Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 Mar 2018 to 13 Sep 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Version / remarks:
1998
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Version / remarks:
1998
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
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
- Solubility and stability of the test substance in the solvent/vehicle: the test item was dissolved in dehydrated and deacifidied corn oil
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not applicable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: 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 as a solution and dosed within 6 hours after completionof the preparation of the formulation
- Preliminary purification step (if any): no
- Final dilution of a dissolved solid, stock liquid or gel: not applicable
- Final preparation of a solid: not applicable

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Han
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 6 weeks old at initiation of dosing
- Weight at study initiation: Animals weight was within 20% of the group mean sex weight. Males weighed between 140 and 177 gram and females weighed between 114 and 144 gram.
- Fasting period before study: no
- Housing: Animals were housed in groups of 5 in polycarbonate cages. During locomotor activity monitoring, animals were housed individually in a Hi-temp polycarbonate cage.
- Diet (e.g. ad libitum): Pelleted rodent diet, at libitum
- Water (e.g. ad libitum): Municipal tap water, ad libitum
- Acclimation period: 8 days

DETAILS OF FOOD AND WATER QUALITY: The feed was analyzed by the supplier for nutritional components and environmental contaminants. Results of the analysis were provided by the supplier and are on file at the Test Facility. Periodic analysis of the water was performed, and results of these analyses are on file at the Test Facility. It was considered that there are no known contaminants in the water that would interfere with the objectives of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 to 22°C
- Humidity (%): 36 to 60%
- Air changes (per hr): 10 ot more per hour
- Photoperiod (hrs dark / hrs light): 12-hour light/12-hour dark cycle

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral 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 as a solution and dosed within 6 hours after completionof the preparation of the formulation

VEHICLE
- Justification for use and choice of vehicle (if other than water): Dehydrated and deacidified corn oil was chosen based on information provided by the sponsor.
- Concentration in vehicle: 4, 20, 150 mg/mL
- Amount of vehicle (if gavage): not specified
- Lot/batch no. (if required): 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 weeks 1, 6 and 13. Concentration analysis was performed to all dose levels, homogeneity analysis was performed to low dose and high dose groups. Stability of the test item formulation was analysed only during week 1 for low and high dose levels.
Duration of treatment / exposure:
maximum 90 days
Frequency of treatment:
daily, 7 days a week
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control group
Dose / conc.:
20 mg/kg bw/day (actual dose received)
Remarks:
Low dose group
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Remarks:
Middle dose group
Dose / conc.:
750 mg/kg bw/day (actual dose received)
Remarks:
High dose group
No. of animals per sex per dose:
Test grups: 10 males and 10 females
Control group: 10 males and 10 females
Recovery animals for control and high dose groups: 5 males and 5 females
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 repeated dose toxicity study with oral exposure of 1,1,1,3,5,5,5-Heptamethyl-3-[(trimethylsilyl)oxy]trisiloxane in rats, Test Facility Study No. 20140240), and in an attempt to produce graded responses to the test item. The high-dose level should produce some toxic effects, but not excessive lethality that would prevent meaningful evaluation. The mid-dose level is expected to produce minimal to moderate toxic effects. The low-dose level should produce no observable indications of toxicity.
- Rationale for animal assignment (if not random): random
- Fasting period before blood sampling for clinical biochemistry: Yes, animals were fasted overnight prior to blood collection.
- Rationale for selecting recovery groups: Recovery groups were included for the control and high dose groups.
- Post-exposure recovery period in recovery groups: 2 weeks following last dose administration
- Section schedule rationale (if not random): random
Positive control:
Not included

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily
- Cage side observations included: general health/mortality and moribundity

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Animals were weighed individually weekly, starting on Day 1. A fasted weight was recorded on the day of necropsy.

FOOD CONSUMPTION: Yes
- Time schedule: Food consumption was quantitatively measured weekly starting on Day 1 and continuing weekly throughout the dosing and recovery period.

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: No. Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: at pretreatment and at the end of the dosing period
- Dose groups that were examined: all test and recovery animals

HAEMATOLOGY: Yes
- Time schedule for collection of blood: end of treatment period and end of recovery period
- Anaesthetic used for blood collection: Yes (isoflurane )
- Animals fasted: Yes
- How many animals: all animals at the end of treatment period, and all animals from control and high dose recovery groups at the end of recovery period
- Parameters checked in table [No.1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: end of treatment period and end of recovery period
- Animals fasted: Yes
- How many animals: all animals at the end of treatment period, and all animals from control and high dose recovery groups at the end of recovery period
- Parameters checked in table [No.1] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: during Week 13
- Dose groups that were examined: on the first 5 animals per sex per group during Week 13
- Battery of functions tested: hearing ability; pupillary reflex; static righting reflex; fore- and hind-limb grip strength; locomotor activity

