Reaction products of [3-(2,3-epoxypropoxy)propyl]trimethoxysilane and triacetoxyvinylsilane.

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:

2010-05-19 to 2010-05-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Method

Target gene:

histidine operon (for the Salmonella strains), tryptophan operon (for the E.coli strain)

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:

phenobarbital/beta-napthoflavone induced rat liver S9

Test concentrations with justification for top dose:

3, 10, 33, 100, 333, 1000, 2500, 5000 μg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: methanol
- Justification for choice of solvent/vehicle: solubility properties and relative nontoxicity to the bacteria

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

methanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA 1535, TA 100 (without metabolic activation)

Controls 2

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

methanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylene-diamine, 4-NOPD

Remarks:

TA 1537, TA 98 (without metabolic activation)

Controls 3

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

methanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

WP2 uvr A (without metabolic activation)

Controls 4

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene, 2-AA

Remarks:

all strains (with metabolic activation)

Details on test system and experimental conditions:

ACTIVATION: The amount of S9 supernatant was 10% v/v in the cultures. Cofactors were added to the S9 mix to reach the following concentrations in the S9 mix:
8 mM MgCl2
33 mM KCl
5 mM Glucose-6-phosphate
4 mM NADP
in 100 mM sodium-ortho-phosphate-buffer, pH 7.4

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
-After solidification, the plates were incubated upside down for 48-72 hours at 37°C in the dark

SELECTION AGENT (mutation assays): histidine-deficient agar (Salmonella strains); tryptophan-deficient agar (E.coli strain)

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY: Toxicity of the test item results in a reduction in the mean number of spontaneous revertants (> 50% reduction) or a reduction of the bacterial background lawn, in comparison to the solvent control.

Evaluation criteria:

A test item is considered as a mutagen is a biologically relevant increase in the mean number of revertants exceeding the threshold of twice (strains TA 98, TA 100 and WP2 uvr A) or thrice (strains TA 1535 and TA 1537) the mean colony count of the corresponding solvent control is observed.

A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.

An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.

Statistics:

No statistical evaluation of the data was required.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

bacteria, other: TA 1535, TA 100, WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

positive

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:

S. typhimurium, other: TA 1537, 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

Any other information on results incl. tables

Table 1: Plate incorporation test: Mean number of revertants per plate (3 plates)

Dose (µg/plate)	TA 1535		TA 1537		TA 98		TA 100		WP2 uvr A
	- MA	+ MA	- MA	+ MA	- MA	+ MA	- MA	+ MA	- MA	+ MA
3	14	29	10	14	26	36	118	124	39	49
10	18	22	9	16	24	41	101	122	42	46
33	20	24	10	16	27	38	118	129	43	52
100	33	33*	8	15*	28	37	123	128	44	50
333	73	72*	9	18*	27	43*	155	132*	45	56*
1000	133	151*	14	18*	25	37*	214	193*	63	57*
2500	242	270*	11	13*	23	35*	374	283*	83	64*
5000	404	407*	14	15*	25	38*	662	511*	129	105*
Untreated	16	19	9	19	27	45	121	127	44	55
Solvent control (methanol)	14	17	9	19	28	40	117	126	40	48
Positive control (NaN3) - 10	1516	-	-	-	-	-	1608	-	-	-
Positive control (4-NOPD) - 10	-	-	-	-	336	-	-	-	-	-
Positive control (4-NOPD) - 50	-	-	66	-	-	-	-	-	-	-
Positive control (MMS) - 3	-	-	-	-	-	-	-	-	749	-
Positive control (2-AA) - 2.5	-	221	-	455	-	2224	-	3198	-	-
Positive control (2-AA) - 10	-	-	-	-	-	-	-	-	-	270

* Reduced background growth

NaN3 = sodium azide

2 -AA = 2 -aminoanthracene

4 -NOPD = 4 -nitro-o-phenylene-diamine

MMS = methyl methane sulfonate

Applicant's summary and conclusion

Conclusions:

In a valid bacterial reverse mutation assay, conducted according to OECD Test Guideline 471 and in compliance with GLP, reaction mass of 3-(2,3-epoxypropoxy)propyltrimethoxysilane and triacetoxyvinylsilane was tested using Salmonella typhimurium TTA 100, TA 1535, TA 98 or TA 1537 and E. coli WP2 uvr A. A substantial and dose dependent increase in the number of revertants was observed with or without metabolic activation in Salmonella typhimurium strains TA 100, TA 1535 an d E. coli WP2 uvr A when tested up to limit concentrations. No evidence of a test-substance related increase in the number of revertants was observed with or without metabolic activation in Salmonel la typhimurium strains TA 98 or TA 1537. Appropriate positive and solvent controls were included and gave the expected results. It is concluded that the test substance is positive for mutagenicity to bacteria under the conditions of the test.