Ethenyltrimethoxysilane was studied in female rats in a single dose oral toxicity test at 2000 mg/kg in the first trial and at 300 mg/kg in second and third trials. Two animals receiving 2000 mg/kg died within 3 days of administration. On observation for general signs, diarrhea, perianal soiling, and reddish urine were noted at 2000 mg/kg, and diarrhea and perianal soiling at 300 mg/kg. At necropsy, small thymi and spleens were noted in the dead animals at 2000 mg/kg. On histopathological examination, atrophy of the cortex of the thymus and the red and white pulp of the spleen were noted. Therefore, the chemical was classified into category 4 of the GHS regarding acute toxicity.

Ethenyltrimethoxysilane was further studied in rats in an OECD combined repeated dose toxicity study with the reproductive/developmental toxicity screening test at oral doses of 0, 62.5, 250 and 1000 mg/kg.

Two males and 1 female died in the 1000 mg/kg group. Soiled hair, decrease in locomotor activity, reddish urine, hypothermia, perioral smudges, perianal soiling, diarrhea, bradypnea, and/or piloerection were noted in the dying animals. Soiled hair and reddish urine were noted in the surviving males and females of the 1000 and 250 mg/kg groups, along with low body weights and food consumption in males and females receiving 1000 mg/kg. On urinalysis, occult blood, epithelial cells, erythrocytes, and leucocytes were noted in both sexes of the 1000 mg/kg group. On hematological examination, low red blood cell count, hemoglobin concentration, hematocrit, MCV, and MCH and high fibrinogen concentration were noted in males of the 1000 mg/kg group, low hematocrit was noted in females of the 1000 and 250 mg/kg groups, and low hemoglobin concentration and high APTT were noted in females of the 1000 mg/kg group. On blood chemical analysis, low total protein, albumin, A/G ratio, and potassium and high g-GTP, urea nitrogen, and creatinine were noted in males of the 1000 mg/kg group, low total protein and triglycerides were noted in females of the 1000 mg/kg group, and a tendency for high g-GTP was noted in females of the 1000 and 250 mg/kg groups. At necropsy, swollen kidneys were evident in males of the 1000 mg/kg group. On organ weight measurement, low absolute thymus weights, high absolute kidney weights and high relative weights of the spleen, kidneys, and adrenals were noted in males of the 1000 mg/kg group, low relative weights of the thymus in the females of the 1000, 250 and 62.5 mg/kg groups, and low absolute thymus weights and high relative liver weights in females of the 1000 mg/kg group. On histopathological examination, hyperplasia of transitional epithelium in the urinary bladder was noted in males of the 1000, 250 and 62.5 mg/kg groups, vacuolization of the lamina propria in the duodenum, jejunum, and/or ileum in males of the 1000 and 250 mg/kg groups, and sinus vacuolization in the mesenteric lymph nodes, hyperplasia of transitional epithelium and pyelonephritis in the kidney, along with hyperplasia of transitional epithelium in the urethra in males of the 1000 mg/kg group. Vacuolization of the lamina propria in the duodenum or jejunum and hyperplasia of transitional epithelium in the urinary bladder were also noted in females of the 1000 and 250 mg/kg groups, and atrophy of cortex and medulla in the thymus, sinus vacuolization in the mesenteric lymph nodes, hyperplasia of transitional epithelium, regeneration of urinary tubules, and pyelitis in the kidney, hyperplasia of transitional epithelium, metaplasia of keratinized stratified squamous epithelium, cellular infiltration, and necrosis of epithelium in the urethra in females of the 1000 mg/kg group.

The NOAEL for repeated dose toxicity was therefore estimated to be less than 62.5 mg/kg/day for both sexes.

Regarding reproductive/developmental toxicity, a low number of estrous cases was noted in the 1000 mg/kg group. No changes attributable to the chemical were noted for the copulation index, number of conceiving days, number of pregnant females, fertility index, gestation length, gestation index, delivery conditions, nursing conditions, number of corpora lutea, number of implantation sites, or the implantation rate.

The NOAELs for reproductive performance of parental animals were estimated to be 1000 mg/kg/day for males and 250 mg/kg/day for females.

Regarding the pups, no changes attributable to the chemical were noted in the number, number of stillbirths, number of pups born, sex ratio, delivery index, birth index, live birth index, general signs, number of live pups on Day 4 of lactation, viability index of pups on Day 4 of lactation, appearance, body weight, or necropsy findings.

The NOAEL for pups was estimated to be 1000 mg/kg/day.

A reverse mutation test using bacteria was performed to examine the mutagenic potential. Ethenyltrimethoxysilane did not induce gene mutations in bacteria under the conditions of this study.

