Dodecamethylcyclohexasiloxane

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

In the oral combined repeated dose toxicity study with the reproduction/developmental toxicity screening test in rats with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 422 and in compliance with GLP (Dow Corning Corporation, 2005), the NOAEL for reproductive toxicity was concluded to be at least 1000 mg/kg bw/day, the highest dose tested, based on no adverse effects.
A non-statistically significant increase in non-gravid sperm-positive females at the top dose was observed. In comparison to the extended one-generation reproductive toxicity test (OECD Test Guideline 443), which requires groups of 20 pregnant females, this combined screening test requires 10 females. Therefore a larger group size might provide better evidence of the significance of this effect and there is an ongoing extended one-generation reproductive toxicity study (EOGRTS) in accordance with an ECHA final decision No PE-D-2114471588-34-01/F. The draft study report is expected in October 2022 with the final report available in March 2023, therefore the substance will be updated with the study results by the end of June 2023.

A preliminary dose range-finding study to assist the dose level selection for the main extended one-generation reproductive toxicity study has been conducted and the general systemic, reproductive and developmental toxic potential of dodecamethylcyclohexasiloxane (D6; CAS 540-97-6) has been assessed (Covance Laboratories Limited, 2020). Based on the results obtained, it was concluded that oral gavage administration of D6 to Sprague-Dawley rats at dose levels up to 1000 mg/kg bw/day was well-tolerated and there were no adverse effects observed on any of the parameters assessed in the P0 or F1 generation.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2005-04-26 to 2006-10-24

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Labs, Raleigh, NC, USA
- Age at study initiation: (P) 9 wks
- Weight at study initiation: (P) Males: 285-386 g; Females: 173-254 g
- Housing: 1/seuspended wire mesh cage
- Diet: standard diet ad libitum
- Water: drinking water ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 67.7-72.5 deg F (approximately 20 degrees celsius)
- Humidity (%): 35-59
- Air changes (per hr): 15.6
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: From 2005-05-12 for 28 days (m) and 46 days (f) in the reproductive portion of this study.

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

Prepared for the whole study. Analysed (GC) 4 times to ensure homogeneity, stability and concentration.

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation: 2 wk
- Proof of pregnancy: vaginal plug or sperm day 0 of pregnancy

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

GC analysis on 4 occasions to ensure homogeneity, stability and concentration.

Duration of treatment / exposure:

males: 28 days (including 14 days prior to mating)
females: 46 days (from 14 days prior to mating to postnatal day 3)

Frequency of treatment:

daily. 7 days/wk

Details on study schedule:

F1 not mated.

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
- Rationale for animal assignment (if not random): weight stratified randomization

Positive control:

None

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily. From day 20 after mating females were checked 3 times daily on weekdays for evidence of parturition.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

OTHER [reported fully under Repeated Dose]

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at sacrifice
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: yes
- How many animals: all toxicity groups

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at sacrifice
- Animals fasted: Yes / No / No data
- How many animals: all toxicity groups

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: FOB at start and completion
- Dose groups that were examined: males and toxicity group females
- Battery of functions tested: cage-side observations, hand-held observations, open field observations, categorical observations, hind and forelimb grip strength, landing foot splay, motor activity.

GROSS PATHOLOGY: Yes
all toxicity groups

HISTOPATHOLOGY: Yes
Microscopic examination of a relatively large range of tissues of all control and hi-dose group animals, and the lungs and thyroid of both sexes and liver of females at the low and mid doses. Examinations included testes, uterus, epididymides, ovaries and prostate.

ORGAN WEIGHTS
weights of a relatively large range of organs determined (including testes, uterus, epididymides, ovaries, prostate)

Oestrous cyclicity (parental animals):

No data

Sperm parameters (parental animals):

Parameters examined in males of the P generation: testis weight, epididymis weight, prostate weight.

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross abnormalities, weight gain (PND0-PND4)

GROSS EXAMINATION OF DEAD PUPS:
yes, for external abnormalities; possible cause of death was not determined for pups born or found dead

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: after 2 wk mating
- Maternal animals: PND4

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera of all adults. For pregnant females the number of corpora lutea and implantation sites were recorded

HISTOPATHOLOGY / ORGAN WEIGHTS
Microscopic examination of a relatively large range of tissues of all control and hi-dose group animals, and the lungs and thyroid of both sexes and liver of females at the low and mid doses. Organ weights of a relatively large range of organs determined . The ovaries, uterus, testes, epididymides and prostate were included in organ weight determination and microscopic examinations.

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring were subjected to postmortem examinations (macroscopic) for external gross abnormalities

Statistics:

ANOVA (analysis of variance): body weights, organ weight, litter size, haematology, clinical chemistry etc
ANCOVA (analysis of covariance): FOB, reproductive prameters
Cochran-Armitage: microscopic findings

Reproductive indices:

mean gestation length
mean litter size
mean live litter size
mean litter weight
mean ratio of live births/litter size
mean implantation sites
mean corpora lutea
mean mating and fertility indices

Offspring viability indices:

vaible pups
average pup weight

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

No effect on reproductive organs. [For effects on other organs see report of this study under repeated dose toxicity.]

Histopathological findings: neoplastic:

not specified

Other effects:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: No clear effects on reproductive toxicity in screening study

Critical effects observed:

no

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not examined

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not examined

Anogenital distance (AGD):

not specified

Nipple retention in male pups:

not specified

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

Other effects:

no effects observed

Description (incidence and severity):

Gross pathology of the offspring - No treatment-related effects.

