1,1,1,3,3,3-hexamethyldisilazane

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 April 2013 to 19 November 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted according to an appropriate guideline and in compliance with GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

2001

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Version / remarks:

1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Hydroxytrimethylsilane

EC Number:

213-914-1

EC Name:

Hydroxytrimethylsilane

Cas Number:

1066-40-6

Molecular formula:

C3H10OSi

IUPAC Name:

Trimethylsilanol

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): Trimethylsilanol

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crl:CD IGS

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Germany
- Age at study initiation: 12 weeks
- Weight at study initiation: 230 to 316g
- Fasting period before study: no
- Housing: In groups of three to five animals in Makrolon type-4 cages with wire mesh tops up to the day of mating and afterwards individually in Makrolon type-3 cages with wire mesh tops with sterilized standard softwood bedding (‘Lignocel’) with paper enrichment (ISO-BLOX).
- Diet (ad libitum): Pelleted standard Harlan Teklad 2018C
- Water (ad libitum): community tap water
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.2 - 23.5
- Humidity (%): 27.2 - 70.3
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 22 April 2013 To: 23 May 2013

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Trimethylsilanol was weighed into a glass beaker on a tared precision balance and the vehicle (corn oil) was added. Using an appropriate homogenizer, a homogeneous suspension was pre-pared. Separate formulations were prepared for each concentration
Dose formulations were devided into daily aliquots.
Homogeneity of the test item in the vehicle was maintained during the daily administration period using a magnetic stirrer.

VEHICLE
- Concentration in vehicle: 0, 10, 30, 90 mg/mL
- Amount of vehicle: 5 mL/kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

On the first treatment day samples from the control group as well as three samples (top, middle and bottom) of about 1 g of each concentration were taken prior to dosing for analysis of concentration and homogeneity. Samples of about 1 g of each concentration were taken from the middle only to confirm the stability (4 hrs at room temperature and 8 days in refrigerator). During the last week of the treatment, samples were taken from the middle to confirm concentration. The aliquots for analysis of dose formulations were frozen (-20 ± 5 °C) and delivered on dry ice to the person responsible for formulation analysis and stored there at -20 ± 5 °C until analysis.

The samples were analyzed by Headspace GC-Method The test item was used as the analytical standard.

Duplicates were taken of all samples and were stored.

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: 1:1
- Length of cohabitation: until evidence of copulation observed
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug or sperm in vaginal smear referred to as day 0 post coitum
- Any other deviations from standard protocol: no

Duration of treatment / exposure:

Day 6 to Day 20 post coitum

Frequency of treatment:

daily

Duration of test:

21 days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (actual dose received)

Remarks:

Group 1 - Control Group

Doses / concentrations 2

Dose / conc.:

50 mg/kg bw/day (actual dose received)

Remarks:

Group 2

Doses / concentrations 3

Dose / conc.:

150 mg/kg bw/day (actual dose received)

Remarks:

Group 3

Doses / concentrations 4

Dose / conc.:

450 mg/kg bw/day (actual dose received)

Remarks:

Group 4

No. of animals per sex per dose:

22 female

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were selected based on a previous dose range-finding toxicity study in Sprague Dawley rats using dose levels of 0, 50, 150 and 600/400 mg/kg bw/day.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 3 times daily during treatment

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION: Yes
- Time schedule for examinations: recoreded at 3-day intervals, days 0-3, 3-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post coitum
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/animal/day: Yes

WATER CONSUMPTION: No
- Time schedule for examinations:

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- liver weight recorded

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: half per litter

Statistics:

The following statistical methods were used to analyze food consumption, body weights, reproduction and skeletal examination data:

• Means and standard deviations of various data were calculated and included in the report.

• The Dunnett-test (many to one t-test) based on a pooled variance estimate was applied if the variables could be assumed to follow a normal distribution for the comparison of the treated groups and the control groups for each sex.

• The Steel-test (many-one rank test) was applied instead of the Dunnett-test when the data could not be assumed to follow a normal distribution.

• Fisher's exact-test was applied if the variables could be dichotomized without loss of information.

Historical control data:

Data from test facility derived during 1992 to 2005 included

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Clinical signs such as uncoordinated movement, decreased activity and/or prostrate appearance and/or ruffled fur occurred after dosing the animals with dose levels of 150 and/or 450 mg/kg bw/day during the treatment period. These were considered to be related to the treatment with the test item. Due to the full recovery of the animals, slight severity grade and frequency at 150 mg/kg bw/day, this dose level was considered to cause treatment-related but not adverse effects in pregnant rats. Due to the nature, frequency and severity of the findings and the observation that one animal did not fully recover from the symptoms, the dose level of 450 mg/kg bw/day was considered to cause treatment-related and adverse effects in pregnant rats.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Description (incidence):

All females survived until the scheduled necropsy.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

At 450 and 150 mg/kg bw/day, mean absolute body weight and body weight gain were decreased statistically significantly and were considered to be related to the treatment with the test item. At 450 mg/kg bw/day, the corrected body weight gain was also affected by treatment with the test item.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

At 450 and 150 mg/kg bw/day, test item-related statistically significant decreases were observed for mean food consumption.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, treatment-related

Description (incidence and severity):

Uncoordinated movement, decreased activity with dose levels of 150 and/or 450 mg/kg bw/day during the treatment period. These were considered to be related to the treatment with the test item.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

No effects on liver weights were noted.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No findings were observed at any dose level during macroscopical examination.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Maternal developmental toxicity

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

The relevant reproduction data (pre-and post-implantation loss and number of fetuses per dam) were not affected by treatment with the test item.
Mean numbers of corpora lutea and implantation sites were similar across all groups.

