Bis(5-oxo-L-prolinato-N1,O2)zinc

Endpoint summary

Description of key information

Skin Irritation

Under the conditions of this study, it was concluded that the test material was not irritating.

Eye Irritation

Under the conditions of this study, since lesions continued even 21 days after exposure, it was concluded that the test material in its original form is classified as a Category 1 eye irritant.

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:

09 April 2003 to 02 June 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 2.8 - 4.4 kg
- Housing: Bracket cages with 1 animal/cage.
- Diet: Each animal was limited to 150 g of feed per day.
- Water: Tap water ad libitum
- Acclimation period: Not specified, but healthy rabbits without island skin were selected.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 3 °C
- Humidity (%): 50 ± 20 %
- Air changes (per hr): All-fresh ventilation of 10 to 15 cycles per hour.
- Photoperiod (hrs dark / hrs light): 12 hours from 7 am to 7 pm; between 200 and 600 lux.

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

water

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 g
- Concentration: The test item was prepared as a paste of the maximum concentration (68 %) by adding distilled water at room temperature.

Duration of treatment / exposure:

4 hours

Observation period:

1 week

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: In order to eliminate bias based on the application site, the patch was applied to different locations for each animal.
- Type of wrap if used: Patches were created by placing cotton lint sized 2.5 cm x 2.5 cm on surgical tape. 0.5 g of the test article was placed on the cloth. The patches were applied to the skin on the clipped site of the animals, and fixed into place using stretchable adhesive tape. The patches were covered occlusively for 4 hours.

REMOVAL OF TEST SUBSTANCE
- Washing: the tape and patch were removed, and the test article was removed from the application area using wet paper towels.
- Time after start of exposure: Four hours after application.

OBSERVATION TIME POINTS
An observation of the application site was made 1 hour after removing the patch.
General observation of symptoms: Observations were conducted once a day, with the exception of holidays, for all animals.
Measurement of body weight: The body weight of all of the animals was measured on the day of treatment and during each reading.

SCORING SYSTEM
- Method of calculation:
A. 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 (beet red) or eschar formation preventing grading of erythema (lesion in deep area)

B. Oedema formation
0 - No oedema
1 - Very slight oedema (barely perceptible)
2 - Slight oedema (edges of area well defined by definite raising)
3 - Moderate oedema (raised approximately 1 mm)
4vSevere oedema (raised more than 1 mm and extending beyond area of exposure)

Observations for changes in scaling, coloration, and corrosion were also conducted.

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Skin irritation
For the 68 % of the test material paste, very slight erythema was observed one hour after after removing the patch in 1/3 of the animals, and skin irritation disappeared 24 hours after removal of the patch. There were no changes observed afterwards in the week after removal of the patch.

Other effects:

General observation of symptoms
During the study period, there were no abnormalities in clinical signs observed.

Measurement of body weight
During the study period, there were no significant changes in body weight.

Table 1: Skin Irritation Scores of the Test Material

Reading Time (Hours)	Animal No.	Erythema	Oedema	Mean Score
				Erythema	Oedema	Total
1	00101 00102 00103	0 1 0	0 0 0	0.3	0.0	0.3
24	00101 00102 00103	0 0 0	0 0 0	0.0	0.0	0.0
48	00101 00102 00103	0 0 0	0 0 0	0.0	0.0	0.0
72	00101 00102 00103	0 0 0	0 0 0	0.0	0.0	0.0
1 week	00101 00102 00103	0 0 0	0 0 0	0.0	0.0	0.0

Interpretation of results:

other: Not classified according to EU critera.

Conclusions:

Under the conditions of this study, it was concluded that the test material was not irritating.

Executive summary:

The skin irritation potential of the test material was investigated in a study which was conducted in accordance with the standardised guidelines OECD 404 (Acute Dermal Irritation/ Corrosion). The study was conducted in accordance with generally accepted scientific principles, possibly with incomplete reporting or methodological deficiencies, which do not affect the quality of the relevant results.

