lithium 3-((3,4-dicyanophenyl)sulfonyl)propane-1-sulfonate)

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Remarks:

Study carried out before the entry into force of amendments to annex VII

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Between 06 June 2012 and 02 July 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP and in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do no effect the quality of the relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

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

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

lithium 3-((3,4-dicyanophenyl)sulfonyl)propane-1-sulfonate)

EC Number:

700-777-2

Cas Number:

667889-11-4

Molecular formula:

C11H9N2O5S2Li

IUPAC Name:

lithium 3-((3,4-dicyanophenyl)sulfonyl)propane-1-sulfonate)

Test material form:

solid: particulate/powder

Details on test material:

Sponsor's identification: S193308
Description : white powder
Batch number : 422 (Bag 3)
Purity : 98.9%
Date received : 02 May 2012
Expiry date : not supplied
Storage conditions: room temperature in the dark
The integrity of supplied data relating to the identity, purity and stability of the test item is the responsibility of the Sponsor.

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Three 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 2.51 to 2.85 kg and were twelve to twenty weeks old. After an acclimatisation 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 Certified 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. Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study. 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.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

A volume of 0.1 ml of the test item, which was found to weigh approximately 72 mg.

Duration of treatment / exposure:

1 hour

Observation period (in vivo):

7 days

Number of animals or in vitro replicates:

3

Details on study design:

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 volume of 0.1 ml of the test item, which was found to weigh approximately 72 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 according to the six point scale shown in appendix 1.

After consideration of the ocular responses produced in the first treated animal, two additional animals were treated. In order to minimise pain on application of the test item, one drop of local anaesthetic (Tetracaine hydrochloride 0.5%, Bausch & Lomb, Surrey, UK) was instilled into both eyes of these animals 1 to 2 minutes before treatment.

Assessment of ocular damage/irritation was made approximately 1 hour and 24, 48 and 72 hours following treatment, according to the numerical evaluation given in apppendix 2 below (from Draize J H (1977) "Dermal and Eye Toxicity Tests" In: Principles and Procedures for Evaluating the Toxicity of Household Substances, National Academy of Sciences, Washington DC p.48 to 49).

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. An additional observation was made on Day 7 to assess the reversibility of the ocular effects. Individual bodyweights were recorded on Day 0 (the day of dosing) and at the end of the observation period.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

conjunctivae score

Remarks:

Redness

Basis:

animal: 72093 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

2

Max. score:

3

Reversibility:

fully reversible within: 72 hours

Results 2

Irritation parameter:

conjunctivae score

Remarks:

Redness

Basis:

animal: 72126 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

2

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 3

Irritation parameter:

conjunctivae score

Remarks:

Redness

Basis:

animal: 72156 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

2

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 4

Irritation parameter:

chemosis score

Basis:

animal: 72093 Male

Time point:

other: Highest score at 1, 24, 48 and 72 hours

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Results 5

Irritation parameter:

chemosis score

Basis:

animal: 72126 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 6

Irritation parameter:

chemosis score

Basis:

animal: 72156 Male

Time point:

other: mean at 24, 48 and 72 hours

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 7

Irritation parameter:

cornea opacity score

Basis:

animal: 72093 Male

Time point:

other: Highest score at 1, 24, 48 and 72 hours

Score:

0

Max. score:

4

Reversibility:

other: No effect noted

Results 8

Irritation parameter:

cornea opacity score

Basis:

animal: 72126 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 9

Irritation parameter:

cornea opacity score

Basis:

animal: 72156 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

0

Max. score:

4

Reversibility:

other: No effect noted

Results 10

Irritation parameter:

iris score

Basis:

animal: 72093

Time point:

other: Highest score at 1, 24, 48 and 72 hours

Score:

0

Max. score:

2

Reversibility:

other: no effect noted

Results 11

Irritation parameter:

iris score

Basis:

animal: 72126 Male

Time point:

other: mean at 24, 48 and 72 hours

Score:

1

Max. score:

2

Reversibility:

fully reversible within: 7 days

Results 12

Irritation parameter:

iris score

Basis:

animal: 72156 Male

Time point:

other: Highest score at 1, 24, 48, 72 hours and 7 days

Score:

0

Max. score:

2

Reversibility:

other: no effect noted

Irritant / corrosive response data:

Individual and group mean scores for ocular irritation are given in Table 1 and Table 2.
Iridial inflammation was noted in one treated eye one hour after treatment. Scattered or diffuse corneal opacity and iridial inflammation were noted in this treated eye at the 24, 48 and 72 Hour observations. No corneal or iridial effects were noted in the two other treated eyes during the study.
Moderate conjunctival irritation was noted in all treated eyes one hour after treatment. Moderate conjunctival irritation was noted in two treated eyes with minimal conjunctival irritation noted in one treated eye at the 24 Hour observation. Moderate conjunctival irritation was noted in one treated eye with minimal conjunctival irritation noted in two treated eyes at the 48 Hour observation. Moderate conjunctival irritation persisted in one treated eye with minimal conjunctival irritation noted in one other treated eye at the 72 Hour observation.
One treated eye appeared normal at the 72 Hour observation and two treated eyes appeared normal at the 7 Day observation.

