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Genetic toxicity: in vitro

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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:
comparable to guideline study

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Salmonella mutagenicity tests: V. Results from the testing of 311 chemicals
Author:
Zeiger E; Anderson B; Haworth S; Lawlor T; Mortelmans K
Year:
1992
Bibliographic source:
Environ. Mol. Mutagen, Vol 19, Suppl. 21, pp. 2-141
Reference Type:
other: NTP web site
Title:
Results for this Salmonella study's detailed data, CAS no. 67-68-5
Author:
NTP
Year:
1987
Bibliographic source:
http://ntp-apps.niehs.nih.gov/ntp_tox/index.cfm?fuseaction=ntpsearch.searchhome&crumbspot=1
Reference Type:
other: NTP web site
Title:
Results for this Salmonella study's detailed data, CAS no. 67-68-5
Author:
NTP
Year:
1988
Bibliographic source:
http://ntp-apps.niehs.nih.gov/ntp_tox/index.cfm?fuseaction=ntpsearch.searchhome&crumbspot=1

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Dimethyl sulfoxide
EC Number:
200-664-3
EC Name:
Dimethyl sulfoxide
Cas Number:
67-68-5
Molecular formula:
C2H6OS
IUPAC Name:
dimethyl sulfoxide
Details on test material:
Test compound: Dimethylsulfoxide
CAS no.: 67-68-5
Source: Burdick and Jackson
Purity: > 99.5%

Method

Target gene:
Histidine operon
Species / strain
Species / strain / cell type:
S. typhimurium, other: Strains TA97, TA98, TA100, TA1535, TA1537
Metabolic activation:
with and without
Metabolic activation system:
S9 fractions of Aroclor 1254-induced male Sprague Dawley rats and male Syrian hamster livers
Test concentrations with justification for top dose:
0, 100, 333, 1000, 3333 and 10000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: no data
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: - Without S9: Sodium azide (TA 1535 and TA100), 9-aminoacridine or ICR-191 (TA 97 and TA1537), 4-nitro-o-phenylenediamine (TA98) - With S9: 2-aminoanthracene (all strains)
Details on test system and experimental conditions:
In the Salmonella assay, a test tube containing a suspension of one strain of Salmonella typhimurium plus S9 mix or plain buffer without S9, is incubated for 20 minutes at 37º C with the test chemical. Control cultures, with all the same ingredients except the test chemical, are also incubated. In addition, positive control cultures are also prepared; these contain the particular bacterial tester strain under investigation, the various culture ingredients, and a known potent mutagen. After 20 minutes, agar is added to the cultures and the contents of the tubes are thoroughly mixed and poured onto the surface of Petri dishes containing standard bacterial culture medium. The plates are incubated for 48 hours and then counted.

Initial testing was without metabolic activation, with 10% rat liver S-9, or with 10% hamster liver S-9. After a negative result was obtained, DMSO was retested without S-9 and with 30% S-9 from rat and hamster.

