Registration Dossier

Administrative data

Description of key information

Not skin irritating

Eye Irrit. 2 (H319), according to CLP Regulation (EC 1272/2008)

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:
June from 9th to 16th, 1993
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Qualifier:
according to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Version / remarks:
1981
Qualifier:
according to
Guideline:
EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
The test material was used as supplied.
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: David Percival Ltd., Moston, Sandbach, Cheshire, U.K.
- Age at study initiation: approximately 12 to 16 weeks old.
- Weight at study initiation: 2.32 - 2.36 kg.
- Housing: animals were individually housed in suspended metal cages.
- Diet: mains food RABMA Rabbit Diet, ad libitum.
- Water: mains drinking water, ad libitum.
- Acclimation period: minimum 5 days.
- Health check: on the day before the test, each rabbit 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.

ENVIRONMENTAL CONDITIONS
- Temperature: 18 - 23 °C
- Humidity: 58 - 74 %. On one occasion the humidity was above the limit specified in the protocol (70 %).
- Air changes: approximately 15 changes per hour.
- Photoperiod: 12 hours light and 12 hours darkness.
Type of coverage:
semiocclusive
Preparation of test site:
clipped
Vehicle:
water
Controls:
yes, concurrent no treatment
Amount / concentration applied:
0.5 g of test material, moistened with 0.5 ml of distilled water
Duration of treatment / exposure:
4 hours
Number of animals:
Two males and one female
Details on study design:
TEST SITE
- Area of exposure: on the day of the test a suitable test site was selected on the back of each rabbit. The test material was introduced under a 2.5 cm × 2.5 cm gauze patch and placed in position on the shorn skin.
- % coverage:
- Type of wrap if used: the patch was secured in position with a strip of surgical adhesive tape (approximate size 2.5 cm × 4.0 cm). To prevent the animals interfering with the patches, the trunk of each rabbit was wrapped in an elasticated corset.

REMOVAL OF TEST SUBSTANCE
- Removal: any residual test material was removed by gentle swabbing with cotton wool soaked in distilled water followed by diethyl ether.
- Time after start of exposure: 4 hours.

SCORING SYSTEM
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 irritatin and scored according to the 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".
An additional observation was made on day 7 to assess the reversibility of skin reactions.
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. If irreversible alteration of the dermal tissue is noted in any rabbit, which may include ulceration and clear necrosis or signs of scar tissue, the test material is classified as corrosive to rabbit skin.

Ervthena and Eschar Formation
No erythena 0
Very stight erythena (barely perceptible) 1
Well-defined erythema 2
Moderate to severe erythena 3
Severe erythena (beet redness) to slight eschar formation (injuries in depth) 4

Oedena Formation
No oedena 1
Very slight oedena (barely perceptible) 1
Slight oedena (edges of area well-defined by definite raising) 2
Moderate oedena (raised approxinately 1 millimetre) 3
Severe oedena (raised more than 1 millimetre and extending beyond the area of exposure) 4
Irritation parameter:
erythema score
Basis:
animal: 3/3
Time point:
24/48/72 h
Score:
< 2.3
Reversibility:
fully reversible within: 7 days
Irritation parameter:
edema score
Basis:
animal: 3/3
Time point:
24/48/72 h
Score:
< 2.3
Reversibility:
fully reversible within: 7 days
Irritant / corrosive response data:
Pink-coloured staining caused by test material was noted at all treated skin sites following patch removal. The staining did not affect evaluation of erythema.
Very slight erythema was noted at all treated skin sites one hour after patch removal. Very slight to well-defined erythema was noted at two treated skin sites at the 24 and 48-hour observations. Well-defined erythema persisted at one treated skin site at the 72-hour observation.
Very slight oedema was noted at two treated skin sites one hour after patch removal with very slight or slight oedema at 24-hour observation. Slight oedema persisted 72-hour observations.
Skin reactions noted at all treated skin sites seven days after treatment were crust formation or desquamation.

