Disodium {sulfonylbis[4,1-phenylenediazene-2,1-diyl(1-ethyl-6-hydroxy-4-methyl-2-oxo-1,2-dihydropyridine-5,3-diyl)]}dimethanesulfonate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental Starting Date: 28-Sep-2010 Experimental Completion Date: 20-Oct-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

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:

Acid Yellow RN 2903

IUPAC Name:

Acid Yellow RN 2903

Details on test material:

Identification: Acid Yellow RN 2903
Description: Yellow to orange powder
Batch Number: Vers. Kilo 6, 26.05.2009
Purity: 65.7 %
Correction Factor for the Purity: No
Stability of Test Item: Stable under storage conditions
Expiry Date: 01-Aug-2015
Storage Conditions: At room temperature (range of 20 ± 5 °C, provided by Harlan Laboratories Ltd.), light protected.
Safety Precautions: Routine hygienic procedures will be used to ensure the health and safety of the personnel.

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Test Animals:
Animals: Young Adult New Zealand White Rabbit, SPF
Rationale: Recognized by international guidelines as a recommended test system.
Breeder: Harlan Laboratories U.K. Ltd., Hillcrest, Dodgeford Lane, Belton, Loughborough, Leics, LE12 9TE /UK
Number of Animals per Test: 3 males
Age (when treated): 13 weeks
Body Weight Range (when treated): 2800 g – 2914 g
Identification: By unique cage number and corresponding ear number.
Randomization: Selected by hand at time of delivery. No computer generated randomization program.
Acclimatization: Five days under laboratory conditions after health examination. Only animals without any visual signs of illness were used for the study.

Environmental Conditions:
Conditions: Standard Laboratory Conditions. Air-conditioned with 10-15 air changes per hour, and continuously monitored environment with a room temperature of 17-23 °C and a relative humidity between 30-70%, automatically controlled light cycle of 12 hours light and 12 hours dark and music played during the daytime light period.
Accommodation: Individually in stainless steel cages equipped with feed hoppers and drinking water bowls.
Diet: Pelleted standard Teklad Global High Fiber Rabbit Diet 2031C (batch no. 25/10 provided by Provimi Kliba AG, 4303 Kaiseraugst / Switzerland) available ad libitum. Results of analyses for contaminants are archived at Harlan Laboratories Ltd. A piece of wood (batch no. 102240, imported by Indulab AG, Gams / Switzerland from ABEDD® - LAB & VET GmbH, 1160 Vienna / Austria) and a haystick 4642 (batch no. 80/09, Provimi Kliba AG, 4303 Kaiseraugst / Switzerland) were provided for environmental enrichment.
Water: Community tap water from Füllinsdorf ad libitum. Results of bacteriological, chemical and contaminant analyses are archived at Harlan Laboratories Ltd.

Test system

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

water

Remarks:

The test item was moistened with 0.5 mL purified water.

Controls:

no

Amount / concentration applied:

The test item was applied as a weight of 0.5 g/animal, the dose specified in the test guidelines for
solid test items.

Duration of treatment / exposure:

4 hours

Observation period:

14 days

Number of animals:

3 males

Details on study design:

Purpose:
The purpose of this primary skin irritation study was to assess the irritation potential when a single dose of Acid Yellow RN 2903 was placed on the skin of rabbits for approximately four hours. This study should provide a rational basis for risk assessment in man as skin contact is one of the possible routes of human exposure.

Preparation of Dose Formulations:
The test item was used as delivered by the Sponsor and was moistened with approximately 0.5 mL of purified water before application. The pH of the test item was specified by the Sponsor as 7.5 to 8.5 at 10 g/L. According to Commission Regulation (EC) No. 440/2008, B.4. and OECD Guidelines 404, a test item does not need to be tested if the pH-value is less than 2 or greater than 11.5, owing to its predictable corrosive properties.

Test Item Administration:
The test item was applied as a weight of 0.5 g/animal, the dose specified in the test guidelines for solid test items.
Four days before treatment, the left flank was clipped with an electric clipper, exposing an area of approximately 100 cm2 (10 cm x 10 cm). The skin of the animals was examined one day before treatment and regrown fur of all animals was clipped again. On the day of treatment, 0.5 g of the test item was moistened with approximately 0.5 mL purified water and placed on a surgical gauze patch (ca. 2.5 cm x 2.5 cm). The patch was held in contact with the skin by means of an adhesive hypoallergenic aerated semi-occlusive dressing and a restrainer bandage wrapped around the trunk. The duration of treatment was 4 hours. Then the dressing was removed and the skin was flushed with lukewarm tap water to clean the application site so that any reactions were clearly visible at that time.
A single animal was treated first. As neither a corrosive effect nor a severe irritant effect was observed after the 4-hour exposure, the test was completed using the two remaining animals for an exposure period of four hours. Rationale: The application form and dose were used to detect an irritating potential of the test item applied.

