Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 September - 04 October, 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
17th December 2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
30 May 2008
Deviations:
no
Qualifier:
according to
Guideline:
other: Regulation (EC) No. 1272/2008
Version / remarks:
1272/2008
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes

Test material

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

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Crl:WI
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Justification of strain: The Wistar rats as a rodent is one of the standard species of acute toxicity studies
- Source: TOXI COOP ZRT. Cserkesz u. 90. 1103 Budapest, Hungary
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 8 weeks
- Weight at study initiation: 186 - 187 g
- Fasting period before study:
- Housing: Group caging (3 animals/cage) in Type II polypropylene/polycarbonate cages.
- Diet: ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany, ad libitum
- Water: tap water from municipal supply, ad libitum
- Acclimation period: 5 days in first step, 6 days in second step
- Hygienic level at arrival: SPF
- Hygienic level during the study: Good conventional
- The diet and drinking water are periodically analysed and are considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): > 10 air exchanges/hour by central air-condition system.
-The temperature and relative humidity were recorded daily during the study.
- Photoperiod (hrs dark / hrs light): (12/12) from 6.00 a.m. to 6.00 p.m.






Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
All doses were formulated in the vehicle. Concentration of formulations was adjusted to maintain a treatment volume of 10 mL/kg bw. The test item was applied in a concentration of 200 mg/mL. The correction factor was taken into consideration in the course of the making of solution. Formulations were prepared just before the administration and were stirred continuously during the treatment.
A single oral administration - followed by a fourteen-day observation period - was performed by gavage. The day before treatment the animals were fasted. The food but not water was withheld overnight. Animals were weighed before the application and the food was given back 3 hours after the treatment.
The acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg/kg bw as the starting dose in three female rats. No animal died in the first step at 2000 mg/kg bw dose level, so treatment with 2000 mg/kg bw was repeated on further three female rats. No animal died in the second step, too, so the test was finished, the stopping criteria of Annex 2d of OECD Guideline No. 423 was met.
Doses:
2000 mg dyestuff/kg bw (corresonding to 2240 mg product/kg bw)
No. of animals per sex per dose:
3 animals/group
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Fasting: The day before treatment. The food but not water was withheld overnight and the food was given back 3 hours after the treatment.
- Frequency of observations and weighing: Animals were observed individually after dosing at least once during the first 30 minutes, then 1 h, 2 h, 3 h, 4 h after the treatment and twice each day for 14 days thereafter (General state, external appearance, behavior and clinical symptoms). The body weights were recorded on day 0 (just before the treatment), on day 7 and on day 15.
- Necropsy of survivors performed: Yes, at the end of the observation period all survivor rats 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.
- Other examinations performed: clinical signs, body weight, organ weights, histopathology, other: Individual observations were performed on the skin and fur, eyes and mucous membranes and also respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
Statistics:
The method used is not intended to allow the calculation of a precise LD50 value. The mean of the body weight and body weight gain were calculated by Excel spreadsheet software

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Remarks on result:
other: corresonding to >2240 mg product/kg bw
Mortality:
No lethality was noted at a single oral dose of 2000 mg/kg bw.
Clinical signs:
After both administrations of the test item, no clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of experimental animals were normal.

Body weight:
The body weight development was undisturbed in all animals.
Gross pathology:
All organs of the animals treated with 2000 mg/kg bw proved to be free of treatment related gross pathological changes.

Any other information on results incl. tables

Dose
(mg/kg bw)

Mortality
(dead/treated)

LD50
(mg/kg bw)

2000

0/6

between 5000 and 2000

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
In an acute oral toxicity study according to OECD guideline 423 in rat, a LD50 of above 2000 mg dyestuff/kg bw was determined.

Executive summary:

In an acute oral toxicity study according to OECD guideline 423 in rat, the acute oral toxicity of the test item was determined. The test item was applied via oral gavage to three female Wistar rats at a starting dose of 2000 mg dyestuff/kg bw (corresponding to 2240 mg product/kg bw). Since no mortality was observed, the administration was repeated in three female rats. Again, no mortality was observed. No clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of experimental animals were normal. The body weight development was undisturbed in all animals.

The method used is not intended to allow the calculation of a precise LD50 value. However, since no animal died after oral adminsitration, a LD50 of above 2000 mg dyestuff/kg bw was determined. The test item is not considered to be classified for acute oral toxic under Regulation (EC) 1272/2008 as amended for the tenth time in Regulation (EC) No 2017/776.