Trisodium bis[3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-2-hydroxy-5-nitrobenzenesulphonato(3-)]chromate(3-)

Endpoint summary

Description of key information

Skin: Not irritant

Eye: Not irritant

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

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:

July 28, 1975

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

other: "Hazardous Substances Regulations" under the U.S. Federal Hazardous. Substances Labelling Act Sect. 191.11 (February 1965).

Deviations:

no

GLP compliance:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 10-13 weeks
- Weight at study initiation: average body weights of 2.73 kg (males) and 2.38 kg (females)
- Housing:The rabbits were caged singly in an experimental room.
- Diet (e.g. ad libitum): a commercial irradiated diet (Styles-Oxoid) was fed ad libitum.
- Water (e.g. ad libitum): sterile filtered water was available at all times.
- Acclimation period: six rabbits were acclimatized in the test area for one week prior to the start of the trial.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 1°
- Humidity (%): 50-70
- Photoperiod (hrs dark / hrs light):Animals were exposed to artificial light for 10 hours daily from 08.00 - 18.00 hours.

Type of coverage:

semiocclusive

Preparation of test site:

abraded

Vehicle:

water

Controls:

not specified

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 10 g of the test compound was mixed with 12 ml of water to make a solution of 15ml 0.75 ml of which was applied to each test- site on a 2.5 cm square gauze pad.

Duration of treatment / exposure:

24 hours

Observation period:

24 hours and 72 hours

Number of animals:

Six rabbits ( 3 male and 3 female)

Details on study design:

TEST SITE
Twenty-four hours prior to the dermal application, the backs of the rabbits were shaved over an area consisting of at least 10% of the total body surface. Two test sites lateral to the mid line of the back were used on each rabbit. Immediately before the application of the test compound, the right hand site was abraded with the point of a sterile hypodermic needle. The abrasions were sufficiently deep to penetrate the stratum corneum but not to damage the dermis. The left hand sits remained intact.

These were covered with aluminum foil secured with "Sleek"* adhesive tape. The test sites were then enclosed by a 6" wide "Cobon” self adhesive bandage the edges of which were fixed to the skin by strips of "Sleek" in order to retain the test substance in close contact with the skin.

SCORING SYSTEM:
Primary irritation score Rating
0 Non irritating
0.1 - 0.5 Minimally irritating
0.6 - 1.5 Slightly irritating
1.6 - 3.0 Mildly irritating
3.1 - 5.0 Moderately irritating
5.1 - 6.5 Severely irritating
6.6 - 8.0 Extremely irritating

Irritation parameter:

erythema score

Basis:

animal: 11, 12, 13, 15, 16

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: no symptoms

Remarks on result:

other: intact skin

Irritation parameter:

erythema score

Basis:

animal: 14

Time point:

24/48/72 h

Score:

ca. 0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: intact skin

Irritation parameter:

edema score

Basis:

animal: 11, 15, 16

Time point:

24/48/72 h

Score:

ca. 1.66

Max. score:

2

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: intact skin

Irritation parameter:

edema score

Basis:

animal: 12

Time point:

24/48/72 h

Score:

ca. 0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: intact skin

Irritation parameter:

edema score

Basis:

animal: 13

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

0

Reversibility:

other: no symptoms

Remarks on result:

other: intact skin

Irritation parameter:

edema score

Basis:

animal: 14

Time point:

24/48/72 h

Score:

ca. 2.66

Max. score:

3

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: intact skin

Erythema Formation:

No./Sex	Intact		Abraded		Total
	24 hrs	72 hrs	24 hrs	72 hrs
11	0	0	0	0
13	0	0	0	0
15	0	0	0	0
12	0	0	1	0
14	1	0	1	0
16	0	0	0	0
Mean	0.17	0	0.33	0	0.5

Oedema Formation

No./Sex	Intact		Abraded		Total
	24 hrs	72 hrs	24 hrs	72 hrs
11	2	0	3	0
13	0	0	1	0
15	2	0	2	0
12	1	0	1	0
14	3	0	2	0
16	2	0	1	0
Mean	1.67	0	1.67	0	3.33

Total Erythema + Oedema Score = 3.83

Interpretation of results:

other: CLP criteria not met

Conclusions:

Not irritant

Executive summary:

Method

The skin irritation potential of the test substance was determined according to the method given in the "Hazardous Substances Regulations" under the U.S. Federal Hazardous. Substances Labelling Act Sect. 191.11 (February 1965).

 

Observations

Slight erythema and very slight to moderate edema was seen on 2/6 and 6/6 rabbits respectively, 24 hours after application of the compound. All sites were normal by 72 hours.

There was no marked difference in the reactions between abraded and intact sites.

 

Conclusion

Not 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:

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

Adequacy of study:

key study

Study period:

July 28, 1975

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

other: "Hazardous Substances Regulations" under the U.S.A. "Federal Hazardous Substances Labelling Act Sect. 191.12 (February 1965), with slight modification.