During the study, animal was kept individually in each cage, and a card indicating the protocol number, animal number, and the name and concentration of the test article was attached to each of the cages. In addition, mix-ups between the animals were prevented, by refraining from opening 2 cages at the same time. The animals were treated with the 0.5 g test material via semi-occlusive adhesion. Patches were created by placing cotton lint sized 2.5 cm x 2.5 cm on surgical tape. 0.5 g of the test article was placed on the cloth. The patches were applied to the skin on the clipped site of the animals, and fixed into place using stretchable adhesive tape. The patches were covered occlusively for 4 hours. In order to eliminate bias based on the application site, the patch was applied to different locations for each animal. Four hours after application, the tape and patch were removed, and the test article was removed from the application area using wet paper towels.

For the 68 % test material paste, very slight erythema was observed one hour after after removing the patch in 1/3 of the animals, and skin irritation disappeared 24 hours after removal of the patch. There were no changes observed afterwards in the week after removal of the patch.

Under the conditions of the study, the test material was determined to be not irritating.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 May 2003 to 07 August 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

GLP compliance:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Age at study initiation:
- Weight at study initiation: 3.0 - 3.24 kg
- Housing: Bracket cages with 1 animal/cage.
- Diet: Each animal was limited to 150 g feed per day.
- Water: Tap water ad libitum.
- Acclimation period: Upon conducting observations and measurements of the following and monitoring the growth status of the animals during the 11-day quarantine/acclimation period, there were no abnormalities found.
1) Clinical signs: An observation of clinical signs based on visual examinations was carried out every day for all animals, with the exception of holidays.
2) Measurement of body weight: The body weights of all of the animals were measured on the day that the quarantine/acclimation period started and on the day that quarantine/acclimation ended.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 3 °C
- Humidity (%): 50 ± 20 %
- Air changes (per hr): All-fresh ventilation of 10 to 15 cycles per hour.
- Photoperiod (hrs dark / hrs light): 12 hours from 7 am to 7 pm; 200 to 500 lux.

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount applied: The rabbits were fixed into place using retention devices. 0.1 g of the test article was ocularly instilled into the right eye after pulling the lower eyelid away from the eyeball into a saclike state. After ocular instillation, the upper and lower eyelids were gently held together for approximately 2 to 3 seconds.

Duration of treatment / exposure:

24 hours

Observation period (in vivo):

21 days

Number of animals or in vitro replicates:

Three

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: The eye was washed for approximately 10 seconds using distilled water for injection that was heated to approximately 30 °C.
- Time after start of exposure: 24 hours after ocular instillation.

SCORING SYSTEM:
(1) Cornea:
Opacity: Degree of opacity (reading taken from the most opaque area)
0 - No ulceration or opacity
1 - Scattered or diffuse area 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 - Nacrous area; no details of iris visible; size of pupil barely discernible
4 - Opaque cornea; iris is not discernible through the opacity
 
(2) Iris:
0 - Normal
1 - Markedly deepened rugae, congestion, swelling, moderate circumcorneal hyperaemia; iris reactive to light (a sluggish reaction is considered to be an effect)
2 - Hemorrhage,gross destriction, or no reaction to light
 
(3) Redness of the conjunctivae (refers to palpebral and bulbar conjunctivae; sxcluding cornea and iris strictest manner by comparing them to the reference eye):
0 - Normal
1- Some blood vessels hyperemic( injected)
2 - Diffuse, crimson color; individual vessels not easily discernible
3 - Diffuse, beefy red
 
4) Chemosis: Lids and/or nictating membranes
0 - Normal
1 - Some swelling above normal
2 -Obvious swelling, with partial extroversion of lids
3 - Swelling with lids about half closed
4 - Swelling with lids more than half closed

Ocular irritation potential was also graded in the same manner at 24, 48, and 72 hours after ocular instillation. As irritation was still present 72 hours after ocular instillation, observations were also conducted 1 week, 10 days, 15 days, and 21 days after ocular instillation.

