Reaction mass of 1,1,1,5,5,5-hexamethyl-3-phenyl-3-((trimethylsilyl)oxy)trisiloxane and 1,1,1,7,7,7-hexamethyl-3,5-diphenyl-3,5-bis[(trimethylsilyl)oxy]tetrasiloxane and 1,1,1,9,9,9-hexamethyl-3,5,7-triphenyl-3,5,7-tris((trimethylsilyl)oxy)pentasiloxane

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:

1995-02-06 - 1995-02-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1995

Report date:

1995

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

OECD not actually specified but mentioned in 'Statement of Compliance'.

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Method

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 pKM 101

Species / strain 3

Species / strain / cell type:

E. coli, other: WP2 pKM101

Metabolic activation:

with and without

Metabolic activation system:

Aroclor induced rat liver S9

Test concentrations with justification for top dose:

100, 333, 1000, 3333 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Acetone
- Justification for choice of solvent/vehicle: Sponsor's request and compatibility with the target cells.
NOTE: the solvent is described as acetone in the method section but in most of the tables of results it is reported to be DMSO. It is thought by the reviewer that this is a typographical error: if DMSO was used rather than acetone the study would still be valid.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene 1.0 µg/plate

Remarks:

TA98, TA100, TA1535, TA1537 with metabolic activation

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene 10 µg/plate

Remarks:

WP2 uvrA (pKM101) with metabolic activation

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: sterigmatocystin 100 µg/plate

Remarks:

WP2 (pKM101) with metabolic activation

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

TA98 without metabolic activation 1.0 µg/plate

Controls 5

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA100, TA1535 without metabolic activation 1.0 µg/plate

Controls 6

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

TA1537 without metabolic activation 75 µg/plate

Controls 7

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

WP2 uvrA (pKM101), WP2 (pKM101) without metabolic activation 1,000 µg/plate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

ACTIVATION: S9 mix contained glucose-6-phosphate and NADP as co-factors. Concentration of S9 in the mix was 10%. 0.5mlk of S9 mix were added to a total volume of 2.65ml, giving a final concentration of approximately 2% S9.

DURATION
- Preincubation period: 60 minutes +/- 2 minutes at 37ºC
- Exposure duration: 48 - 72 hours at 37ºC


SELECTION AGENT (mutation assays): histidine deficient agar

NUMBER OF REPLICATIONS: triplicate plates

DETERMINATION OF CYTOTOXICITY
- Method: other: condition of background lawn

Evaluation criteria:

For the test to be evaluated positive, it must cause a dose-related increase in the mean revertants per plate of at least one tester strain with a minimum of two increasing concentrations of test article. TA1535 and TA1537 were judged positive if the increase in mean revertants is equal to or greater than three times the mean vehicle control value. TA98, TA100, WP2 uvrA (pKM101) and WP2 (pKM101) were judged positive in the increase in mean revertants is equal to or greater than two times the mean vehicle control value.

Statistics:

None stated in report

Results and discussion

Test results

Test results 1

Key result

Species / strain:

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

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A pKM 101

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

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

E. coli, other: WP2 (pKM101)

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

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS

- Precipitation: Precipitation was observed at 333 µg/plate and above

ADDITIONAL INFORMATION ON CYTOTOXICITY: Cytotoxicity was not evident at any concentration

Remarks on result:

other: No mutagenic potential

Any other information on results incl. tables

Summary of results – Experiment B1 revertants per plate (mean of 3 plates)

Dose µg/plate	+/- metabolic activation	Average revertants per plate
		TA98	TA100	TA1535	TA1537	WP2 uvrA (pKM101)	WP2 (pKM101)
Solvent control	-	21	143	11	7	210	44
100	-	19	158	11	6	212	52
333	-	24	141	11	7	199	50
1000	-	20	136	15	6	232	48
3333	-	20	142	14	4	163	53
5000	-	22	153	14	6	169	46
Positive control	-	804	675	467	156	1963	205
Solvent control	+	28	157	15	9	276	58
100	+	27	159	15	9	294	61
333	+	29	188	10	8	242	50
1000	+	30	182	12	8	254	52
3333	+	30	157	14	7	287	55
5000	+	28	180	12	7	270	58
Positive control	+	1363	1241	94	179	1395	2018

NOTE: the solvent is described as acetone in the method section but in most of the tables of results it is reported to be DMSO. It is thought by the reviewer that this is a typographical error, however if DMSO was used rather than acetone the study would still be valid.

Applicant's summary and conclusion

Conclusions:

Silsesquioxanes, phenyl has been tested for mutagenicity to bacteria, in a study which was conducted according to a protocol that was similar to OECD 471, and in compliance with GLP. No evidence of a test substance related increase in the number of revertants was observed with or without activation in the experiment. 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.