Other effects:

Bodyweight
Individual bodyweights and bodyweight changes are given in Table 3.
All animals showed expected gain in bodyweight during the study.

Any other information on results incl. tables

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), J. Soc. Cosmet. Chem.13, 281‑289 (seeAppendix3) 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 interpreted according to EU labelling regulations Commission Directive 2001/59/EC for classification and labelling of dangerous substances andRegulation (EC) No 1272/2008, relating to the Classification, Packaging and Labelling of Dangerous Substances.

Table 1              Individual Scores and Individual Total Scores for Ocular Irritation

Rabbit Number and Sex	72093Male				72126Male					72156Male
	IPR= 3				IPR = 0+					IPR = 0+
Time After Treatment	1 Hour	24 Hours	48 Hours	72 Hours	1 Hour	24 Hours	48 Hours	72 Hours	7 Days	1 Hour	24 Hours	48 Hours	72 Hours	7 Days
CORNEA
E = Degree of Opacity	0	0	0	0	0	1	1	1	0	0	0	0	0	0
F = Area of Cornea Involved	0	0	0	0	0	2	2	1	0	0	0	0	0	0
Score (E x F) x 5	0	0	0	0	0	10	10	5	0	0	0	0	0	0
IRIS
D	0	0	0	0	1	1	1	1	0	0	0	0	0	0
Score (D x 5)	0	0	0	0	5	5	5	5	0	0	0	0	0	0
CONJUNCTIVAE
A = Redness	2	2	1	0	2	2	2	2	0	2	2	2	1	0
B = Chemosis	1	1	1	0	2	2	2	2	0	1	1	1	1	0
C = Discharge	2	1	0	0	2	2	1	1	0	2	0	0	0	0
Score (A + B + C) x 2	10	8	4	0	12	12	10	10	0	10	6	6	4	0
Total Score	10	8	4	0	17	27	25	20	0	10	6	6	4	0

IPR=  Initial pain reaction

+=     One drop of local anaesthetic instilled into both eyes 1 to 2 minutes before treatment

Table 2              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	7 Days
72093Male	10	8	4	0	-
72126Male	17	27	25	20	0
72156Male	10	6	6	4	0
Group Total	37	41	35	24	0
Group Mean Score	12.3	13.7	11.7	8.0	0.0

- =     Observation not required - considered to be zero for calculation of Group Mean Score

Table 3              Individual Bodyweights and Bodyweight Changes

Rabbit Number and Sex	Individual Bodyweight (kg)		Bodyweight Change (kg)
72093Male	Day 0	Day 3	0.05
	2.85	2.90
72126Male	Day 0	Day 7	0.08
	2.79	2.87
72156Male	Day 0	Day 7	0.14
	2.51	2.65

Applicant's summary and conclusion

Interpretation of results:

other: mild irritant (Class 4 on a 1 to 8 scale)

Remarks:

Criteria used for interpretation of results: other: modified Kay and Calandra classification system

Conclusions:

The test item produced a maximum group mean score of 13.7 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. The test item did not meet the criteria for classification as irritant or corrosive according to EU labelling regulations Commission Directive 2001/59/EC or the criteria for classification according to Regulation (EC) No 1272/2008, relating to the Classification, Packaging and Labelling of Dangerous Substances.

Executive summary:

Introduction. The study was performed to assess the irritancy potential of the test item to the eye of the New Zealand White rabbit. The method was designed to be compatible with the following:

OECD Guidelines for the Testing of Chemicals No. 405 "Acute Eye Irritation/Corrosion" (adopted 24 April 2002)

Method B5 Acute Toxicity (Eye Irritation) of Commission Regulation (EC) No. 440/2008

Result. A single application of the test item to the non-irrigated eye of three rabbits produced scattered or diffuse corneal opacity, iridial inflammation and moderate conjunctival irritation. One treated eye appeared normal at the 72‑Hour observation and two treated eyes appeared normal at the 7‑Day observation.

Conclusion. The test item produced a maximum group mean score of 13.7 and was classified as a mild irritant (Class4on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system.

The test item did not meet the criteria for classification as irritant or corrosive according to EU labelling regulations Commission Directive 2001/59/EC or thecriteria for classification according to Regulation (EC) No 1272/2008, relating to the Classification, Packaging and Labelling of Dangerous Substances.