Reaction mass of trisodium 4-amino-3-[[4-[[4-[(2-amino-4-hydroxyphenyl)azo]phenyl]amino]-3-sulphonatophenyl]azo]-5-hydroxy-6-(phenylazo)naphthalene-2,7-disulphonate and trisodium 4-amino-3-[[4-[[4-[(4-amino-2-hydroxyphenyl)azo]phenyl]amino]-3-sulphonatophenyl]azo]-5-hydroxy-6-(phenylazo)naphthalene-2,7-disulphonate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vivo

Type of information:

other: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

October from 5th to 12th, 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

The test was conducted by means of Read Across approach. The reliability of the source study report is 1. Further information was attached at section 13

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

adopted May 12th, 1981

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Direct Black 168:1 - Similar Substance 01

IUPAC Name:

Direct Black 168:1 - Similar Substance 01

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: BRL, Biological Research Laboratories Ltd.
- Age at study initiation: 15 weeks males, 16 weeks females.
- Weight at study initiation: 2.6 kg males, 2.8 kg females.
- Housing: individually in stainless steel cages equipped with an automatic cleaning and drinking system.
- Diet: pelleted standard Kliba 341, ad libitum.
- Water: community tap water from Itingen, ad libitum.
- Acclimation: four days under test conditions after veterinary examination.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 3 °C.
- Humidity: 40 - 70 %.
- Air changes: 10-15 air changes per hour.
- Photoperiod: 12 hours artificial fluorescent light/12 hours dark.
- Other: music during the light period, air-conditioned room.

Test system

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

water

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Preparation: the test article was applied moistened with bi-distilled water.
- Amount applied: 0.5 g.

Duration of treatment / exposure:

4 hours

Observation period:

72 hours

Number of animals:

2 males and 1 female.

Details on study design:

TEST SITE
- Area of exposure: approximately 24 hours before treatment, the dorsal fur was clipped with an electric clipper, exposing an area of approximately 100 square centimeters (10 cm x 10 cm). The skin of the animals was examined once, 1 day prior to test article administration. Only those animals with no signs of skin injury or irritation were used in the test. On test day the test article was applied to approx. 6 cm2 of the intact skin of the clipped area.
- Type of wrap if used: treated area was covered with a 3.0 cm x 3.0 cm patch of surgical gauze. The gauze was covered with a semi-occlusive dressing. The dressing was wrapped around the abdomen and anchored with an elastic bandage.

REMOVAL OF TEST SUBSTANCE
- Washing: the skin was flushed with lukewarm tap water.
- Time after start of exposure: 4 hours.

OBSERVATIONS
- Viability/Mortality and clinical signs: daily.
- Body Weights: at the start of acclimatization, day 1 of test (application day) and at termination of observation.

SCORING SYSTEM
The skin reaction was assessed at 1, 24, 48 and 72 hours after the removal of the dressing, gauze patch and test article. The corrosive properties of the test article and the staining of the treated skin area related to the test article were described and recorded.
The dermal irritation scores were evaluated in conjunction with the nature and reversibility or otherwise of the responses observed. The individual scores do not represent an absolute standard for the irritant properties of a material, and they are viewed as reference values.

ERYTHEMA AND ESCHAR FORMATION
No erythema.......................................................................................................... 0
Very slight erythema (barely perceptible)............................................................. 1
Well defined erythema........................................................................................... 2
Moderate to severe erythema................................................................................ 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth).... 4
Maximum possible score 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 mm)........................................................ 3
Severe edema (raised more than 1 mm and extending beyond area of exposure).4
Maximum possible score 4
Maximum cumulative score: 8

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

erythema score

Basis:

animal: 3/3

Time point:

24/48/72 h

Score:

< 2.3

Remarks on result:

no indication of irritation

Results 2

Irritation parameter:

edema score

Basis:

animal: 3/3

Time point:

24/48/72 h

Score:

< 2.3

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Test item showed a primary irritation score of 0.00 when applied to intact rabbit skin. Local signs (mean values from 1 hour to 72 hours) consisted of grade 0.00 erythema and grade 0.00 edema.
A black coloration on the back was observed in all animals from 1 to 72 hours.
No destructions or irreversible alterations of the treated skin were observed; no corrosive effect was evident on the skin.

Other effects:

CLINICAL SIGNS AND VIABILITY / MORTALITY
No acute clinical symptoms were observed in the animals during the test period, and no mortality occurred. Body weight gain of all rabbits was similar. A black coloration on the back was observed in all animals from 1 to 72 hours.

Any other information on results incl. tables

The skin irritation scores for each animal are presented below.

Time period	Animal	Sex	Erythema	Edema	Cumulative score	Mean cumulative score
1 hr	69	M	0	0	0	0.00
	70	M	0	0	0
	71	F	0	0	0
24 hrs	69	M	0	0	0	0.00
	70	M	0	0	0
	71	F	0	0	0
48 hrs	69	M	0	0	0	0.00
	70	M	0	0	0
	71	F	0	0	0
72 hrs	69	M	0	0	0	0.00
	70	M	0	0	0
	71	F	0	0	0

The mean scores for skin irritation as a mean of 24, 48 and 72 hours are presented below.

Animal	Sex	Erythema	Edema	Cumulative  score	Mean Cumul. Score (24/48/72)
69	M	0.00	0.00	0.00	0.00
70	M	0.00	0.00	0.00
71	F	0.00	0.00	0.00

Applicant's summary and conclusion

Interpretation of results:

other: not classified, according to the CLP Regulation (EC) No 1272/2008

Conclusions:

The substance is not a skin irritant.

Executive summary:

The irritancy potential of the test material to the skin of New Zeland white rabbit was assessed, according to the OECD Guideline 404. The intact skin of the rabbits was exposed for 4 hours to the substance, in a semi-occlusive dressing. Animals were examined for signs of erythema and edema and the responses were scored at 1 hour following the patch and substance removal/washing, and then 24, 48 and 72 hours later.

No acute clinical symptoms were observed and no mortality occurred. A black coloration on the back was observed in all animals from 1 to 72 hours, but no signs of irritations were observed. Test item showed a primary irritation score of 0.00 when applied to intact rabbit skin. Local signs (mean values from 1 hour to 72 hours) consisted of grade 0.00 erythema and grade 0.00 edema.

Conclusion

The substance is not a skin irritant.