Deviations:

no

GLP compliance:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Age at study initiation: 10 - 13 weeks
- Weight at study initiation: average body weights of 2.73 kg (males) and 2.38 kg (females)
- Housing:The rabbits were caged singly in an experimental room.
- Diet (e.g. ad libitum):A commercial irradiated diet (Styles-Oxoid) was fed ad libitum.
- Water (e.g. ad libitum):Sterile filtered water was available at all times.
- Acclimation period: Six rabbits were acclimatised in the test area for one week prior to the start of the trial

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 1 °C
- Humidity (%): 50 - 70
- Photoperiod (hrs dark / hrs light):Animals were exposed to artificial light for 10 hours daily'from 08.00 - 18.00 hours

Vehicle:

unchanged (no vehicle)

Controls:

other: The right eye served as a control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit) : 100 mg.

Duration of treatment / exposure:

30 seconds

Observation period (in vivo):

1, 24, 48 and 72 hours

Number of animals or in vitro replicates:

Six rabbits (3 males and 3 females)

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): after 30 seconds the compound was, as far as possible, flushed out of the eyes' of three of the rabbits with warm water for 1 minute.

SCORING SYSTEM: the ocular reactions were scored by the method described in "Appraisal of the Safety of Chemicals in Food Drugs and Cosmetics" page 51, published by the Association of Food and Drug Officials of the U.S.A.

Irritation parameter:

conjunctivae score

Remarks:

redeness

Basis:

animal: 15

Time point:

24/48/72 h

Score:

ca. 0.33

Max. score:

1

Reversibility:

fully reversible

Remarks on result:

other: washed eyes

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal: 14, 16

Time point:

24/48/72 h

Score:

ca. 0.66

Max. score:

1

Reversibility:

fully reversible

Remarks on result:

other: washed eyes

Irritation parameter:

chemosis score

Basis:

animal: 14, 15, 16

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

0

Reversibility:

other: no symptoms

Remarks on result:

other: washed eyes

Irritant / corrosive response data:

A slight to moderate conjunctival reaction was seen in all treated eyes one hour after application of the compound. This had subsided slightly by 24 hours and thereafter continued to do so until all eyes were normal by 48 hours (1/3 washed) 72 hours (2/3 unwashed 2/3 washed) and 6 days (1/3 unwashed). Staining of the conjunctivae and cornea by the compound was seen in the unwashed eyes for up to 40 hours.

Other effects:

None

Rabbit eye irritation scores-reference procedure:

		1 hour							6 hours							1 Day							2 Days							3 Days							6 Days
	Rabbit No	11	13	15	12	14	16		11	13	15	12	14	16		11	13	15	12	14	16		11	13	15	12	14	16		11	13	15	12	14	16		11	13	15	12	14	16
	Conjuctivae
A	Redness	2	2	2	2	1	1		-	-	-	-	-	-		1	2	1	2	1	1		1	1	-	1	1	1		-	1	-	-	-	-		-	-	-	-	-	-
B	Chemosis	2	2	1	2	1	2		-	-	-	-	-	-		1	1	-	1	-	-		1	1	-	1	-	-		-	-	-	-	-	-		-	-	-	-	-	-
C	Discharge	2	1	-	3	-	-		-	-	-	-	-	-		-	-	-	-	-	-		-	-	-	-	-	-		-	-	-	-	-	-		-	-	-	-	-	-
c=(A+B+C) X2	(max. 20)	12	10	6	14	4	6		-	-	-	-	-	-		4	6	2	6	2	2		4	4	-	4	2	2		-	2	-	-	-	-		-	-	-	-	-	-
Total	(Max.110)	12	10	6	14	4	6		-	-	-	-	-	-		4	6	2	6	2	2		4	4	-	4	2	2		-	2	-	-	-	-		-	-	-	-	-	-

Eyes washed in Nos. 14, 15 & 16 hours

Interpretation of results:

other: CLP criteria not met

Conclusions:

Not irritant

Executive summary:

Method

The study was performed to determine the eye irriattion potential of the test substance in rabbits according to the procedure set out in the "Hazardous Substances Regulations" under the U.S.A. "Federal Hazardous Substances Labelling Act Sect. 191.12 (February 1965), with slight modification.

Conclusion

Not irritant

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

No studies on the "Irritation/Corrosion" are available for the substance in itself nevertheless, studies were conducted with an analogue molecule (Similar Substance 01). Further information are reported in the Read Across justification attached to section 13.

Skin Irritation

A skin irritation potential of the test substance was determined in the key study according to the method given in the "Hazardous Substances Regulations" under the U.S. Federal Hazardous. Substances Labelling Act Sect. 191.11 (February 1965). Slight erythema and very slight to moderate edema was seen on 2/6 and 6/6 rabbits respectively, 24 hours after application of the compound. All sites were normal by 72 hours. There was no marked difference in the reactions between abraded and intact sites. The primary irritation score was 1.0.

