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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2020/07/01 - 2021/02/01
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2021

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Triethoxy(3-thiocyanatopropyl)silane
EC Number:
252-161-3
EC Name:
Triethoxy(3-thiocyanatopropyl)silane
Cas Number:
34708-08-2
Molecular formula:
C10H21NO3SSi
IUPAC Name:
3-(triethoxysilyl)propyl thiocyanate
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 Deutschland, Sulzfeld, Germany
- Females: Based on age, nulliparous and non-pregnant
- Age at study initiation: 6-7 weeks
- Weight at study initiation: Males weighed 153-206 g and females weighed 88-153 g
- Fasting period before study: No
- Housing: Up to 5 animals of same sex / dose group, in Polycarbonate cages (Makrolon type IV or Makrolon type 2000P)
- Diet: SM R/M-Z from SSNIFF, ad libitum
- Water: Municipal tap water, ad libitum
- Acclimation period: At least 5 days

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

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22
- Humidity (%): 46-77
- Air changes (per hr): 10 or more
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2020/07/28 (initiation of dosing) To: 2020/11/25

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Remarks:
Dried and deacidified arachis oil, specific gravity 0.885
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 preparation of the formulation.

No adjustment was made for specific gravity of the test item. No correction was made for the purity/composition of the test item.

Test item dosing formulations were kept at room temperature until dosing. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing.

VEHICLE
- Justification for use and choice of vehicle: Trial preparations were performed to establish a suitable formulation procedure, not part of this study
- Concentration in vehicle: 0, 7.5, 25, 75 mg/mL
- Amount of vehicle: Dosing solution volume, 4 mL/kg bw
- Lot/batch no.: 1911106
- Purity: Not specified
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose formulation samples were collected in weeks 1, 6 and 12, with analysis performed using a validated analytical procedure to assess concentration and homogeneity
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
7 d/wk
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
30 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10M/10F in main design (all dose levels), 5M/5F for recovery groups (0, 300 mg/kg bw/day groups)
Control animals:
yes
Details on study design:
- Dose selection rationale: Results of the dose range finding study
- Rationale for animal assignment: Random
- Fasting period before blood sampling for clinical biochemistry: Yes, overnight with a maximum of 24 hours
- Rationale for selecting satellite groups: To assess recovery from or delayed toxicity
- Post-exposure recovery period in satellite groups: 28 days
- Section schedule rationale: Not specified

Examinations

Observations and examinations performed and frequency:
MORTALITY: Yes
All animals, at least twice daily beginning upon arrival through termination.

CAGE SIDE OBSERVATIONS: Yes
All main study and recovery animals, at least twice daily. From Day 1, 0-15 minutes post dose, and 45-75 minutes post dose.

DETAILED CLINICAL OBSERVATIONS: Yes
All main study and recovery animals, once pretreatment and weekly during treatment and recovery periods; and on the day of necropsy.

BODY WEIGHT: Yes
All main study and recovery animals, weekly from at least Day 1 and throughout the study.

FOOD CONSUMPTION Yes
All main study and recovery animals, weekly from at least Day 1 and throughout the study, quantitative per cage.

FOOD EFFICIENCY: No

WATER CONSUMPTION: Yes
All main study and recovery animals, regular basis throughout the study, visual inspection of water bottles.

OPHTHALMOSCOPIC EXAMINATION: Yes
Pretreatment: All main study and recovery animals once (including spare animals). Dosing Period: All 0 and 300 mg/kg bw/day animals during Week 13.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At end of treatment period
- Anaesthetic used for blood collection: Yes, isoflurane
- Animals fasted: Yes, overnight with a maximum of 24 hours
- How many animals: All main study and recovery animals
- See Table 1 below for list of analyzed parameters

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At end of treatment period
- Animals fasted: Yes, overnight with a maximum of 24 hours
- How many animals: All main study and recovery animals
- See Table 2 for list of analyzed clinical chemistry parameters.
- In addition, coagulation was assessed (prothrombin Time (PT) and activated partial thromboplastin time (APTT).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Once during the Dosing Period. The first 5 animals per sex per group during Weeks 12-13
- Dose groups that were examined: All animals
- Battery of functions tested: hearing ability, pupillary reflex, static righting reflex, fore- and hind-limb grip strength, locomotor activity (total movements and ambulations)

IMMUNOLOGY: No

OTHER: ESTROUS STAGE DETERMINATION
All main study females, on the day of necropsy at end of treatment, vaginal smear to determine the stage of estrus. All recovery females, on the day of necropsy.
Sacrifice and pathology:
SACRIFICE:
Main study and recovery animals surviving until scheduled euthanasia were weighed, and euthanized using isoflurane, followed by exsanguination. Animals were fasted (overnight with a maximum of 24 hours) before their scheduled necropsy.

