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Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP - Guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): docosanoic acid
- Physical state: white powder
- Analytical purity: 85.9%
- Lot/batch No.: 60805X
- Impurities (identity and concentrations): C14-C20 fatty acids: 10.9%, C24 fatty acids: 2.3%
- Storage condition of test material: at room temperature in the dark

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Japan, Hino, Tokyo, Japan
- Age at study initiation: 8 weeks
- Weight at study initiation: 312.1 - 363.7 g (males); 205.3 - 230.8 g (females)
- Housing: metal wire floor cages
- Diet (ad libitum): CE-2, CLEA Japan
- Water (ad libitum): tap water
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 ± 1
- Humidity (%): 50 - 65
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: dosing solutions were prepared more frequently than once a week; aliquots of each concentration were kept refrigerated in airtight conditions.

VEHICLE
- Justification for use and choice of vehicle (if other than water): due to the insolubility in water, corn oil was selected as appropriate vehicle
- Amount of vehicle (if gavage): 5 mL/kg bw
- Lot/batch no. (if required): V6H2050
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
- males: 42 days
- females: 14 days prior to mating to day 3 of lactation
Frequency of treatment:
daily
Doses / concentrations
Remarks:
Doses / Concentrations:
100, 300, 1000 mg/kg bw/d
Basis:
actual ingested
No. of animals per sex per dose:
13
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on a preliminary 14 day-repeated dose toxicity study, where no signs of toxicity were found

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least once a day

BODY WEIGHT: Yes
- Time schedule for examinations: males: day 1, 8, 15, 22, 29, 36 and 42; females: premating day 1, 8 and 15; pregnancy day 0, 7, 14 and 20; lactation day 0 and 4

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: 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

HAEMATOLOGY: Yes (males only)
- Time schedule for collection of blood: prior to autopsy
- Anaesthetic used for blood collection: Yes (identity): pentobarbital sodium
- Animals fasted: 18 - 24 h before sacrifice
- How many animals: all surviving animals
- Parameters checked: red blood cell count (RBC), white blood cell count (WBC), haemoglobin content (Hb), mean corpuscular volume (MCV), haematocrit (Ht), mean corpuscular haemoglobin content (MCH), mean corpuscular haemoglobin concentration (MCHC)

CLINICAL CHEMISTRY: Yes (males only)
- Animals fasted: 18 - 24 h before sacrifice
- How many animals: all surviving animals
- Parameters checked: total protein, albumin, total cholesterol concentration, glucose, urea nitrogen, creatinine, alkaline phosphatase activity (ALP), glutamate-oxalacetate-transaminase (GOT), glutamate pyruvate transaminase (GPT), gamma-glutamyl transpeptidase (γ-GTP), triglyceride concentration, inorganic phosphorus, total bilirubin, calcium, sodium, potassium, chlorine, albumin/globulin ratio (A/G ratio)

URINALYSIS: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
Organ weights:
male: heart, liver, kidneys, thymus, testes, epididymides
female: heart, liver, kidneys, thymus

HISTOPATHOLOGY: Yes (control and 1000 mg/kg bw /day group):
male: heart, liver, spleen, kidneys, adrenal, testes, epididymis, brain, thymus, bladder
female: brain, liver, spleen, thymus, kidneys, adrenals, bladder, heart, ovaries
Statistics:
Yates-test, Mann-Whitney-U-test, Fisher exact-test, Bartlett-test, Dunnett-test, Scheffe-test, Kruskal-Wallis-test

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
100 mg/kg bw/d: increased body weight of males (non-adverse)
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
100 mg/kg bw/day (females): significant reduced food consumption during lactation (non-adverse); 100 mg/kg bw/day (males): increased food consumption
Food efficiency:
no effects observed
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):
300 and 1000 mg/kg bw/d: significant decrease of mean corpuscular haemoglobin concentration in males (non-adverse)
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
all dose groups: sporadic effects on glucose, total protein, calcium, ALP (non-adverse)
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
100 mg/kg bw/d: significant liver weight increase in males and signficant decrease in kidney weights in females (non-adervse)
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
all dose groups: sporadic effects on adrenal gland, thyroids, thymus, lung, liver, kidney and spleen in males and females (non-adverse)
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
1000 mg/kg bw/d: sporadic findings in brain, heart, liver, spleen, kidney, adrenals, thymus and testes in males and females (non-adverse)
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
No mortality and no clinical signs were observed.

BODY WEIGHT AND WEIGHT GAIN
Body weight gain in male animals of the 100 mg/kg bw/day group was higher when compared to the 300 and 1000 mg/kg bw/day dose groups and to the control group (10-33%) over the whole treatment period. Since this effect showed no dose-relation and no other effects of toxicity were observed in this dose group, it was regarded as non-adverse.

FOOD CONSUMPTION
Food consumption of males of the 100 mg/kg bw dose group was about 4-10% increased when compared to controls. A significant decreased food consumption was observed in females of the same dose group during lactation. Since this effect was not dose-related, it was regarded to be not treatment related.

HAEMATOLOGY
A significant decrease of 3% of mean corpuscular haemoglobin concentration in males of the 300 and 1000 mg/kg bw/day dose groups was observed after 42 days of treatment. Since this effect showed no clear dose-relation, it was regarded as non-adverse.

CLINICAL CHEMISTRY
A significant decrease of the glucose level (22%) in the 1000 mg/kg bw/day dose group males was observed. Additionally a decrease in total protein (4%) and calcium (5%) in the 300 mg/kg bw/day dose group males and in ALP (16-18%) in males of all dose groups was found. As these effects occured sporadically and showed no dose-relationship, they were regarded as non-adverse.

ORGAN WEIGHTS
In the 100 mg/kg bw/day dose group, a significant liver weight increase (12% (absolute weight), 5% (relative weight)) in males and a signficant decrease in kidney weights in females (8% (absolute weight), 3% (non-adervse)) was observed. As there was no dose relationship and no effects were observed at histopathology, they were regarded as non-adverse.

GROSS PATHOLOGY
In males of the 100 mg/kg bw/dose group, diminishment of thymus (1 animal), accessory spleen (1 animal) and liver changes (yellowish colouring (2 animals), pale spot (1 animal), accentuation of lobular pattern (1 animals)) were observed. In males of the 300 mg/kg bw/dose group, enlargment of thyroid (1 animal), pale lung (1 animal), diminishment of thymus (1 animal) and liver changes ((pale area (2 animals), pale spot (1 animal)) were observed. In males of the 1000 mg/kg bw/dose group, diminishment of thymus (1 animal), small and enlarged thymus (1 animal each), pale lung (1 animal) and yellowish colouring of the liver (2 animals) was observed.
Focal constriction of the spleen was observed in two high-dose group females and adrenal enlargement in one female animal of the 100 mg/kg bw/day dose group. Since the observed effects in male and female animals were found without any dose-relationship and only in few animals, they were regarded as not treatment related.

HISTOPATHOLOGY: NON-NEOPLASTIC
Mycardial degeneration, liver fatty change, changes in spleen and kidney were found to the same extent in few male animals of the highest dose group as compared to males of the control group. One male animal of the highest dose group showed slight fibrosis of the adrenal gland and another one very slight changes of the testes.
The only change differing from controls in females of the high dose group was a thalamus mineralisation in the brain found in one animal. Thus, the effects were regarded as not treatment related.

Effect levels

Dose descriptor:
NOAEL
Remarks:
(repeated dose toxicity)
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no substance-related findings were observed

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion