Triethoxy(methyl)silane

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:

07 Nov 2011 - 22 Mar 2012

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:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Triethoxy(methyl)silane

EC Number:

217-983-9

EC Name:

Triethoxy(methyl)silane

Cas Number:

2031-67-6

Molecular formula:

C7H18O3Si

IUPAC Name:

triethoxy(methyl)silane

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Keep the substance in cool and dark place; tightly closed, in refrigerator (1 - 10 °C); fill nitrogen after opening of package.
- Stability under test conditions: The test substance is stable. The stability is confirmed at the end of exposure using IR spectroscopy.
- Solubility and stability of the test substance in the solvent/vehicle: soluble in DMSO at 50 mg/mL
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No reactivity


TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Final preparation of a solid: Test material was measured and diluted in DMSO at 50 mg/mL. This solution was diluted sequencly in the appropriate concentration.

Method

Target gene:

trp operon and his operon

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

S9 mix, prepared from the livers of rats treated with phenobarbital and 5-6-benzoflavone

Test concentrations with justification for top dose:

Experiment 1 and 2: 39.1, 78.1, 156, 313, 652, and 1250 μg/mL were tested for TA1535 (up to the cytotoxic concentration)
Experiment 1 and 2: 156, 313, 652, 1250, 2500 and 5000 μg/mL were tested for TA100 , WP2uvrA, TA98 and TA1537

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: due to the limited solubility of the test substance in water, DMSO was selected as the vehicle

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

benzo(a)pyrene

furylfuramide

other: 6-chloro-9-[3-(2-chloroethylamino)propylamino]-2-methoxyacridine dihydrochloride for TA1537 without S9 mix, 2-amonoanthracene for TA1535 and WP2uvrA with S9 mix

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 minutes
- Exposure duration: 49.5 h for the first experiment and 48 h for the second experiment

NUMBER OF REPLICATIONS:
3 replication each in 2 independent experiments

NUMBER OF CELLS EVALUATED: all cells

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

Evaluation criteria:

A test substance was considered positive in the Ames test if:
a) a more than 2-fold increase in the number of revertants was observed
b) the positive result was reproducible inspite of dose-related increase.

Statistics:

Mean values and standard deviations were calculated.

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:

cytotoxicity

Remarks:

at 1250 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

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:

cytotoxicity

Remarks:

≥ 2500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

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:

cytotoxicity

Remarks:

≥ 2500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

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:

cytotoxicity

Remarks:

≥ 2500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

≥ 2500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
Preliminary test was performed with 19.5, 78.1, 313, 1250 and 5000 μg/mL with/without S9.
There was no reverse mutation observed in all species at all concentration. Furthermore, cytotoxicity was observed at 1250 μg/mL and more (TA1535) and at 5000 μg/mL (WP2urvA, TA98, TA100 and TA1537) with and without metabolic activation.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: number of revertant colonies

Any other information on results incl. tables

Table 1. Test results of the experiment 1 with or without S9 mix for triethoxy(methyl)silane

With or without S9 mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (Mean of 3 replicate ± SD)
		Base-pair substitution type				Frameshift type
		TA100	TA1535		WP2urvA	TA98	TA1537
-	0	84 ± 17.6	6 2.0±		20± 3.5	12± 3.8	5± 2.0
	39.1	-	7± 1.5		-	-	-
	78.1	-	6± 1.0		-	-	-
	156	72± 12.1	8± 0.6		16± 4.9	13± 7.6	5± 0.6
	313	87± 26.2	10± 3.5		17± 2.1	11± 4.6	5± 1.0
	625	71± 10.2	7± 3.5		13± 2.0	9± 3.2	4± 1.2
	1250	67± 7.4	7± 2.6		14± 5.1	14± 7.0	5± 1.5
	2500	0± 0.0*	NT		10± 0.6*	0± 0.0*	0± 0.0*
	5000	0± 0.0*	NT		5± 2.3*	0± 0.0*	0± 0.0*
Positive controls, – S9 mix	Name	AF2	NaN3		AF2	AF2	ICR-191
	Concentration (µg/plate)	0.01	0.5		0.01	0.1	1.0
	Mean No. of colonies/plate ± SD	764± 8.1	350± 11.0		54± 21.5	365± 60.1	908± 69.8
+	0	80± 15.5		6± 1.2	15± 6.1	18± 4.9	10± 1.0
	39.1	NT		8± 1.5	NT	NT	NT
	78.1	NT		8± 2.5	NT	NT	NT
	156	83± 12.1		8± 3.1	17± 3.1	15± 6.5	5± 1.5
	313	93± 10.0		8± 1.5	13± 2.3	22± 7.0	6± 1.0
	625	85± 9.0		8± 0.6	18± 5.0	14± 6.4	4± 2.3
	1250	84± 8.2		6± 2.6*	11± 1.5	15± 5.7	6± 1.5
	2500	51± 8.1*		NT	12± 3.5*	13± 4.7*	0 ± 0*
	5000	0 ± 0*		NT	5 ± 2.0*	0 ± 0*	0 ± 0*
Positive controls, + S9 mix	Positive controls	B[a]P		2AA	2AA	B[a]P	B[a]P
	Concentration (µg/plate)	5.0		2.0	10.0	5.0	5.0
	Mean No. of colonies/plate ± SD	957 ± 93.6		254± 16.5	812± 66.4	250± 44.3	67± 3.1

