4,4,13,13-tetraethoxy-3,14-dioxa-8,9-dithia-4,13-disilahexadecane

Toxicity to reproduction

Administrative data

Endpoint:

extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2 Dec 2020 (study initiation) to 9 Jul 2021 (in-life completion)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 443 (Extended One-Generation Reproductive Toxicity Study)

GLP compliance:

yes

Limit test:

no

Justification for study design:

- Premating exposure duration for parental (F0) animals: 10 weeks per OECD Test Guideline 443
- Basis for dose level selection: Based on a previous OECD Test Guideline 414 dose range finder in which the maternal and foetal NOAELs were at least 1000 mg/kg bw/day
- Inclusion of F1 Cohort 1B: Primarily to primary assess endocrine and reproductive tissues at necropsy, with tissues process through paraffin blocks for possible histopathology depending on the F1 Cohort 1A findings
- Neurotoxicity F1 Cohorts 1A and 1B (treated PNDs 21-90 or 97) Changes in gait and posture, presence of clonic or tonic movements, stereotypy or bizarre behaviour were assessed as part of the detailed clinical observations
- Immunotoxicity F1 Cohort 1A (treated PNDs 21-90): Spleen immunophenotyping was performed at necropsy
- Route of administration: Oral gavage selected since expected route of human exposure
- Species / strain: Han Wistar rat chosen since this strain is an accepted rodent species for non-clinical toxicity testing by regulatory agencies
- Vehicle: Justification not specifically specified, but the test item was shown to be stable and homogeneous in olive oil at 21 °C
- Number of animals: At (F1 cohorts, 20/sex/dose) or slightly higher than (F0 parents, 25/sex/dose) than specified in OECD Test Guideline 443

Test material

Test material information

Constituent 1

Reference substance name:

Reaction Mass of 4,4,15,15-tetraethoxy-3,16-dioxa-8,9,10,11-tetrathia-4,15-disilaoctadecane and 4,4,14,14-tetraethoxy-3,15-dioxa-8,9,10-trithia-4,14-disilaheptadecane and 4,4,13,13-tetraethoxy-3,14-dioxa-8,9-dithia-4,13-disilahexadecane

EC Number:

915-673-4

Cas Number:

211519-85-6

Molecular formula:

For the substance, C18H42O6SnSi2 (n = 2 – 4) Constituent 1 (S2): C18H42O6S2Si2 Constituent 2 (S3): C18H42O6S3Si2 Constituent 3 (S4): C18H42O6S4Si2

IUPAC Name:

Reaction Mass of 4,4,15,15-tetraethoxy-3,16-dioxa-8,9,10,11-tetrathia-4,15-disilaoctadecane and 4,4,14,14-tetraethoxy-3,15-dioxa-8,9,10-trithia-4,14-disilaheptadecane and 4,4,13,13-tetraethoxy-3,14-dioxa-8,9-dithia-4,13-disilahexadecane

Test material form:

liquid

Test animals

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI(Han)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River UK, Margate, Kent, UK
- Females: F0 sexually mature, F1 Cohort 1A and 1B aged PND 21 (assessed for sexual maturation)
- Age at study initiation (dosing): F0 males: 8-9 weeks, F0 females: 7-8 weeks; F1 Cohorts 1A and 1B: PND 21
- Weight at study initiation (dosing): F0 males: 228-342 g, F0 females: 142-204 g; F1 Cohort 1A males: 39-66 g, F1 Cohort 1A females: 33-68 g; F1 Cohort 1B males: 37-69 g, F1 Cohort 1B females: 32-63 g
- Fasting period before study: No
- Housing: For all animals, appropriately sized polycarbonate cages with solid bottoms, appropriate bedding provided
> F0, Cohort 1A, 1B animals: Up to 3/sex/dose together
> F0 males few days prior to mating: Transferred to individual solid bottomed cages
> F0 females for mating: Transferred to male individual cages
> F0 males after mating: Re-housed with their original cages mates until termination from Study Day (SD) 114.
> F0 females after either positive mating sign or at end mating period: Transferred to individual solid bottomed cages and retained in this type of cage until termination LDs 22-24. Appropriate nesting material provided
> F1 pups: Housed with their dams until litter culling on PND 4 (culled pups terminated), with F1 pups selected for F1 Cohorts 1A and 1B on PND 21 housed
- Diet, F0, F1 Cohorts 1A and 1B: Special Diet Services VRF-1, ad libitum except during designated procedures
- Water F0, F1 Cohorts 1A and 1B: Public supply tap water, ad libitum via water bottles except during designated procedures
- F1 pups: Nursing PNDs 0-21
- Acclimation period: Two weeks prior to start of dosing

ENVIRONMENTAL CONDITiONS
- Temperature (°C): 17-36
- Humidity (%): 30-129, excursions outside 30-70 transient, did not affect integrity or outcome of study
- Air changes (per hr): Ten (10) or more
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 1 Dec 2020 To: 9 Jul 2021

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Dose formulations were prepared at least weekly, with the aliquots removed from the freezer and allowed to warm to room temperature and stirred for at least 30 minutes before dosing. The doses were given via syringe with an attached gavage cannula. If a female was found to be in the process of littering or had recently littered at the time of scheduled dose administration, the animal was not dosed on that day.

