Registration Dossier

Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October - November 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: crystalline
Details on test material:
- Physical state: see above
- Appearance: see above
Specific details on test material used for the study:
TEST MATERIAL
- Substance identification/name in the report: LZ 596
- Molecular formula: C21H30N2
- Batch no.: 1229
- Analysis date: August 12, 2014
- Date of production: August 11, 2014

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage conditions: Controlled room temperature (15-25ºC, <70 RH%), protected from light and humidity.
- Stability under test conditions: stable for min. 3 years from manufacturing date
- Expiry date: August 10, 2017
- Safety precautions: Routine safety precautions (lab coat, gloves, safety glasses, face mask) for unknown materials were applied to assure personnel health and safety.

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS:
- Species / Strain: Crl:(WI)BR rats
- Source: Toxi-Coop Zrt. 1103 Budapest, Cserkesz u. 90.
- Hygienic level: SPF at arrival and kept in a good conventional environment during the study
- Number of animals: preliminary study 2 female animals/dose / main study: 5 animals/dose
- Age of animals: Young adult rats, females were nulliparous and nonpregnant
- Housing: single housing (main study)
- Sex: female and male
- Body weight range: in preliminary study at starting: 252 - 285 g / in main study at starting: males 239 - 248 g / females 254 - 287 g
- Feeding: ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany
- Water supply: tap water from municipal source
- Acclimatization time: in preliminary study: 47 days / in main study: 61 days in females and 5 days in malesTEST ANIMALS

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 30 - 70 %
- Air changes: 10-15 air exchanges/hour by central air-condition system
- Photoperiod (hrs dark / hrs light): Artificial light, from 6 a.m. to 6 p.m.

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
water
Remarks:
The test substance was moistened sufficiently with water to ensure good contact with the skin.
Details on dermal exposure:
The back of animals was shaven (approximately 10 % area of the total body surface) 24 hours prior to the treatment. The test item was applied in a single 2000 mg/kg bw dose uniformly over the shaved skin throughout a 24-hour exposure period. Sterile gauze pads were placed on the skin of rats. These gauzes were kept in contact with the skin by a patch with adhesive hypoallergenic plaster. The entire trunk of the animal was wrapped with semi
occlusive plastic wrap for 24 hours. At the end of the 24-hour exposure period any residual test item was removed using body temperature water
Duration of exposure:
24 hours
Doses:
- Preliminary test: 5, 50, 300, 2000 mg/kg
- Main test: 2000 mg/kg
No. of animals per sex per dose:
- Preliminary test: 2 females/dose
- Main test: 5 males and 5 females
Control animals:
no
Details on study design:
OBSERVATION:
A single administration was performed by dermal route and was followed by a 7-day observation period in preliminary study and 14-day observation period in main study.

MORTALITY:
Inspection for signs of morbidity and mortality were made twice daily at the beginning and end of the working day.

CLINICAL OBSERVATIONS:
Careful clinical observation was made at the following intervals: 1h, 5h after the treatment and once each day for 7 days thereafter in preliminary study and for 14 days thereafter in main study. Individual observations included the skin and fur, eyes and mucous membranes, and also respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern as well. Particular attention was directed to the observation of tremors,
convulsions, salivation, diarrhoea, lethargy, sleep and coma.

BODY WEIGHTS:
The body weight was recorded on day 0 (shortly before the treatment) in preliminary study and on day 0 (just before the treatment), on day 7 and on day 15 with a precision of 1 g in main study.

PATHOLOGY:
At the end of the observation period all rats of main study were sacrificed under isofluran anaesthesia. After examination of the external appearance, the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed, and any abnormality was recorded with details of its location, colour, shape and size. Animals of preliminary study were humanely sacrificed on Day 7.
Statistics:
no statistics perfomed

Results and discussion

Preliminary study:
There were no deaths at 5, 50, 300 and 2000 mg/kg bw dose levels. Therefore, in the main test animals were administered the limit dose of 2000 mg/kg bw.
Effect levels
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality were observed over a 14-day period following a 24-hour dermal exposure to the test item in male and female rats.
Clinical signs:
No behavioural changes, symptoms of dermal irritation and corrosion or signs of systemic toxity were noted during the study.
Body weight:
The mean body weight of the male animals corresponded to their species and age throughout the study. A body weight loss was observed in two females treated with 2000 mg/kg bw test item between Day 0 and Day 7. Both body weight losses were very slight (approx. 0.4 % in both cases) and the body weight of animals exceeded the original body weight by the end of study, thus it can be evaluated as an individual variation without toxicological meaning.
Gross pathology:
All animals survived until the scheduled necropsy on Day 15. Pale coloured kidneys as an internal necropsy finding were found in one male.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Under the experimental conditions, the acute dermal LD50 value of the test item proved to be greater than 2000 mg/kg bw in male and female WISTAR rats.
Executive summary:

An acute dermal toxicity study was performed with the test item in Crl:(WI)BR rats, in accordance with testing methods EC B.3 and OECD 402. A limit test was carried out. Single groups of male and female animals (n=5 animals/sex) were exposed to the test item at 2000 mg/kg bw by dermal route. The test item was applied in original form and left in contact with the skin for 24 hours, followed by a 14-day observation period. No mortality occurred during the study. Neither male nor female animals treated at 2000 mg/kg bw showed behavioural changes and no signs of systemic toxicity were observed during the study. No signs of irritation and corrosion were noted. The body weight development was undisturbed in male animals and slight body weight loss was observed in two females on first week, but it could not be evaluated as a toxic effect of test item. No macroscopic alterations of organs referred to the systemic toxic effect of the test item were seen during necropsy.

Under the experimental conditions the acute dermal LD50 value of the test item LZ 596 proved to be greater than 2000 mg/kg bw in male and female Crl:(WI)BR rats.