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Key value for chemical safety assessment

Effects on fertility

Description of key information

In the key fertility study (combined repeated dose oral toxicity study with the reproduction / developmental toxicity screening test) with the registered substance 3-(trimethoxysilyl)propane-1-thiol (CAS No. 4420-74-0, EC No. 224-588-5), conducted according to OECD Test Guideline 422 and in compliance with GLP (Charles River Laboratories, 2021b, reliability score 1), the NOAEL for reproductive effects was concluded to be equal to or higher than 600 mg/kg bw/day based on no adverse effects on reproduction parameters. The parental systemic NOAEL was considered to be 300 mg/kg bw/day based on adverse clinical signs, decreased body weight, body weight gain, and/or food consumption in both sexes at 600 mg/kg bw/day, with bladder urothelial hyperplasia combined with degenerative bladder histopathology in a single female at this dose.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
16 Dec 2020 (experiment start) - final report date when available
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)
Version / remarks:
2016
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EPA OPPTS 870.3650, Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test
Version / remarks:
2000
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The Wistar Han rat was chosen as the animal model for this study as it is an accepted rodent species for toxicity testing by regulatory agencies. Charles River Den Bosch has general and reproduction/developmental historical data in this species from the same strain and source. This animal model has been proven to be susceptible to the effects of reproductive toxicants.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: 10-11 weeks (males) ; 13-14 weeks (females); (F1) x wks
- Weight at study initiation: (F1) Males: x-x g; Females: x-x g
- Fasting period before study: not specified
- Housing: polycarbonate cages (Makrolon, MIV/MIII type, height 18 cm). The cages contained appropriate bedding (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany). Animals were separated during designated procedures/activities. Main males and females were cohabitated on a 1:1 basis during the mating phase.
Diet (e.g. ad libitum): Pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany), ad libitum (no access to food during motor activity measurements for a max. of 2 hours)
- Water (e.g. ad libitum): Municipal tap water via water bottles, ad libitum (no access during motor activity measurments for a max. of 2 hours)
- Acclimation period: 6 days prior to start of the pretest period (females) or 7 days before the commencement of dosing (males).
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C
- Humidity (%): 40 to 70%
- Air changes (per hr): Ten or greater air changes per hour with 100% fresh air (no air recirculation)
- Photoperiod (hrs dark / hrs light): 12 hour light/12 hour dark




Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Test item dosing formulations (w/w) were homogenized to visually acceptable levels at appropriate concentrations to meet dose level requirements. The dosing formulations were prepared daily and dosed within 6 hours after adding the vehicle to the test item. Test item dosing formulations were kept at room temperature until dosing. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing. Adjustment was made for specific gravity of the vehicle and test item. No correction was made for the purity/composition of the test item.


VEHICLE
- Justification for use and choice of vehicle (if other than water): Trial preparations were performed to select the suitable vehicle and to establish a suitable formulation procedure. These trials were not performed as part of this study and these preparations were not used for dosing.
- Concentration in vehicle: 0, 20, 60, 120 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg bw
- Purity: not specified


Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: maximum of 14 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 post-coitum
- After successful mating each pregnant female was caged (how): individually, in Makrolon plastic cages (MIII type, height 18 cm).
- Any other deviations from standard protocol: no
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Duplicate sets of samples (approximately 500 mg) for each sampling time point were sent to the analytical laboratory. Concentration results were considered acceptable if mean sample concentration results were within or equal to ± 10% for solutions of target concentration. Homogeneity results were considered acceptable if the coefficient of variation (CV) of concentrations was <= 10%.
Duration of treatment / exposure:
At least 29 days. Main females were treated for 51-65 days; 14 days prior to mating and at least 13 days after delivery, up to and including the day before scheduled necropsy. Main females which failed to deliver were treated for 41-52 days. Recovery females were treated during the same period as Main females, until at least the day before the first scheduled necropsy of Main females. Main males and Recovery males were treated for 29 days, including a minimum of 14 days prior to mating and during the mating period for Main males. For both Main and Recovery males, treatment ended one day before scheduled necropsy of Main males.
Frequency of treatment:
Daily, 7 days a week
Details on study schedule:
N/A screening study
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Group 1 = Control group
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
Group 2
Dose / conc.:
300 mg/kg bw/day (nominal)
Remarks:
Group 3
Dose / conc.:
600 mg/kg bw/day (nominal)
Remarks:
Group 4
No. of animals per sex per dose:
10 animals/sex/group in the main groups and 5/animals/sex/group in the recovery groups
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected based on the results of a 14-Day Dose Range Finder (DRF) with oral administration of 3-(trimethoxysilyl)propane-1-thiol (CAS 4420-74-0) in rats (Test Facility Reference No. 20244995), and a prematurely terminated OECD 422 study (Test Facility Study No. 20244996) in rats, and in an attempt to produce graded responses to the test item. See mode details under details of results.
- Rationale for animal assignment (if not random): random
- Fasting period before blood sampling for clinical biochemistry: yes
- Rationale for selecting satellite groups: not specified
Positive control:
No
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule:
Animals were observed for general health/mortality and moribundity twice daily, in the morning and at the end of the working day. Animals were not removed from the cage during observation, unless necessary for identification or confirmation of possible findings.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:
Once daily, beginning during the first administration of the test item and lasting throughout the dosing and recovery periods up to the day prior to necropsy.

Clinical observations included changes in gait, spontaneous activity, response to handling as well as the presence of clonic or tonic movements, stereotypes, difficult or prolonged parturition or bizarre behaviour, 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.

BODY WEIGHT: Yes
- Time schedule for examinations:
Animals were weighed individually on the first day of treatment (prior to dosing), and weekly thereafter. Mated main females were weighed on days 0, 4, 7, 11, 14, 17, and 20 post-coitum and during lactation on PND 1, 4, 7, and 13. A terminal weight was recorded on the day of scheduled necropsy

FOOD CONSUMPTION AND COMPOUND INTAKE : Yes
- Time schedule: Food consumption was quantitatively measured weekly, except for Main males and Main females which were housed together for mating and for Main females without evidence of mating. Food consumption of mated Main females was measured on days 0, 4, 7, 11, 14, 17, and 20 post-coitum and during lactation on PND 1, 4, 7, and 13

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule: Water consumption was monitored on regular basis throughout the study by visual inspection of the water bottles. No quantitative investigation was introduced as no effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of the treatment period or end of the recovery period on the day of scheduled necropsy
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: All animals
- Parameters checked in Table 1 below were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the end of the treatment period or end of the recovery period on the day of scheduled necropsy
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: All animals
- Parameters checked in Table 1 below were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations:
Functional tests performed after dosing, after completion of clinical observations. During week 4 of treatment for 5 selected main males and all recovery males, and during last week of lactation for 5 selected main females and all recovery females. As potential treatment-related findings were noted in locomotor activity in recovery females at the end of the treatment period, locomotor activity measurements were also conducted for all Recovery females at the end of the recovery period
- Battery of functions tested: hearing ability, pupillary reflex, static righting reflex, fore- and hind-limb grip strength, locomotor activity (total movements and ambulations).

