Disodium 2,2'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[3-bromo-5-butyltoluene-4-sulphonate]

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 August 1993 to 25 August 1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

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

Reference substance name:

Disodium 2,2'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[3-bromo-5-butyltoluene-4-sulphonate]

EC Number:

277-429-7

EC Name:

Disodium 2,2'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[3-bromo-5-butyltoluene-4-sulphonate]

Cas Number:

73398-29-5

Molecular formula:

C36H34Br2N2O8S2.2Na

IUPAC Name:

disodium 2-bromo-3-({4-[(2-bromo-4-butyl-6-methyl-3-sulfonatophenyl)amino]-9,10-dioxo-9,10-dihydroanthracen-1-yl}amino)-6-butyl-4-methylbenzene-1-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

- Appearance: dark blue powder
- Storage conditions: room temperature in the dark

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: twelve to sixteen weeks old
- Weight at study initiation: 2.32 - 2.47 kg
- Housing: The animals were individually housed in suspended metal cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: minimum of five days

ENVIRONMENTAL CONDITIONS
- Temperature: 18 - 24 °C
- Humidity: 56 - 66 %
- Air changes: approximately 15 changes per hour
- Photoperiod: the lighting was controlled by a time switch to give continuous 12 hours light and 12 hours darkness

Test system

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Controls:

not specified

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.5g of the test material, moistened with 0.5 mL of distilled water

Duration of treatment / exposure:

Four hours

Observation period:

72 hours

Number of animals:

Three

Details on study design:

TEST SITE
- Area of exposure: dorsal flank area
- On the day before the test each of a group of three rabbits was clipped free of fur from the dorsal flank area using veterinary clippers. Only animals with a healthy intact epidermis by gross observation were selected for the study.
- On the day of the test a suitable test site was selected on the back of each rabbit. A quantity of 0.5 g of the test material, moistened with 0.5 mL of distilled water, was introduced under a 2.5 cm x 2.5 cm gauze patch and placed in position on the shorn skin. The patch was secured in position with a strip of surgical adhesive tape (BLENDERM: approximate size 2.5 cm x 4.0 cm). To prevent the animals interfering with the patches, the trunk of each rabbit was wrapped in an elasticated corset (TUBIGRIP) and the animals were returned to their cages for the duration of the exposure period.

REMOVAL OF TEST SUBSTANCE
- Four hours after application the corset and patches were removed from each animal and any residual test material removed by gentle swabbing with cotton wool soaked in distilled water followed by diethyl ether.

OBSERVATION TIME POINTS
- Approximately one hour following the removal of the patches, and 24, 48 and 72 hours later, the test sites were examined for evidence of primary irritation and scored according to scale from Draize J.H. (1959) Association of Food and Drug Officials of the United States, Austin, Texas, "The Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics".

SCORING SYSTEM:
EVALUATION OF SKIN REACTIONS
- Erythema and Eschar Formation Value:
No erythema = 0
Very slight erythema (barely perceptible) = 1
We 11-defi ned erythema = 2
Moderate to severe erythema = 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth) = 4

- Oedema Formation Value:
No oedema = 0
Very slight oedema (barely perceptible) = 1
Slight oedema (edges of area well-defined by definite raising) = 2
Moderate oedema (raised approximately 1 millimetre) = 3
Severe oedema (raised more than 1 millimetre and extending beyond the area of exposure) = 4

- Any other skin reactions, if present, were also recorded.
- An additional observation was made on day 7 to assess the reversibility of skin reactions.

