Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
17 March 2017 to 21 September 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF, the Japanese Ministry of Agriculture, Forestry and Fisheries, Notification of 12 NohSan-8147, Guideline 2-1-18, Teratogenicity study, 24 November 2000.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Details on test animals and environmental conditions:
Species and Sex: Rats, time-mated females
Strain and Justification: Crl:CD(SD) rats were selected because of their general acceptance and suitability for toxicity testing, the availability of historical background data, and the reliability of the commercial supplier.
Supplier and Location: Charles River Laboratories (CRL) (Raleigh, North Carolina, USA)
Age and Weight at Study Start: Sexually mature adult, weighing approximately 200-250 g

Health Status and Acclimation: Upon arrival all animals were acclimated to the laboratory for at least four days prior to the start of test material administration. During the acclimation period, each animal was evaluated by a veterinarian trained in the field of Laboratory Animal Medicine, or a trained animal/toxicology technician, to determine the general health status and acceptability for study purposes. The Toxicology and Environmental Research and Consulting Laboratory is fully accredited by the Association for Assessment and Accreditation of Laboratory Animal Care International (AAALAC International).

Housing: Upon arrival animals were housed one per cage in stainless steel cages.

Environmental conditions:
Temperature: 22°C with a range of 20°C-26°C
Humidity: 50% with a range of 30-70%
Air Changes: 10-15 times/hour (average)
Photoperiod: 12-hour light/dark (on at 6:00 a.m. and off at 6:00 p.m.)
Enrichment: From the day of arrival until necropsy. Enrichment included paper nesting material and open areas on the cage sides for visualization of other rats.
Randomization and Identification: Animals were stratified by GD 0 body weight and then randomly assigned to treatment groups using a computer program designed to increase the probability of uniform mean group weights and standard deviations at the start of the study.
Animals that were placed on study were uniquely identified via subcutaneously implanted transponders (BioMedic Data Systems, Seaford, Delaware) that were correlated to unique alphanumeric identification numbers.

Feed and water: LabDiet Certified Rodent Diet #5002 (PMI Nutrition International, St. Louis, Missouri) in meal form. Feed and municipal water were provided ad libitum.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Remarks:
(dried and deacidified corn oil)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

- Octamethyltrisiloxane was prepared in dried and deacidified corn oil at dose levels of 0, 75, 250, or 750 mg/kg/day.

VEHICLE
- dried and deacidified corn oil

The rats were ordered and mated in six different replicates from the supplier to stagger cesarean sections over a period of two weeks.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration Verification and Homogeneity:
Dose solutions were prepared for analysis. Dosing solutions were shipped and stored at room temperature. Dose confirmation analyses of all dose levels, plus control, were determined from the first mix prior to exposure. The homogeneity of the low-dose and the high-dose test solutions was determined concurrent with dose confirmation. Additional samples of the low-dose and mid-dose concentrations were sent to the Lab to be analysed for dose confirmation and homogeneity following the start of dosing.
The initial analytical dose check for the low-dose and mid-dose solutions were below the acceptable range of targeted concentration from the first mix; therefore, no solutions from this mix were used to dose the animals and samples from the low-dose and mid-dose of the second mix were analysed. The method used for analysing the test material in dried and deacidified corn oil was gas chromatography with flame ionization detector (GC/FID) (Vogel, 2016) and followed the Lab's Standard Operating Procedures.
Octamethyltrisiloxane in dried and deacidified corn oil has been shown to be stable for up to 14 days at concentrations ranging from 12.5 - 250 mg/mL (Vogel, 2016). Dose solutions were prepared and used within these ranges; therefore, additional stability analyses were not conducted.
Retainer Samples. A sample of each lot number of the bulk test material was retained. No samples of dose solutions were retained.
Details on mating procedure:
The rats were ordered and mated in six different replicates from the supplier to stagger caesarean sections over a period of two weeks.
Duration of treatment / exposure:
GD 6-20
Frequency of treatment:
Daily
Duration of test:
Test material administration began on March 20, 2017 and the last group of rats were necropsied on April 19, 2017.
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day
Dose / conc.:
75 mg/kg bw/day
Dose / conc.:
250 mg/kg bw/day
Dose / conc.:
750 mg/kg bw/day
No. of animals per sex per dose:
24
Control animals:
yes, concurrent vehicle
Details on study design:
Groups of 24 Crl:CD(SD) rats were administered octamethyltrisiloxane in dried and deacidified corn oil by gavage at dose levels of 0, 75, 250, or 750 mg/kg/day on GD 6-20. The rats were ordered and mated in six different replicates from the supplier to stagger cesarean sections over a period of two weeks.

