Registration Dossier

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1996-11.11 to 1996-12-20
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report Date:
1997

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Qualifier:
according to
Guideline:
EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
Principles of method if other than guideline:
Method: other: OECD Guideline 407 (1981) and Directive 92/69/EEC, B.7
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
cyclooctene produced by Hüls AG in March 1996. Purity 96.5 %; ID No. 0637/81782

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ORGANISMS
- Strain: Wistar (Hsd/Win:WU)
- Source: Harlan Winkelmann GmbH, 33176 Borchen (Germany)
- Age: 6-8 weeks
- Weight at study initiation: females mean 123 g, males mean 134 g
- Number of animals: 5 per dose group and sex, total 60 including  satellite groups
- Diet: Ssniff R 10 diet ad libitum
- Water: tab-water ad libitum:
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3 °C
- Humidity (%): 30 - 70 %
- Photoperiod (hrs dark / hrs light): artificial light from 7.00 a.m. to 7.00 p.m.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
ADMINISTRATION / EXPOSURE 
- Vehicle: corn oil
- Concentration in vehicle: 0 / 25 / 75 / 250 g/l
- Total volume applied: 2 ml/(kg bw * day)
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The homogeneity, the concentration and the stability of the test substance formulations were determined. Samples of all dose levels were taken
repeatedly and were analyzed for concentration of cyclooctene and were within 10% of the nominal concentration. Stability and homogeneity were also demonstrated.
Duration of treatment / exposure:
4 weeks, 28 days
Frequency of treatment:
daily including weekends
Doses / concentrations
Remarks:
Doses / Concentrations:
50, 150, and 500 mg/(kg bw * d)
Basis:
actual ingested
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Details on study design:
Post-exposure period: 14 days (satellite groups) / 2 days (other groups)

SATELLITE GROUPS AND REASONS THEY WERE ADDED: Control and high-dose  satellite groups for recovery studies
Positive control:
no

