Y-513

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:

10.08.2007 - 30.10.2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was performed according to the internationally recognized guidelines and according to GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: 28-day repeated dose toxicity study in mammalian species prescribed in 'Concerning testing methods relating to the new chemical substances' (No 1121002, Pharmaceutical and Food Safety Bureau, MHLW

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): Y-513
- Substance type: organic
- Physical state: Yellow powder
- Purity test date: no data
- Expiration date of the lot: unknown
- Stability under test conditions: stable
- Storage condition of test material: dark place at room temperature (tolerance temperature 10-30°C)

Test animals

Species:

rat

Strain:

other: Crl: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Japan
- Age at study initiation: 5 weeks
- Weight at study initiation: 147.4 - 167.9g (males); 121.9 - 144.1g (females)
- Housing: stainless steel hanger cages with wire-mesh floor, individually housed
- Diet: MF pellet diet, ad libitum
- Water: dechlorinated tap water, ad libitum
- Acclimation period: 10 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.4 - 24.0
- Humidity (%): 48.1 - 63.0
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Gum arabic in water (5.0 w/v%)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The test substance and gum arabic were weighted accurately (the final concentration of gum arabic in the formulation was 5.0 w/v%) and suspended in purified water into a mortar to make 10.0 w/v% formulation. The lower concentrations of 2.5 and 0.5 w/v% suspensions were diluted from 10.0 w/v% formulation.

- Rate of preparation of dosing solution (frequency): Once per 7 days
- Storage temperature of dosing solution: stored in the dark and cold place


VEHICLE
- Justification for use and choice of vehicle (if other than water): the vehicle was chosen in a 14-day pilot study
- Concentration in vehicle: 0, 0.5, 2.5 and 10%
- Amount of vehicle (if gavage): 10 mL/ kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The homogeneity and stability analyses were performed. The formulations were stable for 9 days and showed good homogeneity.
Analytical verification of test substance in the formulations was done by HPLC. The relative values to the setting values of all the formulations were in the range 100 +/- 10%.

Duration of treatment / exposure:

28 days

Frequency of treatment:

daily

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 50, 250, 1000 mg/kg bw/ day
Basis:
actual ingested

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

- Dose selection rationale:
A 14-days oral repeated dose toxicity study was performed with exposure at 25, 250, 500 and 1000 mg/kg bw/ day. Based on absence of adverse effects in the animals (males and females), the highest dose was chosen at 1000 mg/kg bw/ day.

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 3 times per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: day 1, 3, 8, 12, 17, 21, 26, 28 exposure period and day 1, 5, 10, 14 recovery period

FOOD CONSUMPTION:
- Time schedule for examinations: days 1,3, 8, 15, 22, 28 dosing period; days 4, 8, 14 recovery period

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the completion of dosing period and of the recovery period
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes
- How many animals: All animals
- Parameters examined:
Red blood cell count
White blood cell count
Hemoglobin
Hematocrit value
Mean corpuscular volume
Mean corpuscular hemoglobin
Mean corpuscular hemoglobin concentration
Platelet count
Reticulocyte count
Prothrombin time
Activate partial thromboplastin time
Differentiation of leukocytes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at sacrifice
- Animals fasted: Yes
- How many animals: All animals
- Parameters examined:
Asparate aminotransferase
Alanine aminotransferase
Alkaline phosphatase
Cholinesterase
y-Glutamyl transpeptidase
Total cholesterol
Trigliceryde
Glucose
Total protein
Albumin
a/g ratio
Blood urea nitrogen
Creatinine
Total bilirubin
Calcium
Inorganic phosphorus
Sodium
Potassium
Chloride

URINALYSIS: Yes
- Time schedule for collection of urine: at the completion of dosing period
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- How many animals: all animals
- Parameters examined:
Urine volume
Color
Turbidity
Specific gravity
pH
Protein
Glucose
Occult blood
Urinary sediments (conducted for Control and 1000 mg/kg bw/ day groups)

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: week 4
- Dose groups that were examined: all animals
- Battery of functions tested: reflex test, grip strength and motor activity

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
Gross necropsy incl body surfaces, all orifices, subcutaneous tissue, cranial, thoracic, abdominal and pelvic cavities;
Organ weights: liver, heart, kidneys, testes, epididymides, ovaries, brain, spleen, thymus, adrenals.