General observation of symptoms
Observations were conducted for all animals once daily, with the exception of holidays.

Measurement of body weight
The body weight of all of the animals was measured when ocularly instilling the test article and during the observations (excluding the observation conducted 1 hour after ocular instillation).

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1.67

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

fully reversible

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.67

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2.33

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.67

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

fully reversible

Irritant / corrosive response data:

1 hour after ocular instillation of the test material in its original form, cornea opacity to the extent where the details of the iris were clearly visible was observed in 1/3 of the animals. Congestion of the iris was observed in all animals, hyperaemia of the conjunctivae was observed in all of the animals, and swelling with lids about half closed was observed in all of the animals.
24 hours after ocular instillation, cornea opacity to the extent where details of the iris were clearly visible was observed in all of the animals, and congestion of the iris was observed in all animals. Redness of the conjunctivae of a crimson colour was observed in 2/3 of the animals and hyperaemia was observed in 1/3 of the animals; swelling with lids about half closed was observed in 2/3 of the animals and obvious swelling was observed in 1/3 of the animals.
48 hours after ocular instillation, cornea opacity to the extent where the details of the iris were slightly obscured was observed in 2/3 of the animals, and to the extent where the details of the iris were clearly visible was observed in 1/3 of the animals. Congestion of the iris was observed in all of the animals. Redness of the conjunctivae of a beefy red colour was observed in 1/3 of the animals and of a crimson colour was observed in 2/3 of the animals; swelling with lids about half closed was observed in 2/3 of the animals and obvious swelling was observed in 1/3 of the animals.
72 hours after ocular instillation, cornea opacity to the extent where the details of the iris were slightly obscured was observed in 2/3 of the animals, and to the extent where the details of the iris were clearly visible was observed in 1/3 of the animals. Congestion of the iris was observed in all animals; redness of the conjunctivae of a beefy red colour was observed in 2/3 of the animals and of a crimson colour in 1/3 of the animals; swelling with lids about half closed was observed in 1/3 of the animals and obvious swelling in 2/3 of the animals.
Since ocular irritation did not disappear 72 hours after ocular instillation, observations were continued. 1 week after ocular instillation, cornea opacity to the extent where the details of the iris were slightly obscured was observed in 2/3 of the animals, and to the extent where the details of the iris could be clearly discerned in 1/3 of the animals; redness of the conjunctivae of a beefy red colour with haemorrhaging, and some swelling above normal were observed in all animals. 10 days after ocular instillation, cornea opacity to the extent where the details of the iris were clearly visible was observed in 2/3 of the animals, redness of the conjunctivae of a beefy red colour was observed in all animals, and some swelling above normal was observed in 2/3 of the animals. 15 days after ocular instillation cornea opacity to the extent where the details of the iris were clearly visible was observed in 1/3 of the animals, redness of the conjunctivae of a beefy red colour was observed in all animals, and some swelling above normal was observed in 2/3 of the animals. 21 days after ocular instillation, redness of the conjunctivae of a beefy red colour was observed in all animals, and some swelling above normal was observed in 2/3 of the animals.

