Registration Dossier

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
07 November 2016 to 09 February 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
1996
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OPPTS 870.3650, Combined Repeated Dose Toxicity Study with the Reproduction/Developm ental Toxicity Screening Test
Version / remarks:
2000
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
1995
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OPPTS 870.3550, Reproduction/Developmental Toxicity Screening Test
Version / remarks:
2000
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents
Version / remarks:
2000
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Octaphenylcyclotetrasiloxane
EC Number:
208-904-9
EC Name:
Octaphenylcyclotetrasiloxane
Cas Number:
546-56-5
Molecular formula:
C48H40O4Si4
IUPAC Name:
octaphenyl-1,3,5,7,2,4,6,8-tetraoxatetrasilocane
Test material form:
other: white powder
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature
- Stability under test conditions: stable until 31 March 2018
- Solubility and stability of the test substance in the solvent/vehicle: Stability in propylene glycol for at least 24 hours at room temperature and for at least 8 days in a refrigerator is confirmed over the concentration range 1 to 200 mg/mL

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Formulations (w/w) were prepared daily within 6 hours prior to dosing and were homogenized to a visually acceptable level. Adjustment was made for specific gravity of the vehicle. No correction was made for the purity/composition of the test item.
- Preliminary purification step (if any): No correction factor required.
- Final dilution of a dissolved solid, stock liquid or gel: not applicable
- Final preparation of a solid: not applicable

FORM AS APPLIED IN THE TEST (if different from that of starting material): diluted in propylene glycol

Test animals

Species:
rat
Strain:
other: Crl:WI(Han)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: males: approximately 10 weeks; females: approximately 11 weeks
- Weight at study initiation: not specified
- Fasting period before study: no
- Housing: Animals were housed in groups of 5 animals/sex/cage in Macrolon plastic cages during premating period. This is also applicable for Recovery animals throughout the complete study. During mating period Main females were caged together with Main males on a one-to-one-basis in Macrolon plastic cages. At post-mating period Main males were housed in their home cage (Macrolon plastic cages, MIV type, height 18 cm) with a maximum of 5 animals/cage. Main females were individually housed in Macrolon plastic cages. During lactation Main females were housed in Macrolon plastic cages (MIII type, height 18 cm). Pups were housed with the dam, except during locomotor activity monitoring of the dams, when the pups were kept warm in their home cage using bottles filled with warm water.
- Diet (e.g. ad libitum): pelleted rodent diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 5 days
DETAILS OF FOOD AND WATER QUALITY: Diet, water, bedding and cage-enrichment/nesting material evaluation for contaminants and/or nutrients was performed according to facility standard procedures. There were no findings that could interfere with the study.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C,
- Humidity (%): 40 to 70%
- Air changes (per hr): 10 room air changes/hour
- Photoperiod (hrs dark / hrs light): 12-hour light/12-hour dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
propylene glycol
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Formulations (w/w) were prepared daily within 6 hours prior to dosing and were homogenized to a visually acceptable level. Adjustment was made for specific gravity of the vehicle. No correction was made for the purity/composition of the test item.
VEHICLE
- Justification for use and choice of vehicle (if other than water): Based on trial formulations performed at Charles River Den Bosch.
- Concentration in vehicle: not specified
- Amount of vehicle (if gavage): not specified
- Lot/batch no. (if required): not specified
- Purity: not specified
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: minimum of 14 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. Not applicable
- Further matings after two unsuccessful attempts: not specified
- After successful mating each pregnant female was caged (how): Main females were individually housed in Macrolon plastic cages.
- Any other deviations from standard protocol: none
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%. Formulations were considered stable if the relative difference before and after storage was maximally 10%.
Duration of treatment / exposure:
Main and Recovery males: 29 to 30 days
Main females with offspring: 41-45 days
Main females that failed to deliver: 41 to 45 days
Recovery females: 44 days
Frequency of treatment:
Once daily for 7 days per week
Details on study schedule:
- F1 animals not mated.
- Selection of parents from F1 generation when pups were [...] days of age. Not applicable
- Age at mating of the mated animals in the study: 12-13 weeks
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
Main males: 10
Main females: 10
Recovery animals: 5
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Based on the results of a 14-day dose range finding study in which no signs of toxicity were observed up to 1000 mg/kg, the dose levels for this combined 28-day oral gavage study with reproduction/developmental toxicity screening test were selected to be 100, 300 and 1000 mg/kg.
- Rationale for animal assignment (if not random): random
Positive control:
Not used.

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Cage side observations checked included: The animals were checked for mortality and viability at least twice daily.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: At least once daily from start of treatment onwards up to the day prior to necropsy, detailed clinical observations were made for all animals. The following functional observations tests were performed on each individual animal of the selected 5 Main animals/sex/group and all Recovery animals: hearing ability, pupillary reflex, fore- and hind-limb grip strength, locomotor activity. The selected Main males and all Recovery males were tested during Week 4 of treatment and the selected Main females were tested once during the last week of lactation (from lactation Day 4 onwards), and all Recovery females were tested on the first day a Main female was tested. Motor activity measurements were also conducted for all Recovery males at the end of the Recovery period since a potential increase in motor activity was recorded for males at 300 and 100 mg/kg at the end of the treatment period.
BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of exposure (prior to first dosing) and weekly thereafter. Mated Main females were weighed on Days 0, 4, 7, 11, 14, 17 and 20 postcoitum and during lactation on PND 1 and 4.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): not applicable
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not applicable
Oestrous cyclicity (parental animals):
Not examined
Sperm parameters (parental animals):
Parameters examined in F0 male parental generations: testis weight, epididymis weight
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

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

GROSS EXAMINATION OF DEAD PUPS: yes, for external and internal abnormalities
ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY: Not examined

ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY: Not examined
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals after a minimum of 28 days were deeply anaesthetized and sacrificed
- Maternal animals: All surviving animals were deeply anaesthetized and sacrificed on lactation days 5-7

GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.]

