Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Initial day of treatment 16 January 2019 Experimental completion date (final macroscopic examination) 05 February 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
712 C-02-190. 2002
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
fixed dose procedure
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder

Test animals

Species:
rat
Strain:
Wistar
Remarks:
RccHan™:WIST albino rats
Sex:
female
Details on test animals and environmental conditions:
The animals were allocated without conscious bias to cages within the treatment groups. They were housed in groups of one or four rats of the same sex.

Each animal was identified uniquely within the study by tail marking. Each cage label was color-coded and was identified uniquely with the study number, dose level and animal mark.

The animals were allowed to acclimatize to the conditions described below for at least six days before treatment. For those animals selected for this study, their body weights were in the range 160 to 169 g and they were approximately eight to twelve weeks of age prior to dosing (Day 1). The body weight variation did not exceed  20% of the mean body weight of any previously treated animals.

Animal Care and Husbandry
Animals were housed inside a limited access rodent facility (Building F21, Room 044/045). The facility was designed and operated to minimize the entry of external biological and chemical agents and to minimize the transference of such agents between rooms.

The animal room was kept at positive pressure with respect to the outside by its own supply of filtered fresh air, which was passed to atmosphere and not re-circulated. The temperature and relative humidity controls were set to maintain the range of 20 to 24C and 40 to 70% respectively. Any minor deviations from these ranges would not have had an adverse effect on the animals and would not affect the integrity or validity of the study. Artificial lighting was controlled to give a cycle of 12 hours continuous light and 12 hours continuous dark per 24 hours. Environmental parameters are archived with the departmental raw data.

Periodic checks were made on the number of air changes in the animal rooms. Temperature and humidity were monitored daily.

Alarms were activated if there was any failure of the ventilation system, or temperature limits were exceeded. A stand-by electricity supply was available to be automatically brought into operation should the public supply fail.

The cages were solid bottomed polycarbonate cages with a stainless steel mesh lid. Each cage contained a quantity of autoclaved softwood bark-free fiber bedding. Cages, food hoppers, water bottles and bedding were changed at appropriate intervals.

The animals were allowed free access to a standard rodent diet (Teklad 2014C Diet), except for overnight prior to and approximately four hours after dosing. This diet contained no added antibiotic or other chemotherapeutic or prophylactic agent.

Potable water taken from the public supply was freely available via polycarbonate bottles fitted with sipper tubes.
Each cage of animals was provided with an Aspen chew blocks for environmental enrichment. Chew blocks were provided throughout the study and were replaced when necessary. Each cage of animals was provided with a plastic shelter for environmental enrichment, which was replaced at the same time as the cages.

Each batch of diet was analyzed routinely by the supplier for various nutritional components and chemical and microbiological contaminants. Supplier’s analytical certificates were scrutinized and approved before any batch of diet was released for use. The quality of the water supply is governed by regulations published by the Department for Environment, Food and Rural Affairs. Certificates of analysis were received routinely from the water supplier. Certificates of analysis were received routinely from the supplier of the chew blocks. Since the results of these various analyses did not provide evidence of contamination that might have prejudiced the study, they are not presented.

No other specific contaminants that were likely to have been present in the diet or water were analyzed, as none that may have interfered with or prejudiced the outcome of the study was known.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Remarks:
Distilled
Details on oral exposure:
Using available information on the toxicity of the test item, 2000 mg/kg body weight was chosen as the starting dose - One single animal was treated.

In the absence of mortality or toxicity at a dose level of 2000 mg/kg body weight, an additional group of four animals animals was treated at 2000 mg/kg.
Doses:
2000 mg/kg
No. of animals per sex per dose:
1 female at initial dosine
4 females in additional dose group
Control animals:
no
Details on study design:
Dose Administration
The appropriate dose volume of the test item was administered to each rat by oral gavage using a plastic syringe and plastic catheter based on the most recent body weight of each animal.
A record of the weight of each formulation dispensed and the amount remaining after dosing was made. The balance of these two weights was compared with the predicted usage as a check that the doses had been administered correctly.
Formulations were stirred before and throughout the dosing procedure.

Results and discussion

Preliminary study:
Absence of mortality and toxicity.
Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Mortality:
There were no deaths during the study.
Clinical signs:
Clinical signs observed were dark blue faeces seen in all females and dark skin colour on the extremities seen in four females dosed at 2000 mg/kg. These signs were seen on Day 2. Recovery of animals was complete by Day 3.
Body weight:
All animals were considered to have achieved satisfactory body weight gains throughout the study.
Gross pathology:
No abnormalities were noted in any animal at the macroscopic examination at study termination on Day 15.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The acute median lethal oral dose (LD50) to rats of the test item was demonstrated to be greater than 2000 mg/kg body weight. The substance is not classified, according to the Globally Harmonised System (GHS). It did not meet the criteria for classification according to Regulation (EC) No. 1272/2008 on the Classification, Labelling and Packaging of Substances and Mixtures
Executive summary:

The study was performed to assess the acute oral toxicity of the test item to the rat.

Fasted female rats received a single oral gavage dose of the test item, formulated in distilled water, at the following dose level:

Sighting investigation: 2000 mg/kg body weight

Main study: Based on the results of the sighting investigation a further four fasted females were similarly dosed at 2000 mg/kg body weight.

During the study, clinical condition, body weight and macropathology investigations were undertaken.

Results

There were no deaths during the study.

Clinical signs observed were dark blue faeces seen in all females and dark skin colour on the extremities seen in four females dosed at 2000 mg/kg. These signs were seen on Day 2. Recovery of animals was complete by Day 3.

All animals were considered to have achieved satisfactory body weight gains throughout the study.

No abnormalities were noted in any animal at the macroscopic examination at study termination on Day 15.

Conclusion

The acute median lethal oral dose (LD50) to rats of the test item was demonstrated to be greater than 2000 mg/kg body weight.