IMMUNOLOGY: No
Oestrous cyclicity (parental animals):
Estrous cycles were evaluated by examining the vaginal cytology of samples obtained by vaginal lavage.
Daily vaginal lavage was performed for all females (Main and Recovery) during 14 days prior to treatment (pretest period) and the first 14 days of treatment. For Main females, daily vaginal lavage was continued during mating until evidence of copulation was observed. On the day of necropsy, a vaginal lavage was also taken from Main females to determine the stage of estrous
Sperm parameters (parental animals):
Parameters examined in male parental generations: testes and epdidymis weight.
For the testes of all selected Main males of Groups 1 and 4, and all Main males that failed to sire, a detailed qualitative examination was made, taking into account the tubular stages of the spermatogenic cycle.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were euthanised by decapitation. Blood samples were collected from two of the surplus pups (if possible, from one male and one female pup) and samples were pooled to one sample per litter. Selective elimination of pups, e.g. based upon body weight or AGD, was not done. All remaining pups were euthanized on PND 14-16.

PARAMETERS EXAMINED
The following parameters were examined in F1] offspring:
Number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, anogenital distance (AGD), presence of nipples/areolae in male pups, T4 levels. Particular attention should be paid to the external reproductive genitals which should be examined for signs of altered development.

GROSS EXAMINATION OF DEAD PUPS:
[yes, for external and internal abnormalities; presence of milk in stomach; and if possible defects or cause of death evaluated]

ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY: No

ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY: No
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals [Following completion of the mating period for Main males which sired or failed to sire; after the recovery period of at least 14 days for Recovery males]
- Female animals: All surviving animals [At PND 14-16 for Main females which delivered; Post-coitum Days 25-26 for Main feamles with evidence of mating which failed to deliver; After the recovery period of at least 14 days for Recovery females]

GROSS NECROPSY / HISTOPATHOLOGY / ORGAN WEIGHTS: Yes
Gross necropsy consisted of external and internal examinations, including examination of the cranial, thoracic and abdominal cavities and their contents, as well as reproductive organs.

A number of organs from all selected main animals and all recovery animales (see Table 2 below) as well as from males that failed to sire and females that failed to deliver (see Table 3 below) were prepared for microscopic examination and weighed.

All tissues indicated in these tables were examined by board-certified toxicological pathologist with training and experience in laboratory animal pathology. For the testes of all selected Main males of Groups 1 and 4, and all Main males that failed to sire, a detailed qualitative examination was made, taking into account the tubular stages of the spermatogenic cycle.
Postmortem examinations (offspring):
SACRIFICE
- Pups younger than 7 days were euthanized by decapitation.
- All remaining pups (PND 14-16), except for the two pups per litter selected for blood collection were euthanized by an intraperitoneal injection of sodium pentobarbital.
- The pups selected for blood collection on PND 14-16 were anesthetized using isoflurane followed by exsanguination.

GROSS NECROPSY / HISTOPATHOLOGY
Sex was determined both externally and internally. Descriptions of all external abnormalities were recorded. Particular attention was paid to the external reproductive genitals to examine signs of altered development.
In addition, blood was collected from two pups per litter, and the thyroid from two pups per litter (if possible one male and one female pup) was preserved in 10% buffered formalin. The pups selected for blood sampling were the same pups as selected for thyroid preservation.
Statistics:
See repeated dose toxicity endpoint 7.8.1.089
Reproductive indices:
Reproductive-type parameters evaluated were evidence of mating, pregnancy and duration of gestation, lenght and regularity of the estrous cycle, and number of uterine implantation sites.
Offspring viability indices:
Pups were examined daily for survival, external abnormalities and clinical signs. Number and sex of the pups, the number of pups alive per litter, number of pups dead were noted. All pups were counted, weighed, sexed and the sex ratio calculated.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Abnormal gait, belly sliding behaviour and back scratching were noted in all males and females at 600 mg/kg bw/day. Lethargy in males and females was noted at 600 mg/kg bw/day.
Uncoordinated movements as well as yellow staining/discoloration of the genital region in females only were observed at 600 mg/kg bw/day.
These effects were considered adverse clinical signs.
Hunched posture, salivation and incidental findings including alopecia, piloerection and scabs were also observed but were not considered not to be toxicologically relevant.


Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No mortality occurred during the study period.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weights and body weight gain was considered to be affected by treatment with the test item in males and main females at 600 mg/kg bw/day.

In males at 600 mg/kg bw/day, mean body weight and mean body weight gain were decreased throughout the treatment period (not always reaching statistical significance), followed by only slight recovery during the 14 days treatment-free Recovery Period. Mean body weights were 5% and 4% lower compared to the concurrent control group at the end of the treatment period and end of the recovery period, respectively.
In females at 600 mg/kg bw/day, mean body weight gain was decreased between post-coitum Days 7-20 (not always reaching statistical significance), followed by values that were similar to the concurrent control group during the lactation period. At the end of the periods of gestation and lactation, mean body weight were 6% lower compared to the concurrent control group. In non-pregnant recovery females, body weight was considered to be unaffected by treatment with the test item up to 600 mg/kg bw/day.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption before or after correction for body weight was considered affected by treatment with the test item in males and main females at 600 mg/kg bw/day.

In males at 600 mg/kg bw/day, a trend towards lower mean absolute and relative food consumption was observed (-19 and -17% of control, respectively; not statistically significant) over Days 1-8 of the premating period which recovered to levels similar to the concurrent control thereafter.
In main females at 600 mg/kg bw/day, mean absolute and relative food consumption were decreased from post-coitum Days 14-17 up to and including Lactation Days 7-13.

During the last interval of the lactation period, mean absolute and relative food consumption were 24 and 21% lower compared to the concurrent control, respectively (not always reaching statistical significance).
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Hematology findings in males comprised of higher white blood cell and neutrophil concentrations (at 600 mg/kg bw/day), and monocyte concentrations (starting at 300 mg/kg bw/day), in Main females of higher reticulocyte concentration and red blood cell distribution width gated (starting at 300 mg/kg bw/day), and lower neutrophil and monocyte concentrations (at 600 mg/kg bw/day), and in Recovery females of higher red blood cell concentration, and lower mean corpuscular volume and mean corpuscular hemoglobin (at 600 mg/kg bw/day). Except for monocyte concentration in males and red blood cell concentration in recovery females, partial recovery was seen for all other differences at the end of the recovery period.
These findings were considered non adverse since these changes were not associated with any adverse pathological alterations.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Clinical biochemistry findings in males comprised of higher alanine aminotransferase and aspartate aminotransferase activity (starting at 300 mg/kg bw/day), and bile acid and urea concentration (at 600 mg/kg bw/day), and in Main females of higher alanine aminotransferase activity and albumin concentration, and lower inorganic phosphate concentration (all at 600 mg/kg bw/day), and in recovery females of higher bile acid concentration and lower total protein concentration, albumin concentration and calcium concentration (all at 600 mg/kg bw/day). Except for bile acid concentration (males), total protein concentration, albumin concentration and calcium concentration (females), (partial) recovery was noted for all other changes at the end of the recovery period. These findings were considered non adverse since these changes were not associated with any adverse pathological alterations.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
In main females, mean total movements and ambulations were increased (1.34x and 1.29x of control, respectively) at 600 mg/kg bw/day.
In recovery females, mean total movements was decreased (0.88x of control mean) at 600 mg/kg bw/day at the end of the treatment period and recovered to control levels at the end of the recovery period. Mean ambulations were considered not affected by treatment with the test item at 600 mg/kg bw/day at the end of the treatment and recovery period.
Motor activity was considered not to be affected by treatment with the test item in males up to 600 mg/kg bw/day. All groups showed a similar motor activity habituation profile with in general a decreasing trend in activity over the duration of the test period. Hearing ability, pupillary reflex, static righting reflex and grip strength were normal in all examined animals up to 600 mg/kg bw/day.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Treatment-related bladder findings were observed in both sexes at 300 and 600 mg/kg bw/day but, with the exception of one female at 600 mg/kg bw/day, are considered non-adverse by the Registrant. In addition, non-adverse mesenteric lymph node findings were seen in males at these two doses.

