Ethoxytrimethylsilane

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Gene mutation (Bacterial reverse mutation assay/ Ames test): negative in S. typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 with and without metabolic activation (according to OECD 471)

Mutagenicity in mammalian cells: negative in mouse lymphoma cells (according to OECD 490)

Cytogenicity in mammalian cells: negative in Chinese hamster V79 fibroblasts (according to OECD 473)

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2002-05-07 to 2002-08-29

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Freie und Hansestadt Hamburg, Behörde für Arbeit, Gesundheit uns Soziales, Germany

Type of assay:

bacterial reverse mutation assay

Target gene:

His operon

Species / strain / cell type:

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

Species / strain / cell type:

S. typhimurium TA 102

Metabolic activation:

with and without

Metabolic activation system:

Aroclor induced rat liver S9

Test concentrations with justification for top dose:

100, 316, 1000, 3160 and 5000 µg/plate with and without metabolic activation

Vehicle / solvent:

- Vehicle/solvent used: ethanol

- Justification for choice of solvent/vehicle: Solubility properties and relative non-toxicity to bacteria

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

cyclophosphamide

methylmethanesulfonate

other: 2-anthracene amide

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION

- Preincubation period: 60 minutes

- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: 3 plates for each test concentration in two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: Background lawn assessment, revertant count

Evaluation criteria:

A result is positive if the number of revertants is significantly increased compared with the solvent control to at least 2-fold of the solvent control for TA 98, TA 100 and TA 102 and 3-fold of the solvent control for TA 1535 and TA 1537 in both experiments.

Positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking on histidine-free agar plates.

Cytotoxicity is defined as a reduction in the number of colonies by >50% compared with the solvent control and/or a sparse background lawn.

Statistics:

MANN and WHITNEY and Spearman’s rank.

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 3160 (only preincubation method) and 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 3160 (only preincubation method) and 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 3160 (only preincubation method) and 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 3160 (only preincubation method) and 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 3160 (only preincubation method) and 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

COMPARISON WITH HISTORICAL CONTROL DATA: Results were within range of historical control data

Table 2: Dose range-finding study Number of revertants per plate (2 plates)

	TA100
Conc. (µg/plate)	Plate 1	Plate 2	Cytotoxic (yes/no)
0*	137	147	No
0.316	133	130	No
1	126	114	No
3.16	118	132	No
10	130	122	No
31.6	153	121	No
100	181	126	No
316	170	175	No
1000	151	172	No
3160	124	118	No
5000	133	123	Yes

*solvent control with Ethanol

Table 3: Experiment 1 Plate incorporation Number of revertants per plate (mean of 3 plates)

	TA98			TA100			TA102
Conc. (µg/plate)	—  MA	+ MA	Cytotoxic (yes/no)	— MA	+ MA	Cytotoxic (yes/no)	—  MA	+ MA	Cytotoxic (yes/no)
0*	41	38.7	No	131.7	128.7	No	292	294.3	No
100	40	31.7	No	172	113	No	266.7	268.3	No
316	45.7	37.7	No	150.7	136.3	No	279	262	No
1000	40.7	34.7	No	154.7	131.7	No	281	246.7	No
3160	45.7	36.7	No	162	148.3	No	278	287	No
5000	37	32.7	Yes	146.3	161.3	Yes	272.7	274.3	Yes
Positive control	1169.3	1177.3	No	1105.7	1134.3	No	1177.7	1095	No

*solvent control with ethanol

Table 3: Experiment 1 Plate incorporation Number of revertants per plate (mean of 3 plates)

	TA1535			TA1537
Conc. (µg/plate)	— MA	+ MA	Cytotoxic (yes/no)	— MA	+ MA	Cytotoxic (yes/no)
0*	15	13.3	No	7.3	7.3	No
100	14	14.7	No	4	7	No
316	16.7	14.7	No	3.7	4.7	No
1000	16.3	15	No	4.3	7	No
3160	13.7	16	No	4	6.3	No
5000	13.7	15.7	Yes	5	6	Yes
Positive control	575.3	584.3	No	579.3	581	No

*solvent control with ethanol

Table 4: Experiment 2 Preincubation Number of revertants per plate (mean of 3 plates)

