Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04 April 2017 - 01 June 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001
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: particulate/powder
Details on test material:
Test Item: Leuco Sulfur Blue 15
Appearance: Dark, near black powder

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals 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): 205 - 211 g
Body weight range
at starting (second step): 216 - 220 g
Acclimatization time: 6 days in first step and 7 days in second step

Housing: Group caging (3 animals/cage)
Cage type: Type III 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.

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.

Vehicle:
Name: Aqua purificata Ph.Hg. VIII.
Batch number: 1702-5502
Date of expiration: 02.08.2017
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. 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 dye/kg bw (corresponding to 2240 mg product/kg bw)
No. of animals per sex per dose:
6 female animals (3 female animals per step)
Control animals:
no
Details on study design:
Animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out 15th day after the treatment.

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 once 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 rats were necropsied.

No death occurred after the single 2000 mg/kg bw oral dose of Leuco Sulfur Blue 15.
There were no toxic clinical signs and any related to the effect 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.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 - < 5 000 mg/kg bw
Based on:
act. ingr.
Mortality:
No death occurred after the single 2000 mg/kg bw oral dose of Leuco Sulfur Blue 15.
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 effect of the test item during the macroscopic examination of animals treated with 2000 mg/kg bw dose.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The oral LD50 was determined to be > 2000 mg dye/kg bw (corresponding to > 2240 mg product/kg bw).
Executive summary:

An acute oral toxicity study according to OECD guideline 423 was performed. The test item was administered oral via garvage. The day before treatment the animals were fasted. Food but not water was withheld overnight. Animals were weighed before the application and food was given back 3 hours after the treatment. The observation period was 14 days. The acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg dye/kg bw (corresponding to 2240 mg product/kg bw) as the starting dose in three female rats. Starting dose was selected on the basis of the available information about the test item. 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. The oral LD50 was determined to be > 2000 mg/kg bw.