An in vitro chromosomal aberration test of ethenyltrimethoxysilane was performed using a fibroblast cell line (CHL/IU) from a Chinese hamster lung. Ethenyltrimethoxysilane induced chromosomal aberrations under the conditions of this study.

1. Single Dose Oral Toxicity 1)

Purity	:	100 %
Test species/strain	:	Rat/Crj:CD(SD)IGS
Test method	:	OECD Test Guideline 423
Route	:	Oral(gavage)
Dosage	:	300, 2000 mg/kg
Number of animals/group	:	Females, 6(300 mg/kg), 3(2000 mg/kg)
Vehicle	:	Corn oil
GLP	:	Yes

　Test results:

Two animals receiving 2000 mg/kg died within 3 days of administration. On observation for general signs, diarrhea, perianal soiling, and reddish urine were noted at 2000 mg/kg, and diarrhea and perianal soiling at 300 mg/kg. At necropsy, small thymi and spleens were noted in the dead animals at 2000 mg/kg. On histopathological examination, atrophy of the cortex of the thymus and of red and white pulp of the spleen were noted in the dead animals at 2000 mg/kg. The chemical was therefore classified into category 4 of the GHS regarding acute toxicity.

2. Repeated Dose and Reproductive/Developmental Toxicity 1)

Purity	:	100 %
Test species/strain	:	Rat/Crj:CD(SD)IGS
Test method	:	OECD Test Guideline 422
Route	:	Oral(gavage)
Dosage	:	0(vehicle), 62.5, 250, 1000 mg/kg
Number of animals/group	:	Males, 6 + 6(recovery group); females, 12 + 6(recovery group)
Vehicle	:	Corn oil
Administration period	:	Males, 42 days Females, from 14 days before mating to day 6 of lactation
Terminal killing	:	Males, 43 days or 57 days Females, day 7 of lactation or 57 days
GLP	:	Yes

　Test results:

<Repeated dose toxicity>

Two males and 1 female died in the 1000 mg/kg group. Soiled hair, decrease in locomotor activity, reddish urine, hypothermia, perioral smudges, perianal soiling, diarrhea, bradypnea, and/or piloerection were noted in the dying animals. Soiled hair and reddish urine were noted in the surviving males and females of the 1000 and 250 mg/kg groups. Low body weights and food consumption were also noted in males and females of the 1000 mg/kg group. On urinalysis, occult blood, epithelial cells, erythrocytes, and leucocytes were noted in both sexes receiving 1000 mg/kg. On hematological examination, low red blood cell counts, hemoglobin concentrations, hematocrit, MCV, and MCH and high fibrinogen concentrations were noted in males of the 1000 mg/kg group, low hematocrit in females of the 1000 and 250 mg/kg groups, and low hemoglobin concentrations and high APTT in females of the 1000 mg/kg group. On blood chemical analysis, low total protein, albumin, A/G ratios, and potassium, and high g-GTP, urea nitrogen, and creatinine were noted in males of the 1000 mg/kg group, low total protein and triglycerides in females of the 1000 mg/kg group, and a tendency for high g-GTP in females of the 1000 and 250 mg/kg groups. At necropsy, swollen kidneys were noted in males of the 1000 mg/kg group. On organ weight measurement, low absolute thymus weights, high absolute kidney weights, and high relative weights of the spleen, kidneys, and adrenals were noted in males of the 1000 mg/kg group, low relative thymus weights in females of the 1000, 250 and 62.5 mg/kg groups, and low absolute thymus weights and high relative liver weights in females of the 1000 mg/kg group. On histopathological examination, hyperplasia of transitional epithelium in the urinary bladder was noted in males of the 1000, 250 and 62.5 mg/kg groups, vacuolization of the lamina propria in the duodenum, jejunum, and/or ileum in males of the 1000 and 250 mg/kg groups, and sinus vacuolization in the mesenteric lymph nodes, hyperplasia of transitional epithelium and pyelonephritis in the kidneys, and hyperplasia of transitional epithelium of the urethra in males of the 1000 mg/kg group. Vacuolization of the lamina propria in the duodenum and/or jejunum and hyperplasia of transitional epithelium in the urinary bladder were noted in females of the 1000 and 250 mg/kg groups, and atrophy of the cortex and medulla in the thymus, sinus vacuolization in the mesenteric lymph nodes, hyperplasia of transitional epithelium, regeneration of urinary tubules, and pyelitis in the kidneys, and hyperplasia of transitional epithelium, metaplasia of keratinized stratified squamous epithelium, cellular infiltration, and necrosis of epithelium in the urethra in females of the 1000 mg/kg group.
The NOAEL for repeated dose toxicity was estimated to be less than 62.5 mg/kg/day for both sexes.