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

no effects observed

Other effects:

not specified

Details on maternal toxic effects:

Statistically significantly decreased mean food consumption, body weight, body weight gain and corrected body weight gain were affected by the treatment with the test
item and were considered to be adverse maternal toxic effects.

Effect levels (maternal animals)

Key result

Dose descriptor:

NOAEL

Remarks:

Maternal General Toxicity

Effect level:

ca. 150 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

clinical signs

food consumption and compound intake

Maternal abnormalities

Key result

Abnormalities:

no effects observed

Results (fetuses)

Fetal body weight changes:

effects observed, treatment-related

Description (incidence and severity):

At 450 mg/kg bw/day, there were statistically significantly decreased fetal body weights both on litter and individual basis and this was related to the treatment with the test item.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Description (incidence and severity):

No test item-related effects on the sex ratio of the fetuses were noted in any group.

Changes in litter size and weights:

effects observed, treatment-related

Description (incidence and severity):

At 450 mg/kg bw/day, there were statistically significantly decreased fetal body weights both on litter and individual basis and this was related to the treatment with the test item.

Changes in postnatal survival:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Description (incidence and severity):

No test item-related findings were noted during skeletal examination of the fetuses. Non-ossified cervical vertebral bodies (1 - 7), some non-ossified caudal vertebrae, incompletely ossified sternebra 5, non-ossified metatarsalia 1 and non-ossified digits on fore and hind limbs were seen in fetuses at 450 mg/kg bw/day. These findings indicated a slight delay in ossification at 450 mg/kg bw/day which correlated with the lower fetal body weight in this dose group and this was related to the treatment with the test item.
At 450 mg/kg bw/day, statistically significantly increased incidence of supernumerary rudimentary ribs, long ventral plate and long or interrupted costal cartilage occurred both on an individual
and litter basis and these indicated a slight disturbance in development.
These observations were only variations with high spontaneous incidence of this strain.

Visceral malformations:

no effects observed

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

The mean fetal body weight was statistically significantly lower at 450 mg/kg bw/day and was considered to be related to the treatment with the test item. In correlation with the lower fetal weights a slight delay in ossification was noted in the high dose group (450 mg/kg bw/day). An increased incidence of long or interrupted costal cartilages or long ventral plate indicated a slight disturbance in development at this dose level.

Effect levels (fetuses)

Key result

Dose descriptor:

NOEL

Effect level:

ca. 150 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

fetal/pup body weight changes

other:

Remarks on result:

other: Effects seen are considered secondary to maternal toxicity or variations only

Fetal abnormalities

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

skeletal: forelimb

skeletal: rib

skeletal: vertebra

skeletal: hindlimb

Description (incidence and severity):

An increased incidence of long or interrupted costal cartilages or long ventral plate indicated a slight disturbance in development at this dose level. However these observations are variations only, and not reported as malformations.

Overall developmental toxicity

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

450 mg/kg bw/day (actual dose received)

Treatment related:

yes

Relation to maternal toxicity:

developmental effects as a secondary non-specific consequence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

not specified

Any other information on results incl. tables

Table 1: Results for Formulation Analysis

Dose Group	Sample taken from	Nominal Conc (mg/mL)	Actual Conc (mg/mL)	Relative to Nominal (%)	Mean (%)
Control	vehicle	0.0	ND	-	-
2	top middle bottom	10.0 10.0 10.0	9.273 9.830 9.721	92.7 98.3 97.2	96.1
3	top middle bottom	30.0 30.0 30.0	31.61 29.51 29.25	105.4 98.4 97.5	100.4
4	top middle bottom	90.0 90.0 90.0	89.91 90.59 90.59	99.9 100.7 100.7	100.4

ND = not detected

In addition, the test item was found to be stable in application formulations when kept four hours at room temperature and eight days at in the refrigerator (5 ± 3 °C) due to recoveries which met the variation limit of 10% from the time-zero (homogeneity) mean value.

Table 2 - Summary of Performance of Mated Females

Group Dose (mg/kg bw/day)	1 (0)	2 (50)	3 (150)	4 (450)
Female numbers	1 - 22	23 - 44	45 - 66	67 - 88
Number of mated females	22	22	22	22
Non-pregnant females (A)	1	0	1	1
Number of pregnant females	21	22	21	21
Number of females with live fetuses at termination*	21	22	21	21

*    Only dams with at least one live fetus at caesarean section were used for the calculations of food consumption, body weight gain and corrected body weight gain data.

(A) Female no. 10 from group 1 (control), female no. 57 from group 3 and female no. 79 from group 4 were not pregnant.

Applicant's summary and conclusion

Conclusions:

In the pre-natal developmental toxicity study conducted according to OECD Test Guideline 414 and in compliance with GLP, administration of trimethylsilanol (CAS 1066-40-6) by oral gavage to Sprague-Dawley rats from Day 6 to 20 of pregnancy at 0, 50, 150 or 450 mg/kg/day resulted in clinical signs and reduced food consumption and body weight gain in dams at 150 and 450 mg/kg bw/day. The effects on food consumption and body weight were considered to be adverse at 450 mg/kg/day, therefore the maternal NOAEL for general toxicity was considered to be 150 mg/kg/day. There was no maternal developmental effects observed but reduced foetal weight, delayed ossification and increased incidence of some cartilaginous variations were noted in foetuses at 450 mg/kg/day. The skeletal findings are reported to be variations only and are not considered to be adverse effects as such. The reduced foetal weight is considered secondary to maternal toxicity. Therefore the developmental NOEL is considered to be 150 mg/kg bw/day based on findings observed in the pups.