INTERPRETATION OF RESULTS
- The scores for erythema and oedema at the 24 and 72-hour readings were totalled for the three test rabbits (12 values) and this total was divided by 6 to give the primary irritation index of the test material. The test material was classified according to the following scheme:
Primary Irritation Index
0 = Non-irritant
> 0 – 2 = Mild irritant
> 2 – 5 = Moderate irritant
> 5 – 8 = Severe irritant
- If irreversible alteration of the dermal tissue is noted in any rabbit, as judged by the Study Director, which may include ulceration and clear necrosis or signs of scar tissue, the test material is classified as corrosive to rabbit skin. Classification according to Draize may, therefore, not be applicable.
- The results were interpreted according to the EEC Commission Directive of 91/325/EEC which adapts Council Directive 67/548/EEC on the regulations relating to the classification, packaging and labelling of dangerous substances as follows:
i) Interpretation according to Annex VI Part II (B): The test material will be classified as irritant or corrosive to the skin according to the following criteria:
a) Corrosion: The test material will be considered to be corrosive and will require the appropriate "C" symbol if it produces full thickness destruction of the skin tissue on at least one animal. If this criterion is not satisfied, the test material will be classified as non-corrosive.
b) Irritation: The test material will be considered to be irritant and will require the appropriate "Xi" symbol if it causes inflammation of the skin, which persists for at least 24 hours, and if either erythema/eschar formation or oedema formation equivalent to a mean value of two or more, calculated for each animal separately is observed in two or more animals. The 24, 48 and 72-hour readings for each animal will be used to calculate the mean values. If these criteria are not satisfied the test material will be classified as non-irritant.
ii) Interpretation according to Annex VI, Part II (D): In addition, the following risk (R) phrases will be assigned to the test material, if appropriate, according to the following criteria:
Corrosive:
R 35 CAUSES SEVERE BURNS: If, when applied to healthy intact animal skin, full thickness destruction of skin tissue occurs as a result of up to three minutes exposure.
R 34 CAUSES BURNS: If, when applied to healthy intact animal skin, full thickness destruction of skin tissue occurs as a result of up to four hours exposure.
Irritant
R 38 IRRITATING TO SKIN: If, when applied to healthy intact skin for up to four hours, significant inflammation is caused which is present 24 hours or more after the end of the exposure period. Inflammation is significant if the mean score for either erythema/eschar formation or oedema formation, observed in two or more animals, is equivalent to a value of two or more. The 24, 48 and 72-hour readings for each animal will be used to calculate the mean scores.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

erythema score

Basis:

animal #1

Remarks:

46 Female

Time point:

24/48/72 h

Score:

0

Max. score:

0

Results 2

Irritation parameter:

erythema score

Basis:

animal #2

Remarks:

50 Female

Time point:

24/48/72 h

Score:

0

Max. score:

0

Results 3

Irritation parameter:

erythema score

Basis:

animal #3

Remarks:

54 Female

Time point:

24/48/72 h

Score:

1

Max. score:

1

Reversibility:

fully reversible within: 7 days

Results 4

Irritation parameter:

edema score

Basis:

animal #1

Remarks:

46 Female

Time point:

24/48/72 h

Score:

0

Max. score:

0

Results 5

Irritation parameter:

edema score

Basis:

animal #2

Remarks:

50 Female

Time point:

24/48/72 h

Score:

0

Max. score:

0

Results 6

Irritation parameter:

edema score

Basis:

animal #3

Remarks:

54 Female

Time point:

24/48/72 h

Score:

0.7

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Irritant / corrosive response data:

- Faint blue-coloured staining caused by the test material was noted at all treated skin sites after patch removal and persisted up to the 72- hour observation. This did not affect evaluation of skin responses.
- Very slight erythema was noted at one treated skin site at the one, 24, 48 and 72-hour observations. Desquamation was noted at two treated skin sites at the 7-day observation.
- Very slight oedema was noted at one treated skin site at the 24 and 48-hour observations.

Applicant's summary and conclusion

Interpretation of results:

other: Not classified in accordance with EU criteria

Conclusions:

Under the conditions of this study the test material was not irritating.

Executive summary:

The skin irritation potential of the test material was investigated in accordance with the standardised guidelines OECD 404 and EU Method B4, under GLP conditions.

On the day before the test each of a group of three female New Zealand White rabbits was clipped free of fur from the dorsal flank area using veterinary clippers. Only animals with a healthy intact epidermis by gross observation were selected for the study. On the day of the test a suitable test site was selected on the back of each rabbit. A quantity of 0.5g of the test material, moistened with 0.5 mL of distilled water, was introduced under a 2.5 cm x 2.5 cm gauze patch and placed in position on the shorn skin. Four hours after application the corset and patches were removed from each animal. Approximately one hour following the removal of the patches, and 24, 48 and 72 hours later, the test sites were examined for evidence of primary irritation.

Faint blue-coloured staining caused by the test material was noted at all treated skin sites after patch removal and persisted up to the 72- hour observation. This did not affect evaluation of skin responses. Very slight erythema was noted at one treated skin site at the one, 24, 48 and 72-hour observations. Desquamation was noted at two treated skin sites at the 7-day observation. Very slight oedema was noted at one treated skin site at the 24 and 48- hour observations.

The test material produced a primary irritation index of 0.5 and was classified as a mild irritant to rabbit skin according to the Draize classification scheme. No corrosive effects were noted.

Under the conditions of this study the test material was not irritating.