Minimal to slight urothelial hyperplasia/hypertrophy was noted in the urinary bladder of males and females treated at 300 and 600 mg/kg bw/day. This finding showed no recovery after the treatment free recovery period. In a single female at 600 mg/kg bw/day, effects of a degenerative nature also were seen and included ulcer of the urothelium, haemorrhage, oedema and subepithelial fibrosis. However, based on a review of all available rat data across a wide range of silicon-based compounds including multiple trimethoxysilanes, the silicon industry considers diffuse urothelial hyperplasia in the bladder without atypia to be an adaptive (non-adverse) response to physical or chemical irritation. Thus, except for the single female with degenerative bladder histopathology at 600 mg/kg bw/day, the bladder urothelial hyperplasia seen at 300 and 600 mg/kg bw/day for 3-(trimethoxysilyl)propane-1-thiol is assessed as non-adverse by the Registrants.

In the mesenteric lymph nodes of males at 300 and 600 mg/kg bw/day, sinus histiocytosis observed was without any degenerative finding and there was partial/ongoing recovery after the treatment-free recovery period. Therefore, this finding was considered non adverse.
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Description (incidence and severity):
No treatment-related toxicologically significant changes were noted in T4 thyroid hormone levels.
Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
Length and regularity of the estrous cycle were considered not to have been affected by treatment with the test item up to 600 mg/kg bw/day. All females had regular cycles of 4 to 5 days, except for Recovery Female No. 96 (600 mg/kg bw/day) which had an irregular cycle during the Treatment Period that was characterized by an extended di estrous period. Given the incidental nature, this finding was considered not related to treatment with the test item.
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Description (incidence and severity):
Mating index was not affected by treatment with the test item up to 600 mg/kg bw/day. All Main females showed evidence of mating, which resulted in a mating index of 100% for all groups.
Precoital time was not affected by treatment with the test item up to 600 mg/kg bw/day. All paired females showed evidence of mating within 5 days, except for Female Nos. 53, 57 (control) and 70 (100 mg/kg bw/day) for which mating took 14, 13 and 14 days, respectively.
During the prematurely terminated OECD 422 study (dose levels: 0, 100, 300 and 1000 mg/kg bw/day), 1/15 males and 1/15 females at 1000 mg/kg bw/day were found dead on treatment Day 6 and all remaining males and females at 1000 mg/kg bw/day were euthanized in extremis due to the severe toxicity observed within one week of dosing. Most animals at 1000 mg/kg bw/day had lost more than 10% (10-19%) of their body weight within 5 days of treatment and/or showed one or a combination of the following clinical symptoms: hunched posture, flat posture, uncoordinated movements, decreased locomotor activity, tremor, lethargy, slow breathing and red discoloration of the urine. Macroscopic examination indicated that the stomach was severely affected at 1000 mg/kg bw/day as it was thickened, had an irregular surface and contained multiple foci.
Salivation, scabs and wounds were noted at 100 and 300 mg/kg bw/day. Body weight (gain) and food consumption was similar to control at 100 and 300 mg/kg bw/day. Macroscopic examination revealed no abnormalities at 300 mg/kg bw/day.

Based on the above described results of the 14-Day DRF with oral gavage administration of 3 (trimethoxysilyl)propane-1-thiol in rats (Test Facility Study No. 20244995) and prematurely terminated OECD 422 study (Test Facility Study No. 20244996), a dose level of 600 mg/kg bw/day was considered to be well tolerated. Therefore, this dose level was selected as the highest dose level for the current OECD 422 study (Test Facility Study No. 20281767). A mid- and low-dose level of 300 and 100 mg/kg bw/day were chosen in order to provide information on possible dose-response relationships.
Key result
Dose descriptor:
NOAEL
Remarks:
reproductive
Effect level:
>= 600 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on the absence of reproductive effects
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
body weight and weight gain
food consumption and compound intake
histopathology: non-neoplastic
Remarks on result:
other: See other information on results
Critical effects observed:
yes
Lowest effective dose / conc.:
600 mg/kg bw/day (nominal)
System:
urinary
Organ:
bladder
Treatment related:
yes
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Less milk, pale appearance and/or cold to touch was recorded in one or two litters at 600 mg/kg bw/day on PND 1, 2, 3 and/or 4. These clinical signs were considered not to be toxicologically relevant as these were only evident for a short duration (i.e. 1 or 2 days) and in a subset of pups from one or two litters at 600 mg/kg bw/day.
The nature and incidence of other clinical signs remained within the range considered normal for pups of this age and were therefore considered not to be related to treatment with the test item.
Dermal irritation (if dermal study):
not examined
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
The live birth indices were 100, 97, 98, 97% for the control, 100, 300 and 600 mg/kg bw/day groups, respectively.
Three pups (Litter No. 67) at 100 mg/kg bw/day, two pups (Litter No. 80) at 300 mg/kg bw/day, and three pups (Litter No. 95) at 600 mg/kg bw/day were found dead at the first litter check. At necropsy, no milk in the stomach was noted for all these dead pups, except for one dead pup from Litter No. 67 which was already partially cannibalized.
No toxicological relevance was attributed to these dead pups since the mortality incidence did not show a dose-related trend and remained within the range considered normal for pups of this age.

Viability indices were 100, 99, 100 and 97% for the control, 100, 300 and 600 mg/kg bw/day groups, respectively.
One pup (Litter No. 72) at 100 mg/kg bw/day and three pups (one in Litter No. 91 and two in Litter No. 95) at 600 mg/kg bw/day were missing on PND 2 or 3. These missing pups were most likely cannibalized.
Previous to death, the pup from Litter No. 72 displayed lethargy and less milk in the stomach at first litter check. The pup from Litter No. 91 presented with less milk in its stomach at PND 1, and both pups from Litter No. 95 were pale at PND 1 and/or 2 of which one contained less milk in its stomach on PND 2.
No toxicological relevance was attributed to these missing pups and clinical signs since the mortality incidence did not show a dose-related trend and remained within the range considered normal for pups of this age.

Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weights of pups were affected by treatment with the test item at 600 mg/kg bw/day, but were considered by the Registrants to be secondary to maternal systemic toxicity.

Mean pup body weights were decreased at 600 mg/kg bw/day in both sexes throughout the lactation period (not statistically significant on PND 1). At PND 13, mean pup body weights were 24, 22 and 23% lower than control in males, females or combined sexes, respectively.
Food consumption and compound intake (if feeding study):
not examined
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
Sexual maturation:
not examined
Anogenital distance (AGD):
no effects observed
Description (incidence and severity):
Anogenital distance (absolute and normalized for body weight) in male and female pups was considered not to be affected by treatment with the test item up to 600 mg/kg bw/day. All values remained within the normal range of biological variation.
Nipple retention in male pups:
no effects observed
Description (incidence and severity):
Treatment with the test item up to 600 mg/kg bw/day had no effect on areola/nipple retention. For none of the examined male pups nipples were observed at PND 13.
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No macroscopic findings were noted among pups that were considered to be related to treatment with the test item up to 600 mg/kg bw/day.
The nature and incidence of macroscopic findings remained within the range considered normal for pups of this age, and were therefore considered not to be related to treatment with the test item.
Histopathological findings:
not examined
Other effects:
no effects observed
Description (incidence and severity):
Serum T4 levels in male and female PND 14-16 pups were considered not to be affected by treatment with the test item up to 600 mg/kg bw/day.
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
Key result
Dose descriptor:
NOAEL
Remarks:
systemic
Generation:
F1
Effect level:
>= 600 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Lower pup body weight throughout the lactation period secondary to maternal systemic toxicity at 600 mg/kg bw/day.
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

As discussed under the histopathology results, the silicon industry considers diffuse urothelial hyperplasia in the bladder without atypia to be an adaptive (non-adverse) response to physical or chemical irritation based on a review of all available rat data across a wide range of silicon-based compounds including multiple triethoxysilanes. Thus, with the exception of the single female with bladder urothelial hyperplasia combined with degenerative bladder histopathology at 600 mg/kg bw/day, the Registrants consider the male and female bladder urothelial hyperplasia at 300 and 600 mg/kg bw/day to be non-adverse.

Conclusions:
In a combined repeated dose oral toxicity study with the reproduction / developmental toxicity screening test, with the registered substance 3-(trimethoxysilyl)propane-1-thiol and conducted according to OECD Test Guideline 422 and in compliance with GLP, the NOAEL for reproductive effects was concluded to be equal to or higher than 600 mg/kg bw/day based on no adverse effects on reproduction parameters. The NOAEL for parental systemic effects was concluded to be 300 mg/kg bw/day based on based on adverse clinical signs, decreased body weight, body weight gain, and/or food consumption in both sexes at 600 mg/kg bw/day. A single female at 600 mg/kg bw/day also had bladder urothelial hyperplasia combined with degenerative bladder histopathology.
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
600 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Reliability 1 study available
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The registered substance 3-(trimethoxysilyl)propane-1-thiol (CAS No. 4420-74-0, EC No. 224-588-5) has been evaluated in the combined repeated dose oral toxicity study with the reproduction/ developmental toxicity screening test, conducted according to OECD Test Guideline 422 and in compliance with GLP (Charles River Laboratories, 2021b, reliability score 1). Male and female Wistar Han rats were exposed to 0, 100, 300, 600 mg/kg bw/day administered daily via oral gavage in corn oil for a minimum of 28 days, followed by a 14-day recovery period.

 

Main and recovery males were treated for 29 days, including 2 weeks prior to mating, during mating, and up to and including the day before scheduled necropsy for main males. Females that delivered offspring were treated for 2 weeks prior to mating, during mating, during post-coitum, and 13-15 days of lactation (for a total of 51-65 days). Females that failed to deliver pups were treated for 41-52 days. Recovery females were treated during the same period as main females, until at least the day before the first scheduled necropsy of main females. The following parameters and endpoints were evaluated in this study: mortality/moribundity, clinical signs, functional observations, body weight and food consumption, oestrous cycle, clinical pathology, measurement of thyroid hormone T4 (F0‑males), gross necropsy findings, organ weights and histopathologic examinations. In addition, the following reproduction/developmental parameters were determined: mating and fertility indices, precoital time, number of implantation sites, gestation index and duration, parturition, maternal care, sex ratio and early postnatal pup development (mortality, clinical signs, body weights, sex, anogenital distance, areola/nipple retention and macroscopy, measurement of thyroid hormone T4 (post-natal day (PND) 14-16 pups)).

 

No reproduction toxicity was observed up to the highest dose level tested. No treatment-related changes were noted in any of the reproductive parameters investigated in this study (i.e., mating and fertility indices, precoital time, number of implantations, oestrous cycle, spermatogenic profiling, and histopathological examination of reproductive organs).

 

In this study with the registered substance 3-(trimethoxysilyl)propane-1-thiol, the reproductive NOAEL was concluded to be equal to or higher than 600 mg/kg bw/day. The parental systemic NOAEL was considered to be 300 mg/kg bw/day based on adverse clinical signs, decreased body weight, body weight gain, and/or food consumption in both sexes at 600 mg/kg bw/day. A single female at this dose also had bladder urothelial hyperplasia combined with degenerative bladder histopathology.

Effects on developmental toxicity

Description of key information

In the key study conducted according to OECD Test Guideline 414 and in compliance with GLP (Charles River Laboratories, 2021c, reliability score 1), pregnant Wistar Han rats were administered the analogue substance 3-(triethoxysilyl)propanethiol (CAS No. 14814-09-6, EC No. 238-883-1) at 0, 100, 300, and 1000 mg/kg bw/day via oral gavage (corn oil vehicle) during days 6 to 20 post-coitum, inclusive. The maternal systemic NOAEL was 300 mg/kg bw/day, based on clinical signs and decreased body weight / food consumption at 1000 mg/kg bw/day. The developmental NOAEL was 100 mg/kg bw/day based on increased post-implantation loss at 300 and 1000 mg/kg bw/day. Test item-related decreased foetal body weight and increased external and skeletal malformations were observed in the presence of maternal systemic effects at 1000 mg/kg bw/day.

 

Although there is an OECD Test Guideline 422 study for the registered substance 3-(trimethoxysilyl)propane-1-thiol (CAS No. 4420-74-0, EC No. 224-588-5), there are no developmental study data for this substance.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2020/08/24 - (in-life completion) 2020/10/22
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Remarks:
Han
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Deutschland, Sulzfeld, Germany
- Age at study initiation: 11-15 weeks
- Weight at study initiation: 192-272 g
- Fasting period before study: No
- Housing: Individually in Macrolon plastic cages (MIII type)
- Diet: Pelleted rodent diet (SM R/M-Z from SSNIFF), ad libitum
- Water: Municipal tap water, ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21
- Humidity (%): 51-65
- Air changes (per hr): 10 or more
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2020/09/29 and 2020/10/01 To: 2020/10/22
Route of administration:
oral: gavage
Vehicle:
corn oil
Remarks:
dried and deacidified
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Test item dosing formulations (w/w) were homogenized to visually acceptable levels at appropriate concentrations to meet dose level requirements. The dosing formulations were prepared at least weekly as a solution, filled out in daily portions and stored in the refrigerator.

