(chloromethyl)triethoxysilane

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:

29 June to 27 December 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(1997)

Deviations:

yes

Remarks:

only 2-aminoanthracene was used as positive control for metabolic activation, however, in pre-tests of S9 mix benzo(a)pyrene was also used to establish functionality of S9 mix

GLP compliance:

yes (incl. QA statement)

Remarks:

Bayerisches Landesamt fuer Gesundheit und Lebensmittelsicherheit, Landesinstitut fuer Arbeitsschutz und Produktsicherheit, Munich, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

(chloromethyl)triethoxysilane

EC Number:

239-311-3

EC Name:

(chloromethyl)triethoxysilane

Cas Number:

15267-95-5

Molecular formula:

C7H17ClO3Si

IUPAC Name:

(chloromethyl)triethoxysilane

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): (Chloromethyl)triethoxysilane
- Physical state: colourless liquid
- Lot/batch No.: KH09561
- Expiration date of the lot/batch: 29. April 2013
- Stability under test conditions: stable
- Storage condition of test material: at room temperature, protected from light

Method

Target gene:

his operon

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Additional strain / cell type characteristics:

other: deep rough mutation (rfa), defect DNA nucleotide excision repair (uvrB-), TA100 and TA98 contain pkM101 plasmid to detect weak mutagens due to enhancing an error-prone DNA repair system

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 102

Additional strain / cell type characteristics:

other: deep rough mutation (rfa), defect DNA nucleotide excision repair (uvrB-), contain pkM101 plasmid to detect weak mutagens due to enhancing an error-prone DNA repair system

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with phenobarbitale and ß-Naphthoflavone

Test concentrations with justification for top dose:

pre-experiment: 3.16, 10.0, 31.6, 100, 316, 1000, 2500, 5000 µg/plate
experiment I: 10.0, 31.6, 100, 316, 1000, 2500, 5000 µg/plate
experiment II: 5.0, 15.8, 50.0, 158, 500, 1580, 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: The solvent was compatible with the survival of the bacteria and the S9 activity.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Remarks:

aqua dest.

Positive controls:

yes

Positive control substance:

other: sodium azide (NaN3, TA100, TA1535 10µg/plate, -S9); 4-nitro-o-phenylene-diamine (4-NOPD, TA98 10µg/plate, TA1537 40µg/plate, -S9); methylmethanesulfonate (MMS, TA102 1µL/plate, -S9); 2-aminoanthracene (2-AA, TA102 10µg/plate, all other - 2.5µg/plate, +S9)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: triplicates each in two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency & relative total growth

Evaluation criteria:

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 TA98, TA100 and TA102 the number of reversions is at least twice as high
- if in tester strains TA1535 and TA1537 the number of reversions is at least three times higher
than the revision rate of the solvent control.
A test item producing neither a dose related increase in the number of revertants nor a reproducible biologically relevant positive response at any of the dose groups is considered to be non-mutagenic in this system.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

exp.I: for all strains: at 316 µg/plate and higher (-S9) and 1000µg/plate and higher (+S9); exp.II: 500 µg/plate and higher (-S9) and 1580 µg/plate and higher (+S9), each time depending on particular tester strain

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

exp.I: for all strains: at 316 µg/plate and higher (-S9) and 1000µg/plate and higher (+S9); exp.II: 500 µg/plate and higher (-S9) and 1580 µg/plate and higher (+S9), each time depending on particular tester strain

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
The pre-experiment was done with strains TA98 and TA100. Cytotoxicity was detected by a clearing or rather diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately equal or less then 0.5 in relation to the solvent control.

COMPARISON WITH HISTORICAL CONTROL DATA:
the resulted are in range with the historical control data (see tables 1-4 under "Any other information on meterials and methods including tables")