Eye Irritation Scores of the Test Material

No.	Sex	Reading time	Cornea		Iris		Conjuctiva
			Cornea	Mean score	Iris	Mean score	Redness	Mean score	Chemosis	Mean score	Secreta
00101 00102 00103	M M M	1 hour	2 2 1	1.7	1 1 1	1.0	1 1 1	1.0	3 3 3	3.0	3 3 3
00101 00102 00103	M M M	24 hours	1 1 1	1.0	1 1 1	1.0	2 2 1	1.7	3 3 2	2.7	0 0 0
00101 00102 00103	M M M	48 hours	2 2 1	1.7	1 1 1	1.0	3 2 2	2.3	3 3 2	2.7	3 3 3
00101 00102 00103	M M M	72 hours	2 2 1	1.7	1 1 1	1.0	3 3 2	2.7	2 3 2	2.3	2 2 2
00101 00102 00103	M M M	1 week	2 2 1	1.7	0 0 0	0.0	3 3 3	3.0	1 1 1	1.0	0 0 0
00101 00102 00103	M M M	10 days	1 1 0	0.7	0 0 0	0.0	3 3 3	3.0	1 1 0	0.7	0 1 0
00101 00102 00103	M M M	15 days	1 1 0	0.3	0 0 0	0.0	3 3 3	3.0	1 1 0	0.7	0 0 0
00101 00102 00103	M M M	21 days	0 0 0	0.0	0 0 0	0.0	3 3 3	3.0	1 1 0	0.7	0 0 0

M = Male

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

Under the conditions of this study, since lesions continued even 21 days after exposure, it was concluded that the test material in its original form is classified as a Category 1 eye irritant.

Executive summary:

The eye irritation potential of the test material was investigated in a study which was in accordance with the standardised guideline OECD 405 (Acute Eye Irritation/Corrosion).

Three male white New Zealand rabbits were used in this study. The rabbits were fixed into place using retention devices. 0.1 g of the test article was ocularly instilled into the right eye after pulling the lower eyelid away from the eyeball into a saclike state. After ocular instillation, the upper and lower eyelids were gently held together for approximately 2 to 3 seconds. The left eye remained untreated and served as the reference control. 24 hours after ocular instillation, the eye was washed for approximately 10 seconds using distilled water for injection that was heated to approximately 30 °C. Observations of clinical signs based on visual examinations were carried out every day for all animals, with the exception of holidays. The body weight of all of the animals was measured when ocularly instilling the test article and during the observations (excluding the observation conducted 1 hour after ocular instillation).

Instillation of undiluted test material into the eyes of the rabbits resulted in ocular irritation. Since ocular irritation did not disappear 72 hours after ocular instillation, observations were continued. 21 days after ocular instillation, redness of the conjunctivae of a beefy red colour was observed in all animals, and some swelling above normal was observed in 2/3 of the animals.

Under the conditions of this study, it was concluded that the test material in its original form has a risk of serious damage to eyes.

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

10 November 2003 to 10 March 2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP in accordance with generally accepted scientific principles, possibly with incomplete reporting or methodological deficiencies, which do not affect the quality of the relevant results.

Qualifier:

no guideline followed

Principles of method if other than guideline:

The primary ocular irritation potential of the test material was investigated using male New Zealand white rabbits. 0.1 mL of 5 % (w/w) and 1 % (w/w) aqueous solution of the test material were placed in the conjunctival sac of rabbit eyes, and ocular reaction was observed periodically. Observations were evaluated according to the Draize method.

GLP compliance:

not specified

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
Preparation was performed immediately before use after purity conversion of the test material. Test article was dissolved at a concentration of 5 % (w/w) and 1 % (w/w), using the distilled water for injection.

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Housing: Individually in stainless steel cages.
- Diet: Animals were allowed 100g/day of pellet diet using stainless-steel automatic feeders.
- Water: Ad libitum
- Acclimation period:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 3 °C
- Humidity (%):50 ± 20 %
- Air changes (per hr): 10 to 15 times per hour.
- Photoperiod (hrs dark / hrs light): 12 hour lighting at 200 – 600 lux.

Vehicle:

water

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL of test article was placed in the conjunctival sac of the right eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about two to three seconds to prevent loss of test article. The left eye remained untreated and served as the reference control.
- Concentration: A concentration of 5 % (w/w) and 1 % (w/w).

Observation period (in vivo):

1 week

Number of animals or in vitro replicates:

3 animals per dose

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: The test material was not removed.

SCORING SYSTEM: Draize.

