Trimethoxysilane

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 with and without activation in all strains tested (OECD TG 471) (Microbiological Associates (1995))

Cytogenicity in mammalian cells: negative without activation, considered to be positive structural, negative numerical with activation (OECD TG 473) (BioReliance (2007)). However the positive effects were observed only in the highest, overtly cytotoxic concentrations so the substance is considered to be negative for cytogenicity.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

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:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

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:

Aroclor 1254-induced rat liver S9 homogenate 

Test concentrations with justification for top dose:

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

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: the solvent was chosen based on solubility of the test article and compatibility with the target cells.

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

TA 98 (without activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA 100, TA 1535 (without activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

TA 1537 (without activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

WP2, WP2uvrA (without activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene

Remarks:

All strains (with activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Sterigmatocystin

Remarks:

All strains (with activation)

Details on test system and experimental conditions:

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

DURATION

- Preincubation period: 60 minutes

- Expression time (cells in growth medium): 48 to 72 hours

NUMBER OF REPLICATIONS: 2 plates per test concentration

DETERMINATION OF CYTOTOXICITY

- Method: relative total growth; background lawn assessment

Evaluation criteria:

The test article is evaluated as positive when it causes a dose-related increase in mean revertants per plate of at least one tester strain with a minimum of two increasing concentrations of test article.

Statistics:

Positive controls and tester strains with revertant counts greater than 100 were counted with a colony counter (Mini Count) and those less than 100 were counted manually.

Key result

Species / strain:

S. typhimurium 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

Remarks:

> 5000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium 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

Remarks:

> 5000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

> 5000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

> 5000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

> 5000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2

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

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: none recorded
- Effects of osmolality: none recorded
- Evaporation from medium: not reported
- Precipitation: Slight precipitation at higher concentrations did not affect assay
- Other confounding effect: In first two tests, a variation in the precipitation pattern was observed, so the tests were repeated. The findings from the initial two experiments are not recorded.

RANGE-FINDING/SCREENING STUDIES:
No toxicity observed

COMPARISON WITH HISTORICAL CONTROL DATA:
The results of the controls lie within the range of the historical control data.


ADDITIONAL INFORMATION ON CYTOTOXICITY: None

Remarks on result:

other: all strains/cell types tested

 Table 1 Preliminary toxicity test

	TA 100			WP2 uvrA (pKM101)
Concentration (µg/Plate)	Plate 1 + MA	Plate 2 - MA	Cytotoxic (Yes/No)	Plate 1 + MA	Plate 2 - MA	Cytotoxic (Yes/No)
0	199	174	no	250	208	no
6.7	180	173	no	234	187	no
10	165	157	no	241	210	no
33	174	163	no	200	198	no
67	179	197	no	228	185	no
100	196	152	no	195	217	no
333	177	170	no	186	184	no
667	175	175	no SP	230	214	no SP
1000	169	156	no SP	138	168	no SP
3333	199	174	no SP	221	169	no SP
5000	174	158	no SP	247	180	no SP

SP Slight precipitate

 

Table 2: Experiment 1 Plate incorporation assay [Number of revertants per plate (mean of 3 plates)]

	TA98			TA100			TA1535
Conc. (µg/plate)	-MA	+ MA	Cytotoxic (yes/no)	-  MA	+ MA	Cytotoxic (yes/no)	- MA	+ MA	Cytotoxic (yes/no)
0*	13	17	no	95	144	no	10	11	no
100	13	17	no	89	127	no	9	11	no
333	13	18	no	86	135	no	8	11	no
1000	14	16	no	85	116	no	9	11	no
3333	17	21	no	97	116	no	6	14	no
5000	13	21	no	95	111	no	7	13	no
Positive Control	1304	1050	-	581	997	-	497	124	-

*solvent control with DMSO

 

Table 2: Experiment 1 Plate incorporation assay [Number of revertants per plate (mean of 3 plates)]