VEHICLE
- Justification for use and choice of vehicle: Stability and homogeneity testing - Dose selection rationale: Based on OECD Test Guideline 414 dose range finder, with a maternal and foetal NOAELs of at least 1000 mg/kg bw/day (See “Justification for study design” above)
- Rationale for animal assignment: Random
- Fasting period before blood sampling for clinical biochemistry: No
- Dosing concentrations: 0, 25, 75, 250 mg/mL
- Amount of vehicle: Amount needed to derive dosing concentrations, dosing volume of 4 mL/kg for all F0 and F1 cohort groups.

Details on mating procedure:

F0 pairing was on a 1 male to 1 female basis. During the evening (after 5 pm) of Day 71 of dosing, females were housed with their allocated co-group male partner for up to 14 days. The day of detection of a copulatory plug in situ and/or of sperm in the lavage was designated Gestation Day 0. If evidence of mating was not observed by the end of the 14-day pairing period, the female was separated from the male the morning following the last night of pairing and treated as if mating had occurred during that night. F1 Cohort 1A and 1B animals were not mated.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For each dose/stratum as relevant, five formulation samples were collected for all groups on Day 1, Week 12 and Week 25, with two samples analysed via ultra-performance liquid chromatography (UPLC) using a validated analytical procedure. The control samples were obtained from the middle stratum, while those for the treated groups were take from the top, middle, and bottom strata.

Duration of treatment / exposure:

- F0, males: 10 weeks prior to mating and continuing throughout and after mating until the day before termination from SD 114
- F0, females: 10 weeks prior to mating and continuing throughout mating and gestation until at least Lactation Day (LD) 21
- F1 pups (PNDs 0-21) were potentially exposed in utero or during lactation
- F1 Cohort 1A: From PND 21 until the day before necropsy (at least PND 90)
- F1 Cohort 1B: From PND 21 until the day before necropsy (at least PND 97)

Frequency of treatment:

Once daily

Details on study schedule:

F1 litter standardisation, PND 4:
- Litter size was standardised to 8 pups per litter (and if possible, comprised of 4 males and 4 females). Any additional pups were selected at random for use in blood sample or necropsy procedures. Culled pups were terminated on PND 4, after (as relevant) any clinical chemistry.

Cohort 1A and 1B selection, F1 PND 21:
- Among the non-culled F1 pups, 40 males and 40 females were selected at random on PND 21 and identified on that day for post-weaning treatment and assessments (F1 Cohorts 1A and 1B), nominally up to 4 males and 4 females from each litter.
- Selected pups were removed from their mother on PND 21 and housed in their new cages
- Unselected pups remained with their dam until maternal (and pup) termination on LDs 22-24.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

- F0: 25/sex/dose
- F1 pups after PND 4 cull, 8 pups//litter, with ideally comprised of 4/sex//litter
- Cohort 1A: 20/sex/dose
- Cohort 1B: 20/sex/dose

Control animals:

yes

Details on study design:

- Dose selection rationale: Based on OECD Test Guideline 414 dose range finder, with a maternal and foetal NOAELs of at least 1000 mg/kg bw/day (See “Justification for study design” above)
- Rationale for animal assignment: Random
- Fasting period before blood sampling for clinical biochemistry: No

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Test item related but non-adverse F0 clinical signs were observed at 300 and 1000 mg/kg bw/day, but not at 100 mg/kg bw/day.

There was an increase in the incidence of:
- Ploughing at 1000 mg/kg bw/day
- Salivation at 300 and 1000 mg/kg bw/day

Both of these signs were predominantly observed immediately post-dose, with only a few instances at the 0-1 hour post-dosing timepoint. These findings were most likely due to the taste of the formulations, and thus were considered non-adverse.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Description (incidence):

There were no unscheduled deaths in F0 animals.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

No test item-related effects on body weight or body weight gain were observed in the F0 animals up to 1000 mg/kg bw/day. Any statistical significance seen was due to individual variation within dose groups.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

No test item-related effects on food consumption were identified in the F0 animals up to 1000 mg/kgbw/day.

