Potassium sodium amino-hydroxy-[(3-{[2-(substituted)ethyl]sulfonyl}phenyl)diazenyl]-[(2-sulfonato-4-{[2-(substituted)ethyl]sulfonyl}phenyl)diazenyl]naphthalene-disulfonate

Eye irritation

Some information in this page has been claimed confidential.

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was conducted between 19 January 2015 and 29 January 2015.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Deviations:

no

Principles of method if other than guideline:

None

GLP compliance:

yes

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

solid: particulate/powder

Details on test material:

None

Specific details on test material used for the study:

None

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Two New Zealand White (Hsdlf:NZW) strain rabbits were supplied by Harlan Laboratories UK Ltd., Leicestershire, UK. At the start of the study the animals weighed 2.54 or 3.01 kg and were twelve to twenty weeks old. After an acclimatization period of at least five days each animal was given a number unique within the study which was written with a black indelible marker pen on the inner surface of the ear and on the cage label.

The animals were individually housed in suspended cages. Free access to mains drinking water and food (2930C Teklad Global Rabbit diet supplied by Harlan Laboratories UK Ltd., Oxon, UK) was allowed throughout the study. The diet and drinking water were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

The temperature and relative humidity were set to achieve limits of 17 to 23 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.

The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Justification
The rabbit is the preferred species of choice as historically used for irritation studies and is specified in the appropriate test guidelines. The number of animals used was the minimum required to achieve the objectives of the study. Testing was conducted in two animals and the response in those animals was such that exposure of a third animal would not affect classification of the test item, therefore, no further testing was needed.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: The left eye remained untreated and was used for control purposes.

Amount / concentration applied:

A volume of 0.1 mL of the test item, which was found to weigh approximately 92 mg (as measured by gently compacting the required volume into an adapted syringe) was used.

Duration of treatment / exposure:

The test item was not removed.

Observation period (in vivo):

72 hours

Number of animals or in vitro replicates:

Two

Details on study design:

Test Item Formulation and Experimental Preparation
For the purpose of the study the test item was used as supplied. The absorption of the test item was not determined.

Procedure
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. Only animals free of ocular damage were used.

Initially, a single rabbit was treated. A subcutaneous injection of buprenorphine 0.01 mg/kg was administered 60 minutes prior to test item application to provide a therapeutic level of systemic analgesia. Five minutes prior to test item application, a pre dose anesthesia of ocular anesthetic (two drops of 0.5% tetracaine hydrochloride) was applied to each eye.

A volume of 0.1 mL of the test item, which was found to weigh approximately 92 mg (as measured by gently compacting the required volume into an adapted syringe) was placed into the conjunctival 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 treatment, to prevent loss of the test item, and then released. The left eye remained untreated and was used for control purposes. Immediately after administration of the test item, an assessment of the initial pain reaction was made.

Eight hours after test item application, a subcutaneous injection of post dose analgesia, buprenorphine 0.01 mg/kg and meloxicam 0.5 mg/kg, was administered to provide a continued therapeutic level of systemic analgesia. The treated animal was checked for signs of pain and suffering approximately 12 hours later. No further analgesia was required.

After consideration of the ocular responses produced in the first treated animal, a second animal was similarly treated.

Assessment of ocular damage/irritation was made approximately 1 hour and 24, 48 and 72 hours following treatment, according to the numerical evaluation (Draize, J.H, 1977). Any other ocular effects were also noted. Examination of the eye was facilitated by the use of the light source from a standard ophthalmoscope.

Any clinical signs of toxicity, if present, were also recorded.

Individual body weights were recorded on Day 0 (the day of dosing) and at the end of the observation period.


Interpretation of Results
The numerical values corresponding to each animal, tissue and observation time were recorded. The data relating to the conjunctivae were designated by the letters A (redness), B (chemosis) and C (discharge), those relating to the iris designated by the letter D and those relating to the cornea by the letters E (degree of opacity) and F (area of cornea involved). For each tissue the score was calculated as follows:

Score for conjunctivae = (A + B + C) x 2
Score for iris = D x 5
Score for cornea = (E x F) x 5

Using the numerical data obtained a modified version of the system described by Kay J.H. and Calandra J.C. (1962) was used to classify the ocular irritancy potential of the test item. This was achieved by adding together the scores for the cornea, iris and conjunctivae for each time point for each rabbit. The group means of the total scores for each observation were calculated. The highest of these group means (the maximum group mean score) together with the persistence of the reactions enabled classification of the eye irritancy potential of the test item.

If evidence of irreversible ocular damage is noted, the test item will be classified as corrosive to the eye.