The positive control plates are counted, and the number of mutant colonies appearing on them must be significantly increased over the spontaneous control number for the test to be considered valid.
Evaluation criteria:
If no increase in mutant colonies is seen after testing several strains under several different culture conditions, the test chemical is considered to be non-mutagenic in the Salmonella test.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535
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 applicable
True negative controls validity:
not applicable
Positive controls validity:
valid
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 applicable
True negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 97
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 applicable
True negative controls validity:
not applicable
Positive controls validity:
valid
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 applicable
True negative controls validity:
not applicable
Positive controls validity:
valid
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 applicable
True negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
The positive control chemicals induced a significant increase of the revertant frequency in all tester strains, either with or without metabolic activation.
DMSO was negative, in the presence and absence of metabolic activation, in all tester strains.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Individual strain data is presented as mean ± standard error.
Abbreviations are noted at bottom of page.
Trial summary calls are shown in parentheses.
Strain: TA1535
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      22 5.2 19 5 16 0.6 14 1 10 1.8 15 2.3
100      19 5.4 24 2.3 12 3.3 14 4.3 12 3.5 14 1.8
333      27 1.7 24 3.2 15 2 14 1.8 11 2.6 15 1
1000      19 1.8 17 2.6 12 1.5 13 0.9 14 1.7 15 1.7
3333      22 2.5 22 2.5 9 1.5 16 3.8 11 1.9 15 0.9
10000      24 1.8 18 2.9 9 3.8 15 2.5 11 0.9 15 4.4
Positive Control 852 11.7 272 16.2 52 3.2 180 6.9 328 17.1 308 4.1
Strain: TA100
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      138 3 113 6.1 105 1.2 142 9.8 113 5 138 3.8
100      133 6.7 107 5.6 110 2.3 145 8.4 121 9.6 140 5
333      155 7.6 106 5.6 111 10.6 155 4.9 119 4.5 147 11.9
1000      139 3.5 110 9.3 105 1.8 159 5.1 116 2.7 128 4
3333      126 10.5 107 7.9 93c 3.5 147 13.5 108 0.9 128 5.8
10000      157 12.3 113 6.9 102 4.2 136 4.8 107 4.7 129 6.1
Positive Control 586 3.5 414 12.2 279 10.3 728 12.7 1440 66.4 1005 15.6
Strain: TA97
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      122 3 118 6 112 3.4 122 5.1 115 7.6 158 11.9
100      141 1.2 114 5 130 4.7 111 5.5 137 3.5 183 2.6
333      131 15 123 3.9 109 5.8 141 3.7 128 2.9 180 11.8
1000      117 3.4 123 12 129 7 136 1.2 131 3.5 155 10.7
3333      150 11.7 131 3.5 118 7.8 132 5.1 119 4.7 167 3.5
10000      138 9.6 118 8 120 2.2 119 8.7 132 6 158 3.9
Positive Control 960 22.6 512 18.1 812 35 1436 48.3 2756 30.8 1606 101.3
Strain: TA98
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      19 0.9 23 1.7 35 2.7 35 1.7 27 3.8 32 1.9
100      13 2.3 21 1.7 36 1.5 34 3.7 27 0.9 31 1.9
333      15 3.5 16 2.1 35 7.2 32 1.7 32 2 31 1
1000      21 4.2 17 3.8 33 4.3 34 4.9 33 4.7 29 1.8
3333      25 2.4 19 1.7 32 2.8 31 6.8 35 3.2 34 4.2
10000      17 3 21 4.2 31 2.9 31 0.7 32 1.7 27 1.2
Positive Control 330 13.1 330 8.4 110 2.6 604 17.9 211 8.2 338 12.6