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table [#] were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offsprings were sacrificed at 5-7 days of age.
- These animals were subjected to postmortem macroscopic examinations.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
Not performed
Statistics:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test based on a pooled variance estimate was applied for the comparison
of the treated groups and the control groups for each sex.
- The Steel-test was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Kruskal-Wallis nonparametric ANOVA test (Ref. 5) was applied to motor activity data to determine intergroup differences. In case intergroup differences were seen, the Wilcoxon test was applied to compare the treated groups to the control group.
Reproductive indices:
Mating index, Fertility index, Gestation index, Duration of gestation,
Offspring viability indices:
Percentage live males at First Litter Check; Percentage live females at First Litter Check; Percentage of postnatal loss; Viability index

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
No treatment-related clinical signs were noted during daily clinical observations or during weekly arena observations.
Clinical signs noted incidentally occurred within the range of background findings to be expected for rats of this age and strain which are housed and treated under the conditions in this study and showed no dose-related trend. At the incidence observed, these were considered to be unrelated to treatment.
Mortality:
no mortality observed
Description (incidence):
No mortality occurred during the study period that was considered to be related to treatment with the test item.
One female at 300 mg/kg (no.85) was euthanized in extremis on Day 3 of the pre-mating period when she showed laboured respiration and gasping. This was not test item-related but due to the gavage procedure as was evident from the presence of a (part of the) dosing tube in the larynx at macroscopic examination.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weight and body weight gain were not considered affected by treatment.
Food efficiency:
no effects observed
Description (incidence and severity):
No treatment-related changes in food consumption before or after allowance for body weight were noted.
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
Haematological parameters were not considered affected by treatment.
Statistically significant variations between treated rats and controls were limited to a lower percentage of eosinophils in 1000 mg/kg females at the end of the recovery period. Based on the minor magnitude of the difference and the absence of a treatment-related change in eosinophils at the end of the treatment
period, this finding was considered to be unrelated to treatment.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
At 1000 mg/kg, higher total protein and glucose was recorded for Recovery females at the end of treatment. Means remained within the range considered normal for rats of this age and strain. Similar but not statistically significant differences were also recorded at the end of the recovery period. These differences were not apparent for Main females at the end of treatment.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
At the end of the treatment period, mean total movements were statistically significantly higher in males at 300 and 1000 mg/kg than in controls. Mean ambulations were higher (not statistically significantly) in males at 1000 mg/kg. Mean values in treated males remained within the normal range for male rats of this strain and age, and showed a normal motor activity habituation profile with a decreasing trend over the duration of the test period. At the end of the recovery period, motor activity was similar in treated males and controls.
Motor activity of females was not considered affected by treatment and showed a normal motor activity habituation profile.
Hearing ability, pupillary reflex and static righting reflex were normal in all examined animals. Grip strength was not affected by treatment.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
There were no test item-related microscopic observations.
Histopathological findings: neoplastic:
not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
Mating index was not affected by treatment. All paired females showed evidence of mating.
Numbers of corpora lutea and implantation sites were not affected by treatment.
Fertility index was not considered to be affected by treatment.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.

Target system / organ toxicity (P0)

Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
No clinical signs occurred among pups that were considered to be related to treatment.
The clinical signs observed incidentally remained within the range considered normal for pups of this age, and were therefore considered to be unrelated to treatment.
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
No mortality occurred among pups that was considered to be related to treatment with the test item.
The incidence of postnatal loss did not show a dose-related trend and remained within the range considered normal for pups of this age. At first litter check, one pup of the control group (litter no. 55) and one pup at 300 mg/kg (litter no. 84) were found dead. One pup of the control group (litter no. 58) and two pups at 1000 mg/kg (litter no. 91) were found missing on PND 3 or PND 7 (presumably cannibalized). This incidental pup mortality was unrelated to treatment with the test item.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weights of pups were not affected by treatment.
Food efficiency:
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
No macroscopic findings were noted among pups that were considered to be related to treatment.
The nature and incidence of the macroscopic findings noted incidentally remained within the range considered normal for pups of this age, and were therefore considered to be unrelated to treatment.
Histopathological findings:
not examined

Developmental neurotoxicity (F1)

Behaviour (functional findings):
not examined

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not examined

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.

Target system / organ toxicity (F1)

Critical effects observed:
no

Overall reproductive toxicity

Reproductive effects observed:
no

Applicant's summary and conclusion

Conclusions:
In the Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test, the reported NOAEL value for octaphenylcyclotetrasiloxane for reproductive toxicity was ≥1000 mg/kg bw/day. No adverse effects were observed in any of the test animals.