Cyclohexyldimethoxymethylsilane

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

not reported

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Reviewer's note: this study is a part of safety evaluation project of the existing chemical substance by the Japanese authority and it is stated clearly in secondary summaries that this study was conducted in compliance with GLP. However, some pages, which may contain the confirmation of GLP compliance, are missing from the published study report. Justification of read-across is given in the Endpoint Summary.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Version / remarks:

OECD 408 compliance is not stated in the report but the report states each examinations/observations are performed in accordance with GLP and OECD guidelines.

Deviations:

yes

Remarks:

There is no description about functional observations. Preservation in the fixation medium of skin was not performed.

Principles of method if other than guideline:

not applicable

GLP compliance:

yes

Remarks:

Certificate is not included in the study report.

Test material

Test material information

Constituent 1

Reference substance name:

Dicyclopentyl(dihydroxy)silane

Cas Number:

211495-85-1

Molecular formula:

C10H20O2Si

IUPAC Name:

Dicyclopentyl(dihydroxy)silane

Test material form:

solid

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

Condition of animal room though quarantine/acclimatization
Temperature: 22±2°C (allowable range: 19~25°C)
Relative humidity: 55±15 % (allowable range: 35~75%)
Ventilation: 12 times/hour supplied with all fresh air
Lighting hour: 12 hour/day (07:00~19:00)

Equipments of animal husbandry
4 or 5 animals (both sex)/cage before the allocation to test groups and 2 animals (both sex)/cage after the allocation to test groups in polycarbonate cages (265W x 426D x 200H mm, Tokiwa Kagaku Kikai Co., Ltd.) lined with sterilized floorcloth for experimental animal (BETA CHIP, Charles River Laboratories Japan Inc.) setting on steel racks (Tokiwa Kagaku Kikai Co., Ltd.). The locations of cages were exchanged weekly.

Food container and Water supply instrument
Sterilized stainless steel pellet-food containers (Tokiwa Kagaku Kikai Co., Ltd.) and sterilized polycarbonate water supply system (700 ml, Tokiwa Kagaku Kikai Co., Ltd.) were used.
They were exchanged weekly.
Pellet food for experimental animals (MF, Oriental Yeast Co., Ltd.) and tap water filtered through 5 μm filter and ultraviolet radiation were used.
Animals were allowed to free access to food and water. They were exchanged weekly.

The concentrations of pesticide residue in the food and floorcloth were conformed to the standard indicated by the study laboratory.
Water examination in accordance with Water Supply Act was performed and it was confirmed that analysed value were conformed to the standard.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 0.1% Tween 80 added 0.5% CMC-Na solution

Details on oral exposure:

Using syringe with feeding tube.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Homogeneity of the test substance in test solution and stability of test solutions in 8 days under refrigerated storage were examined before administration. And test solution of all the lots for each dose level was examined and the concentration was within +/- 10% of nominal concentration.

However, corresponding documentation is not included in the report.

Duration of treatment / exposure:

90 days

Frequency of treatment:

Once a day in the morning

Doses / concentrations

Remarks:

Doses / Concentrations:
40, 200 and 1000 mg/kg bw/day (m/f)
Basis:
actual ingested

No. of animals per sex per dose:

10 male and 10 female in each dose group

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Highest dose was set at 1000 mg/kg as instructed in OECD guideline .

A 4-week recovery period was included for groups of 10 males and 10 females for the control and 1000 mg/kg bw/day dose level.

Positive control:

No

Examinations

Observations and examinations performed and frequency:

The following items were examined. Here, the first day of dosing is indicated as day 1, and days 1 to 7 are indicated as 1 week. And the duration after day 91 was set as the recovery period.

1. Clinical observations
Observations were done twice a day (before and after dosing) during the dosing period, and once a day in the morning during the rest of the period.

2. Body weight
Body weights of all animals were weighed weekly using an electronic balance (EB-5000: Shimadzu Corporation). Also, it was weighed at the day of necropsy and the previous day.

3. Food intake
All individual food with tare was weighed using an electronic balance (EB-5000: Shimadzu Corporation) weekly and mean daily food intake per one animal was calculated.

4. Haematology
At the time of scheduled necropsy on days 91 and 119, all animals were fasted for more than 20 hours from the evening of the previous day, and anesthetized by intraperitoneal injection of thiopental sodium (Labonal®, Mitsubishi Tanabe Pharma Corporation). Blood was withdrawn from the inferior vena cava and the parameters indicated below were analyzed. For the analysis of parameters (9) and (10) indicated below, 3.2 w/v% sodium tri-citrate solution was used as anticoagulant, and plasma obtained by centrifugation was used for the analysis. For the analysis of other parameters, blood treated with EDTA-2K as anticoagulant was used.