IMMUNOLOGY: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table No. 2)
HISTOPATHOLOGY: Yes (see table No. 2)
Statistics:
All statistical tests was conducted at the 5% significance level. All pairwise comparisons were conducted using two sided tests and were reported at the 1% and 5% levels.
Numerical data collected on scheduled occasions for the listed variables were analyzed as indicated according to sex and occasion. Descriptive statistics number, mean and standard deviation (or %CV or SE when deemed appropriate) were reported whenever possible. Inferential statistics were performed according to the matrix below when possible, but excluded semi-quantitative data, and any group with less than 3 observations.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
No toxicologically relevant clinical signs were noted during daily detailed clinical observations.
Mortality:
no mortality observed
Description (incidence):
No mortality occurred during the study period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weights and body weight gain of treated animals remained in the same range as controls over the study period.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Food consumption before or after correction for body weight was similar to the control level over the study period.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No ophthalmology findings were noted that were considered to be related to treatment.
Haematological findings:
no effects observed
Description (incidence and severity):
No toxicologically relevant changes were noted in hematology parameters.
Lower reticulocytes counts and lower red blood cell distribution width were noted in treated males. Based on the magnitude of the change, absence of a clear dose-response and/or absence of corroborative findings in the opposite sex and other red blood cell parameters, these changes were considered not toxicologically relevant.
In females treated at 100 and 750 mg/kg/day higher neutrophil counts, and lower mean corpuscular volume (MCV) and mean corpuscular haemoglobin levels were noted at end of treatment. At end of recovery, lower haemoglobin concentration, haematocrit levels and MCV were noted in females treated at 750 mg/kg/day. Based on the magnitude of the change, absence of corroborative findings in the opposite sex, and/or slightly high control values, these changes were considered to be of no toxicological significance.
No toxicologically relevant changes were noted in coagulation parameters. A shorter prothrombin time was noted for males treated at 20 and 100 mg/kg/day at end of treatment and for females at all dose groups at end of treatment and at end of recovery. Based on absence of biological significance, absence of a clear dose-response and/or slightly high control values, these changes were considered not toxicologically relevant.
A shorter activated partial thromboplastin time was noted for males at end of recovery and for females treated at 20 and 100 mg/kg/day. In absence of a clear dose-response and biological significance these changes were considered not toxicologically relevant.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Lower total bilirubin levels were noted females at 750 mg/kg/day at end of treatment. The toxicological relevance is uncertain.
Higher sodium and chloride levels were noted in males and higher cholesterol and potassium levels were noted in females at all dose levels at the end of treatment. Higher total protein and calcium levels were noted in females treated at 100 and 750 mg/kg/day both at end of treatment and end of recovery. Based on the absence of a clear dose-response, absence of corroborative findings in the opposite sex, magnitude of the change and/or absence of these changes at end of recovery, those changes were considered not toxicologically relevant.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
No findings were noted during weekly arena observations. Hearing ability, pupillary reflex and static righting reflex were normal in all examined males. Grip strength was similar between control and high dose animals (males and females). Motor activity was similar between treated and control groups. All groups (males and females) showed a similar motor activity habituation profile with a decreasing trend in activity over the duration of the test period.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Test item-related higher liver weights (absolute and relative to body weights) were present in main group males and females starting at 100 mg/kg bw/day and in recovery group females (relative to body weight only) at 750 mg/kg bw/day. Higher thyroid gland weights (absolute and relative to body weights) were noted in the 750 mg/kg bw/day females group.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Test item-related increased accentuated lobular pattern was present in the liver of 7/10 main males treated at 750 mg/kg bw/day, compared to 2/10 males treated at 20 mg/kg bw/day and 1/10 males treated at 100 mg/kg bw/day. The microscopic correlate was hepatocellular hypertrophy.
The remainder of the recorded macroscopic findings were within the range of background gross observations encountered in rats of this age and strain.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Test item-related microscopic findings after treatment were noted in the liver and thyroid glands of the males and females and are summarized in Results Tables 3 and 4.
Pigment deposition in the bile duct was present in the liver in main group males starting at 100 mg/kg bw/day up to moderate degree. After the 14-day recovery period pigment deposition in the bile duct was present at 750 mg/kg bw/day up to moderate degree. The pigment depositions were in some cases bright red using polarizing light and many showed Maltese cross formation.
Vacuolation hepatocellular was present in the liver of main group males and females starting at 100 mg/kg bw/day up to slight to moderate degree. After the 14-day recovery period this finding was present in a single male at 750 mg/kg bw/day at slight degree and had completely recovered in females.
Hepatocellular hypertrophy was present in the liver in main group males and females starting at 100 mg/kg bw/day up to slight degree. After the 14-day recovery period this finding was present in a single male and female at minimal degree at 750 mg/kg bw/day suggesting partial ongoing recovery. This correlated with the increased liver weights and the macroscopic accentuated lobular pattern.
Follicular cell hypertrophy was present at increased incidence and severity in the thyroid gland of main group males at 750 mg/kg bw/day and main group females starting at 100 mg/kg bw/day up to slight degree. After the 14-day recovery period this finding was present at background incidence and severity, suggesting complete recovery. This correlated with the increased organ weight.
Colloid alteration was present in the thyroid gland in main group males and females starting at 100 mg/kg bw/day up to slight degree. After the 14-day recovery period this finding was present at minimal degree in males and females, suggesting partial recovery.
The remainder of the recorded microscopic findings were within the range of background pathology encountered in rats of this age and strain. There was no test item related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations.
Histopathological findings: neoplastic:
not examined