Behaviour (functional findings):

not specified

Developmental immunotoxicity:

not specified

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed.

Critical effects observed:

no

Reproductive effects observed:

no

Conclusions:

A well reported oral combined repeated dose/reproductive and developmental toxicity study in the rat, conducted according to the current guideline and in accordance with GLP, identified an NOAEL of at least 1000 mg/kg bw/day for reproductive effects. This screening study found that the increased number of non-gravid females at this dose was not statistically significant. There was no evidence of this effect at the lower dose of 330 mg/kg bw/day.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

A well reported oral combined repeated dose/reproductive and developmental toxicity study in rats with dodecamethylcyclohexasiloxane (D6), conducted according to the current OECD Test Guideline 422 and in compliance with GLP, identified a NOAEL of at least 1000 mg/kg bw/day for reproductive effects, based on no adverse effects (Dow Corning Corporation, 2005). This screening study found that the increased number of non-gravid females at this dose was not statistically significant. There was no evidence of this effect at the lower dose of 330 mg/kg bw/day.

In a preliminary dose range-finding study to assist the dose level selection in the planned extended one-generation reproductive toxicity study, the general systemic, reproductive and developmental toxic potential of dodecamethylcyclohexasiloxane (D6; CAS 540-97-6) has been assessed (Covance Laboratories Limited, 2020).

In the study, the P0 generation, consisting of three groups of ten male and ten female rats received neat D6 at dose levels of 300, 600 or 1000 mg/kg bw/day. Males were treated for two weeks before pairing, up to necropsy after litters were weaned. Females were treated for two weeks before pairing, throughout pairing up to necropsy on Day 21 of lactation. In the F1 generation, ten males and ten females were treated from weaning to Day 34 of age at the same dose levels and volume dose as the P0 generation. A similarly constituted control group received purified water at the same volume dose as the highest dose group.

During the study, for the P0 generation, clinical condition, body weight, food consumption, pre coital interval, mating performance, fertility, gestation length, organ weights and macroscopic pathology investigations were undertaken. For the F1 generation, clinical condition, body weight, food consumption, organ weights and macroscopic pathology investigations were undertaken.

The clinical condition, litter size and survival, sex ratio, body weight, organ weights and macropathology for all offspring were also assessed.

Treatment with D6 at dose levels up to 1000 mg/kg bw/day was well-tolerated. There were no treatment-related premature deaths. Amongst the P0 generation and their offspring there were no signs at physical examination related to treatment and no signs associated with dose administration.  There were no treatment related macroscopic findings at scheduled termination of the P0 generation and no clear effect of treatment on liver or kidney weights.  There were no treatment-related macroscopic findings amongst their unselected offspring or at scheduled termination of the selected F1 generation.  There was no adverse effect of treatment on body weight gain or food intake in the P0 generation and no effect of treatment on reproductive performance, as assessed by pre-coital interval, mating performance and fertility, gestation length and index, litter size, survival indices and offspring sex ratio.

On Day 1 of age the group mean body weight of male and female offspring derived from all groups of treated parents was slightly lower than control with statistical significance attained at 1000 mg/kg bw/day for males and females.  Overall group mean body weight gain from Day 1 to 21 of age of male and female offspring derived from all groups of treated parents was marginally lower than control but with no dose-response apparent.  Group mean terminal body weight for male unselected offspring at scheduled termination on Day 21 of age was statistically significantly lower than control at 1000 and 600 mg/kg bw/day.  Absolute and body weight relative group mean liver weights for unselected offspring derived from all treated groups were slightly lower than control with statistical significance attained at 1000 mg/kg bw/day for male absolute weights.  The extent of these changes was not considered to be adverse.

Group mean body weight gain of all groups of selected F1 males was slightly lower than control, with the greatest effect and statistical significance attained on occasion at 1000 mg/kg bw/day.  Following the commencement of treatment on Day 21 of age food intake for all groups of treated F1 animals was slightly lower than control with the greatest effect generally observed at 1000 mg/kg bw/day. When compared to control, absolute and body weight adjusted liver weights for all groups of treated selected F1 females and body weight adjusted liver weights for selected F1 males at 1000 mg/kg bw/day were slightly high with statistical significance attained at 1000 mg/kg bw/day for male and female body weight adjusted liver weights. The extent of these changes was not considered to be adverse.

Based on the results obtained in this preliminary study of reproductive performance, it was concluded that oral gavage administration of D6 to Sprague-Dawley rats at dose levels up to 1000 mg/kg bw/day may be suitable for investigation in the main extended one-generation reproductive toxicity study because there were no adverse effects of treatment on any of the parameters assessed in the P0 or F1 generation.

Effects on developmental toxicity

Description of key information

In the key developmental toxicity study with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 414 and in compliance with GLP, (Charles River, 2017), no maternal or developmental toxicity was observed in rats at doses up to 1000 mg/kg bw/day. The maternal and developmental NOAEL for D6 were concluded to be at least 1000 mg/kg bw/day based on no adverse effects. 

In the key developmental toxicity study with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 414 and in compliance with GLP (The Dow Chemical Company, 2018), no maternal or developmental toxicity was observed in rabbits at doses up to 1000 mg/kg bw/day. The maternal and developmental NOAELs for D6 were concluded to be at least 1000 mg/kg bw/day based on no adverse effects.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

January to March 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.31 (Prenatal Developmental Toxicity Study)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

yes

Species:

rat

Strain:

other: RccHan:WIST

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories B.V., The Netherlands
- Age at study initiation: approximately 10 to 14 weeks
- Weight at study initiation: not stated
- Fasting period before study: no
- Housing: individually in Macrolon plastic cages (MIII type)
- Diet (ad libitum): SM R/M-Z (SSNIFF Spezialdiaten GmbH, Germany)
- Water (ad libitum): tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24
- Humidity (%): 40 to 70
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 16 February 2015 To: 12 March 2015

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: dosed undiluted

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

No chemical analysis was performed because the test substance was dosed undiluted.(i.e. without the use of a vehicle).