Food efficiency:

not examined

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

no effects observed

Description (incidence and severity):

Water consumption by F0 animals was monitored by visual inspection of the water bottles. No inter-group differences were noted.

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

There were no effects on haematology parameters in the F0 animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the control group.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

There were no effects on clinical chemistry parameters in the F0 animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the control group.

Endocrine findings:

no effects observed

Description (incidence and severity):

No test item-related effects on T4 concentrations in plasma were observed in the F0 animals up to 1000 mg/kg bw/day. The F0 TSH findings will be added when available.

Urinalysis findings:

no effects observed

Description (incidence and severity):

There were no effects on urinalysis parameters in the F0 animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the control group.

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

For the F0 animals, there were no behaviour changes mentioned in the study results for these parameters: gait and posture, presence of clonic or tonic movements, stereotypy, or bizarre behaviour up 1000 mg/kg bw/day, as assessed as part of the detailed clinical examinations.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

In the F0 animals, there were test item-related and adverse effects in the kidney at 1000 mg/kg bw/day in both sexes and at 300 mg/kg bw/day in females, primarily based on the tubular basophilia. The collective renal findings across both doses included tubular basophilia (minimal to moderate), tubular dilatation (minimal to mild), mononuclear cell infiltration (minimal to mild), papillary cellular debris (minimal), mineralisation (minimal to mild), and (in males only and not relevant to humans) minimal to mild accumulation of hyaline droplets in cortical tubules.

Tubular basophilia comprised cortical tubules lined by epithelial cells with cytoplasm that was reduced in amount and/or more basophilic than normal, with nuclei that were larger than normal with dispersed chromatin and with occasional degenerate cells. There also was fibrosis in the basement membranes and surrounding interstitial tissue. Some foci included interstitial mononuclear cell infiltration and/or dilatation of basophilic tubules, that were recorded as separate findings. Papillary cellular debris comprised amorphous or globular, slightly eosinophilic deposits in the lumen of papillary tubules, and/or focal increased cellularity of the epithelium of individual papillary tubules.

Tubular basophilia in the kidney at the mild or moderate grades (considered adverse at mild or higher) occurred only at 1000 mg/kg bw/day in both sexes and at 300 mg/kg bw/day in females. Noting, the average grade of tubular basophilia at 300 and 1000 mg/kg bw/day was higher in females than in males, and papillary cellular debris was more prevalent in females at these doses.

Histopathological findings: neoplastic:

not examined

Other effects:

no effects observed

Description (incidence and severity):

Coagulation findings:
There were no effects on coagulation parameters in the F0 animals up to 1000 mg/kg bw/day. All values were considered to be within normal biological variation when compared with the control group.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

No test item-related effects on maternal oestrous cycles were identified for the F0 animals up to 1000 mg/kg bw/day.

Reproductive function: sperm measures:

no effects observed

Description (incidence and severity):

No test item-related effects on sperm motility, sperm or spermatid counts, or sperm morphology were seen in the F0 males up to 1000 mg/kg bw/day.

Reproductive performance:

no effects observed

Description (incidence and severity):

No test item-related effects on maternal reproduction performance or maternal care were observed in the F0 animals up to 1000 mg/kg bw/day, as based on male and female reproductive (mating, fertility, and pregnancy) indices, gestation length, female post-implantation loss, and on observed deficiencies in maternal care.

Effect levels (P0)

1

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

300 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

read-across polysulphides

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

Remarks on result:

other: Test item-related and adverse renal histopathology at 1000 mg/kg bw/day

2

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysuphides read across

Sex:

female

Basis for effect level:

histopathology: non-neoplastic

Remarks on result:

other: Test item-related and adverse renal histopathology from 300 mg/kg bw/day

3

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: Reproductive function and performance (males / females treated 10 weeks pre-mating until SD 114+ / LD 21)

Remarks on result:

other: Lack of test item-related effects up to 1000 mg/kg bw/day

Target system / organ toxicity (P0)

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

300 mg/kg bw/day (nominal)

System:

urinary

Organ:

kidney

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

F1 PUPS:
In the absence of mention in the study report results, the reviewer considers that milk was present in the F1 pup stomachs and there were no external gross abnormalities, based on daily observations through PND 4 and then PNDs 7, 14, and 21 up to 1000 mg/kg bw/day of parental F0 treatment.

F1 COHORTS:
Test item-related but non-adverse clinical signs were observed in the Cohorts 1A and 1B animals at 300 and 1000 mg/kg bw/day, but not at 100 mg/kg bw/day.

There was an increase in the incidence of salivation at 300 and 1000 mg/kg/day in the Cohort 1A and Cohort 1B animals.