	TA1537			WP2 uvrA (pKM101)			WP2 (pKM101)
Conc. (µg/plate)	-MA	+ MA	Cytotoxic (yes/no)	-  MA	+ MA	Cytotoxic (yes/no)	- MA	+ MA	Cytotoxic (yes/no)
0*	4	6	no	226	263	no	65	64	no
100	4	10	no	236	295	no	54	64	no
333	3	5	no	239	258	no	87	60	no
1000	6	8	no	210	294	no	62	53	no
3333	5	8	no	213	273	no	67	64	no
5000	5	6	no	197	247	no	67	61	no
Positive Control	215	130	-	1554	1262	-	1476	499	-

 

 *solvent control with DMSO

 

Table 3: Experiment 2 Pre incubation assay [Number of revertants per plate (mean of 3 plates)]

	TA98			TA100			TA1535
Conc. (µg/plate)	— MA	+ MA	Cytotoxic (yes/no)	—  MA	+ MA	Cytotoxic (yes/no)	— MA	+ MA	Cytotoxic (yes/no)
0*	15	24	no	102	129	no	11	10	no
50	26	24	no	112	122	no	10	11	no
160	21	20	no	105	116	no	11	9	no
500	24	14	no	101	120	no	12	10	no
1600	23	17	no	110	128	no	9	12	no
5000	19	18	no	100	123	no	10	11	no
Positive Control	764	909	-	527	902	-	433	117	-

*solvent control with DMSO

 

Table 3: Experiment 2 Pre incubation assay [Number of revertants per plate (mean of 3 plates)]

	TA1537			WP2 uvrA (pKM101)			WP2 (pKM101)
Conc. (µg/plate)	-MA	+ MA	Cytotoxic (yes/no)	-  MA	+ MA	Cytotoxic (yes/no)	- MA	+ MA	Cytotoxic (yes/no)
0*	5	7	no	204	334	no	23	21	no
100	5	5	no	274	278	no	25	21	no
333	5	5	no	173	316	no	26	25	no
1000	5	6	no	208	296	no	26	22	no
3333	5	4	no	206	285	no	31	28	no
5000	5	6	no	214	283	no	28	24	no
Positive Control	250	119	-	2225	1304	-	358	1222	-

 

*solvent control with DMSO

 

Conclusions:

Trimethoxysilane has been tested according to OECD Test Guideline 471 and in compliance with GLP. No increase in the number of revertants was observed with or without metabolic activation in either the initial or the repeat assay using Salmonella typhimurium strains and E. coli WP2 uvrA, using the preincubation method. Appropriate solvent and positive controls were included and gave expected results. It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of this study.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromos ome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2006-08-16 to 2006-09-17

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)

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Metabolic activation system:

Aroclor induced rat liver S9

Test concentrations with justification for top dose:

152.5, 305, 610, 1220 µg/ml

In the preliminary toxicity assay, the maximum dose tested was 1220 µg/ml (10 mM). The test article was soluble in DMSO and in the treatment medium at all dose levels tested at the beginning of the treatment period. At the conclusion of the treatment period, visible precipitate was observed in treatment medium at 1220 µg/ml and dose levels = 366 µg/ml were soluble in treatment medium.

Substantial toxicity (at least 50% cell growth inhibition relative to the solvent control) was observed at 1220 µg/ml in both the non-activated and S9-activated 4-hour exposure groups. Substantial toxicity was not observed at any dose level in the non-activated 20-hour exposure group. Based on these findings, the doses chosen for the chromosome aberration assay ranged from 152.5 to 1220 µg/ml for all three exposure groups.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: solubility of the test article and compatibility with target cells.

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

(without activation)

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

(with activation)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION

- Preincubation period: 16 - 24 hours

- Exposure duration: 4 - 20 hours (-MA), 4 hours (+MA)

NUMBER OF REPLICATIONS: 2 flasks per concentration

DETERMINATION OF CYTOTOXICITY

- Method: Cell growth inhibition relative to the solvent control

Evaluation criteria:

Toxicity based on cell growth inhibition relative to solvent control.