Details on mating procedure:

- Impregnation procedure: mated at supplier
- M/F ratio per cage: not stated
- Length of cohabitation: not stated
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy

Duration of treatment / exposure:

Day 6 to Day 20 post-cotum, inclusive

Frequency of treatment:

Daily

Duration of test:

Day 0 to Day 21 post-coitum

Dose / conc.:

0 mg/kg bw/day

Remarks:

Group 1

Dose / conc.:

100 mg/kg bw/day

Remarks:

Group 2

Dose / conc.:

330 mg/kg bw/day

Remarks:

Group 3

Dose / conc.:

1 000 mg/kg bw/day

Remarks:

Group 4

No. of animals per sex per dose:

22 Females

Control animals:

other: yes, water

Details on study design:

- Dose selection rationale: Dose levels were selected based on results of the dose range finding study (Project 507522) in which no remarkable findings were noted at doses of 0, 100, 330 or 1000 mg/kg/day.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: Days 1, 3, 6, 9, 12, 15, 18 and 21 post-coitum

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Days 1, 3, 6, 9, 12, 15, 18 and 21 post-coitum
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: Yes
- Time schedule for examinations: Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatment related effect was suspected.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: macroscopic examination, liver weighed and retained

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
- Other: In case implantations were not macroscopically visible, the uterus was stained using the Salewski technique

Fetal examinations:

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

Statistics:

The following statistical methods were used to analyze the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test based on a pooled variance estimate was applied for the comparison of the treated groups and the control group. The mean of fetal weights per litter and mean of total litter weight was also analyzed using this method.
- The Steel-test was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Mann Whitney test was used to compare mean litter proportions (percent of litter) of the number of viable and dead fetuses, early and late resorptions, total resorptions, pre- and postimplantation loss, and sex distribution.
- Mean litter proportions (percent per litter) of total fetal malformations and developmental variations (external, visceral and skeletal), and each particular external, visceral and skeletal malformation or variation were subjected to the Kruskal-Wallis nonparametric ANOVA test to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunn’s test was used to compare the compound-treated groups to the control group.

All tests were two-sided and in all cases p<0.05 was accepted as the lowest level of significance. Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances.

No statistics were applied for data on maternal survival, pregnancy status, group mean numbers of dead fetuses, early and late resorptions, and pre- and post-implantation loss.

Indices:

Pre-implantation loss (%) = ((number of corpora lutea - number of implantation sites) / number of corpora lutea) x 100

Post-implantation loss (%) = ((number of implantation sites - number of live fetuses) / number of implantation sites) x 100

The fetal developmental findings were summarized by: 1) presenting the incidence of a given finding both as the number of fetuses and the number of litters available for examination in the group; and 2) considering the litter as the basic unit for comparison, calculating the number of affected fetuses as a mean litter proportion on a total group basis, where:

Viable fetuses affected/litter (%) = (number of viable fetuses affected per litter / number of viable fetuses per litter) x 100

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

not examined

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

At 1000 mg/kg/day bw, maternal animals had higher absolute and relative liver weights. The difference from controls was slight but statistically significant. There were no test item-related microscopic findings observed during the histopathological evaluation therefore, the increases in liver weights were not considered toxicologically relevant.

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Number of abortions:

not specified

Pre- and post-implantation loss:

not specified

Total litter losses by resorption:

not specified

Early or late resorptions:

not specified

Dead fetuses:

not specified

Changes in pregnancy duration:

not specified

Changes in number of pregnant:

not specified

Other effects:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

not specified

Changes in litter size and weights:

no effects observed

Changes in postnatal survival:

not specified

External malformations:

no effects observed

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

Based on the initial description of findings (severely malaligned sternebrae, fused sternebrae, vertebral centra anomaly and bent limb bones), an external peer review was performed which concluded that there were no toxicologically relevant effects on skeletal morphology following treatment up to 1000 mg/kg bw/day.

Skeletal malformations only occurred in two fetuses only. At 1000 mg/kg bw/day, 1 fetus had a short rib and since this occurred once only and was seen previously in historical control fetuses, it was considered to not be test substance related. The other malformation was noted in a control fetus with polydactyly (not detected externally) and as such was not considered related to treatment.

Skeletal variations occurred for 33.5%, 31.6%, 38.7% and 50.4% of the fetuses per litter in the control, 100, 330 and 1000 mg/kg bw/day groups, respectively.
Animals at 1000 mg/kg bw/day had a significantly higher percent of combined skeletal variations per litter than controls. None of the individual findings was significantly higher for animals at 1000 mg/kg bw/day though findings like (slight or moderate) malaligned sternebra(e), 14th rudimentary rib(s), caudal shift of the pelvic girdle, unossified metacarpal(s) and/or metatarsal(s), and sternebra(e) #1, 2, 3, 4, 5 and/or #6 unossified were seen at slightly higher incidences than controls. Taken together, these culminated in a statistically significant increase in skeletal variations overall, but was not regarded toxicologically relevant.