Strain: TA1535
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      33 3.3 22 3.5 11 1.7 13 1.3 12 3.8 15 0.6
100      27 2 21 4.3 12 3.2 15 1.8 11 4.3 12 1.2
333      30c 1.5 26 3.2 12 3.2 13 2.2 10 1.2 12 0
1000      28 4.4 15 1.2 8 2 12 1.2 8 1.5 14 2.3
3333      21 2.3 15 2 10 0.7 12 1.2 9 1.8 14 2.2
10000      26 2.6 20 3.5 9 0.9 14 1.8 10 0.7 11 2.1
Positive Control 448 9 232 10.2 154 28.2 569 15.6 153 8.9 195 12.9
Strain: TA100
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      134 4.4 159 6.4 175 1.9 128 4.3 163 7.5 125 13.4
100      126 8.4 149 5.8 137 9.8 123 8.1 139 6.8 127 6.2
333      139 2.6 144 8.2 166 5.5 139 6.4 159 3.9 130 7.8
1000      128 5.8 140c 3 152 14.2 121 11.2 160 9.8 135 14.8
3333      121 2.8 159 4.2 167 4 135 11.2 140 17.9 152 6.1
10000      132 2.8 152 4.6 151 13.7 128 4.3 144 24.1 134 8
Positive Control 370 18.4 384 57.2 712 46 558 32.7 566 12.1 383 13.7
Strain: TA97
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Equivocal) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      153 6.4 178 4.8 204 1.9 198 8.2 203 2.6 222 1.5
100      162 5.8 182 7.6 194 5.5 208 12.7 212 5.5 214 4.4
333      165 6.8 197 6.7 218 2.1 212 6.1 216 2.7 215 10.1
1000      196 10.1 195 9.4 199 7.8 212 3.2 229 7.9 198 4.5
3333      196 4.3 198 11.3 215 2.5 211 4.8 222 7.5 234 7.5
10000      182 7.9 207 9.9 212 6.2 193 2.2 229 3.2 217 8.8
Positive Control 635 28.5 613 34.8 556 35.6 516 8.5 376 27 460 34.5
Strain: TA98
Dose No Activation No Activation 10% HLI 30% HLI 10% RLI 30% RLI
(Negative) (Negative) (Negative) (Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM Mean ± SEM
0      16 3.2 16 2.3 28 2 34 3.6 39 3.2 39 4.9
100      16 1.2 17 1.3 25 2.3 28 1 31 3.6 31 3.3
333      18 3.8 18 3 30 4 36 1.5 34 5.6 36 2.9
1000      19 3.3 13 3.2 34 2.6 33 1.9 29 5.2 33 7.7
3333      15 1.3 21 2.7 33 1.5 33 2.5 23 3.6 27 0.7
10000      17 2.1 17 4.2 26 4.7 28 1.7 35 4 31 4.2
Positive Control 540 11.6 745 51.8 771 3.7 438 12.6 534 4 122 5.8
Strain: TA1537
Dose No Activation 30% HLI 30% RLI
(Negative) (Negative) (Negative)
ug/Plate Mean ± SEM Mean ± SEM Mean ± SEM
0      11 1.5 10 1.7 14 0.6
100      12 1 14 1.2 15 2.3
333      12 2.8 11 0.3 13 2.6
1000      9 1.9 9 1.5 10 1.2
3333      12 0.9 13 0.9 11 1.9
10000      14 2.3 10 0.9 12 2.1
Positive Control 310 30.5 46 8.9 55 4.6
Abbreviations:
RLI = induced male Sprague Dawley rat liver S9
HLI = induced male Syrian hamster liver S9
s = Slight Toxicity; p = Precipitate; x = Slight Toxicity and Precipitate; T = Toxic; c = Contamination

Applicant's summary and conclusion

Conclusions:
Under these experimental conditions, Dimethylsulfoxide is considered as non genotoxic.
Executive summary:

The potential of Dimethylsulfoxide (DMSO) to induce reverse mutation in Salmonella typhimurium (strains: TA97, TA98, TA100, TA1535, TA1537) was evaluated according to a protocol comparable to the OECD guidelines 471.

DMSO was tested in two consecutive experiments, with and without a metabolic activation system, according to the preincubation method (20 min at 37°).

Concentrations of DMSO (100, 333, 1000, 3333, and 10,000 µg), overnight culture of S. typhimurium (0.05-0.10 ml) and S-9 mix or buffer were incubated without shaking for 20 minutes. The top agar was added and the contents of the tubes were mixed and poured onto the surfaces of Petri dishes. His+ (histidine dependent) colonies arising on plates were machine-counted after two days incubation at 37°C.

Initial testing was without metabolic activation, with 10% rat liver S-9, or with 10% hamster liver S-9. After a negative result was obtained, DMSO was retested without S-9 and with 30% S-9 from rat and hamster.

The positive control chemicals induced a significant increase of the revertant frequency in all tester strains, either with or without metabolic activation. DMSO was negative, in the presence and absence of metabolic activation, in all tester strains.