The number and types of aberrations found, % aberrant cells in the total population of cells examined, and mean aberrations per cell were calculated and reported for each treatment group.

The test article was considered to induce a positive response when the percentage of cells with aberrations is increased in a dose-responsive manner with one or more concentrations being statistically significant (p=0.05).

Statistics:

Fisher's exact test was used to compare pairwise the percent aberrant cells of each treatment group with that of the solvent control. In the event of a positive Fisher's Exact test at any test article dose level, the Cochran-Armitage test was used to measure dose-responsiveness.

Key result

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with

Genotoxicity:

other: considered positive structural, negative numerical

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1220 µg/ml (4 hour group)

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1220 µg/ml (4 hour group)

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Table 1: Preliminary toxicity test using Trimethoxysilane in CHO cells with and without S9 metabolic activation (4 hour treatment / 16 hour recovery) 

 

	- MA			+ MA			- MA
	4 hour treatment / 16 hour recovery						20 hour continuous treatment)
Treatment µg/ml	Cell Viability (%)	Cell Growth Index (%) *	Cell Growth Inhibition (%) **	Cell Viability (%)	Cell Growth Index (%) *	Cell Growth Inhibition (%) **	Cell Viability (%)	Cell Growth Index (%) *	Cell Growth Inhibition (%) **
DMSO	98	100	-	98	100	-	100	100	-
0.122	99	95	5	100	89	11	99	86	14
0.366	99	79	21	100	96	4	99	92	8
1.22	100	82	18	97	97	3	100	85	15
3.66	97	76	24	99	80	20	97	87	13
12.2	99	78	22	97	81	19	95	86	14
36.6	98	67	33	98	86	14	96	99	1
122	98	63	37	100	98	2	97	94	6
366	99	68	32	99	78	22	98	87	13
1220	93	41	59	95	42	58	91	67	33

* Cell Growth Index = (cells per flask treated group/cells per flask control group), expressed as a percentage

** Cell Growth Inhibition = 100 % - % cell growth index; not calculated for negative controls 

Table 2: Cytogenetic analysis of CHO cells in the absence of metabolic activation (4 hour treatment, 16 hour recovery period)

 

		Control*		152.5 µg/ml		305 µg/ml		610 µg/ml		Positive control
Flask		A	B	A	B	A	B	A	B	A	B
Cytotoxicity		no	no	no	no	no	no	yes	yes	no	no
Chromatid aberrations***	Breaks	0	0	0	0	0	0	1	0	10	7
	Interchanges	0	0	0	0	0	0	0	0	0	1
Chromosome aberrations***	Gaps**	0	0	1	0	0	0	0	0	1	0
	Breaks	0	0	0	0	0	0	0	0	0	0
	Dicentric	0	0	0	0	0	0	0	0	0	0
	Rings	0	0	0	0	0	0	0	0	0	0
% aberrant cells	Numerical	2	3	5	5	4	5	3	3	1	2
	Structural	0	0	0	0	0	0	1	0	28	24
Mitotic index		11.0	11.4	10.8	10.4	10.0	10.4	9.6	8.8	7.4	6.8

* Solvent control with DMSO

** Total gaps

*** Total number of aberrations

Mitotic Index = number of mitotic figures x 100/500 cells counted

% Aberrant Cells: numerical cells include polyploid and endo reduplicated cells; structural cells exclude cells with only gaps

 

Table 3: Cytogenetic analysis of CHO cells in the presence of metabolic activation (4 hour treatment, 16 hour recovery period)

 

