Reaction mass of N1,N1'-(methanediyldibenzene-4,1 -diyl)diaryldiamine and 2-{4-[(4-aminophenyl)amino]benzyl}-N4-(4-{4-[(4-aminophenyl)amino]benzyl}phenyl)aryldiamine

Acute Toxicity: dermal

Administrative data

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 September 2014 to 16 October 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Test material

Test material information

Constituent 1

Test material form:

solid

Details on test material:

- Appearance/physical state: Black solid
- Storage conditions: Room temperature, protected from light under nitrogen

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

ANIMALS AND ANIMAL HUSBANDRY
- Five male and five female Wistar (RccHan:WIST) strain rats were supplied by Harlan Laboratories UK Ltd, Oxon, UK.
- On receipt the animals were randomly allocated to cages.
- Female rats were nulliparous and non-pregnant.
- After an acclimatisation period of at least five 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.
- 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 Harlan Laboratories Ltd, Oxon, UK) was allowed throughout the study.
- Diet, drinking water and bedding were routinely analysed and were not considered to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
- Temperature and relative humidity were set to achieve limits of 19 to °C and 30 to 70 % respectively.
- Rate of air exchange was at least 15 changes per hour.
- Lighting was controlled by a time switch to give 12 hours continuous light (06:00 to 18:00) and 12 hours darkness.
- The animals were provided with environmental enrichment items which were not considered to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

arachis oil

Details on dermal exposure:

TEST ITEM FORMULATION AND EXPERIMENTAL PREPARATION
- The test item was weighed out according to each animal's individual body weight and moistened with arachis oil BP prior to application.
- Absorption of the test item was not determined.

PROCEDURE
- 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 single group of animals was treated with the test item at a dose level of 2000 mg/kg.
- The appropriate amount of test item, moistened with arachis oil, 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.
- 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 were wiped with cotton wool moistened with arachis oil BP to remove any residual test item.
- As no mortalities were noted a further group of animals (four males and four females) was similarly treated with the test item at a dose level of 2000 mg/kg body weight to give a total of five males and five females. After the 24-Hour contact period the bandages were carefully removed and the treated skin and surrounding hair wiped with cotton wool moistened with arachis oil BP to remove any residual test item. These animals were returned to group housing for the remainder of the test period.
- The animals were observed for deaths or overt signs of toxicity 0.5, 1, 2 and 4 hours after dosing and subsequently once daily for fourteen days.
- After removal of the dressings and subsequently once daily for fourteen days, the test sites were
examined for evidence of primary irritation and scored according to the scale shown below. Any other skin reactions 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.

Duration of exposure:

24 hours

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

Five males and five females

Control animals:

no

Statistics:

DATA EVALUATION
- Data evaluations included the relationship, if any, between the exposure of the animal to the test item and the incidence and severity of all abnormalities including behavioural and clinical observations, gross lesions, body weight changes, mortality and any other toxicological effects.
- Using the mortality data from the study, an estimate of the acute dermal median lethal dose (LD50) was made for the test item.

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

- Individual mortality data are given in Table 1 (attached).
- No unplanned animal deaths took place during the study.

Clinical signs:

- Individual clinical observations are given in Table 1 (attached).
- No signs of systemic toxicity were noted during the observation period.

Body weight:

- Individual body weights and body weight changes are given in Tables 4 (attached).
- Animals showed expected gains in body weight, except for one female which showed body weight loss during the first week but expected gain in body weight during the second week.

Gross pathology:

- Individual necropsy findings are given in Table 5 (attached).
- No abnormalities were noted at necropsy.

Other findings:

- Individual dermal reactions are given in Tables 2 and 3 (attached).
- Very slight erythema and very slight edema were noted at the test sites of the initial two treated animals up to two days after dosing. There were no signs of dermal irritation noted at the test sites of the additional eight treated animals.

Applicant's summary and conclusion

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 > 2000 mg/kg bw.

Executive summary:

GUIDELINE

The study was performed to assess the acute dermal toxicity of the test item in the Wistar strain rat in compliance with the OECD Guideline for the Testing of Chemicals No 402 “Acute Dermal Toxicity” (adopted 24 February 1987) and Method B.3 Acute Toxicity (Dermal) of Commission Regulation (EC) No 440/2008.

 

METHODS

Initially, two animals (one male and one female) were given a single, 24 hour, semi-occluded dermal application of the test item to intact skin at a dose level of 2000 mg/kg body weight. Based on the results of the initial test, a further group of eight animals (four males and four females) was similarly treated. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

 

RESULTS

No animal deaths took place during the study and there were no signs of systemic toxicity. Very slight erythema and very slight edema were noted at the test sites of the initial two treated animals up to two days after dosing. There were no signs of dermal irritation noted at the test sites of the additional eight treated animals. Animals showed expected gains in body weight, except for one female which showed body weight loss during the first week but expected gain in body weight during the second week. 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 > 2000 mg/kg bw.