Examinations

Observations and examinations performed and frequency:
CLINICAL OBSERVATIONS AND FREQUENCY: 
- Clinical signs: recorded daily approximately 1 hour after application  (which was the peak period of the clinical symptoms); additionally  observed 
(noting remarkable changes) each afternoon excluding weekends;  detailed clinical examination once a week.
- Mortality: twice (weekends: once) daily - Body weight: at weekly intervals from prior to first dose until days of  necropsy.
- Food consumption: cagewise (= groupwise, i.e. for 5 animals of same sex  and dose level) at weekly intervals
- Water consumption: observed cagewise, daily
- Ophthalmoscopic examination: during acclimatization and before terminal  bleeding in the high-dose and control animals. An ophthalmoscope 
was used  and one eye was treated with Mydriaticum Stulln(R) prior to investigation.
- Hematology: end of treatment or recovery period: Red blood cell count  (RBC); total white blood cell count (WBC); platelet count (PLT);  
hemoglobin (HGB); hematocrit (HCT); erythrocyte mean corpuscular volume  (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular 
hemoglobin  concentration (MCHC); differential white blood cellcount.
- Biochemistry: end of treatment or recovery period: Sodium; potassium;  calcium; aspartate aminotransferase (AST); alanine aminotransferase  
(ALT); alkaline phosphatase (AP); glucose (GLUC); triglycerides (TRIG);  cholesterol (CHOL); total bilirubin (TBIL); blood urea nitrogen (BUN);  
creatinine (CREA); total protein (TPROT); albumin (ALB).
- Urinalysis: end of treatment or recovery period: Volume (VOL); specific  gravity (SPGR); pH; color.   Semiquantitative: Protein (PROT); 
glucose (GLUC); keton; urobilinogen  (UBG); blood ingredients.    Microscopical urine sediment analysis: Leucocytes (LEUCO); erythrocytes  (ERY); 
bacteria (BAC); epithelial cells, squamous (ECSQ); oxalate  crystals (OXA); triple phosphate crystals (TRIP); urate crystals (URA).
Sacrifice and pathology:
ORGANS EXAMINED AT NECROPSY (MACROSCOPIC AND MICROSCOPIC): 
- Macroscopic: Complete autopsy incuding macropathological examination on  all animals including satellite groups.   
Weights of adrenals, kidneys, liver, spleen, testes.
- Microscopic: preparation of samples of adrenals, aorta (thoracic),  anus, brain, cecum, coagulation gland, colon, concha (tattooed),  duodenum, 
epididymides, eyes, exorbital lacrimal glands, gross lesions,  heart, (not: ileum), sternum, jejunum, kidneys, larynx, liver, lungs,  lymph nodes 
(skin, cervical & mesenteric), mammary gland, muscle  (skeletal), ovaries, esophagus, pancreas, pituitary, prostate, rectum,  salivary glands, 
sciatic nerve, seminal vesicles, skin, spinal cord  (cervical), spleen, stomach, testes, thymus, thyroid / parathyroid,  tongue, trachea, urinary bladder,  uterus / vagina;   examination for selected high-dose and control animals + examination of livers (= target organ) of all other animals
Statistics:
STATISTICAL METHODS: 
- Kruskal Wallis non parametric analysis of variance and in case of  significance Wilcoxon, Mann, and Whitney U-test: absolute body weights  and their changes; absolute and relative organ weights; differential  blood count; urinalysis data.
- One way analysis of variance (ANOVA) incorporating a Bartlett's test  for homogeneity of variance and in case of heterogeneous variances a  Kruskal Wallis test / in case of significant ANOVA a Scheffe test:  Hematological data (except differential blood count) and serum clinical  chemistry data.
- Group means and standard deviations or medians where appropriate.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
effects observed, treatment-related
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
Female and male animals of the high dose groups (including high dose recovery groups) showed mild to moderate clinical symptoms which were most intense about one hour after dosing. No clinical signs were observed in the afternoon. The predominant clinical symptoms were hyperactivity, increased irritability and from the beginning of the second dosing week salivation. Individual animals showed on some days signs like piloerection, gait abnormality and tremor. No clinical signs were observed in the recovery period.

BODY WEIGHT AND WEIGHT GAIN
Affected (p < 0.05) in the male high dose group (total gain -42.0 %) and high dose recovery group (total gain -29.3%). During the recovery period the body weight gain of the dosed males was above that of the controls, but the overall weight gain remained significantly below that of the controls. These findings were considered to be substance related. No significant effect was observed in females.

FOOD CONSUMPTION AND FOOD EFFICIENCY
During the treatment period the food consumption of the male high dose (-11.3 %) and high dose recovery groups (-22.4 %) was reduced and the food conversion rate of these groups was increased (test group: +53.0 %, 4 weeks; recovery group +21.1 %, 6 weeks) when compared with control. Both effects were considered to be substance-related. Statistical significance is not indicated.
No comparable effects were found in females. During the recovery period the food consumption of treated animals (males and females) and control animals was comparatively similar.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
No overt intergroup differences in water consumption were noted.

OPHTHALMOSCOPIC EXAMINATION
No signs of test substance related effects on lenses or cornea were observed in either dose group investigated.

HAEMATOLOGY
Statistically significant differences from controls in treated male and female groups were minor and within the normal range of
historical background data. Individual values of only one male animal fell below the range of the historical control data. Individual values of all the
other animals were within the normal range of historical background data. Therefore these findings were considered to be of minor toxicological
importance.


CLINICAL CHEMISTRY
Statistically significant differences from controls in treated male and female groups were minor and within the normal range of historical backgroud data. A clear pattern in these changes was lacking and examination of the two sexes gave contradictory results. Evidence of reversibility for changes observed at the end of the treatment period were found in the recovery groups. Therefore all these differences were considered to be of minor toxicological importance


URINALYSIS
No findings of toxicological importance were noted.