Effect levels

open allclose all
Key result
Dose descriptor:
NOAEL
Effect level:
>= 750 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: No adverse effects observed in females.
Remarks on result:
other:
Remarks:
Hepatocellular hypertrophy and hepatocellular vacuolation in females were considered non-adverse based on the incidence and complete or partial recovery after the 14-day recovery period. Hypertrophy of the follicular epithelium of the thyroid glands were considered secondary effect to hepatocellular hypertrophy.At current severities and in absence of any other adverse pathologic findings these findings in the thyroid were considered to be non-adverse.
Key result
Dose descriptor:
NOAEL
Effect level:
20 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
histopathology: non-neoplastic
Remarks on result:
other:
Remarks:
Hepatocellular hypertrophy and hepatocellular vacuolation in males were considered non-adverse based on the incidence and complete or partial recovery after the 14-day recovery period. Hypertrophy of the follicular epithelium of the thyroid glands were considered secondary effect to hepatocellular hypertrophy.At current severities and in absence of any other adverse pathologic findings these findings in the thyroid were considered to be non-adverse.

Target system / organ toxicity

Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
100 mg/kg bw/day (actual dose received)
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified

Any other information on results incl. tables

Table 1: Mean Percent Organ Weight Differences from Control Groups - Males

 

Main Males

Main Males

Main Males

Recovery Males

Dose level (mg/kg/day):

20

100

750

750

 

 

 

 

 

LIVER

 

 

 

 

              Absolute

-2

12*

19**

4

              Relative to body weight

-1

12**

19**

5

 

 

 

 

 

THYROID GLANDS

 

 

 

 

              Absolute

11

11

6

12

              Relative to body weight

25

25

25

25

 

 

 

 

 

Table 2: Mean Percent Organ Weight Differences from Control Groups - Females

 

Main Females

Main Females

Main Females

Recovery Females

Dose level (mg/kg/day):

20

100

750

750

 

 

 

 

 

LIVER

 

 

 

 

              Absolute

14

28**

54**

26

              Relative to body weight

9

24**

43**

36**

 

 

 

 

 

THYROID GLANDS

 

 

 

 

              Absolute

25

25

42**

7

              Relative to body weight

17

17

33**

17

 

 

 

 

 

*: P<0.05, **: P<0.01

Table 3: Summary Test Item-Related Microscopic Findings – Males

 

Main Males

Main Males

Main Males

Main Males

Recovery Males

Recovery Males

Dose level (mg/kg/day):

0

20

100

750

0

750

 

 

 

 

 

 

 

LIVERa

10

10

10

10

5

5

    Pigment deposition bile duct

 

 

 

 

 

 

      Minimal

-

-

-

1

-

-

      Slight

-

-

1

1

-

2

      Moderate

-

-

1

3

-

1

    Vacuolation hepatocellular

 

 

 

 

 

 

      Minimal

-

-

-

2

-

-

      Slight

-

-

2

5

-

1

      Moderate

-

-

1

-

-

-

    Hypertrophy hepatocellular

 

 

 

 

 

 

      Minimal

-

-

4

8

-

1

      Slight

-

-

-

2

-

-

THYROID GLANDSa

10

10

10

10

5

5

   Hypertrophy follicular cell

 

 

 

 

 

 

      Minimal

5

4

2

7

-

2

      Slight

-

-

-

3

-

-

   Colloid alteration

 

 

 

 

 

 

      Minimal

-

-

2

3

-

3

      Slight

-

-

-

1

-

-

Table 4: Summary Test Item-Related Microscopic Findings – Females

 

Main Females

Main Females

Main Females

Main Females

Recovery Females

Recovery Females

Dose level (mg/kg/day):

0

20

100

750

0

750

 

 

 

 

 

 

 

LIVERa

10

10

10

10

5

5

    Vacuolation hepatocellular

 

 

 

 

 

 

      Minimal

-

-

4

7

-

-

      Slight

-

-

-

1

-

-

    Hypertrophy hepatocellular

 

 

 

 

 

 

      Minimal

-

-

6

9

-

1

      Slight

-

-

-

1

-

-

THYROID GLANDSa

10

10

10

10

5

5

   Hypertrophy follicular cell

 

 

 

 

 

 

      Minimal

1

2

5

3

-

1

      Slight

-

-

1

7

-

-

   Colloid alteration

 

 

 

 

 

 

      Minimal

-

-

2

8

-

2

      Slight

-

-

-

1

-

-

a = Number of tissues examined from each group.

Applicant's summary and conclusion

Conclusions:
In the 90-day repeated oral toxicity study, conducted according to OECD TG 408 and in compliance with GLP, the concluded NOAEL for males was 20 mg/kg bw/day based on pigment deposition in bile duct at higher doses. No such effect was seen in females and the concluded NOAEL for females was the highest dose tested, 750 mg/kg bw/day.