The results were also interpreted according to the Globally Harmonized System of Classification and Labelling of Chemicals

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Remarks:

Male 74909

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

other: Not classified

Results 2

Irritation parameter:

cornea opacity score

Remarks:

Male 74917

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

other: Not classified

Results 3

Irritation parameter:

iris score

Remarks:

Male 74909

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0

Max. score:

2

Reversibility:

other: Not applicable

Remarks on result:

other: Not classified

Results 4

Irritation parameter:

iris score

Remarks:

Male 74917

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0

Max. score:

2

Reversibility:

other: Not applicable

Remarks on result:

other: Not classified

Results 5

Irritation parameter:

conjunctivae score

Remarks:

Male 74909

Basis:

mean

Remarks:

Redness

Time point:

other: 24, 48 and 72 hours

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: Not classified

Results 6

Irritation parameter:

conjunctivae score

Remarks:

Male 74917

Basis:

mean

Remarks:

Redness

Time point:

other: 24, 48 and 72 hours

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: Not classified

Results 7

Irritation parameter:

chemosis score

Remarks:

Male 74909

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0.66

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: Not classified

Results 8

Irritation parameter:

chemosis score

Remarks:

Male 74917

Basis:

mean

Time point:

other: 24, 48 and 72 hours

Score:

0.66

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: Not classified

Results 9

Irritation parameter:

maximum mean total score (MMTS)

Basis:

mean

Time point:

other: 1 hour

Score:

10

Max. score:

110

Reversibility:

fully reversible within: 72 hours

Results 10

Irritation parameter:

maximum mean total score (MMTS)

Basis:

mean

Time point:

other: 24 hours

Score:

9

Max. score:

110

Reversibility:

fully reversible within: 72 hours

Results 11

Irritation parameter:

maximum mean total score (MMTS)

Basis:

mean

Time point:

other: 48 hours

Score:

4

Max. score:

110

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: Classed as a mild irritant according to a modified Kay and Calandra classification system; not classified according to the Globally Harmonized System of Classification and Labelling of Chemicals.

Irritant / corrosive response data:

Blue colored staining of the fur around the treated eye was noted in both animals during the study.

No corneal or iridial effects were noted during the study.

Moderate conjunctival irritation was noted in both treated eyes 1 and 24 hours after treatment with minimal conjunctival irritation noted at the 48 Hour observation.

Both treated eyes appeared normal at the 72 Hour observation.

Other effects:

Body Weight
No body weight gain was noted in one animal and the other animal showed expected gain in body weight during the study.

Any other information on results incl. tables

Individual Scores and Individual Total Scores for Ocular Irritation

Rabbit Number and Sex	74909Male				74917Male
	IPR= 0				IPR = 0
Time After Treatment	1 Hour	24 Hours	48 Hours	72 Hours	1 Hour	24 Hours	48 Hours	72 Hours
CORNEA
E = Degree of Opacity	0	0	0	0	0	0	0	0
F = Area of Cornea Involved	0	0	0	0	0	0	0	0
Score (E x F) x 5	0	0	0	0	0	0	0	0
IRIS
D	0	0	0	0	0	0	0	0
Score (D x 5)	0	0	0	0	0	0	0	0
CONJUNCTIVAE
A = Redness	2	2	1	0	2	2	1	0
B = Chemosis	1	1	1	0	1	1	1	0
C = Discharge	2Sf	1	0	0Sf	2Sf	2Sf	0Sf	0Sf
Score (A + B + C) x 2	10	8	4	0	10	10	4	0
Total Score	10	8	4	0	10	10	4	0

IPR=Initial pain reaction

Sf = Blue colored staining of the fur around the treated eye

Individual Total Scores and Group Mean Scores for Ocular Irritation

Rabbit Number and Sex		Individual Total Scores At:
		1 Hour		24 Hours		48 Hours		72 Hours
74909Male		10		8		4		0
74917Male		10		10		4		0
Group Total		20		18		8		0
Group Mean Score		10.0		9.0		4.0		0.0

 

Individual Body Weights and Body Weight Change

Rabbit Number and Sex	Individual Body Weight (kg)		Body Weight Change (kg)
	Day 0	Day 3
74909 Male	3.01	3.01	0.00
74917 Male	2.54	2.56	0.02

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item can be considered as not-irritating to the eyes of rabbits.

Executive summary:

The irritancy potential of the test item to the eye of the New Zealand White rabbit was evaluated in a study conducted according to OECD Guideline 405 and EU Method B.5. A single application of the test item to the non-irrigated eye of two rabbits produced moderate conjunctival irritation. Both treated eyes appeared normal at the 72‑Hour observation. The test item produced a maximum group mean score of 10.0 and was classified as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system. However, the test item does not meet the criteria for classification according to the CLP (Regulation 1272/2008) criteria. Hence, can be considered as non-irritating to the eyes of rabbits.