NEUROBEHAVIOUR
Not examined

ORGAN WEIGHTS
No differences between treated and control of toxicological importance or statistical significance were noted in female animals.
Some statistically significant (p < 0.05) differences to the control occurred in the male groups:
Spleen mid-dose -15.1 %, high-dose -34.2 % (recovery +6.4 %, not significant)
Adrenals mid-dose +9.5 %, high-dose +42.9 % (recovery -2.5 %, not significant)
Gonads high-dose +39.1 % (recovery +18.5 %, p < 0.05)
The differences in organ weights between male dose groups and controls were considered to be of minor toxicological importance.

GROSS PATHOLOGY
No macroscopical lesions were considered to be related to the test item.

HISTOPATHOLOGY: NON-NEOPLASTIC
No histopathological lesions were considered to be related to treatment with cyclooctene. Individual animals in both control and high dose groups showed a mild to moderate chronic pneumonia characterized by multifocal interstitial lymphocyte infiltrates. The lesions found in the lung were most likely caused as part of a virus infection. There was also pigmentation in the mandibular lymph nodes caused by tattooing ears. The other changes described were considered part of the normal background in the rat.


HISTOPATHOLOGY: NEOPLASTIC (if applicable)
none

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
150 mg/kg bw/day (nominal)
Sex:
male/female
Dose descriptor:
LOAEL
Effect level:
500 mg/kg bw/day (nominal)
Sex:
male/female

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The reduced food consumption and the increased food conversion rate noticed especially in high dose males during treatment period was considered to be substance-related. However, evidence of the reversibility of these findings were found at the end of the recovery period. The no-observed-adverse-effect level (NOAEL) is 150 mg/kg bw/day.

At the end of the treatment period the mean bodyweight (BWN)
of the male high dose group was reduced by 19.57 % when
compared with the control. Statistical evaluation of this
data revealed a significant difference between control and
high dose group. The effects on bodyweight and bodyweight
change at the end of the treatment period in high dose
animals were considered to be substance related. At the end
of the recovery period the mean bodyweight (BWN) of the male
high dose recovery group showed no statistically significant
differences when compared with the control. In comparison
with the control the weekly bodyweight change of the high
dose male group was increased during the recovery period,
thus denoting reversibility of the bodyweight loss noticed
during the treatment period. But the recovery period was too
short for the male high dose recovery group to reach the
weight of the control group. Therefore the absolute
bodyweight change (BWCHABS) of the high dose male group at
the end of the recovery period showed still a statistically
significant reduction when compared with the control.

The reduced weight of the high dose male group at the end of
the treatment and recovery period is the explanation for the
increased relative organ weights in these groups.
Histopathological changes of these tissues were not found.

The clinical signs noticed after treatment with test
substance in high dose male and female groups (incl. high
dose recovery groups) were considered to be of toxicological
importance. During the recovery period no clinical signs
were observed.
Therefore, in our experimental conditions, the
no-observed-adverse-effect level (NOAEL) is 150 mg/kg
bw/day.
Executive summary:

In a GLP-study according to TG OECD 407 groups of five Wistar rats per sex received daily doses of 50, 150 and 500 mg Cyclooctene /kg bw/d in corn oil by gavage for a treatment period of 28 days. A control and a high dose recovery group of 5 rats per sex each were observed for 14 days after the end of treatment.

There were no mortalities. Clinical signs, water consumption, urinalysis, ophthalmoscopy terminal blood chemistry and haematology, gross and histopathology were without treatment related effects.

No effects were noted on body weight and food consumption in females.

A statistically significant reduction in body weight and food conversion was noted in the high dose group at the end of the treatment period. This effect was considered to be substance related but proved to be reversible as body weights improved during the recovery period.

Therefore, the no-observed-adverse-effect level (NOAEL) is 150 mg/kg bw/day.