The terminal procedures are summarized in Table 3 below.

ORGAN WEIGHT: Yes
All main study and recovery animals.
- See Table 4 below for list weighed organs

GROSS PATHOLOGY: Yes
All main study and recovery animals, as follows:
- Full list of tissues
- See Tables 3 and 5 below for terminal procedures and full list of tissues collected

HISTOPATHOLOGY: Yes
All main study and recovery animals, as follows:
- 0 and 300 mg/kg bw/day dose groups: Full list of tissues
- 30 and 100 mg/kg bw/day dose groups: Gross lesions and target tissues
- Target tissues: Stomach and kidneys: All males and females in main study 30 and 100 mg/kg bw/day groups, and 0 and 300 mg/kg bw/day recovery groups. Thymus: All males in main study 30 and 100 mg/kg bw/day groups, and 0 and 300 mg/kg bw/day recovery groups.
- See Tables 3 and 5 below for terminal procedures and full list of tissues collected

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Hunched posture and/or erected fur were observed at 300 mg/kg bw/day in seven males, and in six females, between Days 2 and 92.

The other clinical findings including the above signs at 30 and 100 mg/kg bw/day were considered not toxicologically relevant.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Test item-related and adverse lower body weight and body weight gain in males at 300 mg/kg bw/day, which recovered during the dosing-free period.
Food consumption and compound intake (if feeding study):
not specified
Description (incidence and severity):
Test item-related and adverse lower food consumption in males at 300 mg/kg bw/day, which partially recovered during the dosing-free period.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
Water consumption was monitored by visual inspection of the water bottles, results not reported.
Ophthalmological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No ophthalmology findings were noted that were considered related to treatment with the test item. Nature and incidence of findings were within the range considered normal for rats of this age and strain.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
At 300 mg/kg bw/day, increased reticulocyte counts were observed in males and females at the end of the dosing period. In addition, females at 300 mg/kg bw/day, showed increased counts of large unstained cells. Both findings normalized following the Recovery Period. In absence of a histopathological correlation, the findings at this dose level were considered not adverse.

Remaining findings were considered of no toxicological significance. No effects reported for 30 or 100 mg/kg bw/day.

See Table 6 below.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Clinical chemistry findings at 100 and/or 300 mg/kg bw/day at the end of the Dosing Period were as follows:
- Increased calcium concentrations in males and females
- Decreased T4 and (at 300 mg/kg bw/day) increased TSH concentrations in males
- Decreased T3 and (at 100 mg/kg bw/day) T4 concentrations, and increased TSH concentrations in females
- Increased triglyceride concentrations (at 300 mg/kg bw/day) in males

There were no corresponding gross- or histopathology findings in the thyroid. In terms of the elevated calcium and triglyceride levels, see the renal histopathology results at 300 mg/kg bw/day as discussed below.

The above findings normalized following the Recovery Period. In absence of a histopathological correlation, the findings at these dose levels were considered not adverse.

Remaining findings were considered not test item-related. No effects reported for 30 mg/kg bw/day.

See Table 7 below.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
All findings were normal compared to controls (hearing ability, hearing ability, pupillary reflex, static righting reflex, male grip strength, female fore-leg grip strength, motor activity), or were considered not toxicologically relevant (female hind-leg grip strength at 100 and 300 mg/kg bw/day). Latter was within range considered normal for rats of this age and strain and lacked a clear dose-related response.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
At 300 mg/kg bw/day, test item-related lower absolute thymus weights in males, higher relative kidney weights in females and higher relative liver weights in males and females were observed.

Changes in kidney weight could be correlated with multifocal bilateral tubular basophilia with mononuclear cell infiltrates and showed no recovery following the treatment-free period. No microscopic correlates were observed for the changes in thymus and liver weights.

Some organ weight differences were statistically significant when compared to the control group but were considered to be the result of a test item-related effect on final body weight or due to a high physiological variability (uterus).

See Table 8 below.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
LOCAL EFFECTS:
Test item-related macroscopic findings were present in the stomach of both sexes. Dark purple/red foci in the glandular stomach, dark grey/purple/red foci in the non-glandular stomach and/or an irregular surface of the non-glandular stomach was seen in one female at 30 mg/kg bw/day, six females at 100 mg/kg bw/day and five males and five females at 300 mg/kg bw/day at the End of Treatment. Microscopic correlates for the abnormalities in the non-glandular stomach consisted of hemorrhage and epithelial hyperplasia, respectively. In addition, two males at 300 mg/kg/day showed enlarged livers (no corresponding histopathology).