Parameter Method
(1) Erythrocyte count(RBC): Sheath flow DC impedance detective method
(2) Haemoglobin concentration(Hb): SLS-haemoglobin method
(3) Haematocrit(Ht): Erythrocyte pulse wave height detection method
(4) Mean corpuscular volume (MCV): Calculated from (1) and (3)
(5) Mean corpuscular hemoglobin (MCV): Calculated from (1) and (2)
(6) Mean corpuscular hemoglobin concentration (MCHC): Calculated from (2) and (3)
(7) Reticulocyte count: Flow cytometry using argon laser
(8) Platelet count: Sheath flow DC impedance detective method
(9) Prothrombin time (PT): Quick one-stage method
(10) Activated partial thromboplastin time (APTT): Activated cephaloplastin method
(11) Leukocyte counts(WBC): RF/DC impedance detective method
(12) Differential white blood cell count: Measured Wright stained sample

Instruments used for measurement
(1)-(3),(8), (11): :NE-4500, Sysmex Corporation
(7): :R-2000, Sysmex Corporation
(9), (10): :KC 10A, Amelung
(12): MICROX HEG-50,Omron

5. Blood biochemistry
A part of blood sample obtained at scheduled necropsy was left at room temperature for more than 30 minutes, centrifuged and the following parameters were determined using the serum obtained. The serum remained after determination was kept in a freezer.

Parameter Method
(1) ASAT (GOT): UV-rate method (improved JSCC method)
(2) ALAT (GPT) : UV-rate method (improved JSCC method)
(3) gammma GT: gamma-glutamil-p- nitroanilido stroma method (improved JSCC method)
(4) ALP: p-nitrophenylphophate stroma method (improved JSCC method)
(5) Total bilirubin: Enzyme method (BOD method)
(6) Urea nitrogen: Enzyme-UVmethod (Urease-LEDH method)
(7) Creatinine: Jaffé method
(8) Glucose: Enzyme-UV method (GlcK-G6PDH method)
(9) Total cholesterol: Enzyme method (CES-CO-POD method)
(10) Triglyceride: Enzyme method (LPL-GK-G3PO-POD method)
(11) Total protein: Biuret method
(12) Albumin: BCG method
(13) A/G ratio: Calculated from (11) and (12)
(14) Calcium: OCPC method
(15) Inorganic phosphate: Enzyme method (PNP-XOD-POD method)
(16) Sodium (Na): Ion selective electrode method
(17) Potassium (K): Ion selective electrode method
(18) Chloride (Cl): Ion selective electrode method
Instrument used for measurement: HITACHI 736-10 type, Hitachi Ltd.

6. Urinalysis
Using fresh urine collected from 10 males at day 87 and 10 females at day 88 of each group, the following parameters were examined.
In the results, changes attributable to dosing of test substance were not observed. Therefore, other parameter and the urinalysis of recovery period were not examined.

Parameter Method
(1) pH: Test paper method (Multisticks, Bayer Sankyo Co.)
(2) Protein: Test paper method (Multisticks, Bayer Sankyo Co.)
(3) Glucose: Test paper method (Multisticks, Bayer Sankyo Co.)
(4) Ketone body: Test paper method (Multisticks, Bayer Sankyo Co.)
(5) Bilirubin: Test paper method (Multisticks, Bayer Sankyo Co.)
(6) Occult blood: Test paper method (Multisticks, Bayer Sankyo Co.)
(7) Urobilinogen: Test paper method (Multisticks, Bayer Sankyo Co.)
Instrument: Clinitech 100, Bayer Sankyo Co.

7. Ophthalmologic examination
Anterior eye, optic media and fundus oculi were investigated using slit lamp (SL-14, Kowa Co.) and fundus camera (GENESIS K9L29, Kowa Co.) for all male at the day before dosing and all female at two days before dosing during quarantine period, and for male at day 88 and female at day 89 of control group and all 1000 mg/kg group during dosing period.
The investigation of optic media and fundus oculi was conducted after ocular instillation of mydriatic (Mydrin-P,Santen Pharmaceutical Co., Ltd.).
In the results, changes attributable to dosing of test substance were not observed. Therefore, the investigation in all male and female of 40 and 200mg/kg group during dosing period and recovery period were not conducted.

Sacrifice and pathology:

(1) Organ weights
Organs indicated below from all animals were weighed using an electronic balance (AEG-120: Shimadzu Corporation). In addition, relative organ weights (ratio to body weight) were calculated based on the body weights at necropsy. Weight of pituitary and thyroid were weighed after formalin fixation.

Brain, heart, liver, kidneys, adrenals, thymus, spleen, testes, epididymis, ovaries, uterus, pituitary, thyroid, lung.