Any other information on results incl. tables

Table 5: Results of experiment I

with or without	Test substance	Mean number of revertant colonies per plate(average of 3 plates Standard deviation)
S9 mix	concentration	Base pair substitutiontype		cross linking type	Frameshift type
	[µg/plate]	TA100	TA1535	TA102	TA98	TA1537
-	acetone	102±12.5	10±4.7	188±6.2	30±10.1	7±1.0
-	10	97±13.0	10±3.1	185±17.2	19±4.7	10±4.5
-	31.6	93±3.2	11±3.8	184±14.6	26±1.7	12±1.5
-	100	96±4.0	6±1.2	188±11.6	27±10.2	8±2.1
-	316	68±4.4 B	8±2.9	152±7.9	21±2.5	8±4.0
-	1000	67±4.2 B	3±1.5 B	89±11.4 B	18±8.7 B	6±1.5 B
-	2500	51±8.5 B	5±1.7 B	89±17.6 B	14±2.5 B	5±1.5 B
-	5000	63±17.7 B	4±2.6 B	82±11.9 B	15±3.2 B	5±1.0 B
neg. control	aqua dest.	113±17.5	11±1.5	195±2.1	21±1.5	17±3.2
pos. control	Name	NaN3	NaN3	MMS	4 -NOPD	4 -NOPD
	µg/plate	10	10	1 µL	10	40
	Mean No. of colonies/plate(average of 3±SD)	974±40.6	984±13.2	1282±141.0	396±20.8	131±8.9
+	acetone	102±11.5	7±2.6	253±14.5	30±6.1	18±3.8
+	10	95±7.5	9±1.7	288±32.0	35±5.3	12±2.0
+	31.6	102±9.5	8±1.5	286±5.1	32±3.8	12±1.0
+	100	105±8.4	8±2.5	261±12.9	44±5.6	20±2.1
+	316	112±16.9	8±2.9	252±10.5	28±7.6	11±2.3
+	1000	75±7.0 B	7±2.5 B	248±41.6	26±8.1 B	6±3.5 B
+	2500	62±5.6 B	4±1.5 B	235±14.5	24±5.3 B	6±0.6 B
+	5000	70±12.3 B	4±0.6 B	253±7.5	21±7.6 B	9±4.4 B
neg. control	aqua dest.	93±7.1	9±5.1	212±9.6	34±7.5	15±5.6
pos. control	Name	2 -AA	2 -AA	2 -AA	2 -AA	2 -AA
	µg/plate	2.5	2.5	10	2.5	2.5
	Mean No. of colonies/plate(average of 3±SD)	1985±133.2	144±5.9	738±74.1	2469±122.3	389±18.6

B:       Background lawn reduced

Table 6: Results of experiment II

with or without	Test substance	Mean number of revertant colonies per plate (average of 3 plates Standard deviation)
S9 mix	concentration	Base pair substitutiontype		cross linkingtype	Frameshifttype
	[µg/plate]	TA100	TA1535	TA102	TA98	TA1537
-	acetone	107 ± 16.1	12 ± 4.9	272 ± 14.0	19 ± 8.7	10 ± 4.4
-	5	119±16.5	13±3.6	266±10.4	19±8.2	9±4.0
-	15.8	112±2.5	15±2.6	235±22.1	22±11.5	7±3.2
-	50	104±7.2	14±4.5	247±8.1	27±5.7	7±2.6
-	158	106±7.6	12±3.5	245±37.9	23±9.8	6±2.1
-	500	96±15.3	8±3.6 B	282±21.7	21±2.1	3±2.3 B
-	1580	51±8.5 B	6±2.6 B	231±22.5	21±3.1	3±2.3 B
-	5000	88±4.6 B	5±0.6 B	189±30.9	21±3.5	3±2.6 B
neg. control	aqua dest.	80±24.4	15±3.5	212±15.7	19±4.0	9±3.1
pos. control	Name	NaN3	NaN3	MMS	4 -NOPD	4 -NOPD
	µg/plate	10	10	1 µL	10	40
	Mean No. of colonies/plate(average of 3±SD)	898±144.4	1289±70.5	1486±379.3	612±122.5	133±4.4
+	acetone	120±13.0	10±3.0	246±18.5	28±5.9	10±2.1
+	5	130±17.2	12±2.0	397±23.2	27±4.6	15±4.0
+	15.8	123±13.0	8±3.1	384±21.2	29±5.5	13±4.0
+	50	134±12.3	8±6.4	317±34.1	35±2.0	14±3.2
+	158	122±9.5	11±2.1	288±46.5	32±4.2	14±5.8
+	500	103±9.9	8±3.1	294±31.3	26±6.9	8±3.2
+	1580	86±21.0 B	4±0.6 B	307±13.2	22±10.0 B	9±0.6 B
+	5000	67±6.1 B	4±1.2 B	286±26.1	21±7.4 B	6±0.6 B
neg. control	aqua dest.	122±3.2	10±1.0	229±18.3	32±4.2	8±1.5
pos. control	Name	2 -AA	2 -AA	2 -AA	2 -AA	2 -AA
	µg/plate	2.5	2.5	10	2.5	2.5
	Mean No. of colonies/plate(average of 3±SD)	2136±261.4	107±14.0	525±7.4	1892±52.5	332±56.6

B: Background lawn reduced

Applicant's summary and conclusion

Conclusions:

Interpretation of results: negative

In conclusion, it can be stated that during the described mutagenicity test, conducted according to OECD 471 and under GLP, and under the experimental conditions reported, (Chloromethyl)triethoxysilane did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used. Therefore, (Chloromethyl)triethoxysilane is considered to be non-mutagenic in this bacterial reverse mutation assay.