CORNEA
Opacity - degree of density (area taken for reading)
0: No ulceration or opacity
1: Scattered or diffuse area; details of iris clearly visible
2: Easily discernible translucent areas; details of iris slightly obscured
3: Opalescent areas of opacity; no details of iris visible; size of pupil barely discernible
4: Opaque; iris invisible

Area of cornea involved:
1: One quarter (or less) but not zero (0 - 1/4)
2: Greater than one quarter less than one half (1/4 - 1/2)
3: Greater than one half less than three quarters (2/1 - 3/4)
4: Greater than three quarters (3/4 - 1)

IRIS
0: Normal
1: Folds above normal, congestion, swelling, circumcorneal injection (any one or all of these or a combination of any thereof), iris still reaction to light (sluggish reaction positive)
2: No reaction to light, haemorrhage, gross destruction (any one or all of these)

CONJUNCTIVAE
Redness (refers to palpebral conjunctiva only)
0: Vessel normal
1: Vessels definitely injected above normal
2: More diffuse, deeper crimson red, individual vessels not easily discernible
3: Diffuse beefy red

Chemosis
0: No swelling
1: Any swelling above normal (includes nictitating membrane)
2: Obvious swelling with partial eversion of the lids
3: Swelling with lids about half closed
4: Swelling with lids about half closed to completely closed

Discharge
0: Normal
1: Any amount different from normal (does not include small amount observed in inner canthus of normal animals)
2: Discharge with moistening of the lids and hairs just adjacent to the lids
3: Discharge with moistening of the lids and considerable area around the eye

For each animal, total irritation scores for three indices in each reading time were calculated according to the following formula.
Total irritation score= Irritation score of cornea+ Irritation score of iris+ Irritation score of conjunctiva And the mean total scores of three animals in each reading time were calculated.

OBSERVATIONS
Clinical signs: Clinical signs of all animals were observed daily from delivery of the animals to the last observation of
the study.
Irritation scores: Ocular irritation was assessed according to the following at 1, 24, 48, 72 and 96 hours, as well as 1 week after administration of the test article.
Body weight measurement: The body weights of all animals was measured at application and at each observation time.
Necropsy: No necropsy was performed in the animals sacrificed at termination of observation.

Irritation parameter:

cornea opacity score

Remarks:

5 % solution

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

4

Irritation parameter:

cornea opacity score

Remarks:

5 % solution

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Irritation parameter:

cornea opacity score

Remarks:

5 % solution

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Remarks:

5 % solution

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Remarks:

5 % solution

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Remarks:

5 % solution

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Remarks:

5 % solution

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Irritation parameter:

conjunctivae score

Remarks:

5 % solution

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

3

Irritation parameter:

conjunctivae score

Remarks:

5 % solution

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Irritation parameter:

chemosis score

Remarks:

5 % solution

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

chemosis score

Remarks:

5 % solution

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

chemosis score

Remarks:

5 % solution

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritant / corrosive response data:

The maximum mean total score of 1 % (w/w) test material was 0.0 at all reading times, and classified as non-irritating.

Other effects:

There were no mortality, nor abnormal clinical signs observed during the study period. The body weights of all animals were considered to be within the normal range of variability

Interpretation of results:

other: Not classified according to EU criteria.

Conclusions:

Under the conditions of the study, it is considered that at 1 % (w/w) and 5 % (w/w) the test material is not irritating to rabbit eye according to the CLP classification and labelling criteria.

Executive summary:

The eye irritation potential of the test material was investigated in a study which was conducted following a method similar to that which is outlined in the Modified Draize method for Primary eye irritation.

The test material was dissolved at a concentration of 5 % (w/w) and 1 % (w/w), using the distilled water for injection. 0.1 mL of test article was placed in the conjunctival sac of the right eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about two to three seconds to prevent loss of test article. The left eye remained untreated and served as the reference control. Clinical signs of all animals were observed daily from delivery of the animals to the last observation of the study. Ocular irritation was assessed according to the Draize evaluation method at 1, 24, 48, 72 and 96 hours, as well as 1 week after administration of the test material.