<Reproductive and developmental toxicity>

A low number of estrous cases was noted in the 1000 mg/kg group. No changes attributable to the chemical were noted in the copulation index, number of conceiving days, number of pregnant females, fertility index, gestation length, gestation index, delivery conditions, nursing conditions, number of corpora lutea, number of implantation sites, or implantation rate.
The NOAELs for reproductive performance of parental animals were estimated to be 1000 mg/kg/day for males and 250 mg/kg/day for females.
No changes attributable to the chemical were noted in the number of pups, number of stillbirths, number of pups born, sex ratio, delivery index, birth index, live birth index, general signs, number of live pups on Day 4 of lactation, viability index of pups on Day 4 of lactation, appearance, body weight, or necropsy findings.
The NOAEL for pups was estimated to be 1000 mg/kg/day.

3. Genetic Toxicity

3-1. Bacterial test 1)

Purity	:	100 %
Test Species/strain	:	Salmonella typhimurium TA100, TA1535, TA98, TA1537, Escherichia coli WP2 uvrA
Test method	:	Guidelines for Screening Mutagenicity Testing of Chemicals (Chemical Substances Control Law of Japan) and OECD Test Guideline 471
Procedures	:	Pre-incubation method
Solvent	:	Dimethyl sulfoxide
Positive controls	:	-S9 mix; 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, TA98, WP2 uvrA), sodium azide (TA1535) and 9-aminoacridin (TA1537) +S9 mix; 2-Aminoanthracene (all strains)
Dosage	:	-S9 mix; 0, 156.3, 312.5, 625, 1250, 2500, 5000 μg/plate (TA100, TA1535, TA1537, TA98), 0, 312.5, 625, 1250, 2500, 5000 μg/plate (WP2 uvrA) +S9 mix; 0, 312.5, 625, 1250, 2500, 5000 μg/plate (TA100, TA1535, WP2 uvrA, TA98), 0, 156.3, 312.5, 625, 1250, 2500, 5000 μg/ plate (TA1537)
S9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone
Plates/test	:	3
Number of replicates	:	2
GLP	:	Yes

　Test results:

This chemical did not induce gene mutations either in S. typhimurium TA100, TA1535, TA98 or TA1537 or in E. coli WP2 uvrA strains with or without S9 mix. Toxicity was observed at 5000 μg/plate for TA100, TA1535, TA1537 and TA98 without S9 mix, and at 5000 μg/plate for TA1537 with S9 mix.

Genotoxic effects:

Salmonella typhimurium TA100, TA1535，TA98，TA1537

	+	?	-
Without metabolic activation:	[ ]	[ ]	[*]
With metabolic activation:	[ ]	[ ]	[*]

Escherichia coli WP2 uvrA

	+	?	-
Without metabolic activation:	[ ]	[ ]	[*]
With metabolic activation:	[ ]	[ ]	[*]

3-2. Non-bacterial in vitro test(chromosomal aberration test) 1)

Purity	:	100 %
Type of cell used	:	Chinese hamster lung (CHL/IU) cells
Test method	:	Guidelines for Screening Mutagenicity Testing of Chemicals (Chemical Substances Control Law of Japan) and OECD Test Guideline 473
Solvent	:	Dimethyl sulfoxide
Positive controls	:	-S9 mix; Mitomycin C +S9 mix; Dimethyl nitrosamine
Dosage	:	-S9 mix (6 hr short treatment method); 0, 93.8, 187.5, 375, 750, 1500 μg/mL +S9 mix (6 hr short treatment method); 0, 93.8, 187.5, 375, 750, 1500 μg/mL -S9 mix (24 hr continuous treatment method); 0, 93.8, 187.5, 375, 750, 1500 μg/mL
S9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone
Plates/test	:	2
GLP	:	Yes

　Test results:

Structural chromosomal aberrations were induced at 750 and 1500 μg/mL with the 6 hr short treatment method and S9 mix (31.5 and 58.0 %, respectively). Polyploidy was not evident.

Lowest concentration producing cytogenetic effects in vitro:
With metabolic activation (6 hr short treatment method): 0.75 mg/mL
(clastogenicity)

Genotoxic effects:

	clastogenicity			polyploidy
	+	?	-	+	?	-
Without metabolic activation:	[ ]	[ ]	[*]	[ ]	[ ]	[*]
With metabolic activation::	[*]	[ ]	[ ]	[ ]	[ ]	[*]

1)	The tests were performed by Hashima Laboratory, Nihon Bioresearch Inc., 6-104, Majima, Fukuju-cho, Hashima, Gifu, 501-6251, Japan. Tel +81-58-392-6222, Fax +81-58-392-1284