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Toxicological information

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
Study period:
2012-01-20 to 2012-02-13
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 guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane
EC Number:
219-863-1
EC Name:
2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane
Cas Number:
2554-06-5
Molecular formula:
C12H24O4Si4
IUPAC Name:
2,4,6,8-tetramethyl-2,4,6,8-tetravinyl-1,3,5,7,2,4,6,8-tetroxatetrasilocane
Test material form:
other: liquid

Method

Target gene:
Histidin operon
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
phenobarbital and beta-naphthoflavone induced rat S9 liver microsomal fraction
Test concentrations with justification for top dose:
Expt I: 31.6, 100, 316, 1000, 2500 and 5000 µg/plate; Expt II: 15.8, 50.0, 158, 500, 1580 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: compatible with the survival of the bacteria and the S9 activity.
Controls
Untreated negative controls:
yes
Remarks:
A. dest., BSL Lot No. 111219, 120105
Negative solvent / vehicle controls:
yes
Remarks:
EtOH, Applichem Lot No. 1R002540
True negative controls:
no
Positive controls:
yes
Positive control substance:
other:
Remarks:
Positive control without metabolic activation: sodium azide for tester strains TA100, TA1535; 4-nitro-o-phenylene-diamine for TA98, TA1537; methylmethanesulfonate for TA102. Positive control with metabolic activation: 2-aminoanthracene for all strains
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)
DURATION
- Exposure duration: at least 48 h; After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.


NUMBER OF REPLICATIONS: For each strain and dose level, including the controls, three plates were used. The experiment was repeated.


DETERMINATION OF CYTOTOXICITY
- Method: clearing or diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately ≤ 0.5 in relation to the solvent control

- ACTIVATION
phenobarbiotol and beta-naphthoflavone induced rat liver S9. S9 mix included 5% S9 and glucose-6-phosphate and NADP as cofactors. 0.5 ml of S9 mix were added to 2.7 ml overlay agar including test substance or control and bacterial suspension, giving a final concentration of S9 of approximately 1%.
Evaluation criteria:
The Mutation Factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).
A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs in at least one tester strain with or without metabolic activation.

A biologically relevant increase is described as follows:
- if in tester strains TA 98, TA 100 and TA 102 the number of reversions is at least twice as high
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher
than the reversion rate of the solvent control.
Statistics:
According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.

Results and discussion

Test results
Key result
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
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:
valid
Positive controls validity:
valid
Additional information on results:
Precipitation was observed in all tester strains used in Experiment I and II (with and without metabolic activation). In Experiment I, precipitation of the test item was noted at dose levels of 2500 µg/plate and higher (without metabolic activation) and 5000 µg/plate (with metabolic activation). In Experiment II, precipitation of the test item was noted at 1580 µg/plate and higher (without metabolic activation) and 5000 µg/plate (with metabolic activation).

Any other information on results incl. tables

Mean number of revertant colonies - Experiment 1

Treatment µg/plate

TA98

TA100

TA1535

TA1537

TA102

-S9

+S9

-S9

+S9

-S9

+S9

-S9

+S9

-S9

+S9

Negative control

22

31

104

120

7

11

5

6

225

311

Solvent control

22

31

87

112

6

12

4

4

244

315

31.6

27

35

102

136

8

11

7

5

236

292

100.0

17

33

106

126

8

15

8

5

227

342

316.0

24

35

92

121

4

15

4

3

248

317

1000

24

35

102

121

4

14

6

5

254

329

2500

32

39

109

114

2

9

3

4

256

295

5000

29

34

97

113

3

14

6

8

250

306

Positive control

372

2860

893

2060

726

123

105

246

1527

763

Mean number of revertant colonies - Experiment 2

Treatment µg/plate

TA98

TA100

TA1535

TA1537

TA102

-S9

+S9

-S9

+S9

-S9

+S9

-S9

+S9

-S9

+S9

Negative control

20

25

132

127

11

7

7

8

230

300

Solvent control

25

30

110

115

13

7

6

7

201

279

15.8

28

35

129

133

12

15

7

10

246

340

50.0

27

37

114

137

9

14

4

7

223

342

158.0

26

30

117

151

6

15

4

8

249

346

500.0

25

30

118

136

9

12

2

4

243

308

1580.0

27

32

130

140

9

13

9

9

300

336

5000.0

30

24

122

138

12

14

5

7

257

350

Positive control

851

2524

855

2380

1212

152

149

188

2054

593

Applicant's summary and conclusion

Conclusions:
2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane has been tested in a reliable bacterial mutagenicity study conducted according to OECD TG 471 and in compliance with GLP. No test-substance related increase in the number of revertants was observed when Salmonella typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 were exposed with and without metabolic activation up to limit concentrations. Appropriate positive and solvent controls were included and gave expected results. It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of the test.
Executive summary:

In a reverse gene mutation assay in bacteria, strains TA 98, TA 100, TA 1535, TA 1537 and TA 102 of S. typhimurium were exposed to 2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane at concentrations of 31.6, 100, 316, 1000, 2500 and 5000 µg/plate (Experiment I) and 15.8, 50.0, 158, 500, 1580 and 5000 µg/plate (Experiment II), in the presence and absence of mammalian metabolic activation according to the plate incorporation method (Experiment I and II).

2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane was tested up to the limit concentration of 5000 µg/plate in all tester strains used.

The positive controls induced the appropriate responses in the corresponding strains.There was no evidence of induced mutant colonies over background.

This study is classified as acceptable. This study satisfies the requirement for Test Guideline OPPTS 870.5100; OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.