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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10/30/91 to 11/28/91
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report date:
1992

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Version / remarks:
Paris Cedex, February 24, 1987
Qualifier:
according to guideline
Guideline:
EU Method B.1 (Acute Toxicity (Oral))
Version / remarks:
Official Journal of the European Communities No.L251196, September 1984
GLP compliance:
yes
Test type:
standard acute method
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
N-cyclohexyl-N-methylcyclohexylamine
EC Number:
231-453-4
EC Name:
N-cyclohexyl-N-methylcyclohexylamine
Cas Number:
7560-83-0
Molecular formula:
C13H25N
IUPAC Name:
N-cyclohexyl-N-methylcyclohexanamine
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 70891CJ
- Expiration date of the lot/batch: not indicated
- Purity: 99%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature in the dark
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: stability in corn oil not indicated by the sponsor
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not indicated

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:
The test substance was dosed undiluted as delivered by the sponsor for administration of animals in groups 1 and 2.
The formulation for group 3 was prepared immediately prior to dosing. The test substance was weighed into a glass flask on an analytical balance and corn oil (w/w) was added. Adjustment was made for specific gravity of test substance and vehicle.
Homogeneity of the test substance in vehicle was obtained by using an electric shaker.

Concentration, stability and homogeneity of test substance in vehicle were not determined by analytical procedures.

- Preliminary purification step (if any): no

FORM AS APPLIED IN THE TEST (if different from that of starting material)
in corn oil as vehicle for group 3

OTHER SPECIFICS:
clear colourless to amber liquid; density 0.9235 g/ml


Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: BRL Ltd., Basel, Switzerland (outbred, SPF-Quality)
- Females (if applicable) nulliparous and non-pregnant: not indicated
- Age at study initiation: 8 - 9 weeks
- Weight at study initiation:
males 183 - 302 g
females 153 - 190 g
- Fasting period before study: Feed was withheld overnight prior to dosing until approximately 3-4 hours after administration of the test substance.
- Housing: group housed, 5 per sex to a cage, using labelled polycarbonate cages containing purified sawdust as bedding material (Woody SPF)
- Diet (e.g. ad libitum): ad libitum (standard pelleted laboratory animal diet Kliba 343)
- Water (e.g. ad libitum): ad libitum (tap-water)
- Acclimation period: at least 5 days before start of treatment, under laboratory conditions


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21°C
- Humidity (%): 55 %
- Air changes (per hr): 15
Occasional fluctuations from these optimal conditions were noted, but were considered not to have affected study integrity.

- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
Treatment/Observation:
October 30, 1991 / 15 days (Group 1)
November 06, 1991 / 2 days (Group 2)
November 14, 1991 / 15 days (Group 3)
Termination: November 28, 1991

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
VEHICLE
- Concentration in vehicle:
Group 3: 200 mg/kg bw
Group 3: 0.4 ml/kg bw (50% in corn oil)

Calculated as follows for group 3:
dose level (g/kg) : dosis concentration (g/ml).

Concentration, stability and homogeneity of test substance in vehicle were not determined by analytical procedures.

- Justification for choice of vehicle: not indicated
- Lot/batch no. (if required): not indicated
- Purity: not indicated
- Specific gravity: 0.92

MAXIMUM DOSE VOLUME APPLIED: 0.866 ml/kg body weight


CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose:

Initially a "Pilot study" was conducted in which three groups, each comprising of 1 male and 1 female rat were orally dosed with POLYCAT 12 at 2000, 1000 or 500 mg/kg body weight. All animals died within 4 hours of dosing. Lethargy, piloerection, ataxia, convulsions and tremors were observed among these animals prior to death. Based on these results, initially one group of animals consisting of five males and five females was selected and dosed at 800 mg/kg body weight. Due to the number of deaths, two further groups were selected for the Full Study and dosed at 400 and 200 mg/kg body weight. As the calculated dose volume of group 3 would have been 0.217 ml/kg body weight (0.2 : 0.9235 = 0.217), a very small volume would have had to have had been administered (approximately 0.05 ml/animal). According to the policy in the testing facility, syringes used are not considered of sufficient accuracy for administration at this level. Therefore the test substance was diluted with corn oil, to a concentration of 50% (w/w).
Doses:
Dose level
Group 1: 800 mg/kg bw
Group 2: 400 mg/kg bw
Group 3: 200 mg/kg bw
Dose volume:
Group 1: 0.866 ml/kg bw
Group 2: 0.433 ml/kg bw
Group 3: 0.4 ml/kg bw (50% in corn oil)

Calculated as follows for groups 1 and 2: dose level (g/kg) : density (0.9235 g/ml).
Calculated as follows for group 3: dose level (g/kg) : dosis concentration (g/ml).
No. of animals per sex per dose:
Group: 5 males and 5 females
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days

- Frequency of observations and weighing
Mortality/Viability: at periodic intervals on the day of dosing (day 1) and twice daily thereafter for 14 days
Body weights: days 1 (pre-administration), 8 and 15 and at death (if found dead after day 1)
Clinical signs: at periodic intervals on the day of dosing (day 1) and once daily thereafter for 14 days

- Necropsy of survivors performed: yes
All animals surviving to the end of the observation period were sacrificed by oxygen/carbon dioxide asphyxiation. All animals assigned to the study were subjected to necropsy and descriptions of all macroscopic abnormalities recorded.