These findings were not seen at the end of recovery.

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):
SYSTEMIC EFFECTS:
Tubular basophilia and mononuclear cell infiltration were ascribed to treatment with the test item only when scored as bilateral and multifocal at 300 mg/kg bw/day. These findings correlated with increased kidney weight in females. These findings were recorded in females at the end of the Dosing Period as well as at the end of the recovery period at severities up to moderate degree and were regarded to be adverse. In the kidney, minimal tubular basophilia and mononuclear cell infiltration are common background pathology findings. Accumulation of hyaline droplets in the kidney in males was also seen, which showed full recovery at the end of the recovery period. Accumulation of hyaline droplets is caused by an increased production of alpha-2-urinary-globulin which is a protein specific for male rats. This is a well-known phenomenon and was in absence of indicators of tubular damage regarded to be non-adverse.

LOCAL EFFECTS:
Histopathological examination in animals treated at 300 mg//kg bw/day findings consisted of hemorrhage, epithelial hyperplasia, ulceration, mixed cell infiltration, edema and hyperkeratosis in the non-glandular stomach in males and females, which was regarded to be adverse. These findings correlated with macroscopically dark grey/purple/red and/or an irregular surface (considered non-adverse) at necropsy.

See Table 9 below.
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
Coagulation parameters of treated rats were considered not affected by treatment up to 300 mg/kg bw/day.

Estrous stage determined, results not reported.

Effect levels

open allclose all
Key result
Dose descriptor:
NOAEL
Remarks:
local effects
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
gross pathology
histopathology: non-neoplastic
Remarks on result:
other: Based on histopathology findings in the non-glandular stomach at 100 and 300 mg/kg bw/day, with corresponding gross pathology
Key result
Dose descriptor:
NOAEL
Remarks:
systemic effects
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
food consumption and compound intake
histopathology: non-neoplastic
organ weights and organ / body weight ratios
Remarks on result:
other:
Remarks:
Based on effects in males (decreased body weight, body weight gain, and food consumption) and females (increased renal organ weight and histopathological alterations) at 300 mg/kg bw/day.

Target system / organ toxicity

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:
no

Any other information on results incl. tables

Table 6. Triethoxy(3-thiocyanatopropyl)silane‑related haematology changes

Dose (mg/kg/day)

300a

Sex

M

F

Reticulocytes

 

 

End of Treatment

1.26/0.941

1.27/0.961

Large unstained cells (LUC)

 

 

End of Treatment

-

2.15/0.291

1 Ratio end of recovery; - : not applicable; x-fold changes compared to concurrent control group are given; changes indicated in bold reached statistical significance

a No toxicologically relevant changes were noted in haematological parameters in males and females up to 100 mg/kg/day

 

Table 7. Triethoxy(3-thiocyanatopropyl)silane‑related clinical chemistry changes

Dose (mg/kg/day)

100a

300

Sex

M

F

M

F

Calcium

 

 

 

 

End of Treatment

1.04

1.06

1.06/

1.021

1.05/

0.981

Triglycerides

 

 

 

 

End of Treatment

1.09

-

1.95/

1.241

-

T3

 

 

 

 

End of Treatment

0.92

1.02

1.03/

0.941

0.81/

1.061

T4

 

 

 

 

End of Treatment

0.73

0.71

0.78/

0.931

0.64/

1.081

TSH

 

 

 

 

End of Treatment

0.96

1.22

2.24/

1.461

1.72/

0.581

1 Ratio end of recovery; - : not applicable; x-fold changes compared to concurrent control group are given; changes indicated in bold reached statistical significance

a No toxicologically relevant changes were noted in males and females up to 30 mg/kg/day

 

Table 8. Mean percent organ weight differences from control groups

Males - Mean Percent Thymus and Liver Weight Differences from Control Groups

Dose level (mg/kg/day):

Main Males

Recovery Males

30

100

300

300

Thymus

 

 

 

 

Absolute

-1

-10

-28

-7

Relative to body weight

3

-3

-15

-1

 

 

 

 

 

Liver

 

 

 

 

Absolute

-1

-3

1

0

Relative to body weight

3

4

19

5

 

 

 

 

 

    Changes indicated in bold reached statistical significance 


Females - Mean Percent Kidney and Liver Weight Differences from Control Groups

Dose level (mg/kg/day):