This clinical sign was predominantly observed immediately post-dose, with only a few instances at the 0-1 hour post-dosing timepoint. The salivation was most likely due to the taste of the formulations, and thus considered non-adverse.

Dermal irritation (if dermal study):

not specified

Mortality / viability:

no mortality observed

Description (incidence and severity):

F1 PUPS AND COHORTS:
There were no unscheduled deaths in F1 pups or F1 Cohorts 1A and 1B animals up to 1000 mg/kg bw/day treatment (parental F0 or direct cohort).

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Test item-related but non-adverse effects on body weight and body weight gain were noted in male F1 pups at 1000 mg/kg bw/day. The effects in males at 100 and 300 mg/kg bw/day and in females up to 1000 mg/kg bw/day were considered unrelated to the test item.

At 1000 mg/kg bw/day, there was a 7% lower body weight gain for the F0 male pups when compared with the control group, and a 5% lower body weight by LD 21. The study report does not indicate this finding is adverse.

F1 COHORTS:
No test item-related effects on body weight or body weight gain were observed in the F1 Cohort 1A or Cohort 1B animals up to 1000 mg/kg bw/day. Any statistical significance seen was due to individual variation within the dose groups.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

F1 COHORTS:
No test item-related effects on food consumption were identified in the F1 Cohorts 1A or 1B animals up to 1000 mg/kg/day.

F1 PUPS:
Not relevant (nursing).

Food efficiency:

not examined

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

no effects observed

Description (incidence and severity):

F1 COHORTS:
Water consumption by F1 Cohort 1A and 1B animals was monitored by visual inspection of the water bottles. No inter-group differences were noted.

F1 PUPS:
Not relevant (nursing).

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

F1 COHORT 1A:
There were no effects on haematology parameters in the F1 Cohort 1A animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the controls.

F1 PUPS AND F1 COHORT 1B:
Not examined.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

There were no effects on clinical chemistry parameters in the F1 Cohort 1A animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the controls.

F1 PUPS AND F1 COHORT 1B:
Not examined.

Urinalysis findings:

no effects observed

Description (incidence and severity):

F1 COHORT 1A:
There were no effects on urinalysis parameters in the F1 Cohort 1A animals up to 1000 mg/kg bw/day. All values were within normal biological variation when compared with the controls.

F1 PUPS AND F1 COHORT 1B:
Not examined.

Sexual maturation:

no effects observed

Description (incidence and severity):

F1 COHORTS:
There were no test-item related effects on sexual maturation or the time to first oestrous cycle in F1 Cohorts 1A or 1B up to 1000 mg/kg bw/day.

F1 PUPS:
Not relevant / examined.

Anogenital distance (AGD):

no effects observed

Description (incidence and severity):

F1 PUPS:
No test item-related effects on AGD in either sex were observed on PND 1 up to 1000 mg/kg bw/day of parental F0 treatment..

F1 COHORTS:
Not relevant / examined for the F1 Cohort 1A and 1B animals.

Nipple retention in male pups:

no effects observed

Description (incidence and severity):

F1 PUPS:
No test item-related effects on nipple retention in male pups were observed on PND 13 up to 1000 mg/kg bw/day of parental F0 exposure.

F1 COHORTS 1A and 1B:
Not relevant / examined for the F1 Cohort 1A and 1B animals.

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

F1 PUPS:
No test item-related mean organ weight differences were seen in the F1 pups up to 1000 mg/kg bw/day of parental F0 treatment, as compared to the control group. The kidneys were not required to be weighed in these pups.

F1 COHORT 1A:
Test item-related but non-adverse mean kidney and liver weight findings were noted in females at 300 and 1000 mg/kg bw/day. The findings in males up to 1000 mg/kg bw/day and in females at 100 mg/kg bw/day were assessed as unrelated to the test item.

For the kidney, the mean weight was higher at 300 and 1000 mg/kg bw/day in females, correlating with the minimal or mild tubular basophilia seen histologically (adverse at a mild grade at 1000 mg/kg bw/day). The maximum difference from controls in weight relative to body weight was +15% in females at 1000 mg/kg bw/day.

The mean liver weight was higher at all doses in males and at 300 and 1000 mg/kg bw/day in females, correlating with the hepatocellular basophilia and hypertrophy seen histologically (examined at 1000 mg/kg bw/day). The maximum difference from controls in weight relative to body weight was +15% in females.

The kidney and liver weight findings are not identified as adverse in the study or pathology reports.

Please see Table 16 below.

