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Toxicological information

Eye irritation

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Administrative data

Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
31 August 2011 to 21 September 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
yes
Remarks:
The relative humidity (max 89%) was out of the target range (30-70%) during the study.
Qualifier:
according to guideline
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
yes
Remarks:
The relative humidity (max 89%) was out of the target range (30-70%) during the study.
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2400 (Acute Eye Irritation)
Deviations:
yes
Remarks:
The relative humidity (max 89%) was out of the target range (30-70%) during the study.
GLP compliance:
yes (incl. QA statement)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
Substance name: Reactive Yellow F01-0555

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
Species and strain: New Zealand White rabbits
Source: S&K-LAP Kft. 2173 Kartal, Császár út 135, Hungary
Justification of strain: The New Zealand White rabbit is one of the standard strains used for acute irritation toxicity studies.
Animal health: Only animals in acceptable health condition were used for the test. Both eyes of each animal provisionally selected for testing were examined approximately one hour before starting the study. Animals showing eye irritation, ocular defects or pre-existing corneal injury were not used.
Number of animals: 3 animals
Age of animals at treatment: ~11 weeks old (adult)
Sex: Male
Body weight range at the
beginning of the life phase: 2754 – 2837 g
end of the life phase: 3152 – 3233 g
Date of receipt: 24 August 2011
Acclimatization time: 7 days
Animal identification: The individual identification was by engraved ear tag. The cages were marked with individual identity cards with information about study code, sex, dose group, cage number and individual animal number.

HUSBANDRY
Animal health: Only healthy animals were used for the test. The veterinarian certified health status.
Number of animal room: 632
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 20 ± 3°C
Relative humidity: 40 - 89 %
Housing/Enrichment: Rabbits were individually housed in AAALAC approved metal wire rabbit cages. Cages were of an open wire structure and cages were placed together to allow some social interaction with rabbit(s) in adjoining cages
Ventilation: 15-20 air exchanges/hour

The environmental parameters were recorded twice daily during the study. Variations from the target humidity (max. 89%) range were observed during the study. These deviations were considered to have no impact on the animal health, as certified by the Clinical Veterinarian, or on the outcome of the study and interpretation of the results.

FOOD AND FEEDING

Animal received PURINA Base – Lap gr. diet for rabbits produced by AGRIBRANDS Europe Hungary PLC, H-5300 Karcag, Madarasi road, Hungary, ad libitum.

WATER SUPPLY AND QUALITY CONTROL OF WATER

The animals received municipal tap water, as for human consumption, ad libitum, from an automatic system.

The quality control analysis is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A.u.36., Hungary). The quality control results are retained in the archives of CiToxLAB Hungary Ltd.

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
A single dose of 0.1 g of the solid test item was administered to each animal.
Duration of treatment / exposure:
1 hour
Observation period (in vivo):
3 weeks
Number of animals or in vitro replicates:
3
Details on study design:
Application of the Test Item

Three male animals in acceptable health condition were selected for the test. Care was taken to select only those animals that had a normal eye condition and any with ocular lesions were rejected.

An initial test was performed using one animal. The test item was instilled into the conjunctival sac of the left eye. The eyelids were held closed for several seconds to prevent the loss of the test item. The contralateral eye served as the control. Immediately after the administration of the test item, an assessment of the initial pain reaction was made.

After consideration of the ocular responses produced in the first animal, two additional animals were treated.

Duration of Exposure

The eyes of the test animals were washed out at 1 hour after application of test item.

OBSERVATIONS AND SCORING

Clinical Observations

The eyes were examined at 1, 24, 48, 72 hours and 1, 2 and 3 weeks after treatment. The duration of the observation period was sufficient to identify reversibility or irreversibility of changes. Any clinical signs of toxicity or signs of ill- health during the study were recorded.

At the end of the observation period, each animal was sacrificed by intramuscular injections of CP-Ketamin 10% and CP-Xylazin 2% followed by iv. Euthasol® 40% anaesthesia. Death was verified by checking pupil and corneal reflex and the absence of respiration.

Scoring and Assessment of Local Reaction

The eye irritation scores were evaluated according to the scoring system by Draize (1977) and OECD 405 (24 April 2002).


Classification of the Test Items

Individual reactions of the animal were recorded at each observation time. The nature, severity and duration of all lesions observed were described.

Results were presented and interpreted according to Regulation (EC) No 1272/2008, as follows:

Irreversible effects on the eye/serious damage to eyes (Category 1)

Substances that have the potential to seriously damage the eyes are classified in Category 1 (irreversible effects on the eye). 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.