(2) Macroscopic pathology
All animals were euthanized by exsanguination after cutting the abdominal aorta, and necropsied.

(3) Histopathology
The following organs, tissues were collected from all animals, fixed and preserved in 10 vol% neutral phosphate-buffered formalin. However, eye balls and harderian gland were fixed in Davidson’s fluid, and testes and epididymis were fixes in Bouin's fluid.

Brain, spinal cord (cervical cord, center part of thoracic spinal cord, lumbar cord), pituitary, thyroid and parathyroid(both), thymus, trachea and lungs, heart, main artery, glandula salivaria (both), liver, spleen, kidney (both),adrenal (both), pancreas, testes (both), epididymis (both), ventral prostate, seminalvesicle, ovary (both),uterus, vagina, skin, gullet, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, bladder, submandibular lymph node, mesenteric lymph node, mammary gland. tailor muscle and sciatic nerve (one side), sternum and bone marrow, eye ball and harderian gland (both), femur (including joint) and sciatic nerve (one side), exorbital lacrimal gland (both), macroscopic lesions.

Haematoxyline and eosin stained sample were prepared routinely on all of the above organs/tissues from the control and all male and female animals of 1000 mg/kg group, lung, liver and kidney from all male and female animals of 40 and 200 mg/kg group, and abnormal sites of all animals including control are examined under microscope.

As a result, further investigation to lung, kidney and pituitary was performed.

Other examinations:

None

Statistics:

The metrical data was analyzed by multiple comparison method for the statistical significance. Namely, the homogeneity of variance was examined by Bartlett test, then, one way analyses of variance were conducted where the variances were shown to be homogenous, or Kruskal-Wallis tests were conducted where the variances were shown to be unequal. Analysis for significant differences by Dunnett’s test or by Dunnett multiple comparison was conducted where significant difference was observed.

The enumeration data was analyzed by Chi-square verification of a x b. Differences between the control group and the dose groups were analyzed by Chi-square verification of Armitage where the significant difference was shown.

The parameters indicated below were statistically analyzed. The results of clinical observations, ophthalmologic examination and the histopathological examination were not analyzed statistically.

Multiple comparisons: body weight, food intake, haematology data, blood biochemistry data, organ weights.

Chi-square test verification: Urinalysis, histopathology

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Transient ataxia of gait and salivation in male and female of 200 and 1000 mg/kg group and a decrease in locomotor activity in male and female of 1000 mg/kg group were observed.

Mortality:

mortality observed, treatment-related

Description (incidence):

Transient ataxia of gait and salivation in male and female of 200 and 1000 mg/kg group and a decrease in locomotor activity in male and female of 1000 mg/kg group were observed.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Decrease in weight in female of 1000 mg/kg at days 78, 85 and 90 was observed.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

There were no changes considered to be attributable to dosing of test substance.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

effects observed, treatment-related

Description (incidence and severity):

There were no changes considered to be attributable to dosing of test substance.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

There were no changes considered to be attributable to dosing of test substance.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Increase in total cholesterol in female of 200 and 1000 mg/kg group, decrease in triglyceride in male of 1000 mg/kg group, in A/G ratio in male of 1000 mg/kg group were observed. Not seen in the investigation at the end of recovery period.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

There were no changes considered to be attributable to dosing of test substance.

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

See the section of "Any other information."

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Enlarged liver in 9 males of 1000 mg/kg group, dark colored liver in 1 male of 1000mg/kg were observed at the end of dosing period. These changes were not seen in the investigation at the end of recovery period.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

See the section of "Any other information."

Histopathological findings: neoplastic:

not specified

Details on results:

Effects such as centrilobular hypertrophy of hepatocytes, diffuse hypertrophy of follicular epithelial cells of thyroid and mild hyaline droplet in proximal tubular epithelial cell of kidney were observed. But centrilobular hypertrophy of hepatocytes is considered an adaptive response, hypertrophy of follicular epithelial cells of thyroid was caused by hypertrophy of hepatocytes and hyaline droplet is known as specific symptom to male rat. Therefore these are not considered as adverse effects.

The frequency of observations of transient ataxia at 200 and 1000 mg/kg/day decreased as the administration period went by.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: Non-reversible increased relative kidney and liver weight at 1000 mg/kg/day. No associated histopathological changes.

2

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Highest dose level tested.

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Organ weights

Increase in absolute and relative liver weight in females of 1000 mg/kg group, increase in relative liver weight in males of 200 and 1000 mg/kg group, relative kidney weight and relativeepididymis weight in males of 1000 mg/kg group were observed at the end of dosing period. In the investigation at the end of recovery period. Among these changes, only increase in relative liver weight in females of 1000 mg/kg group was observed, but it was reduced compared to the value at the end of dosing period. Further to this, increase in relative kidney weight was observed at the end of recovery period.