The dosing formulations were removed from the refrigerator and stirred at room temperature for at least 30 minutes before dosing and dosed within 24 hours after removal from the refrigerator. Test item dosing formulations were kept at room temperature until dosing. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing.

Adjustment was made for specific gravity of the test item and vehicle. No correction was made for the purity/composition of the test item.

VEHICLE
- Justification for use and choice of vehicle: Trial preparations were performed at the Test Facility to select the suitable vehicle
- Concentration in vehicle: 0, 25, 75, 250 mg/ml
- Amount of vehicle (if gavage): 4 ml/kg bw dose volume
- Lot/batch no.: Not specified
- Purity: Not specified
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose formulation samples were collected for analysis during week 1, with samples for all dose groups analysed for concentration and samples for group 2 (100 mg/kg bw/day) and group 4 (1000 mg/kg bw/day) analysed for homogeneity. Duplicate sets of samples (approximately 500 mg) were sent to the analytical laboratory. Analyses were performed using a validated analytical procedure.
Details on mating procedure:
The female animals arrived at the Test Facility on day 0 or day 1 post-coitum, with day 0 post-coitum defined as the day of successful mating.
Duration of treatment / exposure:
Day 6 to 20 post-coitum, inclusive
Frequency of treatment:
Once daily
Duration of test:
Day 0 or 1 post-coitum through sacrifice on day 21 post-coitium
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
22F
Control animals:
yes
Details on study design:
- Dose selection rationale: Based on the results of the dose range finder (7.5.141)
- Rationale for animal assignment: Random
Maternal examinations:
MORTALITY: Yes
- Observed for general health/mortality and moribundity twice daily, in the morning and at the end of the working day

CAGE SIDE OBSERVATIONS: Yes
- Once daily, beginning on day 6 post-coitum up to the day prior to necropsy

DETAILED CLINICAL OBSERVATIONS: Yes
- Weekly, out of cage, beginning during pre-treatment period, and on the day of necropsy

BODY WEIGHT: Yes
- Animals were weighed individually on days 2, 6, 9, 12, 15, 18 and 21 post-coitum

FOOD CONSUMPTION: Yes
- Quantitatively measured for days 2-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post-coitum

WATER CONSUMPTION: Yes
- Monitored on regular basis throughout the study by visual inspection of the water bottles/containers

HAEMATOLOGY: No

CLINICAL CHEMISTRY: Yes
- All F0 animals (except those sacrificed in extremis or that delivered their offspring early), analysis for triiodothyronine (T3), thyroxine (T4), and thyroid-Stimulating Hormone (TSH), on the day of scheduled necropsy, animals were not fasted

SACRIFICE
- Animals surviving until scheduled euthanasia or those sacrificed in extremis were euthanized by a carbon dioxide inhalation (gradual fill) procedure
- Terminal procedures are summarized in Table 1 below

ORGAN WEIGHT: Yes
- Thyroid gland (paired) was weighed at necropsy for all animals surviving to scheduled euthanasia
- Ratios of organ to body weight (day 21 post-coitum) were calculated

GROSS PATHOLOGY: Yes
- All animals (except those sacrificed early or who delivered early) were subjected to an external, thoracic and abdominal examination, with special attention being paid to the reproductive organs
- Any macroscopic abnormalities were recorded, collected and fixed in 10% buffered formalin (neutral phosphate buffered 4% formaldehyde solution)

HISTOPATHOLOGY: Yes
- Thyroid glands of all animals were embedded in paraffin, sectioned, mounted on glass slides, and stained with hematoxylin and eosin. Thyroid glands that were supposed to be microscopically evaluated per protocol but were not available on the slide (and therefore not evaluated) are listed in the pathology report as not present. These missing tissues did not affect the outcome or interpretation of the pathology portion of the study because enough tissues were available.
- No macroscopic abnormalities were identified, thus no further tissues were subject to histopathology
Ovaries and uterine content:
Ovaries and uterine horn of all animals were dissected and content examined as quickly as possible after termination. Examinations included:
- Gravid uterus weight: Yes, for animals surviving to scheduled euthanasia
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Number of live and dead foetuses: Yes
Fetal examinations:
SACRIFICE
- Live foetuses euthanized by administration of sodium pentobarbital into the oral cavity using a small metal feeding tube
- Pups of females that delivered early (female no. 25) or required early sacrifice (female no. 88) were euthanized by decapitation

FOETAL EXAMINATION: Yes
- Litters of females surviving to scheduled necropsy: Detailed external, visceral and skeletal examinations, recorded as variations or malformations
- Pups of females that delivered early or required early sacrifice: Gross external exam

EXTERNAL EXAMINATION: Yes
- Each viable foetus was sexed, examined in detail to detect macroscopic visible abnormalities and their weight (not foetuses of female no. 88 sacrificed before planned necropsy) was determined
- Anogenital distance (AGD) was measured for all viable foetuses, normalized to the cube root of foetal body weight
- For late resorptions, a gross external examination was performed

VISCERAL EXAMINATION: Yes
- Sex of all foetuses was confirmed by internal examination and approximately one-half of the foetuses in each litter (all groups) were examined for visceral anomalies by dissection in the fresh (non-fixed) state.
- Thoracic and abdominal cavities were opened and dissected using a technique described by Stuckhardt and Poppe. This examination included the heart and major vessels. Foetal kidneys were also examined and graded for renal papillae development as described by Woo and Hoar.
- Abnormalities were collected and fixed in 10% buffered formalin at discretion of the Study Director
- Heads were removed from this one-half of the foetuses in each litter and placed in Bouin's solution for soft-tissue examination using the Wilson sectioning technique
- After examination, tissues with variations or malformations were stored in 10% formalin

SKELETAL EXAMINATION: Yes
- All foetuses were eviscerated, followed by fixation in 96% aqueous ethanol, and maceration in potassium hydroxide. Thereafter, they were stained with Alizarin Red S by a method similar to Dawson.
- Skeletal examination was done for one-half of the foetuses (i.e. the foetuses with heads)
- All specimens were archived in glycerin with bronopol as preservative
- A few bones were not available for skeletal examination because they were accidentally damaged or lost during processing, listed in the raw data, no influence on skeletal or overall study findings
Statistics:
Varied by endpoint / dataset evaluated and included:
-Descriptive: Means, standard deviations (or % coefficient of variation or standard error, when deemed appropriate), ratio, percentages, numbers, and/or incidences were reported as appropriate by dataset
- Parametric, non-parametric, incidence methods: Collectively, Levene’s test, one-way ANOVA F-test, Kruskal-Wallis test, Dunnett’s or Dunn’s test, analysis of covariance (ANCOVA), and Fisher’s exact test
- All statistical tests conducted at the 5% significance level
- All pairwise comparisons conducted using two sided tests and were reported at the 1% or 5% levels. Pairwise comparisons were made for: group 2 (100 mg/kg bw/day), group 3 (300 mg/kg bw/day), and group 4 (1000 mg/kg bw/day), each versus group 1 (control).
Indices:
MATERNAL:
- Pregnancy Rate (%): No. of pregnant females x 100 / No. of mated females