Fetuses at 1000 mg/kg bw/day (and at 330 mg/kg bw/day) had a higher incidence of slightly to moderately malaligned sternebra(e) (12.5% and 10.7%, respectively versus control value 8.4% per litter). Besides, three fetuses at the high dose level showed severe malaligned sternebra(e) (0.0% for controls versus 1.2% at 1000 mg/kg bw/d). However, the difference of both severities of malaligned sternebrae from controls was not statistically significant and only slightly higher than controls. Moreover, values of slightly to moderately malaligned sternebra(e) remained far below the historical control upper limit (21.3% per litter). As such, this was not considered to be adverse.

There were slightly higher incidences of unossified metacarpal(s) and metatarsal(s) seen for fetuses at 330 and 1000 mg/kg bw/day (6.4% and 8.3% per litter, respectively, versus control value 2.7% per litter). These were not considered to be toxicologically relevant as there were no other indications of developmental delay seen like decreases in other related ossification parameters and fetal body weights remained in the same range as controls.

Visceral malformations:

no effects observed

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed.

Abnormalities:

no effects observed

Developmental effects observed:

no

Table 1 - Summary of Maternal Survival and Pregnancy Status

Dose (mg/kg/day)	0	100	330	1000
	Number (%)	Number (%)	Number (%)	Number (%)
Females on study	22	22	22	22
Females that aborted or delivered	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)
Females that died females that aborted - nongravid - gravid	0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)
Females that were euthanized - nongravid - gravid	0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0)	0 (0.0) 0 (0.0) 0 (0.0)
Females examined at scheduled - necropsy - nongravid - gravid - with resorptions only - with viable fetuses	22 (100.0) 0 (0.0) 22 (100.0) 1 (4.5) 21 (95.5)	22 (100.0) 0 (0.0) 22 (100.0) 0 (0.0) 22 (100.0)	22 (100.0) 0 (0.0) 22 (100.0) 0 (0.0) 22 (100.0)	22 (100.0) 0 (0.0) 22 (100.0) 1 (4.5) 21 (95.5)
Total females gravid	22 (100.0)	22 (100.0)	22 (100.0)	22 (100.0)

Table 2 - Summary of Fetal Data at Scheduled Necropsy

Dose (mg/kg/day)		Sex		Viable fetuses	Dead fetuses	Resorptions		Post implantation loss	Implantation sites	Corpora lutea	Pre implantation loss	Fetal weight  (g)	No. of gravid females
		M	F			early	late
0	Total mean S.D.	122 5.5 2.22	107 4.9 2.08	229 10.4 3.36	0 0.0 0.00	16 0.7 0.77	0 0.0 0.00	16 0.7 0.77	245 11.1 3.20	268 12.2 1.99	23 1.0 1.59	NA 5.3 0.25	22
100	Total mean S.D.	114 5.2 2.46	124 5.6 1.84	238 10.8 2.86	0 0.0 0.00	10 0.5 0.86	0 0.0 0.00	10 0.5 0.86	248 11.3 2.73	271 12.3 2.46	23 1.0 1.33	NA 5.3 0.29	22
330	Total mean S.D.	120 5.5 1.84	123 5.6 1.71	243 11.0 2.26	0 0.0 0.00	15 0.7 0.99	0 0.0 0.00	15 0.7 0.99	258 11.7 2.05	275 12.5 1.63	17 0.8 1.31	NA 5.3 0.35	22
1000	Total mean S.D.	125 5.7 2.97	117 5.3 2.08	242 11.0 3.59	0 0.0 0.00	13 0.6 0.80	0 0.0 0.00	13 0.6 0.80	255 11.6 3.11	279 12.7 2.63	24 1.1 1.87	NA 5.3 0.41	22

Table 3 - Summary of Fetuses amd Litters Data

	Fetuses				Litters
Dose (mg/kg/day)	0	100	330	1000	0	100	330	1000
Number examined externally Number with findings	229 0	238 0	243 0	242 0	21 0	22 0	22 0	21 0
Number examined viscerally Number with findings	113 0	119 0	120 0	121 0	21 0	22 0	22 0	21 0
Number examined skeletally - polydactyly - rib anomaly - sternebra, malaligned (slight/moderate) - sternebra, malaligned (severe) - ossification , reduced skull - vertebral centra, unossified - bent limb bones	229 1 0 18 0 0 0 0	238 0 0 14 1 1 0 0	243 0 0 27 0 1 0 1	242 0 1 30 3 0 1 1	21 1 0 12 0 0 0 0	22 0 0 11 1 1 0 0	22 0 0 15 0 1 0 1	21 0 1 14 3 0 1 1
Total number with malformations - external - soft tissue - skeletal	0 0 1	0 0 0	0 0 0	0 0 1	0 0 1	0 0 0	0 0 0	0 0 1

Conclusions:

In the prenatal developmental toxicity study with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 414 and in compliance with GLP, no maternal or developmental toxicity was observed in rats at doses up to 1000 mg/kg bw/day. The maternal and developmental NOAEL for D6 were concluded to be at least 1000 mg/kg bw/day based on no adverse effects.

Executive summary:

A prenatal developmental toxicity study (OECD 414) of Dodecamethylcyclohexasiloxane (D6) was carried out in rats by oral gavage. Eighty-eight mated female Wistar Han rats were assigned to four dose groups. The test item was administered undiluted once daily by oral gavage from Days 6 to 20 post-coitum at doses of 0, 100, 330 and 1000 mg/kg bw/day (Groups 2, 3 and 4 respectively). The rats of the control group received water at the same dose volume as Group 4 animals. Females were checked daily for the presence of clinical signs. Food consumption and body weight were determined at periodic intervals.

