Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
25 June 2018
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Bayrisches Landesamt für Gesundheit und Lebensmittelsicherheit
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
- Name of test material (as cited in study report): Methyl [3-(trimethoxysilyl)propyl]carbamate
- Physical state: liquid
- Storage condition of test material: storage at 2-8 °C, in a dry and ventilated place, protected from light and moisture (overlaid with argon after opening)

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, 97633 Sulzfeld, Germany
- Age at study initiation: males - between 12 weeks and not older than 24 weeks; females - approx. 11-12 weeks old
- Weight at study initiation: males: 328 – 424 g; females: 177 - 239 g
- Housing: individually housed in type III H, polysulphone cages
- Diet: Altromin 1324 maintenance diet for rats and mice provided ad libitum
- Water: tap water, sulphur acidified to a pH of approximately 2.8 provided ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): 10 x
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was weighed and the vehicle (corn oil) was added to give the appropriate final concentration of the test item. The formulations were vortexed and/or stirred until visual homogeneity was achieved.

VEHICLE
- Concentration in vehicle: undiluted
- Amount of vehicle: The application volume for all groups was 4 mL/kg bw.
- Lot/batch no. (if required): MKCG3257
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The mean recoveries observed in low-, mid- and high-dose groups were 96.5%, 95.0%, and 96.7% of the nominal concentration, respectively. Nominal concentrations were confirmed for all dose groups, as measured concentrations were within acceptance criterion of 10%.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1:2 (male to female)
- Length of cohabitation: Females were paired for cohabitation in batches in order to control the number of animals for terminal sacrifice on a particular day. After getting 100 sperm positive females, the remaining females and males were discarded without any observations.
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
Gestation days 5 through 19
Frequency of treatment:
Daily
Duration of test:
until gestation day 20
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 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:
25 females/group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels are based on a dose range finding study. In this study pregnant Wistar rats were treated with the test item at doses of 100, 300 and 1000 mg/kg bw/day from GD 5 to 19. No test item-related toxicological effects in terms of clinical signs, morbidity/mortality, body weight development, food consumption and gross pathological observations of pregnant females were observed in the test item-treated groups when compared with the controls. No signs of foetal toxicity were seen in terms of effect on prenatal data, litter data and foetal external findings up to the highest dose level of 1000 mg/kg bw/day.
- Rationale for animal assignment: Mated females were assigned in an unbiased manner to the control and treatment groups ensuring that group mean body weights were comparable with each other. Each animal was assigned a unique identification number.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Clinical observations were made at least once a day, preferably at the same time each day. Twice daily all animals were observed for morbidity and mortality except on weekends and public holidays when observations were made once daily.
- Clinical observations included spontaneous activity, lethargy, recumbent position, convulsions, tremors, apnoea, asphyxia, vocalisation, diarrhoea, changes in the skin and fur, eyes and mucous membranes (salivation, discharge), piloerection and pupil size. Changes in gait, posture, response to handling as well as the presence of clonic or tonic movements, stereotypes or bizarre behaviour were recorded.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Clinical observations were made at least once a day, preferably at the same time each day.

BODY WEIGHT: Yes
- Time schedule for examinations: All animals were weighed once before initiation of pairing to ensure that the body weights are within ±20 % variation. The sperm positive females were weighed on GDs 0, 5, 8, 11, 14, 17 and 20. Males were not weighed in this study except once before initiation of pairing.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE: No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: At the time of termination or death during the study, the dam (presumed pregnant female) was examined macroscopically for any structural abnormalities or pathological changes which may have influenced the pregnancy. Immediately after the termination or as soon as possible after death, the uteri were removed and the pregnancy status of the dams was confirmed.
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:
A statistical assessment of the results of the body weight and food consumption was performed by comparing values of dosed animals with control animals using a one-way ANOVA and a post-hoc Dunnett Test. Foetal evaluation parameters like external, visceral, craniofacial and skeletal parameters were analysed using Fisher’s exact test.
Indices:
Percent preimplantation loss; percent pregnant; percent early and late resorptions
Historical control data:
Historical control range data were provided for: uterine data; litter weight data; fetal external examination; fetal visceral examination; fetal craniofacial examination; and fetal skeletal examination