		Control*		152.5 µg/ml		305 µg/ml		1220 µg/ml		Positive control
Flask		A	B	A	B	A	B	A	B	A	B
Cytotoxicity		no	no	no	no	no	no	yes	yes	yes	yes
Chromatid aberrations***	Breaks	0	0	0	1	0	2	1	8	2	7
	Interchanges	0	0	1	1	3	1	7	4	8	6
Chromosome aberrations***	Gaps**	1	0	1	0	2	0	2	3	0	1
	Breaks	0	0	0	0	0	0	0	0	0	0
	Dicentric	0	1	0	0	0	0	1	1	0	0
	Rings	0	0	0	0	0	0	0	0	2	1
% aberrant cells	Numerical	5	4	5	5	5	4	3	4	5	4
	Structural	0	1	1	2	3	3	8	9	16	18
Mitotic index		11.6	11.6	11.2	11.4	10.6	11.2	5.4	6.4	4.0	3.8

* Solvent control with DMSO

** Total gaps

*** Total number of aberrations

Mitotic Index = number of mitotic figures x 100/500 cells counted

% Aberrant Cells: numerical cells include polyploid and endo reduplicated cells; structural cells exclude cells with only gaps

 

Table 4: Cytogenetic analysis of CHO cells in the absence of metabolic activation (20 hour continuous treatment)

 

		Control*		152.5 µg/ml		305 µg/ml		610 µg/ml		Positive control
Flask		A	B	A	B	A	B	A	B	A	B
Cytotoxicity		no	no	no	no	no	no	yes	yes	no	no
Chromatid aberrations***	Breaks	0	0	0	0	0	1	0	0	4	3
	Interchanges	0	0	0	0	0	0	0	0	2	3
Chromosome aberrations***	Gaps**	0	0	0	0	0	0	0	1	1	0
	Breaks	0	0	0	0	0	0	0	0	0	0
	Dicentric	0	0	0	0	0	0	0	0	0	0
	Rings	0	0	0	0	0	0	0	0	0	0
% aberrant cells	Numerical	3	4	4	4	4	3	4	4	4	3
	Structural	0	0	0	0	0	1	0	0	24	20
Mitotic index		10.4	10.0	10.2	10.4	9.4	10.0	5.2	4.8	6.6	6.0

* Solvent control with DMSO

** Total gaps

*** Total number of aberrations

Mitotic Index = number of mitotic figures x 100/500 cells counted

% Aberrant Cells: numerical cells include polyploid and endo reduplicated cells; structural cells exclude cells with only gaps

 

 

Conclusions:

In a reliable study conducted in accordance with OECD Test Guideline 473 and in compliance with GLP, trimethoxysilane was concluded to be positive for the induction of structural and negative for the induction of numerical chromosome aberrations in CHO cells in the S9-activated test system at the highest dose tested. However these effects were observed only in the highest, overtly cytotoxic concentration, so the substance is considered to be negative for cytogenicity. Appropriate solvent and positive controls were included and gave expected results.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Micronucleus assay inhalation study in rat: negative (similar to OECD TG 474) (Dow Corning Corporation (1982))

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1982-04-21 to 1982-05-14

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was conducted according to an appropriate OECD test guideline with minor deviations. It was conducted in compliance with GLP. The deviations are that only 1000 cells were scored compared with 2000 in current guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

1000 cells scored

Principles of method if other than guideline:

Micronucleus test in vivo: Matter and Schmid, 1971, Mut. Res. 12: 417-425.

GLP compliance:

yes

Type of assay:

micronucleus assay

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS

- Source: Spartan Research Laboratories, Inc. Harlet, MI

- Weight at study initiation: 100 to 175 grams

- Housing: Animals housed individually

- Diet (e.g. ad libitum): PURINA Rodent Laboratory Chow (ad libitum)

- Water (e.g. ad libitum): ad libitum

Route of administration:

inhalation

Vehicle:

- Vehicle(s)/solvent(s) used: air

- Concentration of test material in vehicle: 100 ppm

Details on exposure:

TYPE OF INHALATION EXPOSURE: nose only

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION

- Exposure apparatus: specially constructed glass chamber

- System of generating particulates/aerosols: vapours were generated by bubbling clean, dry air through the liquid test material

Duration of treatment / exposure:

4 hour(s)

Frequency of treatment:

Single 4 hour exposure

Post exposure period:

30 hours

Remarks:

Doses / Concentrations:
100 ppm
Basis:
nominal conc.