HISTOPATHOLOGY: Yes
Trachea, lungs, stomach, intestines, liver, heart, kidneys, urinary bladder, testes, epididymides, prostate, seminal vesicles, ovaries, uterus, vagina, brain, spinal cord, sciatic nerve, bone marrow, axillary and mesentreic lymph nodes, spleen, thymus, pituitary glands, thyroid, adrenals, eye balls

Other examinations:

14 days recovery groups were additionally included (0 and 1000 mg/kg bw)

Statistics:

Data regarding body weights (excluding those at the time of necropsy), food intake, items of hematological examinations, items of blood chemical examinations, urine volume and specific gravity, organ weights, grip strength and locomotor activity count were analyzed using the Bartlett's test for homogeneity of variance. If the variances were homogeneous at a significance level of 5%, the difference between the vehicle control group and each of the treatment groups was analyzed by the Dunnett's test.
If the variances were not homogeneous, the difference between the vehicle control group and each of the treatment group was analyzed by the nonparametric Dunnett's test.
For defecation (number of feces) and urination (number of urine pools), the difference between the vehicle control group and each of the treatment groups was analyzed by the nonparametric Dunnett's test.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
No mortality occurred in males or females.

Yellow stool was observed in both sexes of the 250 mg/kg bw/day or more. This was not observed 3 days after termination of dosing period in both sexes of the 1000 mg/kg bw/day recovery group.
In the vehicle control group, loss of hair and scab formation of the neck in one male were observed as accidental changes.

No abnormalities attributable to the test substance were observed in males. The number of urinations was significantly increased in one female of the 1000 mg/kg bw/day in week 1 of the dosing period.

BODY WEIGHT AND WEIGHT GAIN
No abnormalities observed in males or females.

FOOD CONSUMPTION
No abnormalities observed in males or females.

HAEMATOLOGY
No abnormalities attributed to the test substance were observed in males or females.
Hemoglobin concentration was significantly increased in males of the 50 mg/kg bw/day at termination of the dosing period. Ratios of neutrophiles and lymphocytes were increased statistically in male of the 1000 mg/kg b.w. at termination of recovery period. There changes were not considered of toxicological significance since they were in the range of historical data of the laboratory.

CLINICAL CHEMISTRY
No abnormalities attributed to the test substance were observed in males or females. Calcium concentration was significantly inreased in males of the 50 mg/kg bw/day, chloride was decreased in males of the 50 mg/kg bw/ day and 250 mg/kg bw/ day at termination of the dosing period. Potassium was increased significantly in males of the 1000 mg/kg bw/day group at termination of the recovery period. There changes were not considered of toxicological significance since they were in the range of historical data of the laboratory.

URINALYSIS
No abnormalities attributed to the test substance were observed in males or females.
Specific gravity was decreased significantly in female of the 1000 mg/kg bw/day group. These changes were not considered of toxicological significance since they were in the range of historical data of the laboratory.

NEUROBEHAVIOUR
No abnormalities were noted in males or females.
Locomotor activities were significantly increased in the measure periods of 40-50 min, 50-60 min and total measurement time 0-60 minutes in the female of 250 mg/kg bw/day, but these changes were not considered to be of toxicological significance.


ORGAN WEIGHTS
No abnormalities were observed in males or females.
Incease in absolute and relative thymus weights was measured in females of the 50 mg/kg bw/day group (increases of appr. 49% and 43% for resp. absolute and relative weight ). In absence of a dose-relationship of this effect, this change was not considered of toxicological significance. Increase in absolute and relative epididymides weight in males of the 1000 mg/kg bw/day was seen after the recovery period (increase of appr. 6% and 12% for absolute and relative weight resp.). These changes were not considered of toxicological significance since they were in range of historical data of the laboratory.

GROSS PATHOLOGY
No substance-related abnormalities were observed in males or females.

HISTOPATHOLOGY
No substance-abnormalities were observed in males or females.

Effect levels

Dose descriptor:

NOEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: No effects seen at highest dose (1000 mg/kg bw/day).

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Based on the results of a 28-day repeated dose toxicity study performed according to OECD guideline 407 and GLP principles, a NOEL of 1000 mg/kg bw/ day is established for Y-513 based on no effects seen at the highest dose level of 1000 mg/kg bw/ day.

Executive summary:

A 28-day repeated dose toxicity study was performed according to OECD guideline 407 and GLP principles. Test substance Y-513 was administered by gavage to male and female rats at 0, 50, 250 or 1000 mg/kg bw/day for 28 days. The study included groups for which the exposure period was followed by a 14-day recovery period (0 and 1000 mg/kg bw/ day). No substance related clinical signs were noted, no changes in body weight gain occurred. No abnormalities were noted in sensorimotor functions, food intake, hematological examinations, blood chemistry, urinalyses or pathological examinations. No differences were observed between the exposed and unexposed groups in the recovery period. Based on the absence of effects, the NOEL of Y-513 in rats for daily exposure during 28 days was established at 1000 mg/kg bw/day.