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Predominant clinical signs observed on a few days during the treatment period of the study included moving the bedding (17/25 in the high dose), increased salivation (1/25 in the low dose), prone position (1/25 in control), reduced spontaneous activity (3/25 in the high dose), abnormal recumbency (1/25 in the high dose)) and hyperthermia (1/25 in control). There were also low incidences of clinical signs like hairless area on various body parts (1/25 each in control, low dose and mid dose), dehydration (1/25 in control) and abnormal breathing (1/25 in the high dose)) observed in isolated females of all groups including the control group.

Therefore all clinical signs observed in terminally sacrificed treatment group females were incidental and considered to be of no toxicological relevance or non-adverse in nature.

Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No mortality was observed during the treatment period and all animals survived until the end of the study. One high-dose dam, which delivered mature pups on designated GD 9, was sacrificed before the scheduled day of terminal sacrifice. This delivery was attributed to missing detection of sperm positive vaginal smear at first instance and therefore this female littered after completion of the gestation period. This premature delivery was not considered to be related with test item treatment.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The mean body weight remained unaffected by treatment with the test item and increased with the progress of the study in the control, low-, mid-, or high-dose groups throughout the study period. No statistical significance was achieved in any treatment groups on any day or interval of body weight measurement and all values in the treatment groups were comparable to the controls. However, mean body weight gain was noted to be marginally but statistically significantly lower (p < 0.01) in the mid-dose group during GD 14-17 (61.75% of controls) when compared to the controls. As this effect on group mean body weight gain was seen in the mid-dose group only on just one occasion and due to lack of dose dependency, it was not considered related to treatment with test item.
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 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:
no effects observed
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Details on results:
No mortality was observed during the treatment period and all animals survived until the end of the study. There were no clinical signs of toxicological relevance observed in the females of any treatment group.

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 examined
Details on maternal toxic effects:
No test item-related effects of toxicological relevance were noted for any prenatal parameters including terminal body weight, adjusted maternal weight (carcass weight), uterus weight, net weight change from GD 0, number of corpora lutea, implantation sites, early and late resorptions, number of live foetuses, number female foetuses, and percent pre- and post-implantation loss in treatment groups when compared to the controls.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: No adverse, treatment-related effects were observed.

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
A marginal but statistically significantly higher mean male (p < 0.05) and female (p < 0.01) foetus weight was observed on an individual basis (sum of weight of all foetuses in group divided by total number of foetuses in respective group) in the low-dose group when compared with the controls. As this difference was not dose dependent and in the light of no significant effect on male and female foetus weight observed based on the litter means, it was not considered to be biologically relevant.
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
effects observed, non-treatment-related
Description (incidence and severity):
A statistically significantly lower (p < 0.01) number of males and sex ratio (% males) was observed in the mid-dose group when compared with the controls. Due to lack of dose dependency and consistency, this effect was not considered to be test item-related.
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Low incidences of localised oedema (1 in the low dose), hooked tail (1 in control) and short tail (1 each in control and mid dose) were noted in isolated foetuses of the control group and/or the dose groups without dose dependency. As these findings were observed mostly in single foetuses, they were considered to be incidental in nature and unrelated to treatment with the test item.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
A statistically significantly lower litter incidence for misaligned ossification of 4th sternebra in high dose (0% in high dose compared to 35% in control) and misaligned ossification of 5th sternebra in mid and high dose (0% each compared to 35% in control) were considered to be incidental as frequencies were even less in the treated groups compared to the controls. Therefore, these findings are considered to be solely spontaneous in nature.