Under the conditions of the study, it is considered that at 1 % (w/w) and 5 % (w/w) the test material is not irritating to rabbit eye according to the CLP classification and labelling criteria.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irreversible damage)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation

The skin irritation potential of the test material was investigated in a study which was conducted in accordance with the standardised guidelines OECD 404 (Acute Dermal Irritation/ Corrosion). The study was conducted in accordance with generally accepted scientific principles, possibly with incomplete reporting or methodological deficiencies, which do not affect the quality of the relevant results; it was therefore assigned a reliability score of 2 in line with the criteria of Klimisch et al. (1997).

During the study, animal was kept individually in each cage, and a card indicating the protocol number, animal number, and the name and concentration of the test article was attached to each of the cages. In addition, mix-ups between the animals were prevented, by refraining from opening 2 cages at the same time. The animals were treated with the 0.5 g test material via semi-occlusive adhesion. Patches were created by placing cotton lint sized 2.5 cm x 2.5 cm on surgical tape. 0.5 g of the test article was placed on the cloth. The patches were applied to the skin on the clipped site of the animals, and fixed into place using stretchable adhesive tape. The patches were covered occlusively for 4 hours. In order to eliminate bias based on the application site, the patch was applied to different locations for each animal. Four hours after application, the tape and patch were removed, and the test article was removed from the application area using wet paper towels.

For the 68 % test material paste, very slight erythema was observed one hour after after removing the patch in 1/3 of the animals, and skin irritation disappeared 24 hours after removal of the patch. There were no changes observed afterwards in the week after removal of the patch.

Under the conditions of the study, the test material was determined to be not irritating.

Eye Irritation

The eye irritation potential of the test material was investigated in a key study which was in accordance with the standardised guideline OECD 405 (Acute Eye Irritation/Corrosion). It was assigned a reliability score of 2 in line with the criteria of Klimisch et al. (1997).

Three male white New Zealand rabbits were used in this study. The rabbits were fixed into place using retention devices. 0.1 g of the test article was ocularly instilled into the right eye after pulling the lower eyelid away from the eyeball into a saclike state. After ocular instillation, the upper and lower eyelids were gently held together for approximately 2 to 3 seconds. The left eye remained untreated and served as the reference control. 24 hours after ocular instillation, the eye was washed for approximately 10 seconds using distilled water for injection that was heated to approximately 30 °C. Observations of clinical signs based on visual examinations were carried out every day for all animals, with the exception of holidays. The body weight of all of the animals was measured when ocularly instilling the test article and during the observations (excluding the observation conducted 1 hour after ocular instillation).

Instillation of undiluted test material into the eyes of the rabbits resulted in ocular irritation. Since ocular irritation did not disappear 72 hours after ocular instillation, observations were continued. 21 days after ocular instillation, redness of the conjunctivae of a beefy red colour was observed in all animals, and some swelling above normal was observed in 2/3 of the animals.

Under the conditions of this study, it was concluded that the test material in its original form has a risk of serious damage to eyes.

The eye irritation potential of the test material was also investigated in a supporting study which was conducted following a method similar to that which is outlined in the Modified Draize method for Primary eye irritation. It was assigned a reliability score of 2 in line with the criteria of Klimisch et al. (1997).

The test material was dissolved at a concentration of 5 % (w/w) and 1 % (w/w), using the distilled water for injection. 0.1 mL of test article was placed in the conjunctival sac of the right eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about two to three seconds to prevent loss of test article. The left eye remained untreated and served as the reference control. Clinical signs of all animals were observed daily from delivery of the animals to the last observation of the study. Ocular irritation was assessed according to the Draize evaluation method at 1, 24, 48, 72 and 96 hours, as well as 1 week after administration of the test material.

Under the conditions of the study, it is considered that at 1 % (w/w) and 5 % (w/w) the test material is not irritating to rabbit eye.

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 is classified as a Category 1 eye irritant. No classification is required for skin irritation.