F1 COHORT 1B:
There were test item-related but non-adverse mean kidney weights in females at 1000 mg/kg bw/day, liver weights in females at all doses, and liver weights in males at 1000 mg/kg bw/day. Male kidney weights up to 1000 mg/kg bw/day and all organ weights, except for the female liver, at 100 and 300 mg/kg bw/day were considered unrelated to the test item.

For the kidney, the mean weight was higher in females at 1000 mg/kg bw/day. The maximum difference from controls in weight relative to body weight was +13% at this dose.

The mean liver weight was higher in females at all doses and in males at 1000 mg/kg bw/day. The maximum difference from controls in weight relative to body weight was +21% in females at 1000 mg/kg bw/day.

These kidney and liver weight findings are not identified as adverse in the study or pathology reports. Noting that, per protocol, F1 Cohort 1B tissue samples were collected and processed to paraffin blocks, but not subject to microscopic examination.

Please see Table 17 below.

Gross pathological findings:

no effects observed

Description (incidence and severity):

F1 PUPS: [no effects observed]
There were no gross abnormalities in the pups culled / necropsied on PND 4 or in the F1 unselected pups necropsied on PNDs 22-24.

F1 COHORTS:
No test item-related gross findings were identified for the F1 Cohort 1A and 1B animals up to 1000 mg/kg bw/day. Any findings were generally of the nature commonly observed in this strain of rat at the ages concerned, most were infrequent, and none had a microscopic correlate.

Histopathological findings:

effects observed, treatment-related

Description (incidence and severity):

F1 COHORT 1A:
In the F1 Cohort 1A animals, there were test item-related and adverse effects in the kidney at 1000 mg/kg bw/day in females, primarily based on minimal to mild tubular basophilia (considered adverse at mild or higher). At this dose, the collective kidney findings across both sexes included tubular basophilia (minimal to mild), tubular dilatation (minimal to mild), mononuclear cell infiltration (minimal), papillary cellular debris (minimal), medullary mineralisation (minimal) and, in males only and not relevant to humans, minimal accumulation of hyaline droplets in cortical tubules.

Test item-related increases in tubular basophilia and medullary mineralisation occurred at 300 and 1000 mg/kg bw/day (in a dose level-related fashion). Tubular basophilia was assigned minimal or mild grade (adverse at mild or higher), indicating this finding was less severe than in the F0 animals. Medullary mineralisation comprised basophilic, non-birefringent amorphous deposits in the epithelium of, or between, tubules of the medulla, occasionally involving the outer papilla. The other kidney findings were increased at 1000 mg/kg bw/day only. Tubular basophilia at the mild grade and papillary cellular debris at the minimal grade occurred only in females at 1000 mg/kg bw/day indicating that the kidney findings were more apparent in females, as also evident in the F0 animals.

F1 PUPS AND F1 COHORT 1B:
Not examined, noting that F1 Cohort tissue samples were collected and processed to paraffin blocks for possible microscopic examination.

Other effects:

no effects observed

Description (incidence and severity):

COAGULATION FINDINGS:
F1 COHORT 1A: [no effects observed]
There were no effects on coagulation parameters in the F1 Cohort 1A animals up to 1000 mg/kg bw/day. All values were considered to be within normal biological variation when compared with the controls.

F1 PUPS AND F1 COHORT 1B:
Not examined.

ENDOCRINE FINDINGS:
F1 PUPS AND COHORTS:
No test item-related effects on T4 concentrations in plasma were observed in the F1 pups or in the Cohorts 1A or 1B animals up to 1000 mg/kg bw/day (parental F0 or cohort). The cohort animal TSH findings will be added when available. Samples for TSH analysis were not collected from the F1 pups.

OESTRUS CYCLE FINDINGS:
F1 COHORT 1A:
There were no test item-related effects on maternal oestrous cycles for the Cohort 1A animals up to 1000 mg/kg bw/day.

F1 AND F1 COHORT 1B:
Not examined.

SPERM FINDINGS:
F1 Cohort 1A:
There were no test item-related effects on sperm motility, sperm or spermatid counts, or sperm morphology in the F1 Cohort 1A males up to 1000 mg/kg bw/day.

F1 PUPS AND F1 COHORT 1B:
Not examined.

OVARIAN FOLLICLE COUNTS:
F1 COHORT 1A:
There were no test item-related effects on the ovarian follicle counts in the F1 Cohort 1A females up to 1000 mg/kg bw/day.

F1 AND F1 COHORT 1B:
Not examined.

Developmental neurotoxicity (F1)

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

F1 COHORTS:
For the F1 Cohort 1A and 1B animals, no behaviour changes were mentioned in the study results for these parameters: gait and posture, presence of clonic or tonic movements, stereotypy, or bizarre behaviour up 1000 mg/kg bw/day, as assessed as part of the detailed clinical examinations.