FOETAL:
- Male Foetuses (%): No. male foetuses x 100 / No. of foetuses
- Female Foetuses (%): No. female foetuses x 100 / No. of foetuses
- Pre-Implantation Loss (%): No. of corpora lutea – No. of implantations x 100 / No. of corpora lutea
- Post-Implantation Loss (%): No. of implants – No. of live foetuses x 100 / No. of implantations
- Litter % of Foetuses with Abnormalities: No. of foetuses in litter with a given finding x 100 / No. of foetuses in litter examined
Historical control data:
Yes, for this endpoint:
- Viable foetuses/dam, pregnant Wistar Han rats (period 2016-2020)
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Test item-related clinical signs that were considered adverse were seen at 1000 mg/kg bw/day. All other findings were considered not related to the test item, or not toxicologically relevant.

At 1000 mg/kg bw/day:
- Erect fur, all animals at 1000 mg/kg bw/day, from day 8 post-coitum onwards
- Hunched posture, 21/22 animals at 1000 mg/kg bw/day, from day 8 post-coitum onwards
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, treatment-related
Description (incidence):
One mortality was considered adverse. Female no. 88 (1000 mg/kg bw/day) was euthanized early on day 20 post-coitum, as an early delivery was suspected. Findings for this female included: red liquid (assumed blood) in vaginal area / cage, clinical signs from day 7 until day prior to euthanasia (erect fur, hunched posture and salivation, lower body weight gain compared to other 1000 mg/kg bw/day animals (- 86.8% on days 15-18 post-coitum), moderately decreased food consumption, and thyroid microscopic findings (minimal ultimobranchial, unilateral cyst and minimal bilateral follicular cell hypertrophy). Uterus content consisted of single live foetus and 12 early resorptions, with no empty implantation sites. No abnormalities seen at necropsy.

The early delivery (and euthanasia) of a second female (no. 25, 100 mg/kg bw/day) on day 21 post-coitum (i.e. the day of scheduled necropsy) was not considered test item-related.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Decreased body weight, body weight gain, and body weight gain corrected for gravid uterine weight were observed at 1000 mg/kg bw/day and considered test item-related and adverse. Corrected body weight gain at 100 and 300 mg/kg bw/day was unaffected by test item treatment.

At 1000 mg/kg bw/day:
- Slight mean body weight loss (-3.5%), days 6-9 post-coitum, with body weight loss (up to -4.8%) in 16/22 individual animals
- Significant lower mean body weight compared to controls, from day 9 post-coitum (up to 20% on day 21 post-coitum)
- Significantly lower mean body weight gain compared to controls, from day 9 post-coitum (up to 61% lower on days 18-21 post coitum)
- Based on body weight gain corrected for gravid uterus weight, significant body weight loss compared with controls (-132.0%)
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Lower food consumption was seen at 1000 mg/kg bw/day and considered test item-related and adverse. Food consumption at 300 and 100 mg/kg bw/day was considered unaffected by treatment.

At 1000 mg/kg bw/day:
- Lower food consumption compared to controls, from days 6-9 post-coitum (up to -41% over days 18-21 post-coitum)
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not specified
Description (incidence and severity):
Water consumption was monitored by visual inspection of water bottles/containers, results not reported
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Lower T3 and T4 serum levels were measured at 1000 mg/kg bw/day. This finding was considered test item-related, but the possible adversity could not be assessed in this type of study. The other T3 and T4 findings were not considered related to treatment. No effect on TSH.

At 1000 mg/kg bw/day:
- Significantly decreased mean total T3 and T4 serum levels compared to controls (0.45x and 0.47x of control, respectively)
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no test item-related alterations in thyroid gland weight.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no test item-related gross observations in the thyroid glands, with these findings in the background range for rats of this age and strain.

Other findings noted among control and/or treated animals were within the range of biological variation for rats of this age and strain and considered of no toxicological significance.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Except for an increase in thyroid follicular cell hypertrophy (considered non-adverse), there were no test item-related microscopic changes in the thyroid. All other thyroid findings were within the background range for rats of this age and strain.

At 1000 mg/kg bw/day:
- Increase incidence of follicular cell hypertrophy of the thyroid gland, all recorded at minimal degree and regarded non-adverse

See summary Table 2 below.
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Number of abortions:
not examined
Pre- and post-implantation loss:
effects observed, treatment-related
Description (incidence and severity):
Due to increased early resorptions, an increased post-implantation loss was estimated for 300 and 1000 mg/kg bw/day (no effect at 100 mg/kg bw/day). No effect on estimated pre-implantation loss.

At 1000 mg/kg bw/day:
- Increased mean post-implantation loss compared to controls (9.35% vs. 4.90% in controls)

At 300 mg/kg bw/day:
- Increased mean post-implantation loss compared to controls (8.51% vs. 4.90% in controls)
Total litter losses by resorption:
not examined
Early or late resorptions:
effects observed, treatment-related
Description (incidence and severity):
A slight test item-related increase in early resorptions was observed at 300 and 1000 mg/kg bw/day, considered adverse (no effect at 100 mg/kg bw/day). No test item effect on late resorptions.

At 1000 mg/kg bw/day:
- Slightly increased mean number of early resorptions (1.2 vs 0.6 in controls), not statistically significant
- Four females had higher number of early resorptions (4, 3, 3, 5)

At 300 mg/kg bw/day:
- Mean number of early resorptions slightly increased (0.9 vs 0.6 in controls), not statistically significant
- Mainly attributed to one animal (all 10 implantations died at a very early stage)
- Possible relation to test item treatment could not be excluded
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
No test item-related effect was observed on the number of pregnant females, with the number of females with viable litters equal to 21, 21, 20, 19 in the control, 100, 300 and 1000 mg/kg bw/day groups, respectively.

At scheduled necropsy, four females were non-gravid (1 control, 1 at 300 mg/kg bw/day, 2 at 1000 mg/kg/day). Two females did not survive to scheduled necropsy (1 at 1000 mg/kg bw/day due to early sacrifice on day 20 post-coitum, and 1 at 100 mg/kg bw/day due to early delivery on day 21 post-coitum. One female at 300 mg/kg bw/day had no live foetuses.

Mean numbers of corpora lutea and implantation sites were similar between the treated and control groups, and in the range of normal biological variation.
Other effects:
no effects observed
Key result
Dose descriptor:
NOAEL
Remarks:
general systemic effects
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
clinical signs
food consumption and compound intake
Remarks on result:
other: Based on clinical signs and decreased body weight / food consumption at 1000 mg/kg bw/day
Key result
Dose descriptor:
NOAEL
Remarks:
developmental
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
pre and post implantation loss
Remarks on result:
other: Based on increased post-implantation loss at 300 and 1000 mg/kg bw/day, used as basis of overall developmental NOAEL
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
uterus
Description (incidence and severity):
Increased post-implantation loss at 300 and 1000 mg/kg bw/day, used as basis of overall developmental NOAEL
Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
A test item-related decrease in foetal body weight was seen at 1000 mg/kg bw/day, considered adverse. No test item effect at 100 and 300 mg/kg bw/day.

