m,m'-[carbonylbis[imino(3-methoxy-p-phenylene)azo]]bis(benzenesulphonic) acid, compound with 2,2'-iminodiethanol (1:2)

Endpoint summary

Description of key information

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2126 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight). There were no deaths.

The acute dermal median lethal dose (LD50) of the test item in the Wistar strain rat was found to be greater than 2240 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).

No acute inhalation toxicity study is available.

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

The study was performed to assess the acute oral toxicity of the test item in the Wistar strain rat.
Following a sighting test at dose levels of 319 mg/kg (equivalent to 300 mg active ingredient/kg) and 2126 mg/kg (equivalent to 2000 mg active ingredient/kg) , a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, at a dose level of 2126 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

GLP compliance:

yes

Test type:

fixed dose procedure

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

Animal Information
Female Wistar (RccHan™:WIST) strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non-pregnant. After an acclimatization period of at least 5 days the animals were selected at random and given a number unique within the study by indelible ink-marking on the tail and a number written on a cage card. At the start of the study the animals were 8 to 12 weeks of age. The body weight variation did not exceed ±20% of the mean body weight of any previously treated animals.

Animal Care and Husbandry
The animals were housed in groups of up to four in suspended solid-floor polypropylene cages furnished with softwood flake bedding. With the exception of an overnight fast immediately before dosing and for approximately 3 to 4 hours after dosing, free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

Purpose
The study was performed to assess the acute oral toxicity of the test item in the Wistar strain rat.

Justification
Rats are the preferred species of choice as they are historically used for safety evaluation studies and are specified in the appropriate test guidelines.

Test Item Preparation and Analysis
For the purpose of the study the test item was freshly prepared, as required, as a solution in distilled water.
The test item was formulated within 2 hours of being applied to the test system. The suitability of the proposed mixing procedures was determined and specimen formulations were analysed to assess the stability and homogeneity of the test substance in the liquid matrix. These investigations demonstrated that formulations in the concentration range 1 to 200 mg/mL were stable following ambient storage (15-25°C) for one day and following refrigerated storage (2-8°C) for 15 days.
No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulations from this study. This is an exception with regard to GLP and has been reflected in the GLP compliance statement.

Following a sighting test at dose levels of 319 mg/kg (equivalent to 300 mg active ingredient/kg) and 2126 mg/kg (equivalent to 2000 mg active ingredient/kg) , a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, at a dose level of 2126 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

A total of five animals were therefore treated at a dose level of 2126 mg/kg in the study.
All animals were dosed once only by gavage, using a metal cannula attached to a graduated syringe. The volume administered to each animal was calculated according to the fasted body weight at the time of dosing. Treatment of animals was sequential. Sufficient time was allowed between each dose group to confirm the survival of the previously dosed animals.
Clinical observations were made 30 minutes, 1, 2, and 4 hours after dosing and then daily for up to 14 days. Morbidity and mortality checks were made twice daily, early and late during normal working days, and once daily at weekends and public holidays.
Individual body weights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.
At the end of the observation period the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Doses:

1 animal: 319 mg/kg bw = equivalent to 300 mg active ingredient/kg
5 animals: 2126 mg/kg bw = equivalent to 2000 mg active ingredient/kg

No. of animals per sex per dose:

1 female animal: 319 mg/kg bw = equivalent to 300 mg active ingredient/kg
5 female animals: 2126 mg/kg bw = equivalent to 2000 mg active ingredient/kg

Control animals:

no

Details on study design:

Following a sighting test at dose levels of 319 mg/kg (equivalent to 300 mg active ingredient/kg) and 2126 mg/kg (equivalent to 2000 mg active ingredient/kg) , a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, at a dose level of 2126 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

Statistics:

Evaluation of data included identification of the number of animals that died during the study (or that were killed for humane reasons), and determination of the nature, severity, onset and duration of the toxic effects. If possible, the signs of evident toxicity were described. Evident toxicity refers to the toxic effects of sufficient severity that administration of the next higher dose level could result in development of severe signs of toxicity and probable mortality. Effects on body weights and abnormalities noted at necropsy were also identified.
Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD50) of the test item was made.

Key result

Sex:

female

Dose descriptor:

discriminating dose

Effect level:

2 000 mg/kg bw

Based on:

act. ingr.

Remarks on result:

other: There were no deaths.

Mortality:

There were no deaths.

Clinical signs:

other: Hunched posture was noted in one treated animal during the day of dosing. The animal appeared normal one day after dosing. No abnormalities were detected in other treated animals.

Gross pathology:

No abnormalities were noted at necropsy.

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2126 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight). There were no deaths.

Interpretation of results:

GHS criteria not met

Conclusions:

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2126 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight). There were no deaths.

Executive summary:

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

Following a sighting test at dose levels of 319 mg/kg (equivalent to 300 mg active ingredient/kg) and 2126 mg/kg (equivalent to 2000 mg active ingredient/kg) , a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, at a dose level of 2126 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

There were no deaths. Hunched posture was noted in one treated animal during the day of dosing. The animal appeared normal one day after dosing. No abnormalities were detected in other treated animals. The animal showed expected gains in body weight over the observation period. No abnormalities were noted at necropsy.

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2126 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

2 000 mg/kg bw

Quality of whole database:

GLP guideline study.