INDIVIDUAL DERMAL REACTIONS

Animal

Reaction Time after treatment Mean 24/48/72 hrs
1 hr 24 hrs 48 hrs 72 hrs 7 dy
Female n. 168 Erythema/Eschar 1 STA 2 STA 2 STA 2 STA 0CfSTA 2.00
Oedema 1 2 2 2 0 2.00
Male n. 169 Erythema/Eschar 1 STA 1 STA 1 STA 0STA 0DSTA 0.66
Oedema 1 1 0 0 0 0.33
Male n. 171 Erythema/Eschar 1 STA 0STA 0STA 0STA 0DSTA 0.00
Oedema 0 0 0 0 0 0.00

STA: pink-coloured staining

Cf: crust formation

D: desquamation

Interpretation of results:
other: not classified, according to the CLP Regulation (EC 1272/2008)
Conclusions:
Not irritating
Executive summary:

The study was performed to assess the irritancy potential of the test material to the skin of the New Zealand White rabbit. The method used that described in the OECD guideline 404. A single 4-hour, semi-occluded application of the test material to the intact skin of three rabbits produced very slight to well-defined erythema and very slight to slight oedema. Skin reactions noted at all treated skin sites seven days after treatment were crust formation and desquamation.

The test material produced a primary irritation index of 1.7 and was classified as a mild irritant to rabbit skin, according to the Draize classification scheme. No corrosive effects were noted.

Conclusion

The mean values from gradings at 24, 48 and 72 hours after patch removal were lower than 2.3 in all animals for both erythema/eschar and oedema reactions, thus the test item does not meet the criteria to be classified as irritating, according to the CLP Regulation (EC 1272/2008).

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:
June from 14th to 29th, 1993
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
1987
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
The test material was used as supplied.
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: David Percival Ltd., Moston, Sandbach, Cheshire, U.K.
- Age at study initiation: approximately 12 to 16 weeks old.
- Weight at study initiation: 2.74 - 2.90 kg.
- Housing: animals were individually housed in suspended metal cages.
- Diet: mains food RABMA Rabbit Diet, ad libitum.
- Water: mains drinking water, ad libitum.
- Acclimation period: minimum 5 days.
- Health check: immediately before the start of the test, both eyes of the provisionally selected test rabbits were examined for evidence of ocular irritation or defect with the aid of a light source from a standard ophthalmoscope. Animals showing evidence of ocular Iesions were rejected and replaced.

ENVIRONMENTAL CONDITIONS
- Temperature: 19 - 23 °C
- Humidity: 56 - 66 %
- Air changes: approximately 15 changes per hour.
- Photoperiod: 12 hours light and 12 hours darkness.
Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
0.1 ml of test material, which was found to weigh approximately 65 mg
Observation period (in vivo):
14 days
Number of animals or in vitro replicates:
Two males and one female
Details on study design:
PROCEDURE
- Proceeding: one rabbit was initially treated. After consideration of the ocular responses produced in the first treated animal, two additional animals were treated.
- Application: test item was placed into the coniunctival sac of the right eye, formed by gently pulling the lower lid away from the eyeball. The upper and lower eyelids were held together for about one second immediately after application, to prevent loss of the test material, and then released.
- Pain prevention: in order to minimise pain on application of the test material, one drop of local anaesthetic ("Ophthaine", 0.5 % proxymetacaine hydrochloride) was instilled into both eyes of animals 1 - 2 minutes before treatment.

TOOL USED TO ASSESS SCORE: examination of the eye was facilitated by use of the light source from a standard ophthalmoscope. Confirmation of corneal opacity was made by using sodium fluorescein B.P., if required.

SCORING SYSTEM
Immediately after administration of the test material, an assessment of the initial pain reaction was made.
Assessment of ocular damage/irritation was made approximately 1 hour and 24, 48 and 72 hours following treatment, according to the numerical evaluation 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".
Any other ocular effects were also noted.
Additional observations were made on days seven and fourteen to assess the reversibility of the ocular effects.

