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

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 October 2008 - 08 November 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study; well documented study report

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2009

Report date:

2009

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Test material form:

solid

Details on test material:

- Appearance/physical state: Black lumps
- Storage conditions: Room temperature in the dark

Test animals

Species:

rabbit

Strain:

New Zealand White

Remarks:

albino; SPF-quality

Details on test animals or test system and environmental conditions:

ANIMALS
- Source: Harlan France SARL, Gannat, France
- Animals used within the study were at least 6 weeks old and body weights were at least 1.0 kg
- Identification: Earmark
- Health inspection: A health inspection was performed prior to commencement of treatment to ensure that the animals were in a good state of health. Special attention was paid to the skin to be treated, which was intact and free from abnormalities.

ANIMAL HUSBANDRY
- Animals were housed in a controlled environment in which optimal conditions were considered to be approximately 15 air changes per hour, a temperature of 21.0 ± 3.0 °C (acutal range 22.2 to 23.2 °C), a relative humidity of 30 to 70 % (actual range 32 to 64 %) and 12 hours artificial fluorescent light and 12 hours darkness per day.

ACCOMODATION
- Animals were individually housed in labelled cages with perforated floors (Ebeco, Germany) dimensions 67 x 62 x 55 cm and shelters (Ebeco, Germany) dimensions 40 x 32 x 23 cm.
- Acclimatisation period was at least 5 days before start of treatment under laboratory conditions.

DIET
- Pelleted diet for rabbits (Global Diet 2030 (irradiated) Mucedola srl, Sellimo Milanese, Italy, supplied by Harlan Netherlands B.V., Horst, The Netherlands approximately 100 g per day plus hay (TecniLab-BMI BV, Someren, The Netherlands) provided at least three times per week.

WATER
- Free access to tap water.

ANALYSIS
- Results of analysis for each batch of diet (nutrients and contaminants), hay and water were assessed and did not reveal any findings that were considered to have affected the integrity of the study.

Test system

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

water

Controls:

other: Adjacent areas of the untreated skin of each animal served as controls

Amount / concentration applied:

TEST MATERIAL
- Test substance was ground to a powder using a pestle and mortar prior to weighing and the test substance was moistened with water (Elix, Millipore S.A.S, Molsheim, France) immediately before application to ensure close contact with the animal's skin.
- Amount(s) applied (volume or weight with unit): 0.5 grams of test substance

Duration of treatment / exposure:

4 hours

Observation period:

1, 24, 48 and 72 hours

Number of animals:

3 males

Details on study design:

STUDY DESIGN
- An in vitro study was considered but a negative result was anticipated and this would need to have been confirmed by an in vivo study.
- The study was performed in a stepwise manner and was started by treatment of a single rabbit (sentinel).
- The two other animals were treated in a similar manner one wek later after considering the degree of skin irritation observed in the first animal.
- No histopathology was performed.

TREATMENT
- Approximately 24 hours before treatment, the dorsal fur was clipped with electric clippers exposing an area of approximately 10 x 15 cm (150 cm2).
- To facilitate scoring, treated skin areas were re-clipped at least 3 hours before the observations.
- The test substance (0.5 g) was moistened with 0.8 mL of the vehicle and applied to the skin of one flank, using a metalline patch of 2 x3 cm.
- The patch was mounted on a Micropore tape, which was wrapped around the abdomen and secured with Coban elastic bandage.
- Four hours after the application, the dressing was removed and the skin cleansed of residual test substance using tap water and watery ethanol (50% v/v).

OBSERVATIONS
- Mortality/viability: Twice daily
- Toxicity: At least once daily
- Body weight: Day of treatment (prior to application) and at termination
- Irritation: The eyes of each animal were examined at approximately 1, 24, 48 and 72 hours after removal of the dressings. Irritation scores according to the attached scheme were recorded together with a description of other (local effects).

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

erythema score

Basis:

animal #1

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 966

Results 2

Irritation parameter:

erythema score

Basis:

animal #2

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 26

Results 3

Irritation parameter:

erythema score

Basis:

animal #3

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 28

Results 4

Irritation parameter:

edema score

Basis:

animal #1

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 966

Results 5

Irritation parameter:

edema score

Basis:

animal #2

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 26

Results 6

Irritation parameter:

edema score

Basis:

animal #3

Remarks:

mean score

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Remarks on result:

other: Animal 28

Irritant / corrosive response data:

- Individual skin irritation scores are presented in Table 1 (attached).
- No skin irritation was caused by exposure to the test item for four hours.
- There was no evidence of a corrosive effect on the skin.

Other effects:

- Grey/brown staining of the treated skin by the test substance was observed on Day 1 which did not hamper the scoring of the skin reactions. No remnants of the test substance were present on the skin.
- No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Applicant's summary and conclusion

Interpretation of results:

not irritating

Conclusions:

The test item was considered to be non-irritating based on a primary irritation index of zero.

Executive summary:

GUIDELINE

The study was conducted in accordance with OECD No 404 (2002) "Acute Dermal Irritation/Corrosion", EU No Method B.4 (2008) "Acute Toxicity: Dermal Irritation/Corrosion", EPA OPPTS 870.2500 (1998) "Acute Dermal Irritation" and JMAFF guidelines (2000) including the most recent revisions.

METHODS

Three rabbits were exposed to test item (0.5 g) moistened with water by application onto clipped skin for 4 hours using a semi-occlusive dressing. Approximately 24 hours before treatment, the dorsal fur was clipped with electric clippers exposing an area of approximately 10 x 15 cm (150 cm2). To facilitate scoring, treated skin areas were re-clipped at least 3 hours before the observations. The test substance (0.5 g) was moistened with 0.8 mL of the vehicle and applied to the skin of one flank, using a metalline patch of 2 x3 cm. The patch was mounted on a Micropore tape, which was wrapped around the abdomen and secured with Coban elastic bandage. Four hours after the application, the dressing was removed and the skin cleansed of residual test substance using tap water and watery ethanol (50% v/v).

RESULTS

No skin irritation was caused by exposure to the test item for four hours. Grey/brown staining of the treated skin by the test substance was observed on Day 1, which did not hamper the scoring of skin reactions. No remnants of the test substance were present on the skin.

CONCLUSION

The test item was considered to be non-irritating based on a primary irritation index of zero.