Table 2. Test results of the experiment 2 with or without S9 mix for triethoxy(methyl)silane

With or without S9 mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate  (Mean of 3 replicate ± SD)
-		Base-pair substitution type			Frameshift type
		TA100	TA1535	WP2urvA	TA98	TA1537
		128 ± 11.5	8 ± 1.5	13 ± 1.2	14 ± 2.5	7 ± 1.2
	39.1	-	4 ± 2.5	-	-	-
	78.1	-	6 ± 0.6	-	-	-
	156	116 ± 7.8	8 ± 4.6	13 ± 2.0	17 ± 3.8	10 ± 4.4
	313	98 ± 6.8	8 ± 6.7	12 ± 2.9	11 ± 4.2	6 ± 1.0
	625	126 ± 14.8	6 ± 3.2	19 ± 3.5	12 ± 1.0	4 ± 0.0
	1250	107 ± 12.5	4 ± 1.5*	12 ± 3.5	12 ± 6.1	6 ± 1.5
	2500	0 ± 0.0*	-	12 ± 2.9*	5 ± 2.3*	0 ± 0.0*
	5000	0 ± 0.0*	-	0 ± 0.0*	0 ± 0.0*	0 ± 0.0*
Positive controls, – S9 mix	Name	AF2	NaN3	AF2	AF2	ICR-191
	Concentration (µg/plate)	0.01	0.5	0.01	0.1	1.0
	Mean No. of colonies/plate ± SD	638 ± 47.9	161 ± 3.2	70 ± 7.6	491 ± 31.2	1232 ± 27.8
+	0	118 ± 9.6	8 ± 1.5	22 ± 5.6	24 ± 9.9	7 ± 1.0
	39.1	NT	7 ± 2.5	NT	NT	NT
	78.1	NT	8 ± 2.5	NT	NT -	NT
	156	127 ± 21.4	8 ± 1.5	14 ± 3.6	19 ± 2.5	7 ± 2.3
	313	119 ± 13.7	8 ± 1.5	18 ± 9.9	23 ± 3.1	6 ± 3.2
	625	123 ± 4.2	8 ± 2.6	15 ± 5.8	25 ± 4.2	8 ± 3.1
	1250	114 ± 8.1	8 ± 2.9*	14 ± 2.1	17 ± 3.5	6 ± 1.0
	2500	52 ± 17.3*	NT	7 ± 2.1	8 ± 1.7*	0 ± 0*
	5000	0 ± 0*	NT	9 ± 2.6*	0 ± 0*	0 ± 0*
Positive controls, + S9 mix	Name	B[a]P	2AA	2AA	B[a]P	B[a]P
	Concentration (µg/plate)	5.0	2.0	10.0	5.0	5.0
	Mean No. of colonies/plate ± SD	937 ± 45.0	191 ± 13.0	776 ± 14.2	311 ± 24.0	97 ± 2.4

 

AF2: Furylfulamide

NaN3: sodium azide

ICR-191: 6-chloro-9-[3-(2-chloroethylamino)propylamino]-2-methoxyacridine dihydrochloride

B[a]P: Benzo(a)pyrene

2AA: 2-aminoanthracene

SD:  standard deviation

*: Growth inhibition

NT: This concentration was not tested.

Applicant's summary and conclusion

Conclusions:

Under the conditions of the Ames test the substance was not mutagenic in any of the five strains (S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvrA) tested with and without metabolic activation up to 1250 µg/plate for TA1535 and 5000 µg/plate for other strains.