CONJUNCTIVAE
(A) Redness (refers to pelpebral and bulbar conjunctivae excluding cornea and iris)
Vessels normal 0
Vessels definitely injected above normal 1
More diffuse, deeper crimson red, individual vessels not easily discernible 2
Diffuse beefy red 3

(B) Chemsis
No swelling 0
Any swelling above normal (includes nictitating membrane) 1
Obvious swelling with partial eversion of lids 2
Swelling with lids about half closed 3
Swelling with lids half closed to comèpletely closed 4

(C) Discharge
No discharge 0
Any amount different from normal (does not include small amounts observed in inner canthus of normal animals) 1
Discharge with moinstening of the lids and hairs just adjacent to lids 2
Discharge with moinstening of the lids and hairs a considerable area around the eye 3

TOTAL SCORE (A + B + C) × 2
MAXIMUM TOTAL: 20

(D) IRIS
Normal 0
Folds above normal, congestion, swelling, circumcorneal injection (any or all of these combination of any thereof) iris still reacting to light (slugging reaction is positive) 1

TOTAL SCORE (D) × 5
MAXIMUM TOTAL: 10

CORNEA
(E) Degree of opacity (most degree area used)
No opacity 0
Scattered or diffuse areas, details of iris clearly visible 1
Easily discernible translucent areas, details of iris slightly obscured 2
Opalescent areas, no details of iris visible, size of pupil barely discernible 3
Opaque, iris invisible 4

(F) Area of cornea involved
One quarter (or less) but not zero 1
Greater than one quarter but less than half 2
Greater than half but less than three quarters 3
Greater than three quarters, up to whole area 4

TOTAL SCORE (E × F) × 5
MAXIMUM TOTAL: 80
Irritation parameter:
cornea opacity score
Basis:
animal: 3/3
Time point:
24/48/72 h
Score:
1
Reversibility:
fully reversible within: 14 days
Irritation parameter:
iris score
Basis:
animal: 2/3
Time point:
24/48/72 h
Score:
1
Reversibility:
fully reversible within: 7 days
Irritation parameter:
conjunctivae score
Basis:
animal: 2/3
Time point:
24/48/72 h
Score:
>= 2
Reversibility:
fully reversible within: 14 days
Irritation parameter:
chemosis score
Basis:
animal: 2/3
Time point:
24/48/72 h
Score:
< 2
Reversibility:
fully reversible within: 14 days
Irritant / corrosive response data:
Residual test material was noted around the treated eye of all animals throughout the study.
Red-coloured staining was noted in all treated eyes one hour after treatment.

Areas of diffuse corneal opacity were noted in all treated eyes one hour after treatment, at the 24, 48 and 72-hour observations and in one treated eye at the 7-day observation. Corneal opacity was confirmed using sodium fluorescein B.P. in all treated eyes one hour after treatment. Vascularisation of the cornea was noted in one treated eye at the 7-day observation.
Red-coloured staining prevented accurate evaluation of the iris in all treated eyes one hour after treatment. Iridial inflammation was noted in all treated eyes at the 24-hour observation and in two treated eyes at the 48 and 72-hour observations. No other iridial effects were noted.

Red-coloured staining prevented accurate evaluation of conjunctival redness in all treated eyes one hour after treatment; slight conjunctival chemosis and severe discharge were noted in all treated eyes at this time. Moderate conjunctival irritation was noted in all treated eyes at the 24-hour observation with minimal to moderate conjunctival irritation at the 48-hour observation. Minimal coniunctival irritation was noted in two treated eyes at the 72-hour observation. The reaction increased to severe conjunctival irritation in one treated eye at the 72-hour observation with minimal coniunctival irritation at the 7-day observation.
No ocular effects were noted in two treated eyes seven days after treatment and in one treated eye fourteen days after treatment.

OVERALL SCORE

Animal Reaction Time after treatment Mean 24/48/72 hrs
1 hr 24 hrs 48 hrs 72 hrs 7 dy 14 dy
Female no. 100 Cornea 1FS 1 1 1 0 0 1.00
Iris ?s 1 1 1 0 0 1.00
Conjunctivae redness ?s 2 2 2 0 0 2.00
Conjunctivae chemosis 2 2 2 1 0 0 1.67
Male no. 110 Cornea 1FS 1 1 1 1V 0 1.00
Iris ?s 1 1 1 0 0 1.00
Conjunctivae redness ?s 2 2 3 1 0 2.33
Conjunctivae chemosis 2 2 2 2 1 0 2.00
Female no. 113 Cornea 1FS 1 1 1 0 0 1.00
Iris ?s 1 0 0 0 0 0.33
Conjunctivae redness ?s 2 2 1 0 0 1.67
Conjunctivae chemosis 2 2 1 1 0 0 1.33