	TA98			TA100			TA102
Conc. (µg/plate)	— MA	+ MA	Cytotoxic (yes/no)	—  MA	+  MA	Cytotoxic (yes/no)	— MA	+  MA	Cytotoxic (yes/no)
0*	38.3	36.3	No	136	165.7	No	295	282.7	No
100	36	38	No	137.7	167	No	291	291.7	No
316	31.7	40.3	No	138.3	164.3	No	303.3	280.7	No
1000	30	37.7	No	133.7	164	No	286	279.3	No
3160	28.7	41.3	Yes	159	155	Yes	287.7	294.3	Yes
5000	29.3	44.7	Yes	143.3	145	Yes	292.3	284.3	Yes
Positive control	1292	1255.7	No	1163.7	1191	No	1137.7	1218	No

*solvent control with ethanol

Table 4: Experiment 2 Preincubation Number of revertants per plate (mean of 3 plates)

	TA1535			TA1537
Conc. (µg/plate)	— MA	+ MA	Cytotoxic (yes/no)	— MA	+ MA	Cytotoxic (yes/no)
0*	15.3	15	No	4.7	5.3	No
100	14	14	No	3	3.7	No
316	14	13	No	4.3	5	No
1000	12	13	No	4	3	No
3160	15	13.3	Yes	3.7	3.7	Yes
5000	14.3	13	Yes	4	5	Yes
Positive control	565.3	566	No	566.3	563	No

*solvent control with ethanol

MA: Metabolic Activation

Conclusions:

In a reliable test, conducted under OECD 471, with GLP, no mutagenic effect was observed for the test substance tested up to cytotoxic concentration in any of the test strains in two independent experiments without and with metabolic activation. The test substance is non-mutagenic in test strains used.

Reference 2

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 490 (In Vitro Mammalian Cell Gene Mutation Tests Using the Thymidine Kinase Gene)

Version / remarks:

29 July 2016

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

OGYÉI National Institute of Pharmacy and Nutrition, Budapest, Hungary

Type of assay:

in vitro mammalian cell gene mutation tests using the thymidine kinase gene

Target gene:

thymidine kinase locus

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

CELLS USED
- Source of cells: American Type Culture Collection (Manassas, Virginia, USA)

MEDIA USED
- Type and identity of media including CO2 concentration if applicable: Growth medium, RPMI-10 medium, consisted of 10% v/v horse serum (heat inactivated); 0.01 mL/mL antibiotic-antimycotic solution; 0.5 mg/mL Pluronic-F68; and 0.2 mg/mL pyruvic acid. Treatment medium RPMI-5 medium, was the same as growth medium except 5% v/v horse serum. At the end of the expression period, RPMI-20 was used and consisted of: 20% v/v horse serum (heat inactivated); 0.01 mL/mL antibiotic-antimycotic solution; and 0.2 mg/mL pyruvic acid.
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes

Metabolic activation:

with and without

Metabolic activation system:

phenobarbital/β-naphtholflavone induced rat liver S9-mix

Test concentrations with justification for top dose:

Assay 1, 3-hour treatment with metabolic activation: 2000, 1000, 500, 250, 125, 62.5, 31.25, 15.625, 7.813 and 3.906 µg/mL,
Assay 1, 3-hour treatment without metabolic activation: 2000, 1000, 500, 250, 125, 62.5, 31.25, 15.625, 7.813 and 3.906 µg/mL,
Assay 2, 3-hour treatment with metabolic activation: 2000, 1000, 500, 250, 125, 62.5, 31.25, 15.625, 7.813 and 3.906 µg/mL,
Assay 2, 24-hour treatment without metabolic activation: 2000, 1000, 500, 250, 125, 62.5, 31.25, 15.625, 7.813 and 3.906 µg/mL.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: Acetone was selected based on the solubility of the test material.

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

cyclophosphamide

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium
- Cell density at seeding: 2x10E5 cells/mL

DURATION
- Exposure duration: 3 or 24 h
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent): 12 days

SELECTION AGENT (mutation assays): trifluorothymidine (TFT)

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: four 96-well plates at a density of approximately 2x10E3 cells per well.

