EC Name:

Dimethyl disulphide

Cas Number:

624-92-0

Molecular formula:

C2H6S2

IUPAC Name:

(methyldisulfanyl)methane

Details on test material:

- Physical state: light yellow liquid
- Analytical purity: 99.2%
- Purity test date: LJuly 21, 2007
- Lot/batch No.: VS060001

Method

Target gene:

Histidine locus (Salmonella) and tryptophane locus (Escherichia)

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Liver S9 homogenate was prepared from rats that have been induced with Arochlor 1254

Test concentrations with justification for top dose:

The dose levels tested were 1.5, 5.0, 15, 50, 150, 500, 1500 and 5000 µg per plate in the initial toxicity-mutation assay and 50, 150, 500, 1500 and 5000 µg per plate in the confirmatory mutagenicity assay.

Vehicle / solvent:

Dimethyl sulphoxide

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: With S9: 2-aminoanthracene, 1 µg/plate for all Salmonella strains, 10µg/plate for E. coli. Without S9: TA 98, 2-nitrofluorene (1 µg/plate); TA100 and TA 1535, sodium azide (1 µg/plate); TA 1537, 9-aminoacridine (75 µg/plate); E. coli, MMS (1000 µg/plate)

Details on test system and experimental conditions:

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth (decrease in the number of revertant colonies and/or a thinning of the bacterial lawn);

EXPERIMENTS
- Initial toxicity-mutation assay: in all strains, with or without S9 mix ; 8 dose-levels (2 plates/dose level)
- Confirmatory mutagenicity assay: in all strains, with or without S9 mix ; 5 dose-levels (3 plates/dose level)

METHOD OF APPLICATION: Direct plate incorporation method: for preliminary both experiments

DURATION
- Exposure duration: 48-72H

Evaluation criteria:

Reproducible increase in the number of revertant colonies (2-fold for TA98/TA100 and WP2 uvrA, 3-fold for TA 1535/TA 1537) compared with vehicle controls in any strain at any dose-level and/or evidence of a dose-relationship.
Reference to historical data and consideration to biological relevance may also be taken into account.

Results and discussion

Test results

Test results 1

Species / strain:

E. coli WP2 uvr A

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:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

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:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

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:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 1537

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:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Remarks:

Tested up to limit concentrations recommended by the test guideline

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING STUDY:
- Results on solubility: dimethyl disulphide formed a soluble and clear solution in dimethyl sulfoxide (DMSO) at approximately 500 mg/mL, the highest concentration tested.
- Results on cytotoxicity: In the initial toxicity-mutation assay, the maximum dose tested was 5000 µg/per plate; this dose was achieved using a concentration of 100 mg/mL and a 50 µL plating aliquot. The dose levels tested were 1.5, 5.0, 15, 50, 150, 500, 1500 and 5000 µg per plate. Neither precipitate nor appreciable toxicity was observed.

Any other information on results incl. tables

Number of revertants per plate (first experiment) (mean of 2 plates)
	TA 98		TA 100		TA 1535		TA 1537		WP2 uvrA
Conc. [µg/plate]	- MA	+MA	- MA	+ MA	- MA	+ MA	- MA	+MA	-MA	+MA
DMSO	17	19	126	127	16	16	9	5	11	12
1.5	21	21	122	131	14	13	6	10	7	13
5.0	15	21	101	137	14	18	7	8	8	13
15	19	21	103	142	14	21	2	7	10	9
50	14	24	103	124	10	14	7	6	10	14
150	13	20	95	120	14	17	4	8	11	9
500	20	32	139	137	20	18	9	2	13	12
1500	17	19	136	131	17	7	4	7	10	10
5000	14	19	103	138	16	25	7	8	12	13
Positive control	170	553	426	604	424	70	389	68	52	87
Table 2: Number of revertants per plate (second experiment) (mean of 3 plates)
	TA 98		TA 100		TA 1535		TA 1537		WP2 uvrA
Conc. [µg/plate]	- MA	+MA	- MA	+ MA	- MA	+ MA	- MA	+ MA	- MA	+ MA
DMSO	24	30	119	134	28	15	8	11	14	16
50	18	29	110	137	24	11	6	6	11	13
150	25	29	118	131	32	16	6	5	14	14
500	22	29	113	122	23	14	6	6	14	18
1500	19	34	107	126	26	15	7	6	15	20
5000	21	29	113	121	30	18	6	7	16	18
Positive control	147	458	628	572	478	78	497	53	96	181

Applicant's summary and conclusion

Conclusions:

The results of the Bacterial Reverse Mutation Assay indicate that, under the conditions of this study, Dimethyl disulphide did not cause a positive response in either the presence or absence of Aroclor-induced rat liver S9.

Executive summary:

Dimethyl disulfide, was tested in the Bacterial Reverse Mutation Assay using Salmonella typhimurium tester strains TA98, TA100, TA1535 and TA1537 and Escherichia coli tester strain WP2 uvrA in the presence and absence of Aroclor-induced rat liver S9. The assay was performed using the plate incorporation method. The dose levels tested were 1.5, 5.0, 15, 50, 150, 500, 1500 and 5000 µg per plate in the initial toxicity-mutation assay and 50, 150, 500, 1500 and 5000 µg per plate in the confirmatory mutagenicity assay. No positive mutagenic response was observed. Neither precipitate nor appreciable toxicity was observed. Dimethyl disulfide was concluded to be negative in the Bacterial Reverse Mutation Assay