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental start date: 06 March 2018 Experimental completion date: 20 March 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Physical state / Appearance: amber coloured solid pieces (Sponsors description: Russet coloured crystalline solid)
Expiry Date: 22 June 2018
Storage Conditions: room temperature in the dark

For the purpose of the study the test item was used as supplied. At the request of the sponsor a correction factor of 1.12 was applied to all doses.
The absorption of the test item was not determined.

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Animal Information
Five male and five female Wistar (RccHan:WIST) strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non pregnant. After an acclimatization period of at least 5 days the animals were selected at random and given a number unique within the study by indelible ink marking on the tail and a number written on a cage card. At the start of the study the animals weighed at least 200 g, and were 8 to 12 weeks of age. The weight variation did not exceed ±20% of the mean weight for each sex.

Animal Care and Husbandry
The animals were housed in suspended solid floor polypropylene cages furnished with woodflakes. The animals were housed individually during the 24 Hour exposure period and in groups of five, by sex, for the remainder of the study. Free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 19 to 25 ºC and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Remarks:

moistened with arachis oil BP

Details on dermal exposure:

On the day before treatment the back and flanks of each animal were clipped free of hair.
Using available information on the toxicity of the test item, a group of five male and five female rats was treated with the test item at a dose level of 2240 mg/kg (equivalent to 2000 mg active ingredient/kg body weight).
The calculated volume of test item, ground to a fine powder, moistened with arachis oil BP, was applied as evenly as possible to an area of shorn skin (approximately 10% of the total body surface area) using a graduated syringe. A piece of surgical gauze was placed over the treatment area and semi occluded with a piece of self adhesive bandage. The animals were caged individually for the 24 Hour exposure period. Shortly after dosing the dressings were examined to ensure that they were securely in place.
After the 24 Hour contact period the bandage was carefully removed and the treated skin and surrounding hair wiped with cotton wool moistened with arachis oil BP to remove any residual test item. The animals were returned to group housing for the remainder of the study period.
The animals were observed for deaths or overt signs of toxicity approximately 30 minutes, 1, 2 and 4 hours after dosing and subsequently once daily for 14 days.

Duration of exposure:

24 hr

Doses:

2240 mg/kg (equivalent to 2000 mg active ingredient/kg body weight)

No. of animals per sex per dose:

5 males and 5 females

Control animals:

no

Details on study design:

EVALUATION OF SKIN REACTIONS
Erythema and Eschar Formation Value
No erythema 0
Very slight erythema (barely perceptible) 1
Well-defined erythema 2
Moderate to severe erythema 3
Severe erythema (beef redness) to slight eschar formation (injuries in depth) 4

Edema Formation
No edema 0
Very slight edema (barely perceptible) 1
Slight edema (edges of area well-defined by definite raising) 2
Moderate edema (raised approximately 1 millimeter) 3
Severe edema (raised more than 1 millimeter and extending beyond the area of exposure) 4
Any other skin reactions, if present were also recorded.


Individual body weights were recorded prior to application of the test item on Day 0 and on Days 7 and 14.
At the end of the study the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 240 mg/kg bw

Based on:

test mat.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

act. ingr.

Mortality:

There were no deaths.

Clinical signs:

other: No signs of systemic toxicity were noted during the observation period.

Gross pathology:

No abnormalities were noted at necropsy.

Other findings:

Dermal reactions
Very slight erythema was noted at the treatment sites of all male animals on Days 1 and 2 and persisted at the treatment sites of two male animals on Day 3. Very slight erythema was noted at the treatment sites of two female animals on Day 1. Treatment sites appeared normal on Days 2, 3 or 4.

Interpretation of results:

GHS criteria not met

Conclusions:

The acute dermal median lethal dose (LD50) of the test item in the Wistar strain rat was found to be greater than 2240 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).

Executive summary:

Introduction

The study was performed to assess the acute dermal toxicity of the test item in the Wistar strain rat.

Methods

A group of ten animals (five males and five females) were given a single, 24 hour, semi‑occluded dermal application of the undiluted test item to intact skin at a dose level of 2240 mg/kg body weight(equivalent to 2000 mg active ingredient/kg body weight). Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

Results

Mortality. There were no deaths.

Clinical Observations. There were no signs of systemic toxicity.

Dermal Irritation. Very slight erythema was noted at the treatment sites of all male animals on Days 1 and 2 and persisted at the treatment sites of two male animals on Day 3. Very slight erythema was noted at the treatment sites of two female animals on Day 1. Treatment sites appeared normal on Days 2, 3 or 4.

Body Weight. All animals showed expected gains in body weight over the observation period except for three females which showed a body weight loss during the first week but expected gain in body weight during the second week.

Necropsy. No abnormalities were noted at necropsy.

Conclusion

The acute dermal median lethal dose (LD₅₀) of the test item in the Wistar strain rat was found to be greater than 2240 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

2 000 mg/kg bw

Quality of whole database:

Guideline study.

Justification for classification or non-classification

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2126 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight). There were no deaths.

The acute dermal median lethal dose (LD50) of the test item in the Wistar strain rat was found to be greater than 2240 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).

According to CLP classification criteria (Regulation (EC) No 1272/2008) a classification is not justified.