Route, Method of Administration, Frequency, Duration, and Justification: Test material was administered daily by oral gavage from GD 6-20. Oral administration is the preferred route of exposure specified in the relevant test guideline.
Dose Levels and Justification: Dose levels for this study were selected based on the 28-day and KMD studies discussed in the Previous Toxicity Information section above. The high-dose of 750 mg/kg/day was expected to induce overt signs of maternal toxicity (i.e., increased liver weights and histopathology). The lower dose levels were selected to provide dose response data for any toxicity observed among the high-dose group rats.

Examinations

Maternal examinations:
Daily Observations: A cage-side examination (including morbidity, mortality, and the availability of feed and water) at least twice daily, at approximately the same time each day.
Clinical Observations: At least once daily.
Body weights: On GD 0 by the supplier and daily from GD 6-21.
Feed Consumption: Recorded and statistically analysed for all animals every 3 days from GD 3-21 by weighing feed containers at the start and end of a measurement cycle.
Necropsy: On GD 21
Histopathology: Histopathological evaluation of the liver on all pregnant animals that survived to the scheduled necropsy.
Ovaries and uterine content:
Gravid uterine weights were recorded, along with the number of corpora lutea, uterine implantations, resorptions, and live/dead foetuses. All foetuses were weighed, sexed, and examined for external alterations. Approximately one half of the foetuses were examined for visceral and craniofacial alterations while skeletal examinations were conducted on the remaining foetuses.
Fetal examinations:
The individual body weight of all viable/dead foetuses and sex of all foetuses was recorded.
Total litter weight was calculated by addition of individual foetal body weights in a litter.
All foetuses were 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 foetuses were euthanized by sublingual oral administration of sodium pentobarbital solution. At least one half of all the foetuses in each litter were chosen randomly via computer for 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 observation of the thymus, trachea, esophagus, lungs, great vessels, heart (external and internal), liver, gastrointestinal tract, pancreas, spleen, kidney (sectioned), adrenal glands, ureters, bladder, and reproductive organs.
The heads of these foetuses were removed, placed in Bouin’s fixative and serially sectioned to allow for inspection of the eyes, brain, nasal passages, and tongue (Wilson, 1965). The foetal body was preserved in neutral, phosphate-buffered 10% formalin (to be discarded approximately six months after completion of the final report).
The remaining foetuses not selected for visceral examination were then skinned, eviscerated, preserved in alcohol, and double stained with Alcian Blue and Alizarin Red S for cartilage and bone according to methods based on Trueman et al. (1999) and Zablotny (2002) and a thorough evaluation of the foetal skeleton was conducted (foetal skeletal bodies were archived following completion of the final report). However, a foetus may have been intentionally changed from one selected for visceral examination to one processed for skeletal examination (and vice versa) if it was deemed that such examination would provide more meaningful data about a suspected abnormality.
All foetal alterations were classified as a variation or malformation. A variation is defined as a divergence beyond the normal range of structural constitution that may not adversely affect survival or health. A malformation is defined as a permanent structural change that may adversely affect survival, development or function and/or which occurs at a relatively low incidence in the specific species/strain.
The maternal necropsy and foetal examinations were conducted such that investigators were blind to treatment group assignment
Statistics:
Maternal body weights, maternal body weight gains, organ weights (absolute and relative with the exception of only absolute weight for gravid uterus), total litter weights, foetal 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.
Frequency of pre- and post-implantation loss (calculated), and foetal alterations (if any) were analysed 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 analysed using the Fisher exact probability test (alpha = 0.05; Siegel, 1956) with Bonferroni’s correction. Foetal sex ratios were analysed using a binomial distribution test. Non-pregnant females, females with resorptions only, or females found to be pregnant after staining of their uteri were excluded from the appropriate analyses. Statistical outliers (alpha = 0.02) were identified by the sequential method of Grubbs (1969) and were routinely excluded from feed consumption only. Other outliers, if excluded, were excluded from analysis for documented, scientifically sound reasons. Both Dunnett’s test and Bonferroni’s correction corrected for multiple comparisons to the control and were reported at the corrected alpha level.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Description (incidence and severity):
Examinations performed on all animals revealed no treatment-related findings throughout the duration of the study.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There was a treatment-related 19.8% decrease in body weight gain of animals in the 750 mg/kg/day group from GD 6-9 (Table 1). This decrease recovered and was similar to control for the remainder of the study.

