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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
05 Nov 2019 to 19 Dec 2019
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2020
Report Date:
2020

Materials and methods

Test guideline
Qualifier:
no guideline followed
GLP compliance:
no
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Remarks:
Crl:CD
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, NC
- Females (if applicable) nulliparous and non-pregnant: [yes/no]
- Age at study initiation: 10 weeks old
- Weight at study initiation: weighed between 200 and 418 g
- Fasting period before study: no
- Housing: individually housed in solid-bottom cages containing appropriate bedding equipped with an automatic watering valve
- Diet (e.g. ad libitum): PMI Nutrition International, LLC Certified Rodent LabDiet 5002, ad libitum
- Water (e.g. ad libitum): Municipal tap water after treatment by reverse osmosis and ultraviolet irradiation, ad libitum
- Acclimation period: at least 5 days

DETAILS OF FOOD AND WATER QUALITY: The feed was analyzed by the supplier for nutritional components and environmental contaminants. Periodic analysis of the water was performed. It was considered that there were no known contaminants in the feed or water that would interfere with the objectives of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-26°C
- Humidity (%): 30-70%
- Air changes (per hr): Ten or greater air changes per hour with 100% fresh air
- Photoperiod (hrs dark / hrs light): 12 hour light/12 hour dark

Administration / exposure

Route of administration:
oral: gavage
Details on route of administration:
Oral administration was the preferred route of exposure specified in the relevant test guidelines.
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: Test substance dosing formulations were prepared at appropriate concentrations to meet dose level requirements. The dosing formulations were prepared approximately weekly and an adequate amount of each formulation was dispensed into daily aliquots, which were stored in a room with controls set to maintain 18°C to 24°C, protected from light, until use.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Not specified
- Concentration in vehicle: 0, 25, 125, 250 mg/mL
- Amount of vehicle (if gavage): 4 mL/kg bw
- Lot/batch no. (if required): 2IH0387
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose formulation samples were collected in duplicate from all groups on day 1 and day 8 for concentration analysis. Analyses were performed by a gas chromatography method. Stability analyses performed previously demonstrated that the test substance is stable in the vehicle when prepared and stored under the same conditions at concentrations bracketing those used the DRF study.
Duration of treatment / exposure:
14 days
Frequency of treatment:
daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control group
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Remarks:
Low dose
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
Middle dose
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
High dose
No. of animals per sex per dose:
5 males and 5 females per dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dosage levels were selected by the Sponsor in consultation with the study director based, in part, on the LD50 value of >5000 mg/kg from an acute oral toxicity study. The high-dosage level represented the limit dose as defined by OECD and EPA. The lower dosage levels were selected at intervals that were predicted to be narrow enough to reveal any dose-related trends. Based on the data through Day 6, the decision was made by the Sponsor to not dose the 5th group.
- Rationale for animal assignment (if not random): random
Positive control:
Not used

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily observations for general health/mortality and moribundity were performed. Cage side observations were performed 0.5 to 2 hours postdose.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Day 1 (prior to dosing), weekly (± 2 days) during the study period, and on the day of the scheduled necropsy.

BODY WEIGHT: Yes
- Time schedule for examinations: Animals were weighed daily during the study period and on the day of the scheduled necropsy. A fasted weight was recorded on the day of the scheduled necropsy.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Food consumption was quantitatively measured daily throughout the dosing period.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:
- Animals fasted: Yes
- How many animals: all animals
- Parameters checked in table [No.1] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes. Main study animals were subjected to a complete necropsy examination, which included evaluation of all external surfaces and orifices; the cranial cavity and external surfaces of the brain; and thoracic, abdominal, and pelvic cavities with their associated organs and tissues. The liver and kidney weights were recorded during necropsy. At the time of necropsy, following organ weight collection, the liver, kidneys, and gross lesions were retained and placed in 10% neutral-buffered formalin for possible histopathology.

HISTOPATHOLOGY: No
Statistics:
Data collected during the predose period were not tabulated, summarised, or statistically analysed. All statistical analyses were performed within the respective study phase, unless otherwise noted. Numerical data collected on scheduled occasions were summarised and statistically analysed according to sex and occasion.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
There were no test substance-related clinical observations. All clinical observations in the test substance-treated groups were noted with similar incidence in the control group, were limited to single animals, were not noted in a dose-related manner, and/or were common findings for laboratory rats of this age and strain.
Mortality:
no mortality observed
Description (incidence):
All animals survived until scheduled necropsy.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Test substance-related lower body weight gains were noted from day 1 to 7 and from day 1 to 14 in the 500 and 1000 mg/kg bw/day group males. These differences led to a lower mean day 14 body weight that was only considered different from concurrent controls for the 1000 mg/kg bw/day males.
There were no other test substance-related effects on body weights.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Test substance-related lower mean food consumption was noted in the 1000 mg/kg bw/day group males over Days 1 to 14, compared to the control group. This correlated to the lower body weight gains observed in the same group.
There were no other test substance-related effects on food consumption.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
Serum chemistry parameters were unaffected by test substance administration.
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Organ weights were unaffected by test substance administration.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no test substance-related macroscopic findings at the scheduled necropsy. The single macroscopic finding noted was considered to be spontaneous and/or incidental in nature and unrelated to test substance administration.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not examined

Effect levels

Dose descriptor:
NOAEL
Remarks on result:
not determinable
Remarks:
Dose range-finder study

Target system / organ toxicity

Critical effects observed:
no

Any other information on results incl. tables

Table 2. Comparison of body weight gains

Sex

Males

Males

Males

Males

Females

Females

Females

Females

Dose level (mg/kg bw/day)

0

100

500

1000

0

100

500

1000

Body weight gain (day 1 to 7)

41.4

39.2

35.6

35.0

11.4

17.8

11.8

14.2

Body weight gain (day 1 to 14)

70.2

67.6

61.6

50.4

25.0

33.4

21.4

27.4

Values in bold are considered to be test substance-related.

Applicant's summary and conclusion

Conclusions:
In the 14-day dose range-finding study, which was not conducted according to a guideline or GLP, a NOAEL was not concluded. Based on the results of this study, oral administration of the test substance to rats at dosage levels of 100, 500, and 1000 mg/kg bw/day for 14 consecutive days resulted in lower mean body weights and/or lower cumulative body weight gains in males at 500 and 1000 mg/kg bw/day. The results of this study are used to select dose levels for the main 90-day oral repeated dose toxicity study.