Tetrasodium 4-amino-5-hydroxy-6-[[2-methoxy-5-[[2-(sulphonatooxy)ethyl]sulphonyl]phenyl]azo]-3-[[4-[[2-(sulphonatooxy)ethyl]sulphonyl]phenyl]azo]naphthalene-2,7-disulphonate

Endpoint summary

Description of key information

No irritating effects observed

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1982

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPP 81-5 (Acute Dermal Irritation)

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

The New Zealand White rabbit , weighing 2.0 - 3 .0 kg and approximately 9-10 weeks of age, was used for this study.

The animals were housed and maintained in compliance with the Animal Welfare Act (Pub. L-94-279) 9 CFR Part 3.
The rabbits were not used in any previous study and were accl imated to the laboratory for at least 5 days prior to dosing.

The animals were randomized using a computer generated list of random numbers. They were individually identified by ear tag.
Each cage was identified with a cage card displaying the animal number, sex, project number, test article , date dosed , dose level, and responsible technician's initials.

Husbandry Conditions:
Temperature - 70°F ~ 2°F
Relative Humidity - 45% ~ 5%
Light - 12 hour light/dark cycle
Diet - Wayne 15% Rabbit
Ration and tap water were provided ad libitum. Based on our current knowledge no contaminants were known to be in this diet or water that might be expected to interfere with the objectives of the study.

Caging - Stainless steel with elevated wire mesh flooring
1 rabbit/cage

Bedding - Techboard
Shepherd Products Company
Kalamazoo, Michigan 49005

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

500 mg

Duration of treatment / exposure:

24 h

Observation period:

examination after removal and 2 days later

Number of animals:

6

Details on study design:

A group of six rabbits was clipped over a wide area on their backs approximately 24 hours prior to application.
One side of the animals' backs was abraded at two sites with a lancet sufficiently deep to penetrate the stratum corneum but not enter the derma to produce bleeding. Two intact skin sites were also selected on the other side of the back.

A 0.5 g portion of the experimental material was applied to two abraded and two intact skin sites on the same rabbit.
Gauze patches were then placed over the treated areas and an impervious material wrapped snugly around the trunks of the animals to hold the patches in place. The wrapping was removed at the end of the twenty-four hour contact period, the sites wiped but not washed, and the treated areas examined.

Readings were also made after seventytwo hours. The Draize method of scoring was employed. All other lesions observed were recorded.

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

No signs of systemic toxicity were observed during the 72 hours of observation

Interpretation of results:

GHS criteria not met

Conclusions:

The material does not appear to be a skin irritant .

Executive summary:

A group of six rabbits was clipped over a wide area on their backs approximately 24 hours prior to application. One side of the animals' backs was abraded at two sites with a lancet sufficiently deep to penetrate the stratum corneum but not enter the derma to produce bleeding. Two intact skin sites were also selected on the other side of the back.

A 0.5 g portion of the experimental material was applied to two abraded and two intact skin sites on the same rabbit. Gauze patches were then placed over the treated areas and an impervious material wrapped snugly around the trunks of the animals to hold the patches in place. The wrapping was removed at the end of the twenty-four hour contact period, the sites wiped but not washed, and the

treated areas examined. Readings were also made after seventytwo hours. The Draize method of scoring was employed. All

other lesions observed were recorded.

Primary Irritation Score: 0

The material does not appear to be a skin irritant .

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1982

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPP 81-4 (Acute Eye Irritation)

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

The New Zealand White rabbit, weighing 2 .0 - 3 .0 kg and approximately 9-10 weeks of age, was used for this study.

Husbandry Conditions:
Temperature - 70°F ~ 2°F
Relative Humidity - 45% ~ 5%
Light - 1:2 hour light/dark cycle

Diet - Wayne 15% Rabbit
Ration and tap water were provided ad libitum. Based on our current knowledge no contaminants were known to be in this diet or water that might be expected to interfere with the objectives of the study.