No. of animals per sex per dose:

5 animals per dose level

Control animals:

yes, concurrent no treatment

Positive control(s):

triethylenemelamine

- Route of administration: split-dose intraperitoneal injection (0 and 24 hours)

- Doses / concentrations: 0.5 mg/kg/dose

Tissues and cell types examined:

Animals were exposed to the test article by acute inhalation. They were sacrificed, the bone marrow is extracted and smear preparations made and stained. Polychromatic erythrocytes are scored for micronuclei under the microscope. Both positive and negative (solvent) controls are used in each experiment.

Details of tissue and slide preparation:

At sacrifice the adhering soft tissue and epiphyses of both tibiae were removed. The marrow was aspirated from the bone and transferred to centrifuge tubes containing 5 ml fetal calf serum (one tube for each animal). Following centrifugation to pellet the tissue, the supernatant was drawn off and portions of the pellet was spread on slides and air-dried. The slides were then stained in May-Gruenwald Solution and Giemsa. A thousand polychromatic erythrocytes (PCEs) per animal were scored. The frequency of micronucleated was expressed as percent micronucleated cells based on the total PCEs present in the scored optic field.

Evaluation criteria:

In the normal animal, the normocytes/PCE's is approximately 2. If an agent inhibits the proliferation of erythroblasts the proportion of PCE's is generally reduced. If the agent promotes chromosome breakage or acts as a spindle poison, generally the proportion of red normocytes increases.

Statistics:

Mean normocyte and polychromatic erythrocytes calculated as well as ratio of normocytes to PCE's

Key result

Sex:

female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY

- Dose range: 100 ppm

- Clinical signs of toxicity in test animals: To ensure that the Micronucleus Assay was performed on animals exposed to a lethal concentration of the test material (Group A), a second group of animals (Group B) was simultaneously exposed to the chemical via inhalation as a positive control for lethality. The data demonstrates that both groups (A&B) was exposed to a lethal concentration of the test material. All five animals in Group B experienced weight loss and died within the 14 day observation period. At autopsy all five animals showed extensive lung damage with hemorrhage and atelectasis. Animals exposed via inhalation to the test material all showed slight to moderate evidence of lung damage in the form of petechial hemorrhage and focal atelectasis. Some evidence of kidney congestion was noted in several animals. No abnormal pathology was evident in either the positive control (C) or negative control (D) groups.

- Evidence of cytotoxicity in tissue analyzed: The ratio of of normocytes to PCE's obtained with both the test material treatment (Group A = 10.64) and the negative control (Group D = 10.28) closely approximate the normal expected ratio and were fairly consistent. The mean normocyte count from the positive control group was elevated (Group C = 19.4) indicating that the animals responded to a known clastogen (chromosome breaking agent). This same trend verified by the increase in percentage of micronucleated PCEs in the positive control group.

- Harvest times: Group A (Treatment group): 30 hours after exposure. Group B (Toxicity Control): 14 days. Group C (Positive Control): 30 hours. Group D (Negative Control): 30 hours.

- High dose with and without activation: 100 ppm

RESULTS OF DEFINITIVE STUDY

- Induction of micronuclei (for Micronucleus assay): Within normal range for all groups

- Ratio of PCE/NCE (for Micronucleus assay): Within normal range for all groups

- Statistical evaluation: Comparison of the study groups by Student T-test using a SAS computer program confirms there is no difference between the mean PCE micronucleus count for study groups A and D (test article and negative control) (<.0001) while the positive control is definitely positive (p>0.5) compared to the former groups.

The following table indicates that the animals responded to the positive control substance.

Both Normocytes/PCE **  ratio and % of micronucleated PCEs were significantly increased in the positive control group when compared to test material-treated group.

