Registration Dossier

Administrative data

Description of key information

Skin irritation

Key study:- Jessup (2003) 'Acute dermal irritation/corrosion study in the rabbit' conducted in line with OECD Guideline 404 and EU Method B.4. The test substance was found to be not irritating.

Eye irritation

Key study: Shepard (2003) 'Acute eye irritating/corrosion study in the rabbit' conducted in line with OECD Guideline 405 and EU Method B.5. The test substance was found to be not irritating.

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:
9 September 2003 to 12 September 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: The rabbits were young adults (at least three months old)
- Weight at study initiation: at least 2000 grams
- Housing: Animals were housed in an Association for Assessment and Accreditation of Laboratory Animal Care International-accredited vivarium in accordance with the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996). The rabbits were singly housed in suspended, stainless-steel mesh cages. Cages and racks were washed once a week. Absorbent paper, used to collect excreta, was changed every other day.
- Diet: Certified High Fiber Rabbit Diet was available ad libitum.
- Water: Water was available ad libitum through an automatic watering system. The source of the water was the local public water system.
- Acclimation period: The animals were isolated upon arrival and allowed to acclimate for a period of 5 days. Animals were judged to be healthy prior to testing.

ENVIRONMENTAL CONDITIONS
- Temperature: 21.4 to 23.0 °C
- Humidity: 45.1 to 61.4 %
- Photoperiod: A photoperiod of 12 hours of light was maintained.
Type of coverage:
occlusive
Preparation of test site:
shaved
Vehicle:
water
Controls:
other: No control substance was used. Adjacent areas of untreated skin of each animal served as control sites for the test areas.
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 gram of the test substance was applied topically to each animal. The test substance was administered as a solid moistened with water.

Duration of treatment / exposure:
4 hours
Observation period:
The site of application was examined at 1, 24, 48, and 72 hours after removal of the occlusive patch. Observations included estimation of erythema, edema, necrosis, eschar formation, scarring, erosion, and staining caused by the test substance as well as general systemic effects.
Number of animals:
3
Details on study design:
TEST SITE
- Area of exposure: Not reported
- % coverage: Not reported
- Type of wrap if used: A fiber pad and an occlusive wrap to hold the test substance in place for four hours.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): At the end of the exposure period, the application site was rinsed with running water.
- Time after start of exposure: 4 hours

SCORING SYSTEM:
The most severely affected area within the test substance application sites were scored for dermal reactions. Corrosion was considered to have resulted if destruction or irreversible alteration of the skin tissue (ulceration and/or necrosis) resulted after an exposure of 4 hours or less to the test substance. Skin irritation reactions were graded and scored as described in below:

Grading Of Skin Reactions*

Erythema and Eschar Formation
0 = No erythema
1 = Very slight erythema (barely perceptible)
2 = Well-defined erythema
3 = Moderate to severe erythema
4 = Severe erythema to slight eschar formation

Edema Formation
0 = No edema
1 = Very slight edema (barely perceptible)
2 = Slight edema (edges of area well defined by definite raising)
3 = Moderate edema (edges raised approximately 1.0 mm)
4 = Severe edema (raised more than 1.0 mm and extending beyond area of exposure)

*Graded as described in OECD Guideline 405 (Annex V., Test B.4) (Grading of Skin Reaction)
Irritation parameter:
erythema score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
0
Reversibility:
other: no effects seen
Irritation parameter:
edema score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
0
Reversibility:
other: no effects seen
Irritant / corrosive response data:
No irritant response or serious lesion was noted during the 72-hour observation period.
Other effects:
No toxic effects were noted during the study.

Observations for Irritation

The application site of each animal was examined for signs of irritation at 1, 24, 48, and 72 hours after termination of exposure to the test substance. Observations for irritation (erythema and edema) are presented in Table 1.

Table 1: Response at application site

Animal number Erythema, edema*
1 hour 24 hours 48 hours  72 hours
745 0,0 0,0 0,0 0,0
746 0,0 0,0 0,0 0,0
747 0,0 0,0 0,0 0,0

*Graded as described in OECD Guideline 405 (Annex V., Test B.4) (Grading of Skin Reaction)

Interpretation of results:
other: Not classified in accordance with EU criteria
Conclusions:
Under the conditions of the test, the substance was determined to be a non-irritant to the skin.
Executive summary:

In a GLP compliant study conducted in line with OECD Guideline 404 and EU Method B.4, the dermal irritation of butanoic acid, 4-[[4-[7-chloro-6-(1,1-dimethylethyl)-3H-pyrazolo[1,5-b][1,2,4]triazol-2-yl]phenyl]amino]-4-oxo, tetradecyl ester was investigated by administering single topical doses of 0.5 gram of the test substance to rabbits. The test substance was left in contact with the skin under an occlusive wrap for four hours. Skin lesions were graded according to OECD Guideline 404 (Annex V., Test B.4).