F1 PUPS:
Not examined, see clinical signs for the F1 pups.

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

no effects observed

Description (incidence and severity):

F1 COHORT 1A:
No test item-related effects on immunophenotypic expression in the spleen were observed in F1 Cohort 1A animals treated up to 1000 mg/kg bw/day, as compared to the control group. No dose- or sex-dependent effects were observed for any immune cell populations, as expressed as a percentage of total lymphocytes.

F1 PUPS AND F1 COHORT 1B:
Not examined.

Effect levels (F1)

1

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Generation:

F1

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: F1 pup systemic (potentially exposed in utero / during lactation PNDs 0-21)

Remarks on result:

other: Lack of test item-related and adverse effects up to 1000 mg/kg bw/day of parental treatment

2

Key result

Dose descriptor:

NOAEL

Remarks:

reproductive / developmental

Generation:

F1

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: F1 pup reproductive function / developmental (potentially exposed in utero / during lactation PNDs 0-21)

Remarks on result:

other: Lack of test item-related effects up to 1000 mg/kg bw/day of parental treatment.

3

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Generation:

F1 (cohort 1A)

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male

Basis for effect level:

other: Systemic (treated PNDs 21-90)

Remarks on result:

other: Lack of test item-related and adverse effects up to 1000 mg/kg bw/day

4

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Generation:

F1 (cohort 1A)

Effect level:

300 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

female

Basis for effect level:

histopathology: non-neoplastic

Remarks on result:

other: Test item-related and adverse renal histopathology at 1000 mg/kg bw/day

5

Key result

Dose descriptor:

NOAEL

Remarks:

reproductive

Generation:

F1 (cohort 1A)

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: Reproductive function (treated PNDs 21-90)

Remarks on result:

other: Lack of test item-related effects up to 1000 mg/kg bw/day

6

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Generation:

F1 (cohort 1B)

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: Systemic (treated PNDs 21-97)

Remarks on result:

other: Lack of test item-related and adverse effects up to 1000 mg/kg bw/day

7

Key result

Dose descriptor:

NOAEL

Remarks:

reproductive

Generation:

F1 (cohort 1B)

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

polysulphides read across

Sex:

male/female

Basis for effect level:

other: Reproductive function (treated PNDs 21-97)

Remarks on result:

other: Lack of test item-related effects up to 1000 mg/kg bw/day

Target system / organ toxicity (F1)

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

1 000 mg/kg bw/day (nominal)

System:

urinary

Organ:

kidney

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Overall reproductive toxicity

Key result

Reproductive effects observed:

no

Any other information on results incl. tables

Table 14. F0: Summary group mean organ weight data at scheduled euthanasia

	Males				Females
Group:	1	2	3	4	1	2	3	4
Dose (mg/kg/day):	0	100	300	1000	0	100	300	1000
No. animals examined:	25	25	25	25	25	25	25	25
Terminal Body Weight (No. weighed)	(25)	(25)	(25)	(25)	(21)	(23)	(23)	(23)
Absolute value (g)	490	499	478	465	284	277	281	287
Kidney (No. weighed)	(25)	(25)	(25)	(25)	(21)	(23)	(23)	(24)
Absolute value (g)	2.82	2.81	2.74	3.14**	2.16	2.08	2.28	2.76++
% of body weight	0.576	0.567	0.576	0.675**	0.758	0.755	0.812**	0.963**
% of brain weight	132	131	129	147	110	107	117	144
Liver (No. weighed)	(25)	(25)	(25)	(25)	(21)	(23)	(23)	(24)
Absolute value (g)	16.9	18.0	17.7	18.4	15.8	15.6	16.1	17.9**
% of body weight	3.43	3.61	3.69*	3.94**	5.56	5.65	5.74	6.21++
% of brain weight	787	841	831	863	806	805	826	930

Anova & Dunnett: * = p ≤ 0.05; ** = p ≤ 0.01

Kruskal-Wallis & Dunn: ++ = p ≤ 0.01

% of brain weight not analysed statistically

Group mean values are rounded to 3 significant figures. Statistical significance is based on unrounded data.