Main Females

Recovery Females

30

100

300

300

Kidney

 

 

 

 

Absolute

2

0

9

2

Relative to body weight

2

7

14

10

 

 

 

 

 

Liver

 

 

 

 

Absolute

3

0

13

-3

Relative to body weight

4

7

18

3

 

 

 

 

 

Changes indicated in bold reached statistical significance 

Table 9. Summary Test Item-Related Microscopic Findings

Males

Dose level (mg/kg/day):

Main Males

Recovery Males

0

30

100

300

0

300

 

 

 

 

 

 

 

Kidneya

10

10

10

10

5

5

Accumulation, hyaline droplets

 

 

 

 

 

 

Minimal

2

2

1

2

3

3

Mild

2

1

6

2

-

1

Moderate

-

-

1

4

-

-

 

 

 

 

 

 

 

Stomach (non-glandular)a

10

10

10

10

5

5

Hemorrhage

 

 

 

 

 

 

Minimal

-

-

-

2

-

-

Mild

-

-

-

1

-

-

 

 

 

 

 

 

 

Hyperplasia, epithelial

 

 

 

 

 

 

Minimal

-

-

1

4

-

-

Mild

-

-

2

4

-

-

Moderate

 

-

-

-

1

-

-

Ulceration

 

 

 

 

 

 

Minimal

-

-

-

2

-

-

 

 

 

 

 

 

 

Infiltration, mixed cell

 

 

 

 

 

 

Minimal

-

-

2

4

-

-

Mild

1

-

1

3

-

-

 

 

 

 

 

 

 

Edema

 

 

 

 

 

 

Minimal

-

-

1

4

-

-

Mild

-

-

-

2

-

-

 

 

 

 

 

 

 

Hyperkeratosis

 

 

 

 

 

 

Minimal

-

-

-

5

-

-

Mild

-

-

1

-

-

-

 

 

 

 

 

 

 

     a = Number of tissues examined from each group; -: not applicable

Females

Dose level (mg/kg/day):

Main Females

Recovery Females

0

30

100

300

0

300

 

 

 

 

 

 

 

Kidneya

10

10

10

10

5

5

Basophilia, tubular, bilateral, multifocal

 

 

 

 

 

 

Minimal

-

-

1

1

-

1

Mild

-

-

-

7

-

2

Moderate

-

-

-

2

-

2

 

 

 

 

 

 

 

Infiltration, mononuclear cell,    bilateral, multifocal

 

 

 

 

 

 

Minimal

-

-

-

9

-

3

Mild

-

-

-

-

-

2

 

 

 

 

 

 

 

Stomach (non-glandular)a

10

10

10

10

5

5

Hemorrhage

 

 

 

 

 

 

Minimal

-

-

-

-

-

-

Mild

-

-

1

-

-

-

 

 

 

 

 

 

 

Hyperplasia, epithelial

 

 

 

 

 

 

Minimal

1

3

1

-

-

-

Mild

-

-

6

4

-

-

Moderate

-

-

-

1

-

-

 

 

 

 

 

 

 

Ulceration

 

 

 

 

 

 

Minimal

-

-

1

2

-

-

Mild

-

-

-

1

-

-

 

 

 

 

 

 

 

Infiltration, mixed cell

 

 

 

 

 

 

Minimal

2

1

7

4

-

-

Mild

-

-

-

1

-

-

 

 

 

 

 

 

 

Edema

 

 

 

 

 

 

Minimal

2

1

-

2

-

-

Mild

-

1

2

-

-

-

 

 

 

 

 

 

 

Hyperkeratosis

 

 

 

 

 

 

Minimal

-

-

5

3

-

-

Mild

-

-

1

1

-

-

 

 

 

 

 

 

 

  a Number of tissues examined from each group; -: not applicable

 

Applicant's summary and conclusion

Conclusions:
In a study conducted according to OECD Test Guideline 408 and in compliance with GLP (reliability score 1), Wistar Han rats were administered triethoxy(3-thiocyanatopropyl)silane at 0, 30, 100, and 300 mg/kg bw/day via oral gavage (arachis oil vehicle) for 7 days/week over 13 weeks. The systemic NOAEL for triethoxy(3-thiocyanatopropyl)silane was 100 mg/kg bw/day, based on effects in males (decreased body weight, body weight gain, and food consumption) and females (increased renal organ weight and histopathological alterations) at 300 mg/kg bw/day. For local effects in the non-glandular stomach, the NOAEL was 30 mg/kg bw/day.