Slightly higher litter incidences, but without achieving statistical significance were observed in the high-dose group for incomplete ossification of 1st sternebra (20% compared to 0% in control), 2nd sternebra (35% compared to 25% in control), 3rd sternebra (20% compared to 0% in control), 4th sternebra (40% compared to 25% in control), 5th sternebra (70% compared to 65% in control), skull maxilla (10% compared to 0% in control), skull mandible (30% compared to 20% in control), parietal skull (left) (25% compared to 15% in controls), squamosal skull (left) (15% compared to 10% in controls), un-ossified forelimb phalanges (95% compared to 75% in controls), pelvic girdle caudal shift (bone) (40% compared to 20% in controls) and wavy ribs (35% compared to 25% in controls).

The observed incomplete ossification without achieving statistical significance of a few bones and a few other skeletal findings in the high-dose group were either marginally higher or within historical control data range. Generally delayed ossification is not regarded to persist postnatally and is not associated with long-term consequences on survival, general growth and development and therefore is not considered to be adverse.

The finding of a caudal shift of the pelvic girdle (bilateral) in the high-dose group was well within the historical control data range (35%) and changes in the position of the pelvic girdle relative to the number of pre-pelvic vertebrae can occasionally be seen in animals of this strain and therefore this finding was not considered to be adverse. Wavy ribs are common findings in rodent studies and are considered to be postnatally reversible. Thus, wavy ribs are classified as variations and were not considered as an adverse effect of the treatment with the test item.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
All litter incidences were statistically insignificant when compared with the control except statistically significantly lower litter incidence for long thymus in the mid- and high-dose groups (5 and 10%, respectively, compared to 50% in control). There were higher litter incidences observed for a few visceral findings like umbilical artery malpositioned (15% in the high dose compared to 10% in control), testes malpositioned (10% in high dose compared to 5% in control), abdomen internal haemorrhage (55% in the high dose compared to 25% in control), neck subcutaneous haemorrhages (100% in high dose compared 95% in control)) and renal pelvis dilated (10% in the high dose compared to 5% in control) in the high-dose group without achieving statistical significance. However, findings were either minor variations or values were well within the historical control data range (47.37% renal pelvis dilated) and therefore not considered to be adverse.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
Craniofacial examination by razor blade serial sectioning technique revealed a few predominant findings (discoloured cerebellar lobe, red material in perimeningeal space, increased perimeningeal space, subdural hematoma (mid brain) at low frequencies generally comparable to or in some cases slightly higher or lower in frequency in the dose groups compared to the controls. These findings were considered to be spontaneous in nature and not related to the treatment with the test item.
Details on embryotoxic / teratogenic effects:
There were no test item-related effects of toxicological relevance observed for the mean foetus weight, male and female foetus weight on a per litter basis (group mean of individual litter mean) in any of the treatment groups when compared with the controls. There were no external abnormalities considered to be of toxicological relevance in any of the dose groups. Visceral findings observed in the dose groups were at frequencies generally comparable to or in some cases slightly higher or lower in frequency compared to controls. Craniofacial examination by razor blade serial sectioning technique revealed a few predominant findings (discoloured cerebellar lobe, red material in perimeningeal space, increased perimeningeal space, subdural hematoma (mid brain) at low frequencies generally comparable to or in some cases slightly higher or lower in frequency in the dose groups compared to the controls. These findings were considered to be spontaneous in nature and not related to the treatment with the test item. There was no statistical significance and no indication of a test item-related trend in the type and/or incidences of other skeletal findings and they were therefore considered to be spontaneous in nature.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
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, treatment-related effects were observed.

Fetal abnormalities

Key result
Abnormalities:
effects observed, non-treatment-related

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
The test item was examined for developmental toxicity in a study that was conducted according to OECD 414 test guidelines and in compliance with GLP. Based on the results, no toxicologically relevant effects of methyl [3-(trimethoxysilyl)propyl]carbamate (CAS 23432-62-4) on females and foetuses were found at dose levels up to 1000 mg/kg bw/day. The NOAEL for both maternal toxicity and foetal toxicity of methyl [3-(trimethoxysilyl)propyl]carbamate in this study is considered to be 1000 mg/kg bw/day.