Table 3 : Mean Results of in vivo micronucleus test with mouse bone marrow

		Test Group (A)	Positive Control (C)	Negative Control (D)
Number of cells evaluated		1000	1000	1000
Sampling time (h)		30h	30h	30h
Number of erythrocytes	Normocytes / Field	10.64	19.4	10.28
	PCE / Field	4.36	4.56	4.36
	Micronuclei / 1000 PCE	6.8	39.2	5.6
Ratio of erythrocytes	Normochromatic / polychromatic	2.52	4.38	2.39
	% Micronucleated PCE	0.68	3.92	0.56

 

Conclusions:

In a reliable study conducted according to a protocol similar to the OECD Test Guideline 474 with minor deviations, trimethoxysilane did not induce chromosome breakage or act as a spindle poison in the rodent micronucleus assay even when animals were exposed to lethal concentrations. There is no evidence from this study that the mechanism of kill at toxic levels of exposure involves genotoxicity. The test substance is non-mutagenic in Sprague-Dawley rats under the conditions of this mammalian micronucleus assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

In vitro studies are available for trimethoxysilane for bacterial mutagenicity and cytogenicity in mammalian cells. The results from bacterial mutation tests are in agreement, with both available tests giving negative results. There was evidence for clastogenicity (causing chromosomal aberrations) in the presence of metabolic activation in vitro. However this evidence was observed only in the highest, overtly cytotoxic concentrations, so the substance is considered to be negative for cytogenicity.

In addition, an in vivo study concluded that the in vitro result does not reflect an ability to cause chromosome aberrations in vivo. In view of the availability of an in vivo study, it is not considered necessary to test for mutagenicity in mammalian cells in vitro.

Trimethoxysilane has been tested according to OECD Test Guideline 471 and in compliance with GLP. No increase in the number of revertants was observed with or without metabolic activation in either the initial or the repeat assay using Salmonella typhimurium strains and E. coli WP2 uvrA using the preincubation method.

It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of this study (Microbiological Associates 1995).

This result is supported by a summary of a reliable study conducted according to OECD Test Guideline 471 and in compliance with GLP. The test substance trimethoxysilane did not demonstrate genetic activity in any of the strains used in this study, both with and without metabolic activation. The results indicate that the test substance was not considered mutagenic under the test conditions of OECD 471. (Dow Corning Corporation 1987).

The potential for induction of damage to chromosomes in mammalian cells was investigated by testing trimethoxysilane in a study conducted in accordance with OECD Test Guideline 473 and in compliance with GLP (BioReliance 2007). Trimethoxysilane was concluded to be positive for the induction of  structural and negative for the induction of numerical chromosome aberrations in CHO cells in the S9 activated test system at the highest dose tested. However the positive effects were observed only in the highest, overtly cytotoxic concentrations, so the substance is considered to be negative for cytogenicity.

Trimethoxysilane has been tested in an in vivo mammalian micronucleus study conducted according to a protocol similar to the OECD Test Guideline 474 (with minor deviations). The test substance did not induce chromosome breakage or act as a spindle poison in the rodent micronucleus assay even when animals were exposed to lethal concentrations. There was no evidence from this study that the mechanism of kill at toxic levels of exposure involves genotoxicity.

Therefore it is concluded that the test substance is non-mutagenic in Sprague-Dawley rats under the conditions of this mammalian micronucleus assay.

Justification for classification or non-classification

The available information for the substance indicates that trimethoxysilane (CAS 2487-90-3) does not induce mutations in bacterial cells. The substance does cause evidence of chromosomal aberrations in vitro in the presence of metabolic activation. However these effects were observed only at the highest, overtly cytotoxic concentrations, so the substance is considered to be negative for cytogenicity. In addition, clastogenic effects have not been observed in an in vivo study, so it is concluded that trimethoxysilane does not require classification as a germ cell mutagem according to Regulation (EC) No. 1272/2008.