Caging - Stainless steel with elevated wire mesh flooring
1 rabbit/cage

Bedding - Techboard
Shepherd Products Company
Kalamazoo, Michigan 49005

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: approx. 100mg

Duration of treatment / exposure:

single exposure
6 rabbits: test material was not removed
3 rabbits: treated eyes were washed after 30 seconds for 1 minute with gently flowing tepid tap water (1000 mL/minute)

Observation period (in vivo):

The treated eyes were examined at one, two, three, four and seven days following instillation of the test material into the eyes.

Number of animals or in vitro replicates:

9

Details on study design:

A 100 mg portion of the experimental material was instilled into the right eyes of the test animals while the other eyes remained untreated to serve as controls. The first six rabbits did not have their eyes washed subsequent to instillation. The remaining three rabbits had the treated eyes washed after 30 seconds for 1 minute with gently flowing tepid tap water (1000 mL/minute).

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

3

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Interpretation of results:

GHS criteria not met

Conclusions:

The subject material does not appear to be an ocular irritant in the unwashed eye.

Executive summary:

The New Zealand White rabbit, weighing 2 .0 - 3 .0 kg and approximately 9- 10 weeks of age, was used for this study.

A 100 mg portion of the experimental material was instilled into the right eyes of the test animals while the other eyes remained untreated to serve as controls. The first six rabbits did not have their eyes washed subsequent to instillation. The remaining three rabbits had the treated eyes washed after 30 seconds for 1 minute with gently flowing tepid tap water (1000 mL/minute).

The treated eyes were examined at one, two, three, four and seven days following instillation of the test material into the eyes. The readings of the eyes were made in accordance with the Draize technique for scoring ocular lesions.

AVERAGE OCULAR IRRITATION SCORE: 0

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Additional information

To test Reactive Blue 250 for skin irritating properties, a group of six rabbits was clipped over a wide area on their backs approximately 24 hours prior to application. One side of the animals' backs was abraded at two sites with a lancet sufficiently deep to penetrate the stratum corneum but not enter the derma to produce bleeding. Two intact skin sites were also selected on the other side of the back.

A 0.5 g portion of the experimental material was applied to two abraded and two intact skin sites on the same rabbit. Gauze patches were then placed over the treated areas and an impervious material wrapped snugly around the trunks of the animals to hold the patches in place. The wrapping was removed at the end of the twenty-four hour contact period, the sites wiped but not washed, and the treated areas examined. Readings were also made after 72 hours. The Draize method of scoring was employed. All

other lesions observed were recorded. No irritating effects were observed at any reading. The Mean Primary Irritation Score was: 0

Hence, Reactive Blue 250 is not a skin irritant. This is also consistent with the results of skin irritating study with the structural analogue Reactive Black 5.

To test Reactive Blue 250 for skin irritating properties, a group of 9 New Zealand White rabbit, weighing 2 .0 - 3 .0 kg and approximately 9- 10 weeks of age, was used for this study.

A 100 mg portion of the experimental material was instilled into the right eyes of the test animals while the other eyes remained untreated to serve as controls. The first six rabbits did not have their eyes washed subsequent to instillation. The remaining three rabbits had the treated eyes washed after 30 seconds for 1 minute with gently flowing tepid tap water (1000 mL/minute).

The treated eyes were examined at one, two, three, four and seven days following instillation of the test material into the eyes. The readings of the eyes were made in accordance with the Draize technique for scoring ocular lesions.

No signs for eye irritating properties were observed throughout the study. The AVERAGE OCULAR IRRITATION SCORE was: 0.

This is also consistent with the results of eye irritating study with the structural analogue Reactive Black 5.

No studies for respiratory irritation are available for Reactive Blue 250. However, two inhalative sensitisation studies have been performed with Reactive Black 5. There were no signs for irritating effects on the respiratory tract noted in those studies.

Justification for classification or non-classification

No irritating effect according to EU classification system - No classification necessary