On Day 21 post-coitum, all animals were subjected to an examination post-mortem and external, thoracic and abdominal macroscopic findings were recorded. Terminal body and liver weights were recorded for each dam and body weights gains were calculated. A laparohysterectomy was performed on each dam of all groups. The uteri, placentae and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Gravid uterine weights were recorded, and corrected body weights (changes) were calculated. The fetuses were weighed, sexed and examined for external, visceral and skeletal malformations and developmental variations. All live fetuses were euthanized. One half of the fetuses were decapitated and the heads were fixed in Bouin’s fixative, these fetuses were dissected and examined for visceral anomalies. All fetuses were fixed in 96% aqueous ethanol and stained with Alizarin Red S and had skeletal examinations performed. Histopathology was performed on the livers from all dams of Groups 1 and 4.

RESULTS

At 1000 mg/kg bw/day, maternal animals had higher absolute and relative liver weights. The difference from controls was slight but statistically significant. There were no test item-related microscopic findings observed during the histopathological evaluation, therefore, the increases in liver weights were not considered toxicologically relevant.

No maternal toxicity was observed in the other parameters examined in this study with treatment up to 1000 mg/kg bw/day (mortality, clinical signs, body weights, food consumption and macroscopic examination). No developmental toxicity was observed up to 1000 mg/kg bw/day groups.

CONCLUSION

The maternal and developmental NOAEL for D6 were established as being at least 1000 mg/kg bw/day.

Reference 2

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 October 2017 - 1 May 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

2001

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Version / remarks:

August, 1998

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.31 (Prenatal Developmental Toxicity Study)

Version / remarks:

May, 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF: The Japanese Ministry of Agriculture, Forestry and Fisheries Notification of 12 NohSan-8147, Guideline 2-1-18, Teratogenicity study

Version / remarks:

November, 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Species and Sex: Rabbits, time-mated females
Strain: NZW rabbits
Supplier: Covance Research Products, Inc. (CRP), Greenfield, Indiana, USA
Age and Weight at Study Start: Sexually mature adults weighing 2800-3200 g.

Health Status and Acclimation:
Upon arrival all animals were acclimated to the laboratory for approximately 5 days prior to the start of test material administration. Upon arrival and once during the acclimation period, each animal was evaluated by trained veterinarian to determine the general health status and acceptability for study purposes.

Housing:
Upon arrival and after assignment, animals were housed one per cage in stainless steel and plastic cages. Cages had perforated plastic floors and were suspended above catch pans with absorbent non-contact bedding. Cages contained a J-type stainless steel feeder and a pressure activated lixit valve-type stainless steel watering system.

The following environmental conditions were targeted in the animal room from the day of arrival until necropsy:

Temperature: 20°C with a range of 16°C-22°C (targeted) 19.9°C with a range of 19°C-21°C (actual)
Humidity: 50% with a range of 40-70% (targeted) 50.3% with a range of 49.2-57.7% (actual)
Air Changes: 10-15 times/hour ( targeted average) 11.6 times/hour (actual average)
Photoperiod: 12-hour light/dark (on at 6:00 a.m. and off at 6:00 p.m.)

Enrichment:
Enrichment for animals was given from the day of arrival until necropsy. The enrichment included an elevated resting platform, a variety of stainless steel objects attached inside the cage, and a cardboard tray for manipulation.

Feed and Water:
A stepwise increase in amount of daily feed allotted was implemented to aid in avoiding gastrointestinal disturbances during the acclimation period. Upon receipt, rabbits received approximately 62.5 g of LabDiet 5325 (PMI Nutrition International, Richmond, Indiana) in pelleted form. The amount of feed was increased up to the full daily amount of approximately 125 g the following day. Analyses of the feed were performed by PMI Nutrition International to confirm the diet provided adequate nutrition and to quantify the levels of selected contaminants. Drinking water obtained from the municipal water source was periodically analyzed for chemical parameters and biological contaminants by the municipal water department. In addition, specific analyses for chemical contaminants were conducted at periodic intervals by an independent testing facility.

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

Test material was administered daily by oral gavage from GD 7-27.

Analytical verification of doses or concentrations:

no

Remarks:

No analyses were performed because the test material was administered neat (undiluted). Stability was not applicable to this study due to utilizing neat test material.

Details on mating procedure:

Sexually mature virgin females were naturally mated with one buck of the same strain at the supplier. The observed day of breeding was considered GD 0. GD 0 body weights and records of mating pairs were provided by the supplier and maintained in the study record. Rabbits arrived in the laboratory on GD 1 or 2.

Duration of treatment / exposure:

Rabbits were administered D6 on GD 7-27

Frequency of treatment:

Test material was administered daily by oral gavage from GD 7-27

Duration of test:

Test material administration began on November 13, 2017 and the last group of rabbits was necropsied on December 13, 2017.

Dose / conc.:

0 mg/kg bw/day

Remarks:

Dose Volume: 1.0204 ml/kg

Dose / conc.:

100 mg/kg bw/day

Remarks:

Dose Volume: 0.1020 ml/kg

Dose / conc.:

300 mg/kg bw/day

Remarks:

Dose Volume: 0.3061 ml/kg

Dose / conc.:

1 000 mg/kg bw/day

Remarks:

Dose Volume: 1.0204 ml/kg

No. of animals per sex per dose:

24 number of rabbits/dose level

Control animals:

yes, concurrent vehicle

Details on study design:

Groups of 24 time-mated female NZW rabbits were administered D6 via gavage at dose levels of 0, 100, 300, or 1000 mg/kg/day on GD 7-27. The rabbits were ordered and mated in six different replicates from the supplier to stagger caesarean sections over a period of two weeks.