Body weight gains of animals in the 250 and 750 mg/kg/day groups were higher than control and statistically identified from GD 15-21 with concomitant increases in feed consumption.

Body weight gains of animals in the 250 mg/kg/day group were higher than control and statistically significant over the GD 6-21 and GD 0-21 measured intervals. These higher body weight gains were considered unrelated to treatment with Octamethyltrisiloxane as the differences were minimal, there was no effect in body weight at any dose level, and corrected body weights on GD 21 were similar to control. Body weights of animals in all dose groups and body weight gains of animals in the 75 mg/kg/day dose group were similar to control throughout the duration of the study.
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 controls. Feed consumption of animals in the 250 and 750 mg/kg/day groups were higher and statistically significant from GD 15-21 with concomitant increases in body weight gains (Table 2). These higher feed consumption values were considered unrelated to treatment with octamethyltrisiloxane. Feed consumption was similar to controls for animals in the 75 mg/kg/day group throughout the duration of the study.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
There were treatment-related, statistically significant increases in mean absolute and relative liver weights at all dose levels tested (Table 3) with correlating histopathological observations. The mean relative liver weights of females given 75, 250, or 750 mg/kg/day were 9.3%, 13.5%, or 29.5% higher than controls, respectively.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no treatment-related gross pathologic observations.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Treatment-related very slight hypertrophy of centrilobular/midzonal hepatocytes was present in 12/24 females given 75 mg/kg/day and in 17/24 females given 250 mg/kg/day (Table 4). Treatment-related slight hypertrophy of centrilobular/midzonal hepatocytes was present in 7/24 females given 250 mg/kg/day and in 24/24 females given 750 mg/kg/day. The hepatocellular hypertrophy corresponded to the dose related increases in liver weights of females given 75, 250 or 750 mg/kg/day. Two females given 750 mg/kg/day had treatment-related very slight multifocal chronic inflammation in periportal regions of the liver, and one of the females with chronic inflammation also had treatment-related very slight increased number of mitotic figures in hepatocytes. Treatment-related pigment deposits were present in the intrahepatic bile ducts, hepatocytes and/or Kupffer cells of six females given 750 mg/kg/day. The pigment deposits were brown when examined with a light microscope and birefringent with some deposits featuring Maltese cross formations when examined with polarized light. All of the treatment-related liver effects were consistent with a previously conducted 28-day oral gavage toxicity study with octamethyltrisiloxane (Braun, 2010). The pigment deposits in animals 471, 475, 478, and 482 were negative for the presence of bile when processed with Fouchet’s bile stain.
Histopathological findings: neoplastic:
no effects observed

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
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:
No effects noted.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: Based on the decreased body weight gain and liver weight increases combined with the chronic inflammation of the liver noted in animals in the 750 mg/kg/day group

Maternal abnormalities

Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
Foetal body weights of males in the 750 mg/kg/day group, females in the 250 and 750 mg/kg/day groups, and sexes combined in the 250 and 750 mg/kg/day groups were higher than control and statistically identified. These higher body weights were considered unrelated to treatment with octamethyltrisiloxane as they were within recent historical control values.
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Details on embryotoxic / teratogenic effects:
No effects noted.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
750 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There was no indication of embryo/fetal toxicity or teratogenicity at any dose level tested.

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
no

Any other information on results incl. tables

Table 1. Body Weight Gains (g) – Selected Intervals

Dose

(mg/kg/day)

Day of Gestation

   6 -9  9 -12  15 -18  18 -21  6 -21  0 -21
0  12.1 19.0   37.0 50.4  139.9  171.2 
 75  14.0 21.4  40.7  55.8  153.7  184.9 
 250  13.2 21.6  43.2*  59.1*  159.3*  190.8* 
 750  9.7 20.6  42.5*  59.2*  152.8  184.9 

Bold type indicates the effect was interpreted to be treatment related.