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: suspension growth (SG; the number of times the cell number increased from the starting cell density); relative total growth (RTG; the product of the relative suspension growth [RTG] and the relative cloning efficiency [RCE])

Evaluation criteria:

The assay was considered valid if all of the following criteria were met:
1. The mutant frequency in the negative (vehicle/solvent) control cultures fell within the normal range (50-170 mutants per 10E6 viable cells).
2. The positive controls met at least one of the following two criteria: (1) The positive control chemical demonstrated an absolute increase in total MF that is, an increase above the spontaneous background MF of at least 300 x 10E-6. At least 40% of the IMF reflected in the small colony MF; and (2) the positive control substance had an increase in the small colony MF of at least 150 x 10E-6 above that seen in the concurrent untreated/vehicle control (a small colony IMF of 150 x 10-6).
3. The cloning efficiency (PEviability) of the negative (vehicle) controls was within the range of 65% to 120% at the end of the expression period.
4. At least four test concentrations were present, where the highest concentration produced approximately 80-90% toxicity (measured by RTG), resulted in precipitation, or it was 2 mg/mL, 2 µL/mL or 0.01 M (whichever was the lowest), or it was the highest practical (achievable) concentration.

The test item was considered to be clearly positive (mutagenic) in this assay if all the following criteria were met:
1. At least one concentration exhibited a statistically significant increase (p < 0.05) compared with the concurrent negative (vehicle) control and the increase was biologically relevant higher than the corresponding negative (vehicle/solvent) control value.
2. The increases in mutation frequency were reproducible between replicate cultures and/or between tests.
3. The increase was concentration-related (p ≤ 0.05) as indicated by the linear trend analysis.

The test item was considered clearly negative (non-mutagenic) in this assay if in all experimental conditions examined there was no concentration related response or, if there is an increase in MF, but it did not exceed the Global Evaluation Factor (GEF).

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: There were no large changes in pH.
- Effects of osmolality: There were no large changes in osmolality .
- Precipitation: No precipitation was detected in the samples.

Conclusions:

In conclusion, no mutagenic effect of ethoxy(trimethyl)silane was observed in the presence or in the absence of metabolic activation system under the conditions of this Mouse Lymphoma Assay.

Reference 3

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

2016

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

OGYÉI National Institute of Pharmacy and Nutrition, Budapest, Hungary

Type of assay:

in vitro mammalian chromosome aberration test

Target gene:

not applicable

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (if applicable):

CELLS USED
- Source of cells: ECACC (European Collection of Authenticated Cells Cultures)
- Suitability of cells: stability of karyotype and morphology makes it suitable for genetic toxicity assays with low background aberrations.
- Cell cycle length, doubling time or proliferation index: doubling time = 12-14 h
- Number of passages: not more than 5 before test started
- Modal number of chromosomes: diploid number, 2n=22

MEDIA USED
- Type and identity of media including CO2 concentration if applicable: Dulbecco’s Modified Eagle’s Medium supplemented with 2 mM L-glutamine, 1% (v/v) Antibiotic-antimycotic solution (standard content: 10000 NE/mL penicillin, 10 mg/mL streptomycin and 25 μg/mL amphotericin-B) and 10% (v/v) heat-inactivated fetal bovine serum (DMEM-10, culture medium); approximately 5%CO2
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically 'cleansed' against high spontaneous background: not specified

Additional strain / cell type characteristics:

not applicable

Cytokinesis block (if used):

Colchicine (0.2 μg/mL)

Metabolic activation:

with and without

Metabolic activation system:

cofactor-supplemented post-mitochondrial S9 fraction prepared from activated (phenoparbital and β-naphthoflavone) rat liver

Test concentrations with justification for top dose:

12.35, 37.04, 111.1, 333.3, and 1000 µg/mL for 3-hour treatment with and without S9 mix
concentration selection was based on cytotoxicity assays performed with ten test concentrations between 2000 and 3.906 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: the test item was insoluble at 200 mg/mL concentration using dimethyl sulfoxide (DMSO) or distilled water as vehicles. However, the 400 mg/mL concentration was achievable using acetone as vehicle (solvent). Acetone was selected for vehicle (solvent) of the study. The vehicle was compatible with the survival of the cells and the S9 activity.

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

ethylmethanesulphonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 3 or 20 hours
- Expression time (cells in growth medium): 20 hours (approximately 1.5 normal cell cycles)
- Fixation time (start of exposure up to fixation or harvest of cells): 20 hours

SPINDLE INHIBITOR (cytogenetic assays): Colchicine (0.2 μg/mL) was added 2-2.5 hours prior to harvesting

STAIN (for cytogenetic assays): 5% Giemsa solution

NUMBER OF REPLICATIONS: 2

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: 2-2.5 hours prior to harvesting, cell cultures were treated with Colchicine (0.2 μg/mL). The cells were swollen with 0.075 M KCl hypotonic solution for 4 minutes, then were washed in fixative (methanol : acetic acid 3 : 1 (v : v) mixture) until the preparation became plasma free (4 washes). Then, a suspension of the fixed cells* was dropped onto clean microscope slides and air-dried. The slides were stained with 5% Giemsa solution, air-dried and coverslips were mounted. Three slides were prepared for each culture.