Dose Levels and Justification
Dose levels for this study (0, 100, 300, or 1000 mg/kg/day on GD 7-27) were selected on the basis of the developmental toxicity probe study discussed previously (Johnson et al., 2018). The high-dose of 1000 mg/kg/day represented a limit dose as defined in the test guideline. The lower dose levels were selected to provide dose response data for any toxicity that may have been observed among the high-dose group rabbits.

Dose Preparation
The test material was administered neat (undiluted). Dose volumes were calculated using the most current body weight. Control animals were gavaged with 1.0204 ml of water/kg body weight, which was the volume equal to the largest gavage volume given treated animals.

Maternal examinations:

Daily Observations:
A cage-side examination was conducted twice daily, approximately at the same time each day. In addition, all animals were observed for morbidity, mortality, and the availability of feed and water at least twice daily. For animals showing indications of premature delivery, the delivered fetuses were counted and examined to the extent possible.

Clinical Observations
Clinical observations were conducted on all animals at least once daily. Animals were observed approximately one hour after dosing. Clinical observations included a careful, hand-held examination of the animal with an evaluation of abnormalities in the eyes, urine, feces, gastrointestinal tract, extremities, movement, posture, reproductive system, respiration, skin/hair-coat, and mucous membranes, as well as an assessment of general behavior, injuries or palpable mass/swellings.

Body Weights/Body Weight Gains
Body weights were recorded on GD 0 by the supplier, GD 4, daily during test material administration, and on GD 28.

Feed Consumption
Daily feed consumption was recorded and statistically analyzed for all animals from GD 4-28.

Ovaries and uterine content:

On GD 28, all surviving females (not fasted) were euthanized via intravenous injection of Beuthanasia-D (Henry Schein Animal Health, Dublin Ohio), and a limited gross pathological examination (necropsy) was performed. The sequence of the maternal necropsies was counterbalanced across groups (e.g., control, high, middle, low) to control for potential confounding influences of timing on fetal growth and skeletal ossification.
A detailed examination of the reproductive tract was performed and the number and position of implantations, viable fetuses, dead fetuses and resorptions were recorded. Resorptions were classified as either “early” or “late” based on the presence (late resorption) or absence (early resorption) of grossly recognizable embryonic/fetal form, while a “dead fetus” indicated a very recent death as evidenced by a lack of external degenerative changes. For females with one or more viable fetuses, the number of ovarian corpora lutea was counted. The uteri of females lacking visible implantations were stained with a 10% aqueous solution of sodium sulfide (based on Kopf et al., 1964) and examined for evidence of early resorptions in order to verify pregnancy status.

Fetal examinations:

The maternal necropsy included an examination of the external tissues and all orifices. The skin was reflected from the carcass, the thoracic and abdominal cavities were opened and the viscera were examined. The stomach, liver (with gallbladder) and kidneys were dissected from the carcass and were incised. Any obvious gross pathologic alterations were recorded, and the weight of the liver (with incised gallbladder), kidneys and gravid uterus were recorded. The ratios of liver and kidney weights to terminal body weight were calculated. Representative sections of liver, kidneys, and gross lesions were preserved in neutral, phosphate-buffered 10% formalin. Microscopic examination of tissues was not conducted. Transponders were removed and placed in bags with the tissues.

All fetuses were weighed, and given an external examination that included observations on body proportions, the head and face (including closure of the palate), abdomen, spine, extremities, genitalia, rectum and tail. All viable fetuses were then given a secondary dose of euthanasia solution via sublingual oral administration of Fatal Plus (Schering Corporation, Kenilworth, New Jersey). All fetuses were given a visceral examination conducted by dissection under a low power stereomicroscope for evidence of visceral alterations (Staples, 1974; Stuckhardt and Poppe, 1984). The visceral examination included observations of the thymus, trachea, esophagus, lungs, great vessels, heart (external and internal), liver, gastrointestinal tract, pancreas, spleen, kidneys (sectioned), adrenal glands, ureters, bladder, and reproductive organs. The fetuses were sexed by examination of the gonads. Approximately one half of the fetuses in each litter were randomly selected for craniofacial examination. The heads of these fetuses were removed, placed in Bouin’s fixative and serially sectioned to allow for inspection of the eyes, brain, nasal passages and tongue (Wilson, 1965). All fetuses were preserved in alcohol, eviscerated and stained with Alizarin Red S in order to visualize ossified bone (Dawson, 1926). After staining, skeletons were cleared and a thorough evaluation of the fetal skeleton was conducted.

Statistics:

The litter, rather than the pup, was considered as the experimental unit. Maternal body weights, maternal body weight gains, organ weights (absolute and relative with the exception of only absolute weight for gravid uterus), fetal body weights and feed consumption were evaluated by Bartlett’s test (alpha = 0.01; Winer, 1971) for homogeneity of variance. Based on the outcome of Bartlett's test, a parametric (Steel and Torrie, 1960) or nonparametric (Hollander and Wolfe, 1973) analysis of variance (ANOVA) was performed. If the ANOVA was significant at alpha = 0.05, analysis by Dunnett's test (alpha = 0.05; Winer, 1971) or the Wilcoxon Rank-Sum test (alpha = 0.05; Hollander and Wolfe, 1973) with Bonferroni's correction (Miller, 1966) was performed, respectively.
Feed consumption values were excluded from analysis if the feed was spilled or scratched.
Statistical analyses were conducted on average percent litter response for pre- and post-implantation loss and fetal alterations. Frequency of pre- and post-implantation loss (calculations shown below), and fetal alterations (if any) were analyzed using a censored Wilcoxon test (Haseman and Hoel, 1974) with Bonferroni’s correction applied when the incidence was greater than 5%.
The number of corpora lutea, implantations, and litter size were evaluated using a nonparametric ANOVA (alpha = 0.05) followed by the Wilcoxon Rank-Sum test (alpha = 0.05) with Bonferroni's correction.
Pregnancy rates were analyzed using the Fisher exact probability test (alpha = 0.05; Siegel, 1956) with Bonferroni’s correction.
Fetal sex ratios were analyzed using a binomial distribution test.
Non-pregnant females were excluded from the appropriate analyses.
Statistical outliers (alpha = 0.02) were identified by the sequential method of Grubbs (1969).
Both Dunnett’s test and Bonferroni’s correction corrected for multiple comparisons to the control and were reported at the corrected alpha level.

Clinical signs:

no effects observed

Description (incidence and severity):

Examinations performed on all animals revealed no treatment-related findings.
One animal in the 300 mg/kg/day group delivered early and had pups present in her cage on the morning of GD 28 which was considered unrelated to treatment due to the isolated occurrence. Normal necropsy and fetal exams were performed for this animal to the extent possible except a gravid uterine weight was not recorded. Another animal in the 300 mg/kg/day group was removed from the study and euthanized on GD 26 due to a mechanical injury sustained from a gavage incident.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

There were no treatment-related differences in the amount of feed consumed by any treated groups when compared to their respective controls. Statistically-identified differences were considered spurious and unrelated to treatment due the minimal difference from control (≤4.2%).

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

no effects observed

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

Percent post-implantation loss in the 1000 mg/kg/day groups was slightly higher than controls and reached statistical significance. This increase in post-implantation loss was deemed spurious and unrelated to treatment as:
1) postimplantation loss in the 1000 mg/kg/day group was similar to recent historical controls,
2) postimplantation loss in the control group was lower than recent historical control
3) the postimplantation loss was represented by litters with single resorptions.

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

Description (incidence and severity):

no effects observed

Changes in number of pregnant:

not specified

Other effects:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: No effects observed

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

not specified

Changes in postnatal survival:

not specified

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Description (incidence and severity):

There were no treatment-related skeletal alterations in any dose group. The incidence of one skeletal variation (DO sternebrae) was higher in the 300 and 1000 mg/kg/day dose groups compared to controls. This finding was deemed to be unrelated to treatment as it was within (litter) or similar (fetuses) the range of recent historical control values. Furthermore, this finding is considered to be a non-adverse variation as it is among the most common skeletal variations encountered in this type of guideline developmental toxicity study (Carney and Kimmel, 2007). In both rodents and rabbits, the sternebrae is an area that ossifies rapidly during late gestation (Carney and Kimmel, 2007; Fritz, 1975). Therefore, variable ossification of these structures is normal when necropsy occurs on GD 28. In addition, high background incidence is common due to the laboratory scoring criteria which dictates that the call of delayed ossification be made when >50% of sternebrae numbers 1,2,3,4 and 6 are unossified or sternebra 5 is completely unossified. Delays in ossification are not expected to persist in the postnatal animal due to high remodeling capacity of the skeleton (Carney and Kimmel, 2007; Holmbeck and Szabova, 2006).
Incidental findings bearing no relationship to treatment included the malformations misaligned thoracic centra, forked ribs, and thoracic hemivertebra and the variations delayed ossification (DO) hyoid, crooked hyoid, DO dentoid, DO sternebrae, fused sternebrae, irregular pattern of ossification sternebrae, DO thoracic centra, and DO pubis. Given that these observations occurred in the control group, at low frequencies, and/or lacked a dose response, these observations were considered spurious and unrelated to treatment.

Visceral malformations:

no effects observed

Description (incidence and severity):

There were no treatment-related or statistically-identified visceral alterations in any dose group. Incidental findings bearing no relationship to treatment included the malformations ventricular wall defect and missing gall bladder, and the variations missing caudal lung lobe, hemorrhage thymus, supernumerary hepatic liver lobule, hypoplastic spleen, particulate material kidney, bifurcated renal vein, retrocaval ureter, and paraovarian cyst. Given that these observations occurred in the control group, at low frequencies, and/or lacked a dose response, these observations were considered spurious and unrelated to treatment.

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

No indication of embryo/fetal toxicity or teratogenicity.

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related effects. The incidence of one skeletal variation (DO sternebrae) at 300 and 1000 mg/kg/day dose groups was deemed to be unrelated to treatment.

Remarks on result:

other: No treatment related effects

Abnormalities:

no effects observed

Key result

Developmental effects observed:

no

Conclusions:

In the prenatal developmental toxicity study with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 414 and in compliance with GLP, no maternal or developmental toxicity was observed in rabbits at doses up to 1000 mg/kg bw/day. The maternal and developmental NOAELs for D6 were concluded to be at least 1000 mg/kg bw/day based on no adverse effects.