At 1000 mg/kg bw/day:
- Significantly decreased mean foetal body weights compared to controls (males 20.5%, females 21.8%, combined foetal 21.5%)
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The male : female ratio was unaffected by treatment up to 1000 mg/kg bw/day.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
There were no test item-related effects on litter size up to 1000 mg/kg bw/day. Litter weight not reported, see foetal weight above.

Mean litter sizes were 11.2, 11.8, 11.1 and 10.3 foetuses/litter for the control, 100, 300, and 1000 mg/kg bw/day groups, respectively. All means remained within the biological variation for rats of this age and strain.
Changes in postnatal survival:
not examined
External malformations:
effects observed, treatment-related
Description (incidence and severity):
Test item-related external malformations occurred at 1000 mg/kg bw/day (considered adverse), but not at 300 or 1000 mg/kg bw/day. No test item-related external variations were noted up to 1000 mg/kg bw/day.

At 1000 mg/kg bw/day (^ considered adverse):
- Three affected foetuses from three litters had cleft palate (with underlying skeletal malformation) ^
- A fourth foetus with a malformation had ectrodactyly and syndactyly of a forepaw, not considered test item-related (chance finding due to the single occurrence)

See summary Tables 3 and 4 below.
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
Test item-related skeletal malformations were observed at 1000 mg/kg bw/day and were considered adverse. The single findings in the control and 300 mg/kg bw/day groups were considered unrelated to treatment. No findings at 100 mg/kg bw/day.

At 1000 mg/kg bw/day (6 foetuses, 6 litters), considered adverse:
- Three foetuses, underlying skeletal malformation (split palatine of an external malformation, cleft palate)
- Three other foetuses, malformations of the thoracic vertebra and/or ribs

At 300 mg/kg bw/day, considered unrelated to treatment:
- One foetus, malformation affecting the thoracic centrum
- Presence of one such malformation in a group is considered a chance finding

See summary Tables 3 and 4 below.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The observed visceral malformations were not considered test item-related.

At 100 and 1000 mg/kg bw/day:
- Two animals had visceral malformations, right-sided aortic arch at 100 mg/kg bw/day and fused liver lobe at 1000 mg/kg bw/day, not considered related to treatment

See summary Tables 3 and 4 below.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
ANOGENITAL DISTANCE:
Decreased anogenital distance at 1000 mg/kg bw/day, not considered direct result of test item treatment. No findings at 100 and 300 mg/kg bw/day.

At 1000 mg/kg bw/day:
- Significantly decreased mean anogenital distance for males compared to controls (6.8%)
- Considered secondary to lower foetal weights and not a direct effect of the test item since mean normalized anogenital distance not significantly changed
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
external malformations
skeletal malformations
Remarks on result:
other: Based on decreased foetal body weight and increased external and skeletal malformations at 1000 mg/kg bw/day. In the presence of maternal systemic effects.
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
external: face
external: thorax
skeletal: skull
skeletal: rib
Description (incidence and severity):
Based on external palate and underlying skeletal malformation of the split palatine bone and malformations of the thoracic vertebra and/or ribs at 1000 mg/kg bw/day. These findings were seen in multiple foetuses and multiple litters. In the presence of maternal systemic effects.
Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
300 mg/kg bw/day (nominal)
Treatment related:
yes
Relation to maternal toxicity:
developmental effects in the absence of maternal toxicity effects
Dose response relationship:
yes

Table 2. Summary test item-related microscopic findings – scheduled euthanasia animals

 

Females

Dose level (mg/kg bw/day):

0

100

300

1000

 

 

 

 

 

THYROID GLANDSa

21

21

21

21

   Follicular cell hypertrophy

 

 

 

 

      Minimal

3

2

4

10

a = Number of tissues examined from each group

 

Table 3. Number of foetuses examined (litters)

Dose level (mg/kg bw/day)

0

100

300

1000

External examination

236 (21)

248 (21)

222 (20)

195 (19)

Visceral examination

117 (20)

123 (21)

110 (19)

96 (19)

Skeletal examination

119 (21)

125 (21)

112 (20)

99 (19)

 

Table 4. Summary of malformations - individual descriptions

Dose Level (mg/kg bw/day)

Female No.

Foetus No.

Malformation(s) #

0

18

R11

Humerus, Bent (S)

100

33

L6

Aortic arch, Right-sided (V)a

300

50

R8

Thoracic centrum, 1 or more, Absent (S)a

1000

71

R9

Palate, Cleft Palate (E)

Palatine, Both, Split (S)

72

R6

Palate, Cleft Palate (E)

Palatine, Both, Split (S)

75

L6

Palate, Cleft Palate (E)

Palatine, Both, Split (S)

76

L2

Forepaw, Ectrodactyly (E)a

Forepaw, Syndactyly (E)a

Liver lobe, Fused (V)a

77

L1

Rib, 1 or more, Fused (S)

Sternebra, 1 or more, Absent (S)

Thoracic arch, 1 or more, Absent (S)

Thoracic arch, 1 or more, Fused (S)

Thoracic centrum, 1 or more, Absent (S)

Thoracic centrum, 1 or more, Fused (S)

79

R6

Thoracic vertebra, 1 or more, Supernumerary (S)

80

L2

Thoracic centrum, 1 or more, Absent (S)

#: Including external (E), visceral (V) and skeletal (S) examinations

a Not considered test item-related

Conclusions:
In the key study conducted according to OECD Test Guideline 414 and in compliance with GLP (reliability score 1), pregnant Wistar Han rats were administered the analogue substance 3-(triethoxysilyl)propanethiol at 0, 100, 300, and 1000 mg/kg bw/day via oral gavage (corn oil vehicle) during days 6 to 20 post-coitum, inclusive. The maternal systemic NOAEL was 300 mg/kg bw/day, based on clinical signs and decreased body weight / food consumption at 1000 mg/kg bw/day. The developmental NOAEL was 100 mg/kg bw/day based on increased percent post-implantation loss at 300 and 1000 mg/kg bw/day. Test item-related decreased foetal body weight and increased external and skeletal malformations were observed in the presence of maternal systemic effects at 1000 mg/kg bw/day.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
100 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Reliability 1 studies available
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

Developmental toxicity data are available from the OECD Test Guideline 414 study with the analogue substance 3-(triethoxysilyl)propanethiol (CAS No. 14814-09-6, EC No. 238-883-1) and the OECD Test Guideline 422 study with the registered substance 3-(trimethoxysilyl)propane-1-thiol (CAS No. 4420-74-0, EC No. 224-588-5).

