2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane

Genetic toxicity: in vitro

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 guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

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

Test material information

Constituent 1

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

Test results 1

Key result

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:

valid

Positive controls validity:

valid

Test results 2

Key result

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:

valid

Positive controls validity:

valid

Test results 3

Key result

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:

valid

Positive controls validity:

valid

Test results 4

Key result

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:

valid

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium 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.