Observations:
Viability / Mortality: Daily from acclimatization of the animals to the termination of test.
Clinical Signs (systemic): Daily from acclimatization of the animals to the termination of test.
Body Weights: At start of acclimatization, on the day of application and at termination of observation.

Pathology:
Necropsy: The animals were not sacrificed. On completion of the observation period all animals were reallocated to the eye irritation study (Harlan Laboratories Study D04516).

Data Compilation:
Viability/mortality was recorded on data sheets.
Body weights were recorded on-line with the ToxControl Computer System.
Clinical signs and skin reactions were compiled into the ToxControl Computer System during recording.
The ToxControl Computer System had been licenced for Harlan Laboratories Ltd. and validated with respect to data collection, storage and retrievability.

Statistical Analysis:
No statistical analysis was performed.

Determination of Skin Irritating Potential:

See attached Appendix 1 and 2 for scoring system)

Observation and Scoring: The skin reaction was assessed according to the numerical scoring system listed in the Commission Regulation (EC) No. 440/2008, B.4, at approximately 1, 24, 48 and 72 hours, as well as 7, 10 and 14 days after exposure (removal of the dressing, gauze patch and test item). To allow further examination of the test site all animals were re-clipped on completion of the 7, 10 and 14 days examination.
Data was summarized in tabular form, showing for each animal the irritation scores for erythema and oedema at all measurement intervals. Any lesions were described, including the degree and nature of irritation, corrosion or any other toxic effects observed, and their reversibility.
The mean score was calculated separately for each animal across 3 scoring times (24, 48 and 72 hours after patch removal) for erythema/eschar grades and for oedema grades, respectively. An animal is positive for irritation when the mean score is 2.3 or greater. The test is positive for irritation when at least 2 animals are positive for the same endpoint (erythema/eschar or oedema).

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

erythema score

Basis:

mean

Time point:

other: Mean score at 24, 48 and 72 hours

Score:

0

Results 2

Irritation parameter:

edema score

Basis:

mean

Time point:

other: Mean at 24, 48 and 72 hours

Score:

0

Irritant / corrosive response data:

The mean score was calculated separately for each animal across 3 scoring times (24, 48 and 72 hours after patch removal) for erythema/eschar grades and for oedema grades, respectively. The individual mean erythema/eschar and oedema scores were 0.00 for all animals. Furthermore, no alterations or corrosive effects were observed on the treated skin.

Other effects:

Slight yellow staining produced by the test item was observed in all animals from the 24-hour reading up to test day 14.

Any other information on results incl. tables

Viability / Mortality: No deaths occurred during the course of the study. Clinical Signs: No clinical signs were observed during the course of the study. Macroscopic Findings: Except for the staining produced by the test item, no signs of irritation were observed on the treated skin following the application of the test item. For ethical reasons and in the interest of animal welfare, the animals were therefore re-used for the eye irritation study (Harlan Laboratories Study D04516) with the same test item.

Applicant's summary and conclusion

Conclusions:

Based on the referred classification criteria (Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008),
Acid Yellow RN 2903 is not classified with respect to skin irritation in rabbits.

Executive summary:

The primary skin irritation potential of Acid Yellow RN 2903 was investigated according to OECD test guideline No. 404 and Commission Regulation (EC) No. 440/2008, B.4. The test item was applied by topical semi-occlusive application of 0.5 g to the intact skin of the left flank of each of three young adult New Zealand White rabbits. The duration of treatment was four hours. The scoring of skin reactions was performed 1, 24, 48 and 72 hours, as well as 7, 10 and 14 days after removal of the dressing.

The mean score was calculated separately for each animal across 3 scoring times (24, 48 and 72 hours after patch removal) for erythema/eschar grades and for oedema grades, respectively.

The individual mean erythema/eschar and oedema scores of the three animals were 0.00, 0.00 and 0.00, respectively. No clinical signs were observed during the course of the study. Slight yellow staining produced by the test item were observed in all animals from the 24-hour reading up to test day 14.

Based on the referred classification criteria (Regulation (EC) No 1272/2008 of the European

Parliament and of the Council of 16 December 2008), Acid Yellow RN 2903 is not classified with respect to skin irritation in rabbits.