(E)-N'-(2-cyano-4-(3-(1-hydroxy-2-methylpropan-2-yl)thioureido)phenyl)-N,N-dimethylformimidamide

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental start date (initial animal arrival) 05 March 2020. Experimental completion date (final macroscopic examination) 31 March 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

E-N'-{2-cyano-4-[3-(2-hydroxy-1,1-dimethylethyl)-thioureido]-phenyl}-N,N-dimethylformamidine

Cas Number:

1429755-57-6

Molecular formula:

C15H12N5OS

IUPAC Name:

E-N'-{2-cyano-4-[3-(2-hydroxy-1,1-dimethylethyl)-thioureido]-phenyl}-N,N-dimethylformamidine

Specific details on test material used for the study:

Test item SPS 5290 (Stage 3)
Intended use Industrial chemical
Appearance Yellow crystalline powder
Storage conditions 10-30°C in the dark
Supplier Sponsor
Batch number 80034242B
Expiry date 05 July 2020
Purity 99.5%

Test animals

Species:

rat

Strain:

other: RccHan™:WIST albino

Sex:

female

Details on test animals or test system and environmental conditions:

Animal Information
Healthy nulliparous and non-pregnant female RccHan™:WIST albino rats were obtained fromEnvigo RMS (UK) Ltd.
The animals were allocated without conscious bias to cages within the treatment groups. They were housed in groups of one or four rats.
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 five days before treatment. For those animals selected for this study, their body weights were in the range 162 to 177 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 Aspen chew blocks or balls for environmental enrichment. Chew blocks or balls 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 or balls. 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:

methylcellulose

Remarks:

1% w/v aqueous

Details on oral exposure:

Formulation
The test item was formulated at concentrations of 30 and 200 mg/mL in the vehicle, by adding the vehicle to the test item and mixing. The formulation was administered at a volume of 10 mL/kg body weight.
The test item formulations were prepared on the day of dosing.
The absorption of the test item was not determined.
Determination of the homogeneity, stability and purity of the test item or test item formulations were not undertaken as part of this study.
Detailed records of test item usage were maintained. The amount of test item necessary to prepare the formulations and the amount actually used were determined on each occasion. The difference between these amounts was checked before the formulations were dispensed.

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.
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.

In the absence of data regarding the toxicity of the test item, 300 mg/kg was chosen as the starting dose.

Doses:

300 and 2000 mg/kg

No. of animals per sex per dose:

1 female at 300 mg/kg
5 females at 2000 mg/kg

Control animals:

no

Details on study design:

Serial Observations
Mortality
Cages of rats were checked at least twice daily for any mortalities.
Clinical Observations
Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1. On subsequent days, animals were observed once in the morning and again at the end of the experimental day (with the exception of Day 15 - morning only). The nature and severity, where appropriate, of any clinical signs and the time were recorded at each observation. All animals were observed for 14 days after dosing.
Body Weight
The weight of each rat was recorded on Days -1 (not reported), 1 (prior to dosing), 8 and 15. Individual weekly body weight changes and group mean body weights were calculated.

Terminal Investigations
Method of Kill
All animals were humanely killed on Day 15 by carbon dioxide asphyxiation and subsequently exsanguinated.
Macroscopic Pathology
All animals were subject to a macroscopic examination which consisted of opening the cranial, thoracic and abdominal cavities. The macroscopic appearance of the brain, cecum, duodenum, heart, kidneys, small and large intestine, liver, lungs and bronchi, spleen, stomach, subcutaneous tissue and urinary bladder was recorded.

Results and discussion

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

There were no deaths during the study.

Clinical signs:

There were no clinical signs of reaction to treatment throughout the study.

Body weight:

A low body weight gain was noted for two females (animal numbers 44 and 45) between Day 8 and Day 15. All other animals were considered to have achieved satisfactory body weight gains throughout the study

Gross pathology:

Macroscopic examination at the sighting study termination on Day 15 revealed congestion (darkened tissue) of the lung and bronchi in one female (animal number 41). No controlabnormalities were revealed in any other animal at the macroscopic examination at this time.

Applicant's summary and conclusion

Interpretation of results:

Category 5 based on GHS criteria

Conclusions:

The acute median lethal oral dose (LD50) to rats of SPS 5290 (Stage 3) was demonstrated to be greater than 2000 mg/kg body weight.
SPS 5290 (Stage 3) is included in Category 5/Unclassified, according to the Globally Harmonised System (GHS)

Executive summary:

The study was performed to assess the acute oral toxicity of SPS 5290 (Stage 3) (an industrial chemical), to the rat.

Fasted female rats received a single oral gavage dose of the test item, formulated in 1% w/v aqueous methylcellulose, at the following dose levels:

Sighting investigations: 300 and 2000 mg/kg body weight

Main study: Based on the results of the sighting investigations 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 or clinical signs observed throughout the study. Two main study animals were considered to have a low body weight gain between Day 8 and

Day 15. All other animals were considered to have achieved satisfactory body weight gains throughout the study.

Congestion (darkened tissue) of the lungs and bronchi was observed in a single animal at the necropsy and macroscopic examination following completion of the sighting study at 300 mg/kg. No abnormalities were noted in any other animal at the necropsy and macroscopic examination at study termination on Day 15.

Conclusion

The acute median lethal oral dose (LD50) to rats of SPS 5290 (Stage 3) was demonstrated to be greater than 2000 mg/kg body weight.

SPS 5290 (Stage 3) is included in Category 5/Unclassified, according to the Globally Harmonised System (GHS),