Executive summary:

The maternal and developmental toxicity of D6 in NZW rabbits following repeated oral gavage administration was evaluated. Groups of 24 time-mated female rabbits were administered 0, 100, 300, or 1000 mg/kg/day on gestation day (GD) 7-27. No analyses of concentration verification, homogeneity, or stability were conducted because the test material was administered neat (undiluted). In-life parameters evaluated for all groups included: clinical observations, body weight, body weight gain, and feed consumption. On GD 28 all surviving rabbits were euthanized and examined for gross pathologic alterations. Liver, kidneys, and gravid uterine weights were recorded, along with the number of corpora lutea, uterine implantations, resorptions, and live/dead fetuses. All fetuses were weighed, sexed, and examined for external and visceral alterations. The heads were examined for craniofacial alterations by serial sectioning in approximately one half of the fetuses in each litter, and skeletal examinations were performed on all fetuses.

Gavage administration of D6, up to and including the limit dose of 1000 mg/kg bw/day, resulted in no treatment-related maternal toxicity and no indication of embryo/fetal toxicity or teratogenicity. Therefore, under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for maternal toxicity and developmental toxicity was at least the limit dose of 1000 mg/kg bw/day.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

other: rat and rabbit

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

In the key developmental toxicity study in rats with dodecamethylcyclohexasiloxane (D6) (Charles River, 2017), conducted according to OECD Test Guideline 414 and in compliance with GLP, D6 was administered to pregnant female Wistar Han rats at doses of 0, 100, 330 or 1000 mg/kg/day by oral gavage daily from Days 6 to 20 of pregnancy. At 1000 mg/kg bw/day maternal animals had higher absolute and relative liver weights with the difference from controls being slight but statistically significant. There were no test item-related microscopic findings observed during the histopathological evaluation so the increases in liver weights were considered not to be toxicologically relevant. No maternal toxicity was observed in any of the other maternal parameters examined in this study. Therefore the maternal no-observed-adverse-effect-level was considered to be at least 1000 mg/kg bw/day.

In the initial observation, effects on skeletal morphology were observed at 1000 mg/kg bw/day. A higher incidence of the skeletal malformations of (severe) malaligned and fused sternebra(e) were seen with four fetuses, one from each of four litters, affected. Of these, two fetuses had both severely malaligned and fused sternebra(e) while one fetus had only fused sternebra(e) and one had only severely malaligned sternebra(e). Based on these initial findings, an external expert peer review (Charles River Report Amendment 2, 2017, Appendix 7) was performed which concluded that fused and malaligned sternebrae are variations rather than malformationsand therefore, there were no toxicologically relevant effects on skeletal morphology following treatment up to 1000 mg/kg bw/day. Skeletal malformations only occurred in two fetuses of this study. At 1000 mg/kg bw/day, 1 fetus had a short rib and since this occurred singly and was seen previously in historical control fetuses, it was considered to be a chance finding. The other malformation was noted in a control fetus with polydactyly (not detected externally) and as such was not considered related to treatment. There was no significant impact on skeletal variations proportion/litter (i.e.changes other than malformations) at 1000 mg/kg bw/day nor on fetal skeletal morphology at 100 or 330 mg/kg bw/day and no effects on fetal external or visceral morphology were seen with treatment up to 1000 mg/kg bw/day. The fetal NOAEL was therefore considered to be at least 1000 mg/kg bw/day.

A dose range-finding (DRF) study was performed prior to the pre-natal developmental toxicity study in rabbits (The Dow Chemical Company, 2017). Groups of five time-mated female NZW rabbits were administered neat dodecamethylcyclohexasiloxane (D6) via oral gavage at dose levels of 0, 50, 150, 450, 750, or 1000 mg/kg bw/day on gestation day (GD) 7-27. No maternal toxicity and no indication of embryo/fetal toxicity were reported. On the basis of this DRF study, 0, 100, 300 and 1000 mg/kg bw/day dose levels were selected for the key pre-natal developmental toxicity in rabbits.

In the second key prenatal developmental toxicity study with dodecamethylcyclohexasiloxane (D6), conducted according to OECD Test Guideline 414 and in compliance with GLP (The Dow Chemical Company, 2018a), D6 was evaluated for the maternal and developmental toxicity in New Zealand White rabbits following repeated oral gavage administration. Groups of 24 time-mated female rabbits were administered 0, 100, 300, or 1000 mg/kg bw/day on gestation day (GD) 7-27. No analyses of concentration verification, homogeneity, or stability were conducted because the test material was administered neat (undiluted). In-life parameters evaluated for all groups included: clinical observations, body weight, body weight gain, and feed consumption. On GD 28 all surviving rabbits were euthanized and examined for gross pathologic alterations. Liver, kidneys, and gravid uterine weights were recorded, along with the number of corpora lutea, uterine implantations, resorptions, and live/dead fetuses. All fetuses were weighed, sexed, and examined for external and visceral alterations. The heads were examined for craniofacial alterations by serial sectioning in approximately one half of the fetuses in each litter, and skeletal examinations were performed on all fetuses. Gavage administration of D6, up to and including the limit dose of 1000 mg/kg bw/day, resulted in no treatment-related maternal toxicity and no indication of embryo/fetal toxicity or teratogenicity. Therefore, under the conditions of this study, NOAEL for maternal toxicity and developmental toxicity was at least the limit dose of 1000 mg/kg bw/day.

In a preceding combined repeated dose toxicity study with the reproduction/developmental toxicity screening test, conducted according to OECD Test Guideline 422 and in compliance with GLP (Dow Corning Corporation, 2005), there were no remarkable findings at oral gavage doses of up to 1000 mg/kg bw/day. The maternal and fetal NOAEL were therefore considered to be at least 1000 mg/kg bw/day.

Justification for classification or non-classification

Based on the available data, D6 (dodecamethylcyclohexasiloxane) does not require classification for reproductive or developmental toxicity according to Regulation (EC) No 1272/2008.