 

Histopathology: non-neoplastic

Centrilobular hypertrophy of hepatocytes in 2 females of 200 mg/kg group, 8 male and all female of 1000 mg/kg group, diffuse hypertrophy of follicular epithelial cells of thyroid in 3 males and 7 females of 1000 mg/kg group, mild hyaline droplet inproximal tubular epithelial cell of kidney in 5 males of control group, 7 males of 40 mg/kg group, 6 males of 200 mg/kg group and 3 males of 1000 mg/kg group, moderate in 1 male of control group, 0 male of 40 mg/kg group, 0 male of 200 mg/kg group and 6 males of 1000 mg/kg group were observed at the end of dosing period.

Among of them, diffuse hypertrophy of follicular epithelial cells of thyroidin 1 male of 1000 mg/kg group and expression of hyaline droplet inproximal tubular epithelial cell of kidney in 7 male of control group and 4 male of 1000mg/kg group were observed at the end of recovery period.

Applicant's summary and conclusion

Conclusions:

In a 90-day repeated dose toxicity study with dicyclopentylsilanediol, conducted according to a protocol equivalent to OECD Test Guideline 408 and in compliance with GLP (certificate not included), the No Observed Effect Level (NOEL) was reported to be 40 mg/kg bw/day based on clinical signs (transient ataxia, decreased locomotor activity), changes in clinical chemistry, organ weights and histopathological findings. Examination of the data indicates that for the purposes of hazard assessment the No Observed Adverse Effect Level (NOAEL) for females was 200 mg/kg bw/day based on increased kidney weight at 1000 mg/kg bw/day which was not completely reversed at the end of a 14-day recovery period. There were no associated histopathological changes in the kidneys in females. The NOAEL for males was 1000 mg/kg bw/day.

Executive summary:

The following evaluation of the pathological findings was prepared by the REACH data submitter:

Blood chemistry:
High and medium dose females showed increased Total Cholesterol and no signicant changes  of Triglyceride at the end of exposure period. Total Cholesterol increase showed a strong tendency for reversibilty. At the end of recovery period no significant differences were found when compared with control group animals.

Male animals showed no significant changes of Total Cholesterol at the end of exposure and recovery period. High dose males showed a decrease of triglyceride of low statistical signifcance at the end of the exposure period which showed tendency for reversibilty at the end of the recovery period. The value was still below the control group but not statistically significant.

Total organ weight
No statistical signifcant changes of organ weight were found in male animals at any dose level.

High dose females showed a statistically significant increase of liver weight at the end of exposure period. There was no significant difference of high dose female liver weight at the end of recovery period when compared with the control group.

Relative organ weight
Relative liver weight was statistically significantly increased in male medium and high dose groups. Relative kidney weight was increased only in the male high dose group. Both effects were reversible in males as there were no significant differences compared to control group animals at the end of the recovery period.

High dose female animals showed statistically signicant increase of liver weight at the end of exposure and recovery period. But there is a clear tendency for reversibility shown by the decrease of liver weight at the end of recovery when compared with the liver weight at the end of exposure.

Relative kidney weight showed an increase of low statistical significance at the of recovery period but not at the end of exposure period.

Histopathological findings
Centrilobular hepatocyte hypertrophy (slight (5 animals) and moderate (3 animals)), and hyaline droplets in proximal tubules (slight (3 animals ) and moderate (6 animals)), were found in high dose males. Also high dose females showed centroilobular hepatocyte hypertrophy (slight (5 animals ) and moderate (5 animals)) degree. No significant kidney changes were found in female animals.

At the end of the recovery period no statistically significant differences were found between the high dose and control group.  


Summary and determination of NOAEL
At the end of the exposure period there were dose-dependent test item-related effects on kidney and liver, reflected especially by an increase of absolute and particularly relative organ weight. These weight changes were associated only in females with changes of total cholesterol in high and medium dose group, and in high dose males with a decrease of triclyceride. Significant histopathological changes of liver were only found in high dose male and female animals and significant histopathological kidney changes were only found in high dose animals.  

All effects observed in the study showed strong tendency for reversibility as they were not found at the end of recovery period. Only high dose female animals showed at the end of the recovery period still a significant increase of relative liver and kidney weight. No increase of absolute and relative liver and kidney weight  was found in medium dose females at the end of exposure period.

Based on the increase of relative kidney and liver weight  (which were not associated with significant histopathological changes) in high dose females at the end of recovery period  the NOAEL is 200 mg/kg bw in female animals. The NOAEL for male animals is 1000 mg/kg bw.