* Statistically different from control mean by Dunnett’s Test, Alpha = 0.05.

Table 2. Feed Consumption (g/day)

 Dose (mg/kg/day)

Day of Gestation  

   6 -9  15 -18  18 -21
 0  17.4 21.0  19.8 
 75  17.7 22.6  21.5 
 250  17.6 24.1*  22.3* 
 750  16.5 24.4*  23.6* 

* Statistically different from control mean by Dunnett’s Test, Alpha = 0.05.

Table 3. Liver Weights

 Dose (mg/kg/day) Final Body Weight (g)   Liver (g) Liver (g/100) 
 0  391.8 13.346  3.407 
 75  405.4 15.122*  3.725* 
 250  411.5 15.928*  3.867* 
 750  405.0 17.835*  4.411* 

* Statistically different from control mean by Dunnett’s Test, Alpha = 0.05.

Table 4. Histopathologic Liver Alterations

 Sex           Females
 Dose level (mg/kg/day)/Liver (number examined)  0/24 75/24 250/24 750/24

Hypertrophy, increased eosinophilia, hepatocyte, centrilobular/midzonal (-very slight/-slight)

 0/0

12/0 17/7 0/24 

Increased Number of Mitotic Figures, hepatocyte, -very slight

 0

Inflammation, chronic, periportal, multifocal -very slight

 0
Pigment, bile duct, focal -very slight  0 1 
Pigment, bile duct, multifocal -very slight  0
Pigment, hepatocyte, multifocal -very slight  0
Pigment, Kupffer cell, multifocal -very slight  0

Bold type indicates the effect was interpreted to be treatment related.

Applicant's summary and conclusion

Conclusions:
In a developmental toxicity study (Dow Corning Corporation, 2017) carried out according to OECD Test Guideline 414, there was no indication of embryo/foetal toxicity or teratogenicity at any dose level tested. The NOAEL for developmental toxicity was determined to be 750 mg/kg/day, the highest dose level tested.
Executive summary:

The maternal and developmental toxicity of octamethyltrisiloxane in Crl:CD(SD) rats was studied following repeated gavage administration in this OECD 414 study. Groups of 24 time-mated female Crl:CD(SD) rats were administered octamethyltrisiloxane in dried and deacidified corn oil by gavage at dose levels of 0, 75, 250, or 750 mg/kg/day on gestation day (GD) 6 through 20. In-life maternal study parameters included clinical observations, body weight, body weight gain, and feed consumption. On GD 21, all dams 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 foetuses. All foetuses were weighed, sexed, and examined for external alterations. Approximately one half of the fetuses were examined for visceral and craniofacial alterations while skeletal examinations were conducted on the remaining fetuses.

Administration of octamethyltrisiloxane via oral gavage produced treatment-related maternal toxicity evidenced by decreased body weight gains from GD 6-9 in the 750 mg/kg/day group (20% compared to control), increased absolute and relative liver weights and liver histopathology at all dose levels. Treatment-related very slight hypertrophy of centrilobular/midzonal hepatocytes was present in 12/24 females given 75 mg/kg/day and in 17/24 females given 250 mg/kg/day. Treatment-related slight hypertrophy of centrilobular/midzonal hepatocytes was present in 7/24 females given 250 mg/kg/day and in 24/24 females given 750 mg/kg/day. The hepatocellular hypertrophy corresponded to dose-related increases in absolute and relative liver weights of females given 75, 250, or 750 mg/kg/day. Two females given 750 mg/kg/day had treatment-related very slight multifocal chronic inflammation in periportal regions of the liver, and one of these females also had treatment-related very slight increased number of mitotic figures in hepatocytes. Treatment-related pigment deposits were present in the intrahepatic bile ducts, hepatocytes, and/or Kupffer cells of six females given 750 mg/kg/day. There were no treatment-related clinical observations and no effects on body weight, feed consumption, or reproductive parameters at any dose level tested. There was no indication of embryo/foetal toxicity or teratogenicity at any dose level tested.

Based on the decreased body weight gain and liver weight increases combined with the chronic inflammation of the liver noted in animals in the 750 mg/kg/day group, the NOAEL for maternal toxicity was determined to be 250 mg/kg/day. For developmental toxicity, the NOAEL was 750 mg/kg/day, the highest dose level tested.