- Other examinations performed:
All signs of reaction to treatment were recorded with particular attention paid to changes in the skin, fur, eyes and mucous membranes, as well as to behaviour pattern, tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
The following data were recorded on data sheets and transcribed for compilation and analysis: signs and symptoms, mortality, body weights, macroscopic findings at necropsy.
Statistics:
The LD50 values and the associated 95% confidence interval, the slope of the dose mortality curve were calculated using the Maximum likelihood method (Finney, D.J. Probit Analyses, 3rd Edn., Cambridge University Press, 1971).

Results and discussion

Preliminary study:
Initially a "Pilot study" was conducted in which three groups, each comprising of 1 male and 1 female rat were orally dosed with POLYCAT 12 at 2000, 1000 or 500 mg/kg body weight. All animals died within 4 hours of dosing. Lethargy, piloerection, ataxia, convulsions and tremors were observed among these animals prior to death.
Effect levelsopen allclose all
Key result
Sex:
male
Dose descriptor:
LD50
Remarks:
95% CL not calculated; G >1
Effect level:
>= 267 mg/kg bw
Based on:
test mat.
Remarks on result:
other: Estimated value, calculations invalid according to Finney.
Sex:
female
Remarks on result:
not determinable because of methodological limitations
Remarks:
Due to the abnormal mortality distribution among the females, no calculation of LD50 value for females alone according to the Maximum likelihood method of Finney was possible.
Key result
Sex:
male/female
Dose descriptor:
LD50
Remarks:
95% CL not calculated; G >1
Effect level:
289 mg/kg bw
Based on:
test mat.
Remarks on result:
other: Estimated value, calculations invalid according to Finney.
Mortality:
For observed mortality rates see table below. All deaths occurred on days 1 and 2.
Clinical signs:
Signs of systemic toxicity observed during the study period in each dose group were as follows:
800 mg/kg body weight: lethargy, convulsions, tremors, piloerection, ataxia, emaciation, hunched posture, less than expected amount of faeces.
400 mg/kg body weight: lethargy, convulsions, prostration, piloerection, ataxia.
200 mg/kg body weight: lethargy, piloerection, ataxia.
All surviving animals dosed at 200 mg/kg body weight had recovered by day 2, while the single surviving male dosed at 800 mg/kg body weight showed clinical signs until day 14.
Body weight:
The single surviving male dosed at 800 mg/kg body weight was noted with severe body weight loss over the first week of observation, and improved body weight gain thereafter. The body weight gain shown by the majority of surviving animals treated at 200 mg/kg body weight, considered to be similar to that expected of normal untreated animals of the same age and strain. However, low body weight gain noted in one female and slight body weight loss noted in another female were seen over the second week of the study.
Gross pathology:
Macroscopic post mortem examination of the animals that died during the study revealed the following toxicological significant abnormalities:
800 mg/kg body weight: pale appearance of non-glandular stomach, dark red appearance of liver.
400 mg/kg body weight: whitish contents in stomach, hardened and yellow/brown discoloured processus papillaris of the liver.
Macroscopic post mortem examination of the surviving animals at termination did not reveal any abnormalities.

Any other information on results incl. tables

The following mortality rates were observed:

Dose level

mg/kg

Males

Females

Sexes

combined

800

4/5

5/5

9/10

400

5/5

5/5

10/10

200

1/5

0/5

1/10

Applicant's summary and conclusion

Conclusions:
Due to the mortality distribution, only estimated oral LD50 values of POLYCAT 12 could be calculated for the sexes combined and for males alone.
These were 289 mg/kg body weight for the sexes combined and 267 mg/kg body weight for males alone. Due to the abnormal mortality distribution among the females, no calculation of LD50 value for females alone according to the Maximum likelihood method of Finney was possible.
Executive summary:

The purpose of this study was to assess the toxicity of POLYCAT 12 when administered to rats as a single oral dose.

The study was carried out in accordance with OECD Guideline No. 401, "Acute Oral Toxicity" and EEC Directive 84/449/EEC, Part B.1, "Acute Toxicity-Oral". POLYCAT 12 was administered by oral gavage to five rats of each sex per group, at 800, 400 or 200 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed at the end of the experimental period.

The incidence of mortality among the sexes combined from high to low dose group, was 9, 10 and 1. All deaths occurred on days 1 and 2.

Clinical signs related to toxicity of POLYCAT 12 included mainly lethargy, piloerection, ataxia, convulsions and tremors. All surviving animals dosed at 200 mg/kg body weight had recovered by day 2.

One surviving male, treated at 800 mg/kg body weight, was noted with severe body weight loss over the first week of study and improved body weight gain thereafter. The body weight gain shown by the majority of surviving animals treated at 200 mg/kg body weight, was considered to be similar to that expected of normal untreated animals of the same age and strain.

Macroscopic post mortem examination of the animals that died during the study revealed whitish contents in stomach, pale appearance of the non-glandular stomach, hardened and yellow/brown discoloured processus papillaris of the liver and dark red appearance of the liver, that were considered toxicological significant.

Macroscopic post mortem examination of the surviving animals at termination did not reveal any abnormalities.

Due to the mortality distribution, only estimated oral LD50values of POLYCAT 12 could be calculated for the sexes combined and for males alone. These were 289 mg/kg body weight for the sexes combined and 267 mg/kg body weight for males alone.

Due to the abnormal mortality distribution among the females, no calculation of LD50value for females alone according to the Maximum likelihood method of Finney was possible.

Based on these results and according to the EEC criteria for classification and labelling requirements for dangerous substances and preparations (EEC Directive 91/325/EEC, Amendment to Annex VI of the EEC Directive 67/548/EEC), POLYCAT 12 should be classified as harmful.