INDIVIDUAL SCORES AND INDIVIDUAL TOTAL SCORES FOR OCULAR IRRITATION

Famale no. 100

Time after treatment 1 hr 24 hrs 48 hrs 72 hrs 7 dy 14 dy
Cornea
E: degree of opacity 1FS 1 1 1 0 0
F: area of opacity 3 3 2 2 0 0
Score (E × F) × 5 15 15 10 10 0 0
Iris
D ?s 1 1 1 0 0
Score (D × 5) 0-10 5 5 5 0 0
Conjunctivae
A: redness ?s 2 2 2 0 0
E: chemosis 2 2 2 1 0 0
E: discharge 3Re 2Re 0Re 0Re 0Re 0Re
Score (A + B + C) × 2 10-16 12 8 6 0 0

Total score

25-41 32 23 21 0 0

F: due to red-coloured staining of the cornea, sodium fluorescein B.P. was used to aid confirmation of corneal opacity

S: red-coloured staining

Re: residual test material around treated eye

?s: red-coloured staining preventing accurate evaluation of ocular effect

V: vascularisation of the cornea

Male no. 110

Time after treatment 1 hr 24 hrs 48 hrs 72 hrs 7 dy 14 dy
Cornea
E: degree of opacity 1FS 1 1 1 1V 0
F: area of opacity 4 4 3 3 1 0
Score (E × F) × 5

20

20

15

15

5

0

Iris

D

?s

1

1

1

0

0

Score (D × 5)

0-10

5

5

5

0

0

Conjunctivae

A: redness

?s

2

2

3

1

0

E: chemosis

2

2

2

2

1

0

E: discharge

3Re

3Re

2Re

3Re

0Re

0Re

Score (A + B + C) × 2

10-16

14

12

16

4

0

Total score

30-46

39

32

36

9

0

F: due to red-coloured staining of the cornea, sodium fluorescein B.P. was used to aid confirmation of corneal opacity

S: red-coloured staining

Re: residual test material around treated eye

?s: red-coloured staining preventing accurate evaluation of ocular effect

V: vascularisation of the cornea

Male no. 113

Time after treatment 1 hr 24 hrs 48 hrs 72 hrs 7 dy 14 dy

Cornea

E: degree of opacity 1FS 1 1 1 0 0
F: area of opacity 3 3 2 1 0 0
Score (E × F) × 5

15

15

10

5

0

0

Iris

D

?s

1

0

0

0

0

Score (D × 5)

0-10

5

0

0

0

0

Conjunctivae

A: redness

?s

2

2

1

0

0

E: chemosis

2

2

1

1

0

0

E: discharge

3Re

2Re

0Re

0Re

0Re

0Re

Score (A + B + C) × 2

10-16

12

6

4

0

0

Total score

25 -41

32

16

9

0

0

F: due to red-coloured staining of the cornea, sodium fluorescein B.P. was used to aid confirmation of corneal opacity

S: red-coloured staining

Re: residual test material around treated eye

?s: red-coloured staining preventing accurate evaluation of ocular effect

V: vascularisation of the cornea

Interpretation of results:
other: Eye. Irrit. 2 (H319), according to the CLP Regulation (EC 1272/2008)
Conclusions:
Moderate irritant
Executive summary:

The study was performed to assess the irritancy potential of test material to the eye of the New Zealand White rabbit. The method used followed OECD guideline 405. A single application of test material to the non-irrigated eye of three rabbits produced areas of diffuse corneal opacity, iridial inflammation and moderate to severe conjunctival irritation.

Vascularisation of the cornea was confined to one treated eye. No ocular effects were noted 7 or 14 days after treatment.

The test material produced a maximum group mean score in the range of 34.3 - 42.7 and was classified as a moderate irritant (class 5 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system.

Conclusion

The mean values from gradings at 24, 48 and 72 hours were equal to 1 for corneal opacity and iritis in all rabbits, equal/higher than 2 for conjunctival redness in two out of three rabbits and lower than 2 for chemosis in two out of three rabbits; all the reactions recorded were fully reversible within 14 days.

The substance can be considered as irritating to eye and a classification in category 2 (Eye. Irrit. 2 (H319) of the CLP Regulation (EC 1272/2008) can be considered as appropriated.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

SKIN IRRITATION

Studies on skin irritating potential of Acid Red 057 are available.

