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Description of key information

The LD50 of the test item in the rat was found to be >2000 mg a.i./kg bw (corresponding to > 3640 mg/kg bw test item).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
December 07, 2016-March 16, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
17th December 2001
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
2008 May 30
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
acute toxic class method
Limit test:
yes
Specific details on test material used for the study:
Storage conditions: room temperature, 20 ± 5 °C
Expiration date: 16.06.2020
Correction factor: 1.82
Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals or test system and environmental conditions:
Species and strain: Crl:WI rats
Source: TOXI COOP ZRT. Cserkesz u. 90.
1103 Budapest, Hungary
Hygienic level at arrival: SPF
Hygienic level during the study: Good conventional
Justification of strain: The Wistar rats as a rodent is one of the standard species of acute toxicity studies
Number of animals: 3 animals/group
Sex: Female, nulliparous and non pregnant animals
Age of animals: Young adult rat, 8 weeks old in first and second step
Body weight range
at starting (first step): 211 - 214 g
Body weight range
at starting (second step): 208 - 213 g
Acclimatization time: 6 days in the first step and 7 days in the second step


Housing: Group caging (3 animals/cage)
Cage type: Type II polypropylene/polycarbonate.
Bedding: Laboratory bedding.
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 22 ± 3 °C
Relative humidity: 30 - 70 %
Ventilation: above 10 air exchanges/hour by central air-condition system.

Animals received ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany and tap water from municipal supply, as for human consumption from bottle ad libitum.


In life phase:January 11-27, 2017


Route of administration:
oral: gavage
Vehicle:
other: Aqua purificata Ph.Hg. VIII.
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.

Vehicle:
Name: Aqua purificata Ph.Hg. VIII.
Batch number: 1608-5511
Date of expiration: 11.02.2017
Produced by: Parma Produkt Kft.

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. The dose refers to the dye content (55 %, correction factor of 1.82) and corresponds to 3640 mg product/kg bw. 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 (presented in Appendix VII) was met.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
2000 mg/kg bw : 6 female animals
Control animals:
no
Details on study design:
Duration of observation period after the treatment: 14 days

Frequency of observations:
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.

Body weight measurement:
The body weights were recorded on day 0 (just before the treatment), on day 7 and on day 15 with a precision of 1 g.

Necropsy:
At the end of the observation period all surviving rats were necropsied.

No death occurred after the single 2000 mg/kg bw oral dose of Brown 3.
There were no toxic clinical signs or any treatment related effects of the test item found in body weights and body weight gains during the study. Autopsy revealed no treatment related pathological changes.
Statistics:
No statistics was used in the study.
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Mortality:
No death occurred at 2000 mg/kg bw single oral dose of Brown 3.
Clinical signs:
No treatment related symptoms were observed throughout the 14-day post-treatment period.
Body weight:
The mean body weight of animals treated with 2000 mg/kg bw dose corresponded to their species and age throughout the study.
Gross pathology:
No pathological changes were found related to the treatment with the test item during the macroscopic examination of animals treated with 2000 mg/kg bw dose.

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 starting dose was selected on the basis of the available information about the test item.

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 were met.

Interpretation of results:
GHS criteria not met
Conclusions:
The LD50 value of the test item after single oral application was found to be >2000 mg a.i./kg bw (corresponding to 3640 mg/kg bw test item).
Executive summary:

To assess the acute oral toxicity of the test item in rats, the acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg a.i./kg bw (corresponding to 3640 mg/kg bw test item) as the starting dose in three female rats. No animal died in the first step at 2000 mg a.i./kg bw dose level, so treatment with 2000 mg a.i.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. The animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out on day 15 after the treatment. No mortality and no clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of the experimental animals were normal. The body weight development was unaffected in all animals. All organs of the animals treated with 2000 mg a.i./kg bw proved to be free of treatment related gross pathological changes. The LD50 was found to be >2000 mg a.i./kg bw corresponding to >3640 mg test item/kg bw.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
Value:
2 000 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Acute toxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Oral:

To assess the acute oral toxicity of the test item in rats, the acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg a.i./kg bw (corresponding to 3640 mg/kg bw test item)as the starting dose in three female rats. No animal died in the first step at 2000 mg a.i./kg bw dose level, so treatment with 2000 mg a.i.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. The animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out on day 15 after the treatment. No mortality and no clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of the experimental animals were normal. The body weight development was unaffected in all animals. All organs of the animals treated with 2000 mg a.i./kg bw proved to be free of treatment related gross pathological changes. The LD50 was found to be >2000 mg a.i./kg bw correspinding to >3640 mg test item/kg bw.

Inhalation:

The study does not need to be conducted as exposure of humans via inhalation is not the appropriate route of exposure. Furthermore, physicochemical and toxicological properties do not suggest a potential for a significant rate of respiratory absorption.

Dermal:

No study was conducted as the physiochemical and toxicological properties suggest no potential for a significant rate of absorption through the skin.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data on acute toxicity, the test item is not classified according to Regulation (EC) No 1272/2008 (CLP), as amended f

or the twelfth time in Regulation (EU) No 2019/521.