Bis(1,2,2,6,6-pentamethyl-4-piperidyl) [[3,5-bis(1,1-dimethylethyl)-4-hydroxyphenyl]methyl]butylmalonate

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011-2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant study. Specific target organ investigations performed as part of an OECD 421 study.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD 421

Deviations:

yes

Remarks:

extra investigation of known endpoints (haematology, histopathology of spleen, liver, intestine), etc

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Bis(1,2,2,6,6-pentamethyl-4-piperidyl) [[3,5-bis(1,1-dimethylethyl)-4-hydroxyphenyl]methyl]butylmalonate

EC Number:

264-513-3

EC Name:

Bis(1,2,2,6,6-pentamethyl-4-piperidyl) [[3,5-bis(1,1-dimethylethyl)-4-hydroxyphenyl]methyl]butylmalonate

Cas Number:

63843-89-0

Molecular formula:

C42H72N2O5

IUPAC Name:

bis(1,2,2,6,6-pentamethylpiperidin-4-yl) 2-butyl-2-[(3,5-di-tert-butyl-4-hydroxyphenyl)methyl]propanedioate

Details on test material:

- Substance type: powder
- Physical state: white solid
- Analytical purity: HPLC: approx. 99 area-% (220 nm)
- Expiration date of the lot/batch: 21 Feb 2014
- Stability under test conditions: stable
- Storage condition of test material: room teperature

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sulzfeld
- Age at study initiation: 10-11 weeks; males/ females
- Weight at study initiation: ca 250 g (females), ca 300g (males)
- Fasting period before study: overnight
- Housing: single (except for mating)
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2011-10-11 To: 2011-12-06

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 1% aqueous CMC and 5% Cremophor EL

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:

To prepare the suspension, the appropriate amount of test substance was weighed out depending on the desired concentration. Then,
drinking water containing 1% carboxymethylcellulose and 5 mg/ 100 mL Cremophor EL was filled up to the desired volume, subsequently released with a magnetic stirrer. During administration of the test substance preparations were kept homogeneous by stirring with a magnetic stirrer. The test substance preparations were produced at least once a week and were stored at room temperature.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

HPLC-method

Duration of treatment / exposure:

28 days (males)

Frequency of treatment:

daily

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 0.5, 2 and 10 mg/kg bw
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on range-finder study

Positive control:

not required

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least once daily for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed clinical observations (DCO; including palpation) were performed in all animals prior to the administration period and thereafter at weekly intervals.

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

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: Yes


OPHTHALMOSCOPIC EXAMINATION: No


HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of the study (males: study day 36, females: study days 45, 49 and 50)
- Anaesthetic used for blood collection: Yes (isoflurane anesthesia)
- Animals fasted: Yes
- How many animals: 10
- Parameters were examined:
Leukocyte count (WBC)
Erythrocyte count (RBC)
Hemoglobin (HGB)
Hematocrit (HCT)
Mean corpuscular volume (MCV)
Mean corpuscular hemoglobin (MCH)
Mean corpuscular hemoglobin concentration (MCHC)
Platelet count (PLT)
Differential blood count

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes
All groups: All gross lesions, jejunum, spleen, liver, Mesenteric lymph nodes, ovaries
Control and high dose group: testes, epididymides

Other examinations:

Organ weights: Epididymides, Testes

Statistics:

yes

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Male animal Nos. 33 and 34 of test group 3 (10.0 mg/kg bw/d) showed unsteady gait and swelling of limbs from study day 35 until sacrifice. Poor general state was noted for male animal Nos. 33, 34 and 36 of test group 3 (10.0 mg/kg bw/d) towards the end of the application period. Piloerection was observed in male animal No. 36 of test group 3 (10.0 mg/kg bw/d) from study day 29 onwards as well as in male animal Nos. 33, 34 and 37 on study day 36. No clinical findings were observed for any female animal in test groups 0-3 (0, 0.5, 2.0 and 10.0 mg/kg bw/d).

Mortality:

mortality observed, treatment-related

Description (incidence):

Male animal Nos. 33 and 34 of test group 3 (10.0 mg/kg bw/d) showed unsteady gait and swelling of limbs from study day 35 until sacrifice. Poor general state was noted for male animal Nos. 33, 34 and 36 of test group 3 (10.0 mg/kg bw/d) towards the end of the application period. Piloerection was observed in male animal No. 36 of test group 3 (10.0 mg/kg bw/d) from study day 29 onwards as well as in male animal Nos. 33, 34 and 37 on study day 36. No clinical findings were observed for any female animal in test groups 0-3 (0, 0.5, 2.0 and 10.0 mg/kg bw/d).

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weight change values were significantly lower in male animals of test group 3 (10.0 mg/kg bw/d) from study week 4 to 5 as well as study week 0 to 5.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

increases in white blood cells at 10 mg/kg bw

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

only investigated for testes and epididymides

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Details on results:

Findings at 10 mg/kg bw:
Thickened Jejunum in 15 of 20 animals
Foci in liver in 6/20 animals
Enlarged mesenteric lymph nodes in 17/20 animals
Two animals with enlarged iliac lymph nodes
Enlarged spleen in 4/20 animals, two with foci
multifocal random liver necrosis in males, granulomatous liver inflammation for both sexes
foam cell aggregates in the lamina propria of intestinal villi, in 3 females granulomatous inflammation in the intestinal wall
mesenteric lymph nodes: granulomatous inflammation with necrosis

Increased total white blood cell counts
Increased absolute and relative neutrophils counts
Increased absolute and relative monocytes counts
Increased absolute eosinophils counts
Decreased HGB
Decreased relative lymphocytes counts



Findings at 2 mg/kg bw:
Thickened jejunum in 2 females and 1 male
accumulation of eosinophilic slightly vacuolated histiocytes mostly in the cortex of the mesenteric lypmh node.


Findings at 0.5 mg/kg bw:
Thickened jejunum in 1 animal

Effect levels

Dose descriptor:

NOAEL

Effect level:

2 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Target system / organ toxicity

Critical effects observed:

not specified