 

Table 15. F0: Summary group test item-related microscopic findings at scheduled euthanasia

	Males				Females
Group:	1	2	3	4	1	2	3	4
Dose (mg/kg/day):	0	100	300	1000	0	100	300	1000
No. animals examined:	25	25	25	25	25	25	25	25
Kidney (No. Examined)	(25)	(25)	(25)	(25)	(25)	(25)	(25)	(25)
Tubular basophilia^a	10	7	20	25	0	2	25	25
Minimal	10	7	20	6	0	2	15	0
Mild	0	0	0	18	0	0	10	17
Moderate	0	0	0	1	0	0	0	8
Mononuclear cell infiltration	13	18	19	25	5	7	25	25
Minimal	13	18	19	24	5	7	25	25
Mild	0	0	0	1	0	0	0	0
Tubular dilatation	6	9	11	18	1	0	3	24
Minimal	5	9	11	16	1	0	3	22
Mild	1	0	0	2	0	0	0	2
Cellular debris, papilla, minimal	0	0	1	5	0	0	17	25
Mineralisation, papilla	1	2	1	8	2	3	5	6
Minimal	1	2	1	7	2	3	5	6
Mild	0	0	0	1	0	0	0	0
Accumulation, cortical, hyaline droplets	0	10	7	17	0	0	0	0
Minimal	0	8	4	9	0	0	0	0
Mild	0	2	3	8	0	0	0	0
Liver (No. Examined)	(25)	(5)	(9)	(25)	(25)	(1)	(1)	(25)
Centrilobular hepatocellular hypertrophy, minimal	0	0	0	4	2	0	0	14
Increased hepatocellular basophilia, minimal	1	0	0	6	2	0	0	12
Jejunum (No. Examined)	(25)	–	–	(25)	(25)	–	–	(25)
Villous vacuolation	0	–	–	21	0	–	–	20
Minimal	0	–	–	12	0	–	–	15
Mild	0	–	–	8	0	–	–	5
Moderate	0	–	–	1	0	–	–	0
Ileum (No. Examined)	(25)	–	–	(25)	(25)	–	–	(25)
Villous vacuolation	0	–	–	2	0	–	–	0
Mild	0	–	–	1	0	–	–	0
Moderate	0	–	–	1	0	–	–	0
Mesenteric lymph node (No. Examined)	(25)	–	–	(25)	(25)	(2)	(2)	(25)
Vacuolation	0	–	–	5	0	0	1	9
Minimal	0	–	–	5	0	0	1	3
Mild	0	–	–	0	0	0	0	5
Moderate	0	–	–	0	0	0	0	1
Gut-associated lymphoid tissue (No. Examined)	(24)	–	–	(23)	(23)	–	–	(23)
Vacuolation, minimal	0	–	–	0	0	–	–	1

^a Reported as adverse for males at 1000 mg/kg bw/day and females from 300 mg/kg bw/day

Table 16. F1 Cohort 1A: Summary group mean organ weight data at scheduled euthanasia

	Males				Females
Group:	1	2	3	4	1	2	3	4
Dose (mg/kg/day):	0	100	300	1000	0	100	300	1000
No. animals examined:	20	20	20	20	20	20	20	20
Terminal Body Weight (No. weighed)	(20)	(19)	(19)	(20)	(20)	(20)	(20)	(20)
Absolute value (g)	414	405	396	384	223	219	220	225
Kidney (No. weighed)	(20)	(20)	(20)	(20)	(20)	(20)	(20)	(20)
Absolute value (g)	2.75	2.64	2.55	2.73	1.61	1.61	1.68	1.87**
% of body weight	0.665	0.656	0.643	0.712	0.723	0.738	0.762*	0.828**
% of brain weight	133	129	124	133	85.9	86.8	89.2	99.2
Liver (No. weighed)	(20)	(20)	(20)	(20)	(20)	(20)	(20)	(20)
Absolute value (g)	16.6	17.3	17.0	17.7	8.92	8.89	9.40	10.4**
% of body weight	4.02	4.25*	4.25*	4.59**	4.01	4.06	4.27*	4.62**
% of brain weight	805	842	825	862	475	478	500	554

Anova & Dunnett: * = p ≤ 0.05; ** = p ≤ 0.01

% of brain weight not analysed statistically

Group mean values are rounded to 3 significant figures. Statistical significance is based on unrounded data.

Table 17. F1 Cohort 1B: Summary group mean organ weight data at scheduled euthanasia

	Males				Females
Group:	1	2	3	4	1	2	3	4
Dose (mg/kg/day):	0	100	300	1000	0	100	300	1000
No. animals examined:	20	20	20	20	20	20	20	20
Terminal Body Weight (No. weighed)	(20)	(20)	(20)	(20)	(20)	(20)	(20)	(19)
Absolute value (g)	414	422	413	387	226	223	226	226
Kidney (No. weighed)	(20)	(20)	(20)	(20)	(20)	(20)	(20)	(20)
Absolute value (g)	2.68	2.60	2.65	2.66	1.62	1.64	1.67	1.84**
% of body weight	0.649	0.617	0.645	0.686	0.718	0.738	0.738	0.816**
% of brain weight	130	127	130	130	86.5	87.5	88.5	98.1
Liver (No. weighed)	(20)	(20)	(20)	(20)	(20)	(20)	(20)	(20)
Absolute value (g)	16.2	17.2	16.8	17.1	8.77	9.41	9.44	10.6**
% of body weight	3.91	4.07	4.08	4.41**	3.87	4.23*	4.16*	4.66**
% of brain weight	785	840	822	839	467	502	500	565

Anova & Dunnett: * = p ≤ 0.05; ** = p ≤ 0.01

% of brain weight not analysed statistically

Group mean values are rounded to 3 significant figures. Statistical significance is based on unrounded data.