No irritant response or serious lesion was noted during the 72-hour observation period. Based on these results, the test substance requires no skin irritation classification in line with current EU requirements.

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:
29 September 2003 to 3 October 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Age at study initiation: The rabbits were young adults (at least three months old).
- Weight at study initiation: Not reported
- Housing: Animals were housed in an Association for Assessment and Accreditation of Laboratory Animal Care International-accredited vivarium in accordance with the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996). The rabbits were singly housed in suspended, stainless-steel mesh cages. Cages and racks were washed once a week. Absorbent paper, used to collect excreta, was changed every other day.
- Diet: Certified High Fiber Rabbit Diet was available ad libitum.
- Water: Water was available ad libitum through an automatic watering system. The source of the water was the local public water system.
- Acclimation period: The animals were isolated upon arrival and allowed to acclimate for a period of 5 days. Animals were judged to be healthy prior to testing.

ENVIRONMENTAL CONDITIONS
- Temperature: 21.3 to 23.0 °C
- Humidity: 46.3 to 56.6 %
- Photoperiod: A photoperiod of 12 hours of light was maintained.
Vehicle:
unchanged (no vehicle)
Controls:
other: No control substance was used. The untreated eye (left) of each animal served as a control for the test.
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 grams
Duration of treatment / exposure:
N/A
Observation period (in vivo):
Eyes were observed immediately after instillation of the test substance and 1, 24, 48, and 72 hours thereafter.
Number of animals or in vitro replicates:
6
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done): The treated eyes of three animals were immediately washed with running distilled water; the eyes of the other three animals were not irrigated.
- Time after start of exposure: Immediately

SCORING SYSTEM
- The eyes were examined and grades of ocular reactions recorded for each animal at all observation periods. Ocular lesions were graded and scored as described below:

Grading Of Ocular Lesions (as described in OECD 405):

Cornea:-
0 = No ulceration or opacity
1 = Scattered or diffuse areas of opacity (other than slight dulling of normal luster), details of iris clearly visible
2 = Easily discernible translucent area, details of iris slightly obscured
3 = Nacreous area, no details of iris visible, size of pupil barely discernible
4 = Opaque cornea, iris not discernible through the opacity

Area of Corneal Opacity:-
1 = One-quarter or less, but not zero
2 = Greater than one-quarter up to one-half
3 = Greater than one-half up to three-quarters
4 = Greater than three quarters up to the whole area

Iris:-
0 = Normal
1 = Markedly deepened folds, congestion, swelling, moderate circumcorneal hyperaemia, or injection, any of these or combinations of any thereof, iris still reacting to light(sluggish reaction is positive)
2 = No reaction to light, hemorrhage, gross destruction (any or all of these)

Conjunctivae:-
Redness: refers to palpebral and bulbar conjunctivae excluding cornea and iris
0 = Blood vessels normal
1 = Some blood vessels definitely injected
2 = Diffuse, crimson red, individual vessels not easily discernible
3 = Diffuse beefy red

Chemosis: lids and/or nictitating membranes
0 = No swelling
1 = Any swelling above normal (including nictitating membranes)
2 = Obvious swelling with partial eversion of lids
3 = Swelling with lids about half closed
4 = Swelling with lids more than half closed

TOOL USED TO ASSESS SCORE
- Observations included indications of immediate sensory irritation and estimations of chemosis and redness of the adnexal structures. Also evaluated were effects on the iris, the presence of corneal opacity, andor discharge from the eye. Eyes were treated with a 2 % ophthalmic solution of fluorescein at 24 hours and observed for staining.
Irritation parameter:
cornea opacity score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
0
Reversibility:
other: no effects seen
Irritation parameter:
iris score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
0
Reversibility:
other: no effects seen
Irritation parameter:
conjunctivae score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
1
Reversibility:
other: no effects seen
Irritation parameter:
chemosis score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
0
Reversibility:
other: no effects seen
Irritant / corrosive response data:
In the in vivo eye irritation study, signs of irritation consisted of redness (grade 1) for all unwashed eyes one hour after administration of the test substance. At the 24-hour examinations, all three unwashed eyes appeared clinically normal. Immediate irrigation of the eyes was slightly palliative. Signs of irritation for the three washed eyes were limited to redness (grade 1) for two of three washed eyes at the one-hour examinations. At the 24-hour examination, all three washed eyes appeared clinically normal. Staining was not evident in unwashed or washed eyes when tested with fluorescein dye.
Other effects:
No other lesions were noted for the unwashed or washed eyes during the 72-hour observation period. Staining was not evident for unwashed or washed eyes when tested with fluorescein dye 24 hours after administration of the test substance.
No toxic effects were noted during the study.