NUMBER OF CELLS EVALUATED: at least 150

NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE (if in vitro cytogenicity study in mammalian cells): At least 150* metaphases with 22 ± 2 chromosomes (centromeres) from each culture (replicate) will be examined for the presence or absence of chromosomal aberrations (approximately 1000x magnification), where possible. Chromatid and chromosome type aberrations (gaps, deletions and exchanges) will be recorded separately.
*Note: The examination of slides from a culture was halted when 25 or more metaphases with aberrations (excluding gaps) have been recorded for that culture.


DETERMINATION OF CYTOTOXICITY
- Method: At the scheduled harvesting time, the number of surviving cells was determined using a haemocytometer. Results are expressed compared to the negative (vehicle) control as RICC (Relative Increase in Cell Counts)

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes

Evaluation criteria:

The assay is considered valid, if the following criteria are met:
- The negative (vehicle) control data are within the laboratory’s normal range for the spontaneous aberration frequency.
- The positive controls induce increases in the aberration frequency, which are significant.

The test item is considered to have shown clastogenic activity in this study if all of the following criteria are met:
- Increases in the frequency of metaphases with aberrant chromosomes are observed at one or more test concentrations (only data without gaps will be considered).
- The increases are reproducible between replicate cultures and between tests (when treatment conditions were the same).
- The increases are statistically significant.
- The increases are not associated with large changes in pH or osmolality of the treated cultures.

The historical control data for this laboratory were also considered in the evaluation. Evidence of a dose-response relationship (if any) was considered to support the conclusion.
The test item is concluded to have given a negative response if no reproducible, statistically significant increases are observed.

Statistics:

For statistical analysis, Fisher’s exact test was used. The parameter evaluated for statistical analysis was the number of cells with one or more chromosomal aberrations excluding gaps.

Key result

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: none
- Effects of osmolality: none
- Evaporation from medium: not relevant
- Water solubility: insolubility (oily film/minimal oily film) was detected at the end of the treatment period in the final treatment medium in the 1000-333.3 μg/mL concentration range with and without metabolic activation
- Precipitation: see above

RANGE-FINDING/SCREENING STUDIES: Based on the available solubility information (trial formulations of the test item performed at the Test Facility), acetone at 400 mg/mL concentrations was selected for vehicle (solvent) of the study. The highest examined concentration in the preliminary test was 2000 μg/mL.
Two Concentration Selection Cytotoxicity Assays (Assay A: 3-hour treatment with and without metabolic activation, 20-hour harvesting time; and Assay B: 20-hour treatment without metabolic activation, 20-hour harvesting time) were performed as part of the study to establish an appropriate concentration range for the Chromosome Aberration Assays. A total of ten test concentrations between 2000 and 3.906 μg/mL were used to evaluate toxicity in the presence and absence of metabolic activation in each cytotoxicity assay. Detailed results of the cytotoxicity assays are presented in the attached Tables 2,
3 and 4. Treatment concentrations for the chromosome aberration assays were selected on the basis of results of the performed Concentration Selection Cytotoxicity Assays according to the OECD guideline instructions (up to the maximum recommended concentration and/or up to the solubility limit).

CYTOKINESIS BLOCK (if used)
- Distribution of mono-, bi- and multi-nucleated cells: not reported


HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: see attached Tables
- Negative (solvent/vehicle) historical control data: see attached Tables

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: RICC see attached Table 5 to 8

Summary tables of the main experiments (table 9 to 12) are attached.