Category for irreversible eye effects:

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:
o corneal opacity ≥ 3 and/or
o iritis > 1.5
calculated as the mean scores following grading at 24, 48 and 72 hours after installation of the test material.

Reversible effects on the eye/irritating to eyes (Category 2):

Substances that have the potential to induce reversible eye irritation are classified in Category 2 (irritating to eyes).

Category for reversible eye effects
If, when applied to the eye of an animal, a substance produces:
— at least in 2 of 3 tested animals, a positive response of:
o corneal opacity ≥ 1 and/or
o iritis ≥ 1, and/or
o conjunctival redness ≥ 2 and/or
o 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

Measurement of Body Weight

Individual body weight was recorded at the beginning and end of the experiment.


Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
mean
Remarks:
all animals
Time point:
24/48/72 h
Score:
0
Max. score:
4
Remarks on result:
no indication of irritation
Irritation parameter:
iris score
Basis:
mean
Remarks:
all animals
Time point:
24/48/72 h
Score:
0
Max. score:
2
Remarks on result:
no indication of irritation
Irritation parameter:
conjunctivae score
Basis:
animal #1
Time point:
24/48/72 h
Score:
0.67
Max. score:
3
Reversibility:
fully reversible within: 72 hours
Irritation parameter:
conjunctivae score
Basis:
animal #2
Time point:
24/48/72 h
Score:
1.33
Max. score:
3
Reversibility:
fully reversible within: 2 weeks
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
24/48/72 h
Score:
1.67
Max. score:
3
Reversibility:
fully reversible within: 2 weeks
Irritation parameter:
chemosis score
Basis:
animal #1
Time point:
24/48/72 h
Score:
0
Max. score:
4
Remarks on result:
no indication of irritation
Irritation parameter:
chemosis score
Basis:
animal #2
Time point:
24/48/72 h
Score:
0.33
Max. score:
4
Reversibility:
fully reversible within: 48 hours
Irritation parameter:
chemosis score
Basis:
animal #3
Time point:
24/48/72 h
Score:
0.33
Max. score:
4
Reversibility:
fully reversible within: 48 hours
Irritant / corrosive response data:
The eyes were examined at 1, 24, 48, 72 hours and 1, 2 and 3 weeks after the application.

Initial Pain Reaction (IPR) (score 2) was observed in all animals after test item administration.

One hour after the application:
Conjunctival redness (score 2) was found in all rabbits, conjunctival chemosis (score 1) were observed in two animals and all rabbits showed conjunctival discharge (score 3).

At 24 hours after treatment:
Conjunctival redness (score 2 or 1) was found in all rabbits, chemosis (score 1).was observed in two animals. One rabbit displayed conjunctival discharge (score 1).

At 48 hours after treatment:
Conjunctival redness (score 1) was observed in two animals and redness (score 2) was noted in one rabbit.

At 72 hours after treatment:
Conjunctival redness (score 1) was observed in two animals.

At 1 week after treatment:
Conjunctival redness (score 1) was observed in 2 rabbits.


At 2 and 3 weeks after treatment:
No signs of eye irritation or other clinical signs were observed.

The study was terminated after the 3-week observation.
Other effects:
The conjunctivae were coloured by the test item during the study up to and including the 3-week observation.

Any other information on results incl. tables

TABLE 1: INDIVIDUAL SCORES FOR OCULAR IRRITATION

Study Code:             11/174-005N                     Species:    NZW Rabbit

Dose:                        0.1g                                  Sex:          Male

Start of Exposure:    31 August 2011                Test Item: Reactive Yellow F01-0555

 

 

Time

Animal No.

Score of irritation

IPR

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

1 hour

00427

2

0

3

0

0

0

0

*

2

00388

2

1

3

0

0

0

0

*

2

00389

2

1

3

0

0

0

0

*

2

Time of Observation: Day 0

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

24 hours

00427

1

0

1

0

0

0

0

*

00388

2

1

0

0

0

0

0

*

00389

2

1

0

0

0

0

0

*

Time of Observation: Day 1

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

48 hours

00427

1

0

0

0

0

0

0

*

00388

1

0

0

0

0

0

0

*

00389

2

0

0

0

0

0

0

*

Time of Observation: Day 2

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

72 hours

00427

0

0

0

0

0

0

0

*

00388

1

0

0

0

0

0

0

*

00389

1

0

0

0

0

0

0

*

Time of Observation: Day 3


 

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

1 week

00427

1

0

0

0

0

0

0

*

00388

0

0

0

0

0

0

0

*

00389

1

0

0

0

0

0

0

*

Time of Observation: Day 7

 

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

2 weeks

00427

0

0

0

0

0

0

0

*

00388

0

0

0

0

0

0

0

*

00389

0

0

0

0

0

0

0

*

Time of Observation: Day 14

 

Time

Animal No.

