Cyclopentane, methoxy-

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 2013-07-06 to 2014-08-25

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study, non GLP, but in compliance with China National Metrology Accreditation, in which the test parameters documented are based on testing guideline.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

no

Remarks:

non GLP, but in compliance with China National Metrology Accreditation

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

445-090-6

EC Name:

-

Cas Number:

5614-37-9

Molecular formula:

C6 H12 O

IUPAC Name:

Cyclopentyl methyl ether

Test material form:

liquid

Specific details on test material used for the study:

Batch No.: 2820246
Purity: 99.9%

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Zhejiang Center of Laboratory animals
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 4-5 weeks old
- Weight at study initiation: 92-130 g
- Fasting period before study:
- Housing: Suspended, wire bottom, stainless steel cages, 2 animals per cage by sex
- Diet (e.g. ad libitum): Conventional laboratory diets, ad libitum
- Water (e.g. ad libitum): Tap water by aseptic filtration, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3°C
- Humidity (%): 30-70%
- Air changes (per hr): 12 air changes/hour
- Photoperiod (hrs dark / hrs light): 12 hours light/dark cycle

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

- PREPARATION OF DOSING SOLUTIONS:
The test substance was emulsified with tween-80 and distilled water by emulsifying machine, and then with distilled water to achieve the required dosages.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

90 days

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (actual dose received)

Doses / concentrations 2

Dose / conc.:

32 mg/kg bw/day (actual dose received)

Doses / concentrations 3

Dose / conc.:

125 mg/kg bw/day (actual dose received)

Doses / concentrations 4

Dose / conc.:

500 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10 per sex per dose

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: based on acute oral LD50 values
- Rationale for animal assignment (if not random): randomly
- Rationale for selecting satellite groups: control group and high dose group
- Post-exposure recovery period in satellite groups: 14 days
- Section schedule rationale (if not random):

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded prior to initiation of study, weekly thereafter, and at death or sacrifice.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

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

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes / No / Not specified
- Time schedule for examinations:

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at end of study
- Anaesthetic used for blood collection: Yes, 10% chloral hydrate
- Animals fasted: Yes, fasted overnight
- How many animals: all animals
- Parameters checked: white blood cell (WBC), red blood cell (RBC), hemoglobin (HGB), hematocrit (HCT), lymphocyte (LYM), granulocyte (GRA), monocyte (MID), red blood cell volume distribution width (RDW), platelet (PLT), prothrombin time (PT), activated partial thromboplastin time (APTT)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at end of study
- Animals fasted: Yes, fasted overnight
- How many animals: all animals
- Parameters checked: total bilitubin (BIL-T), alanine aminotransferase (ALT), aspartate aminotransferase (AST), total protein (TP), albumin (ALB), globulin (GLB), alkaline phosphatase (ALP), urea nitrogen (BUN), creatinine (CREA), glucose (GLU), potassium (K), natrium (Na), chlorine (Cl)

URINALYSIS: Yes
- Time schedule for collection of urine: at end of study
- Parameters checked: appearance (color and clarity), glucose (uGLU), bilirubin (uBIL), ketone (ket), occult blood (RBC), acidity (PH), protein (PRO), urobilinogen (uBG), nitrite (NIT), and leukocytes (LEU)

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
All animals in the study was subjected to a full, detailed gross necropsy which includes careful examination of the external surface of the body, all orifices, the cranial, thoracic and abdominal cavities and their contents. The liver, kidneys, adrenals, testes, epididymis, uterus, ovaries, thymus, spleen, brain and heart of all animals were trimmed of any adherent tissue, and their wet weight taken as soon as possible.

HISTOPATHOLOGY: Yes
Tissues and organs (all gross lesion, brain, spinal cord, stomach, thyroid, thymus, small and large intestines, pancreas, liver, kidneys, adrenals, spleen, heart, trachea and lungs, gonads, uterus, accessory sex organs, prostate, urinary bladder, lymph nodes, peripheral nerve, bone marrow) samples from control and treated rats were cut into thin slices, and then fixed into 4% formaldehyde for more than one week. The tissues and organs were then processed through a graded series of ethanol and xylene, and embedded in paraffin. Organs and tissues sections were stained with hematoxylin and eosin for histopathological examination under a microscope and recorded the abnormal condition.
Full histopathology should be carried out on the preserved organs and tissues of all animals in the control and high dose groups. These examinations should be extended to animals of all other dosage groups, if treatment-related changes are observed in the high dose group.

Statistics:

A parametric or non-parametric test was selected based on the results of normality test and homogeneity of variance test. One-way analysis of variance and Dunnett's t test were used in parameter test. Kruskal-Wallis rank sum test and Wilcoxon-Wilcox rank sum test were used in non-parameteric test. Student's t test was adopted by comparing data between additional high dose and control group, or nonparametric Wilcoxon test. Fisher's exact probability test was used for enumeration data.

Results and discussion

Results of examinations

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

During exposure period and additional 14-day observation, obvious clinical sign of nostril flow fluid was observed in part of high dose group animals (3 females and 1 male) and additional high group (1 female and 1 male), obvious clinical sign of fluffy coat was observed in part of high dose group animals (3 females and 1 male), compared with control or additional control group, there were no statistically significant difference.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

For male rats, body weight gain and body weight changes in high dose group were lower than control group, the differences were statistical significance.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Food consumption per day: for male rats, the average food consumption per day in high dose group were lower than control group.
Relative food consumption: for male and female rats, the average relative food consumption in low, intermediate, high dose groups were closer to control groups.

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

For male rats, total food efficiency in high dose groups were lower than control group.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

For female rats, RBC of intermediate and high dose groups were lower than control group, PT and HCT of high dose group were lower than control group, WBC of intermediate and high dose groups were higher than control group, MO and RDW of high dose group were higher than control group, PT of additional dose group were lower than additional control group, RDW of additional high dose group were higher than additional control group.
For male rats, WBC of intermediate dose group was higher than control group, LYM of high dose group was lower than control group, MO and PT of high dose group were higher than control group, PLT of additional high dose group were lower than additional control group.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

For female rats, A/G of intemediate dose group and high dose groups were lower than control group, BIL-T of high dose group were higher than control group, ALP of additional high dose group were higher than additional control group.
For male rats, GLB of additional high dose group were lower than additional control group, A/G of additional high dose group were higher than additional control group.

Urinalysis findings:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

For female rats, compared with control group, the coefficients of brain were increased in low dose group but no dose-effect relationship, the coefficients of kidney were increased in intermediate dose group, the coefficients of heart, liver, kidney and ovaries were increased in high dose group. The coefficient of liver and kidney were increased in additional high dose group compared with additional control group.
For male rats, compared with control group, the coefficients of brain, liver and testes were increased in high dose group, the coefficients of adrenals were increased in additional high dose group compared with additional control group.

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Effect levels

Dose descriptor:

NOAEL

Effect level:

32 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

food consumption and compound intake

haematology

organ weights and organ / body weight ratios

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

In this repeated dose 90-day oral toxicity study, male and female rats with varying degrees of toxicity effects in intermediate and high dose groups. Obvious toxicity effects were not observed for male and female rats in low dose group. In additional 14 days of observation, no delayed toxicity effects were found in additional high dose group. The NOAEL was estimated to be 32 mg/kg/day in female and male rats in accordance with sample intake of rats.