It can be noted that abrasion on skin causes epidermal injury and further expected to cause the skin irritation and adverse effects to epidermis. This reaction could be referred to the damage caused due to abrasion rather than due to the test substance application. In general abrasion is an elevated testing condition and results normally do not have to be taken into account for hazard assessment. Hence, the test substance can be considered as non-irritating to rabbit’s skin.

 

In the supporting study the test was to determine the primary-irritation index, which serves as a measure of the acute irritation, provoked by the compound on the skin of the rabbits. The procedure adopted was the patch-test technique described in the "Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics" (1959) of the US Association of Food and Drug Officials (AFDO). The primary irritation index as the measure of the acute irritation to the skin of rabbits was found to be 0. Therefore the test substance is to be considered as non-irritant to the skin of rabbits.

 

Considering the results from key and supporting study it can be concluded that the test item is non-irritating to rabbit’s skin.

Eye Irritation

A key study was performed to determine the eye irritation potential of the test substance in rabbits according to the procedure set out in the "Hazardous Substances Regulations" under the U.S.A. "Federal Hazardous Substances Labelling Act Sect. 191.12 (February 1965), with slight modification.

The test recorded only the value related to the conjuctiva redness and the chemosis, which resulted to be over the limit for classification.

To support this study a second study was performed to determine the eye irritation potential of the test substance in rabbits according to the technique described in the "Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics (1959) of the US Association of Food and Drug Officials (AFDO).

The primary-irritation index, which serves as a measure of the acute irritation provoked by the substance, was found to be 0 for the cornea, 0 for the iris and o for the conjunctivae.

Therefore the test item is to be considered non-irritant to the eye of rabbits.

Justification for classification or non-classification

SKIN IRRITATION/CORROSION

A corrosive substance is a substance that produces destruction of skin tissue, namely, visible necrosis through the epidermis and into the dermis, in at least 1 tested animal after exposure up to 4 hour duration.

Three subcategories are provided within the corrosive category 1:

Subcategory 1A where responses are noted following up to 3 minutes exposure and up to 1 hour observation;

Subcategory 1B where responses are described following exposure between 3 minutes and 1 hour and observations up to 14 days;

Subcategory 1C where responses occur after exposures between 1 hour and 4 hours and observations up to 14 days

To classify a substance as irritant, the major criterion is that at least 2 of 3 tested animals have a mean score of ≥ 2,3 - ≤ 4,0.

Category 2:

- Mean value of ≥ 2,3 - ≤ 4,0 for erythema/ eschar or for oedema in at least 2 of 3 tested animals from gradings at 24, 48 and 72 hours after patch removal or, if reactions are delayed, from grades on 3 consecutive days after the onset of skin reactions; or

- Inflammation that persists to the end of the observation period normally 14 days in at least 2 animals, particularly taking into account alopecia (limited area), hyperkeratosis, hyperplasia, and scaling; or

- In some cases where there is pronounced variability of response among animals, with very definite positive effects related to chemical eposure in a single animal but less than the criteria above.

 

EYE IRRITATION

Serious eye damage means the production of tissue damage in the eye, or serious physical decay of vision, following application of a test substance to the anterior surface of the eye, which is not fully reversible within 21 days of application.

Eye irritation means the production of changes in the eye following the application of test substance to the anterior surface of the eye, which are fully reversible within 21 days of application.

Irreversible effects on the eye (Category 1):

If, when applied to the eye of an animal, a substance produces:

- at least in one animal effects on the cornea, iris or conjunctiva that are not expected to reverse or have not fully reversed within an observation period of normally 21 days; and/ or

- at least in 2 of 3 tested animals, a positive response of corneal opacity ≥ 3 and/or iritis > 1,5 calculated as the mean scores following grading at 24, 48 and 72 hours after installation of the test material.

These observations include animals with grade 4 cornea lesions and other severe reactions (e.g., destruction of cornea) observed at any time during the test, as well as persistent corneal opacity, discoloration of the cornea by a dye substance, adhesion, pannus, and interference with the function of the iris or other effects that impair sight. In this context, persistent lesions are considered those which are not fully reversible within an observation period of normally 21 days. Substances are also classified in Category 1 if they fulfil the criteria of corneal opacity ≥ 3 or iritis > 1,5 detected in a Draize eye test with rabbits, recognising that such severe lesions usually do not reverse within a 21-day observation period.

The substance does not meet the requirements to be classified in Category 1.

Irritating to eyes (Category 2)

when applied to the eye of an animal, a substance produces:

- at least in 2 of 3 tested animals, a positive response of: corneal opacity ≥ 1 and/or iritis ≥ 1, and/or conjunctival redness ≥ 2 and/or conjunctival oedema (chemosis) ≥ 2

calculated as the mean scores following grading at 24, 48 and 72 hours after installation of the test material, and which fully reverses within an observation period of 21 days.

Based on the results obtained in the skin and eye irritation studies, the test substance does not need to be classified for skin and eye irritation according to the CLP Regulation (EC) No. 1272/2008.