 

The analogue substance 3-(triethoxysilyl)propanethiol has been evaluated in a study conducted according to OECD Test Guideline 414 and in compliance with GLP (Charles River Laboratories, 2021c, reliability score 1). Pregnant Wistar Han rats were administered 3-(triethoxysilyl)propanethiol at 0, 100, 300, and 1000 mg/kg bw/day via oral gavage (corn oil vehicle) during days 6 to 20 post-coitum inclusive. Observations and examinations were performed according to the guideline and, for the maternal (F0) animals included: cage side observations, detailed clinical signs, body weights, food consumption, thyroid hormone levels (triiodothyronine (T3), thyroxine (T4), thyroid-stimulating hormone (TSH)), thyroid and gravid uterine weights, macroscopic findings, histopathologic examination (thyroid gland), and uterine contents (number corpora lutea, number implantation sites, pre-/post-implantation loss, early/late resorptions, number/distribution of live/dead foetuses). In addition, these parameters were determined for the foetal (F1) generation: litter size, foetal body weights, sex ratio, anogenital distance (AGD), and malformations and variations (external, visceral and skeletal).

 

The findings considered test item-related and adverse are discussed below.

 

One maternal mortality at 1000 mg/kg bw/day (early sacrifice on post-coitum day 20 due to suspected early delivery) was considered test item-related. Clinical signs (erect fur, hunched posture) in this female were noted from day 7 until the day prior to sacrifice, with lower body weight / food consumption observed.

 

Test item-related and adverse maternal clinical signs (erect fur, hunched posture) were seen at 1000 mg/kg bw/day.

 

Decreased body weight, body weight gain, and body weight gain corrected for gravid uterine weight were observed at 1000 mg/kg bw/day and considered test item-related and adverse:

- Slight mean body weight loss (-3.5%), days 6-9 post-coitum, with body weight loss (up to 4.8% lower than controls) in individual animals

- Significant lower mean body weight compared to controls, from day 9 post-coitum (up to 20% on day 21 post-coitum)

- Significantly lower mean body weight gain compared to controls, from day 9 post-coitum (up to 61% lower on days 18-21 post coitum)

- Based on body weight gain corrected for gravid uterine weight, significant body weight loss compared with controls (-132.0%)

 

Mean food consumption was likewise lower at 1000 mg/kg bw/day, from days 6-9 post-coitum (up to -41% over days 18-21 post-coitum), with this effect considered test item-related and adverse.

 

Maternal clinical chemistry identified significantly decreased mean total T3 and T4 serum levels at 1000 mg/kg bw/day compared to controls (0.45x and 0.47x of control, respectively). This finding was considered test item-related, but the possible adversity could not be assessed in this type of study.

 

Test item-related and adverse maternal developmental findings were observed at 300 and 1000 mg/kg bw/day and included:

- Increased mean post-implantation loss compared to controls (8.51 and 9.35%, respectively, versus 4.90% in controls)

- Slightly increased mean number of early resorptions at 300 and 1000 mg/kg bw/day (0.9 and 1.2, respectively, versus 0.6 in controls), with 3 to 5 early resorptions for four individual animals

 

Test item-related and adverse findings in foetuses occurred at 1000 mg/kg bw/day (the maternal LOAEL) and consisted of:

- Significantly decreased mean foetal body weights compared to controls (males 20.5%, females 21.8%, combined foetal 21.5%)

- External malformations: cleft palate, with underlying skeletal malformation

- Skeletal malformations: split palatine of an external malformation (cleft palate) and thoracic and/or rib

 

In this study with the analogue substance 3-(triethoxysilyl)propanethiol, the maternal systemic NOAEL was 300 mg/kg bw/day, based on clinical signs and decreased body weight / food consumption at 1000 mg/kg bw/day. The developmental NOAEL was 100 mg/kg bw/day based on increased post-implantation loss at 300 and 1000 mg/kg bw/day. Test item-related decreased foetal body weight and increased external and skeletal malformations were observed in the presence of maternal systemic effects at 1000 mg/kg bw/day.

 

In the supporting study with the registered substance 3-(trimethoxysilyl)propane-1-thiol (combined repeated dose oral toxicity study with the reproduction/ developmental toxicity screening test), conducted according to OECD Test Guideline 422 and in compliance with GLP (Charles River Laboratories, 2021b, reliability score 1), male and female Wistar Han rats were exposed to 0, 100, 300, 600 mg/kg bw/day administered daily via oral gavage in corn oil for a minimum of 28 days, followed by a 14-day recovery period.

 

Main and recovery males were treated for 29 days, including 2 weeks prior to mating, during mating, and up to and including the day before scheduled necropsy for main males. Females that delivered offspring were treated for 2 weeks prior to mating, during mating, during post-coitum, and 13-15 days of lactation (for a total of 51-65 days). Females that failed to deliver pups were treated for 41-52 days. Recovery females were treated during the same period as main females, until at least the day before the first scheduled necropsy of main females. The following parameters and endpoints were evaluated in this study: mortality/moribundity, clinical signs, functional observations, body weight and food consumption, oestrous cycle, clinical pathology, measurement of thyroid hormone T4 (F0‑males), gross necropsy findings, organ weights and histopathologic examinations. In addition, the following reproduction/developmental parameters were determined: mating and fertility indices, precoital time, number of implantation sites, gestation index and duration, parturition, maternal care, sex ratio and early postnatal pup development (mortality, clinical signs, body weights, sex, anogenital distance, areola/nipple retention and macroscopy, measurement of thyroid hormone T4 (post-natal day (PND) 14-16 pups)).

Developmental findings were observed at 600 mg/kg bw/day but are considered by the Registrants to be secondary to the maternal toxicity at this dose. A lower mean post-implantation survival index and a trend towards smaller litter sizes were seen at 600 mg/kg bw/day in 3/9 litters. In addition, lower body weight of pups at 600 mg/kg bw/day occurred throughout the lactation, becoming more severe at the end of the lactation period compared to PND 1. No toxicologically significant changes were noted in any of the other developmental parameters investigated in this study (i.e., gestation, viability and lactation indices, duration of gestation, parturition, sex ratio, maternal care, and early postnatal pup development consisting of mortality, clinical signs, anogenital distance, areola/nipple retention, T4 thyroid hormone levels and macroscopic examination). In parental animals, adverse clinical signs, decreased body weight, body weight gain, and/or food consumption were seen in both sexes at 600 mg/kg bw/day, with a single female at 600 mg/kg bw/day having bladder urothelial hyperplasia combined with degenerative bladder histopathology.

 

Based on the study findings, the Registrants consider the developmental NOAEL for the registered substance 3-(trimethoxysilyl)propane-1-thiol to be greater than or equal to 600 mg/kg bw/day, with the developmental findings as secondary to the maternal systemic toxicity at this dose. Adverse parental effects at 600 mg/kg bw/day included clinical signs, decreased body weight, body weight gain, and/or food consumption, with a single female with bladder urothelial hyperplasia combined with degenerative bladder histopathology.

Justification for classification or non-classification

The conclusion for reproductive (developmental) toxicity classification will be confirmed once the final OECD Test Guideline 414 study report for the analogue substance 3-(triethoxysilyl)propanethiol and final OECD Test Guideline 422 study report for the registered substance 3-(triethoxysilyl)propane-1-thiol are available and in accordance with Regulation (EC) No. 1272/2008.

Additional information