Score of irritation

Conjunctivae

Opacity of cornea

Iris

Control eye

Other sign

R

CH

D

OD

OE

R

3 weeks

00427

0

0

0

0

0

0

0

*

00388

0

0

0

0

0

0

0

*

00389

0

0

0

0

0

0

0

*

Time of Observation: Day 21

 

Abbreviations:   R    = Redness                                OD =   Opacity degree of density

                           CH = Chemosis                               OE =   Extent of opaque area

D    = Discharge                              

*: The conjunctivae were coloured by the test item.

TABLE 2: MEAN VALUES OF EYE IRRITATION (24, 48, 72 hour reading)

Study Code:             11/174-005N                    Species:    NZW Rabbit

Dose:          0.1g                                   Sex:          Male

Start of Exposure:     31 August 2011                Test Item: Reactive Yellow F01-0555

 

  

Animal Number

Cornea Opacity

Iris

Conjunctivae

Redness

Chemosis

Discharge

00427

0.00

0.00

0.67

0.00

0.33

00388

0.00

0.00

1.33

0.33

0.00

00389

0.00

0.00

1.67

0.33

0.00

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The test item Reactive Yellow F01-0555, applied to rabbit eye mucosa, caused significant conjunctival irritant effects at one hour which were reduced at 24 hours after application. There were no corneal effects observed at any timepoint, all conjunctival irritation effects were completely reversed after the 2-week observation period. Some discoloration of the conjuctivae remained at the observation period of 3 weeks, but this is not considered as classifiable effect under CLP or GHS guidelines.
Reactive Yellow F01-0555 is “not classified” according to Regulation (EC) No 1272/2008.

Executive summary:

An acute eye irritation study of the test item Reactive Yellow F01-0555 was performed in New Zealand White rabbits. The irritation effects of the test item were evaluated according to the Draize method (OECD No.: 405, 2002).

The test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. An amount ofof the test item was administered as a single dose.

 

Individual body weight was recorded at the beginning and end of the study. Morbidity and clinical signs of toxicity were checked daily. The eyes were examined at 1, 24, 48, 72 hours and 1, 2 and 3 weeks after the application.

 

No adverse effects on body weight development were noted during the study period. The general state and behavior of animals were normal throughout the study period.

 

Eye examination:

 

During the study, no signs of eye irritation were observed in the control eye of all animals.

 

Initial Pain Reaction(IPR) (score 2) was observed in all animals after test item administration.

 

One hour after the application:

Conjunctival redness (score 2) was found in all rabbits, conjunctival chemosis (score 1) were observed in two animals and all rabbits showed conjunctival discharge (score 3).

At 24 hours after treatment:

Conjunctival redness (score 2 or 1) was found in all rabbits, chemosis (score 1).was observed in two animals. One rabbit displayed conjunctival discharge (score 1).

At 48 hours after treatment:

Conjunctival redness (score 1) was observed in two animals and redness (score 2) was noted in one rabbit.

 

At 72 hours after treatment:

Conjunctival redness (score 1) was observed in two animals.

 

At 1 week after treatment:

Conjunctival redness (score 1) was observed in 2 rabbits.

 

At 2 and 3 weeks after treatment:

No signs of eye irritation or other clinical signs were observed.


The conjunctivae were coloured by the test item during the study up to and including the 3-week observation.

 

The study was terminated after the 3-week observation.

 

The animal’s individual mean scores (considering readings at 24, 48 and 72 hours after the treatment) were as follows:

Animal Number

Cornea Opacity

Iris

Conjunctivae

Redness

Chemosis

Discharge

00427

0.00

0.00

0.67

0.00

0.33

00388

0.00

0.00

1.33

0.33

0.00

00389

0.00

0.00

1.67

0.33

0.00

The test item Reactive Yellow F01-0555, applied to rabbit eye mucosa, caused significant conjunctival irritant effects at one hour which were reduced at 24 hours after application. There were no corneal effects observed at any timepoint, all conjunctival irritation effects were completely reversed after the 2-week observation period. Some discoloration of the conjuctivae remained at the observation period of 3 weeks, but this is not considered as classifiable effect under CLP or GHS guidelines.

Reactive Yellow F01-0555 is “not classified” according to Regulation (EC) No 1272/2008.