Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 05 - October 04, 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 guideline
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
17th December, 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 Red 14
Appearance: light red solid
CAS No: 68585-53-5
EC No: 271-559-8

Test animals

Species:
rat
Strain:
Wistar
Remarks:
SPF
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: TOXI COOP ZRT. Cserkesz u. 90. 1103 Budapest, Hungary
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: Young adult rat, 8 weeks old in first and second step
- Weight at study initiation: at starting (first step): 176 - 177 g; at starting (second step): 180 - 182 g
- Fasting period before study: The day before treatment, the food was given back 3 hours after the treatment.
- Housing: Group caging (3 animals/cage) in Type II polypropylene/polycarbonate.
- Diet: Ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, 59494 Soest, Germany; ad libitum.
- Water: Tap water from municipal supply, as for human consumption from bottle ad libitum.
- Acclimation period: 5 days in first step and 6 days in second step
- 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.
- Animal health: Only healthy animals were used for the study. Health status was certified by the study director.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): above 10 air exchanges/hour by central air-condition system.
- Photoperiod (hrs dark / hrs light): 12/12 hours daily, from 6.00 a.m. to 6.00 p.m.




Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
VEHICLE: Aqua purificata Ph.Hg. VIII., Parma Produkt Kft.
- Batch no.: 1606-5508,

CLASS METHOD
- Rationale for the selection of the starting dose: 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 was met.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
3 females/dose/step
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- 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. 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. 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 of survivors performed: yes
- Sacrifice: At the end of the observation period all survivor rats were sacrificed under isofluran anaesthesia.
- Other examinations performed: 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.
Statistics:
The method used is not intended to allow statistical evaluation and the calculation of a precise LD50 value.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Mortality:
No mortality was observed at 2000 mg/kg bw single oral dose of the test item. All female rats in step 1 and step 2 survived until the end of the 14-day observation period.
Clinical signs:
In group 1 and group 2, treated with 2000 mg/kg bw dose, 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:
All animals treated with 2000 mg/kg bw dose survived until the scheduled necropsy on Day 15. Internal necropsy found a colon full of gas in one animal. This alteration could not be related to the test item toxic effect, but was regarded as an individual variation. Moderate hydrometra was found in another animal. Severe hydrometra was detected in two animals. Hydrometra is a physiological finding and connected to the cycle of the animals. 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.

Any other information on results incl. tables

Table 1: Summary of the results

Dose
(mg/kg bw)

Mortality
(dead/treated)

LD50
(mg/kg bw)

GHS/CLP
category

2000

0/6

between 5000 and 2000

No Category

 

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
The method used, was not intended for calculation of a precise LD50 value.
Conclusions:
In an acute oral toxicity study according to OECD guideline 423 a LD50 of above 2000 mg/kg bw was determined.
Executive summary:

The acute toxic class method according to OECD guideline 423 was carried out involving a stepwise procedure with the use of 2000 mg/kg bw as the starting dose in three young adult (8 weeks old) female Wistar 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.

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.

No mortality was observed at single oral dose of 2000 mg/kg bw. In both treatment steps, 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. All organs of the animals treated with 2000 mg/kg bw proved to be free of treatment related gross pathological changes.

In conclusion, an LD50 of above 2000 mg/kg bw was determined.