Observations for Irritation

The treated eye of each animal was examined for signs of irritation at 1, 24, 48, and 72 hours after instillation of the test substance. Observations of corneal opacity, iris effects, redness, and chemosis are presented in Table 1.

Table 1: Ocular Irritation

Animal Number Irrigated Corneal Opacity, Iris effects, Redness and Chemosis*
1 hour 24 hours 48 hours 72 hours
751 No 0,0,1,0 0,0,0,0 0,0,0,0 0,0,0,0
752 No 0,0,1,0 0,0,0,0 0,0,0,0 0,0,0,0
753 No 0,0,1,0 0,0,0,0 0,0,0,0 0,0,0,0
754 Yes 0,0,1,0 0,0,0,0 0,0,0,0 0,0,0,0
755 Yes 0,0,0,0 0,0,0,0 0,0,0,0 0,0,0,0
756 Yes 0,0,1,0 0,0,0,0 0,0,0,0 0,0,0,0

*Graded as described in OECD GuideIine 405 (Annex V., Test B.5) (Grading for Ocular Lesions)

Interpretation of results:
other: Not classified in accordance with EU criteria
Conclusions:
Under the conditions of the test, the substance was determined to be a non-irritant to the eye.
Executive summary:

In a GLP compliant study conducted in line with OECD Guideline 405 and EU Method B.5, the eye irritation of butanoic acid, 4-[[4-[7-chloro-6-(1,1-dimethylethyl)-3H-pyrazolo[1,5-b][1,2,4]triazol-2-yl]phenyl]amino]-4-oxo, tetradecyl ester was investigated in a total of six rabbits. A single dose of 0.1 gram of the test substance was administered into the conjunctival sac of one eye of each animal. Immediately after administration, the treated eyes of three rabbits were washed with distilled water. The remaining eyes were not irrigated. Ocular lesions were graded according to OECD Guideline 405 (Annex V., Test B.5) (Grades for Ocular Lesions).

For the unwashed eyes, signs of irritation consisted of redness (grade 1) for all unwashed eyes one hour after administration of the test substance. At the 24-hour examinations, all three unwashed eyes appeared clinically normal. Staining was not evident in unwashed eyes when fluorescein dye was applied. Immediate irrigation of the eyes was slightly palliative. Signs of irritation for the three washed eyes were limited to redness (grade 1) for two of three washed eyes at the one-hour examinations. At the 24-hour examination, all three washed eyes appeared clinically normal. Staining was not evident in washed eyes when tested with fluorescein dye. Based on these results, the test substance is not classified for eye irritation in line with current EU requirements.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Additional information

Skin irritation

The key study (Jessup, 2003) was performed in compliance with GLP and to OECD Guideline 404 and EU Method B.4 with a sufficient level of detail to assess the quality of the presented data. The study was performed to a good standard in line with accepted, standardised guidelines and was assigned a reliability score of 1 using the principles for assessing data quality as set out in Klimisch et al. (1997). The test substance was found to be not irritating.

Eye irritation

The key study (Shepard, 2003) was performed in compliance with GLP and to OECD Guideline 405 and EU Method B.5 with a sufficient level of detail to assess the quality of the presented data. The study was performed to a good standard in line with accepted, standardised guidelines and was assigned a reliability score of 1 using the principles for assessing data quality as set out in Klimisch et al. (1997). The test substance was found to be not irritating.

The supporting study (Emmons, 2003) was performed in compliance with GLP and to good scientific principles but not in line with a standard, recognised in vitro guideline. On this basis the study was assigned a reliability score of 2 using the principles for assessing data quality as set out in Klimisch et al. (1997). It was found that the test substance may have the potential to produce, at most, minimal eye irritation.

Justification for classification or non-classification

In accordance with the criteria for classification as defined in Annex I, Regulation (EC) No 1272/2008, the substance does not require classification with respect to skin and eye irritation.