Conclusions:

In conclusion, the test substance did not induce a significant level of chromosome aberrations in Chinese hamster V79 cells in the performed experiments with and without metabolic activation. Therefore, the test substance was considered as not clastogenic in this test system.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Bacterial mutagenicity

The Ames study on ethoxy(trimethyl)silane was conducted in compliance with GLP and according to OECD 471. Ethoxy(trimethyl)silane was examined in 5 Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and TA 102 in two independent experiments, each carried out without and with metabolic activation. The first experiment was carried out as a plate incorporation test and the second as a pre-incubation test. Five concentrations ranging from 100 to 5000 µg/plate of the test substance were applied. No increase in the number of revertants was observed in any strain and in any test. In the plate incorporation test cytotoxicity was noted at the top concentration of 5000 µg/plate in all test strains carried out without and with metabolic activation. Whereas in the pre-incubation test without metabolic activation cytotoxicity was noted at concentration of 3160 µg/plate ethoxy(trimethyl)silane in test strains TA 100, TA 102, TA 1535 and TA 1537 and at the top concentration of 5000 µg/plate in all test strains. In the experiment with metabolic activation cytotoxicity was noted at concentration of 3160 and 5000 µg/plate ethoxy(trimethyl)silane in all test strains. Appropriate solvent and positive controls were included into the test and gave the expected results. Hence, the test item was considered to be not mutagenic to bacteria under the conditions of the test.

 

 

Mammalian mutagenicity

Ethoxy(trimethyl)silane has been tested for mutagenicity to mammalian cells in a study conducted according to OECD 490 and in compliance with GLP (Citoxlab, 2018). An in vitro mammalian cell assay was performed in mouse lymphoma L5178Y TK+/- cells at the tk locus to test the potential of ethoxy(trimethyl)silane to cause gene mutation and/or chromosome damage. Treatment was performed for 3 hours with and without metabolic activation (±S9 mix) and for 24 hours without metabolic activation (-S9 mix). Acetone was used as vehicle. Concentrations up to 2000 µg/mL were tested. No precipitation was detected in the samples and there were no large changes in pH or osmolality after treatment. Following a 3-hour treatment with and without metabolic activation, no marked cytotoxicity of the test item was observed. No statistically significant or biologically relevant increase in the mutation frequency was noted at any of the evaluated concentrations. No dose-response relationship was observed. The same results were found following a 24 -hour treatment without metabolic activation. The experiments were performed using appropriate untreated, negative (vehicle/solvent) and positive control samples in all cases. The spontaneous mutation frequency of the negative (vehicle/solvent) controls was in the appropriate range. The positive controls gave the anticipated increases in mutation frequency. The plating efficiencies for the negative (vehicle) controls at the end of the expression period were acceptable in all assays. The evaluated concentration ranges were considered to be adequate. The number of test concentrations met the acceptance criteria. Therefore, the study was considered to be valid.

In conclusion, no mutagenic effect of ethoxy(trimethyl)silane was observed in the presence or in the absence of metabolic activation system under the conditions of this Mouse Lymphoma Assay.

 

 

Mammalian cytogenicity

Ethoxy(trimethyl)silane was tested for cytogenicity to mammalian cells in a study conducted according to OECD 473 and in compliance with GLP (Citoxlab, 2018). No test-substance related increase in the number of cells with aberrations was observed when Chinese hamster V79 lung fibroblasts were treated with 12.35, 37.04, 111.1, 333.3, and 1000 μg/mL of test substance in the presence and absence of metabolic activation. Insolubility (oily film/minimal oily film) was detected at the end of the treatment period in the final treatment medium in the 1000-333.3 μg/mL concentration range with and without metabolic activation. There were no large changes in the pH and osmolality. No cytotoxicity was observed in any sample. None of the treatment concentrations caused a biologically or statistically significant increase in the number of cells with structural chromosome aberrations in either assay with or without metabolic activation when compared to the appropriate negative (vehicle) control values. Polyploid metaphases (1-8) were found in some cases in the negative (vehicle) control, positive control or test item treated samples in the performed experiments. No endoreduplicated metaphases were detected in the performed experiments. The negative (vehicle) control data were within the acceptable range for the spontaneous aberration frequency, the positive control substances caused a statistically significant increase in the number of structural aberrations excluding gaps in the experiments with or without metabolic activation demonstrating the sensitivity of the test system.

In conclusion, ethoxy(trimethyl)silane did not induce a significant level of chromosome aberrations in Chinese hamster V79 cells in the performed experiments with and without metabolic activation. Therefore, ethoxy(trimethyl)silane was considered as not clastogenic in this test system.

 

Based on the above results the test substance is considered to have no genetic toxicity potential.

Justification for classification or non-classification

The available data on genetic toxicity of the test substance do not meet the criteria for classification according to Regulation (EC) No 1272/2008, and are therefore conclusive but not sufficient for classification.