Table 18. F1 Cohort 1A: Summary test item-related microscopic findings at scheduled euthanasia

	Males				Females
Group	1	2	3	4	1	2	3	4
Dose (mg/kg/day)	0	100	300	1000	0	100	300	1000
No. animals examined	25	25	25	25	25	25	25	25
Kidney (No. Examined)	(25)	(25)	(25)	(25)	(25)	(25)	(25)	(25)
Tubular basophilia^a	10	7	20	25	0	2	25	25
Minimal	10	7	20	6	0	2	15	0
Mild	0	0	0	18	0	0	10	17
Moderate	0	0	0	1	0	0	0	8
Mononuclear cell infiltration	13	18	19	25	5	7	25	25
Minimal	13	18	19	24	5	7	25	25
Mild	0	0	0	1	0	0	0	0
Tubular dilatation	6	9	11	18	1	0	3	24
Minimal	5	9	11	16	1	0	3	22
Mild	1	0	0	2	0	0	0	2
Cellular debris, papilla, minimal	0	0	1	5	0	0	17	25
Mineralisation, papilla	1	2	1	8	2	3	5	6
Minimal	1	2	1	7	2	3	5	6
Mild	0	0	0	1	0	0	0	0
Accumulation, cortical, hyaline droplets	0	10	7	17	0	0	0	0
Minimal	0	8	4	9	0	0	0	0
Mild	0	2	3	8	0	0	0	0
Liver (No. Examined)	(25)	(5)	(9)	(25)	(25)	(1)	(1)	(25)
Centrilobular hepatocellular hypertrophy, minimal	0	0	0	4	2	0	0	14
Increased hepatocellular basophilia, minimal	1	0	0	6	2	0	0	12
Jejunum (No. Examined)	(25)	–	–	(25)	(25)	–	–	(25)
Villous vacuolation	0	–	–	21	0	–	–	20
Minimal	0	–	–	12	0	–	–	15
Mild	0	–	–	8	0	–	–	5
Moderate	0	–	–	1	0	–	–	0
Ileum (No. Examined)	(25)	–	–	(25)	(25)	–	–	(25)
Villous vacuolation	0	–	–	2	0	–	–	0
Mild	0	–	–	1	0	–	–	0
Moderate	0	–	–	1	0	–	–	0
Mesenteric lymph node (No. Examined)	(25)	–	–	(25)	(25)	(2)	(2)	(25)
Vacuolation	0	–	–	5	0	0	1	9
Minimal	0	–	–	5	0	0	1	3
Mild	0	–	–	0	0	0	0	5
Moderate	0	–	–	0	0	0	0	1
Gut-associated lymphoid tissue (No. Examined)	(24)	–	–	(23)	(23)	–	–	(23)
Vacuolation, minimal	0	–	–	0	0	–	–	1

^a Reported as adverse for females at 1000 mg/kg bw/day

Applicant's summary and conclusion

Conclusions:

In the read-across EOGRTS for bis[3-(triethoxysilyl)propyl]polysulfides conducted according to OECD Test Guideline 443 and in compliance with GLP, four groups of Wistar Han rats were investigated: F0 (parental), F1 pups, F1 Cohort 1A, and F1 Cohort 1B, with the latter added to primarily assess endocrine and reproductive tissues via necropsy and organ weights. The F0 and cohort groups were administered the test substance at 0, 100, 300, and 1000 mg/kg bw/day (corn oil vehicle), with the F1 pups potentially exposed in utero or during lactation. Based on the findings, the overall study NOAELs were identified as follows:
- Systemic, males: 300 mg/kg bw/day, based on test item-related and adverse kidney histopathology in F0 males at 1000 mg/kg bw/day
- Systemic, females: 100 mg/kg bw/day, based on test item-related and adverse kidney histopathology in F0 females at 300 mg/kg bw/day
- Reproduction: at least 1000 mg/kg bw/day, based on the lack of test item-related: reproductive effects in the F0, F1 Cohort 1A, and F1 Cohort 1B animals, and reproductive / developmental effects in the F1 pups.