The key study was performed using New Zealand White rabbit, according to the OECD guideline 404. Application of test material on intact skin produced very slight to well-defined erythema and very slight to slight oedema. Skin reactions noted at all treated skin sites seven days after treatment were crust formation and desquamation. No corrosive effects were noted. The mean values from gradings at 24, 48 and 72 hours after patch removal were lower than 2.3 in all animals for both erythema/eschar and oedema reactions.

In an older test, primary irritation to the skin was measured by a patch test technique on abraded and intact skin of rabbits. The primary irritation index was calculated to be 0.3, mildly irritating. No observable response was seen in any of the six tested rabbits or at the 24 hours reading. Very slight erythema with or without very slight oedema was observed in three intact and two abraded sites at the 72 hours reading. Three animals did not show any observable response to treatment throughout the 72 hours observation period.

Furthermore, there are information about two additional experiments run according to the methods indicated in the ETAD Regulation (14.02.1975). Unfortunately, in both cases only a sheet of results is available, thus there are no details about testing procedures. In both cases, test results suggest that the substance can be considered as not irritating or just slightly irritating.

EYE IRRITATION

The irritancy potential of the test material to the eye was investigated in some studies.

The key study was performed in accorcance with OECD guideline 405. A single application of test material to the non-irrigated eye of three rabbits produced areas of diffuse corneal opacity, iridial inflammation and moderate to severe conjunctival irritation. Vascularisation of the cornea was confined to one treated eye. No ocular effects were noted 7 or 14 days after treatment. Test material produced a maximum group mean score in the range of 34.3 - 42.7 and was classified as a moderate irritant (class 5 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system (Allen, 1993). The mean values from gradings at 24, 48 and 72 hours were equal to 1 for corneal opacity and iritis in all rabbits, equal/higher than 2 for conjunctival redness in 2 out of 3 rabbits and lower than 2 for chemosis in 2 out of 3 rabbits; all recorded reactions were fully reversible within 14 days.

The second study available identified the substance as not irritating. Test item was instilled into the eyes of three rabbits without rinsing and the eyes of the three remaining rabbits were irrigated with 20 ml of lukewarm (approx. 37 °C) water 5 seconds after instillation. The eyes are examined and the grade of ocular reaction is recorded at 24, 48 and 72 hours and at the 4th and 7th day. In case of animals of which eyes were not rinsed, none showed posltive reactions. Staining of the cornea by the dye was observed in all three animals. Temporary mild conjunctival reactions only were observed. None of the animals of which eyes were rinsed showed positive reactions; temporary mild conjunctival reactions only were observed.

There are information about two additional experiments performed according to the methods indicated in the ETAD Regulation (14.02.1975). Unfortunately, in both cases only a sheet of results is available, thus there are no details about testing procedures. The scoring system used in the ETAD method is the same of that reported into the OECD guideline, therefore it is possible to evaluate the results, because the raw tables are included into the study report. In both cases, test results suggest that the substance can be considered as eye irritating.

Justification for classification or non-classification

According to the CLP Regulation (EC 1272/2008), 3.2 Skin corrosion/irritation section, skin irritation means the production of reversible damage to the skin following the application of a test substance for up to 4 hours.

In the key study, the mean values from gradings at 24, 48 and 72 hours after patch removal were lower than 2.3 in all animals for both erythema/eschar and oedema reactions, thus test item does not meet the criteria to be classified as irritating.

According to the CLP Regulation (EC) No 1272/2008 substances that have the potential to induce reversible eye irritation are classified in Category 2 (irritating to eyes) if, 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.

In the key study, mean values from gradings at 24, 48 and 72 hours were equal to 1 for corneal opacity and iritis in all rabbits, equal/higher than 2 for conjunctival redness in two out of three rabbits and lower than 2 for chemosis in two out of three rabbits; all the reactions recorded were fully reversible within 14 days. The substance can be considered as irritating to eye and a classification in category 2 (Eye. Irrit. 2 H319) can be considered as appropriated.

In conclusion, the substance is not classified for skin irritation; however, it is classified for eye irritation, category 2 (Eye Irrit. 2, H319), according to the CLP Regulation (EC 1272/2008).