2-Naphthalenesulfonic acid, 7-amino-4-hydroxy-8-[2-[2-sulfo-4-[[2-(sulfooxy)ethyl]sulfonyl]phenyl]diazenyl]-, potassium sodium salt (1:?:?), coupled with diazotized 2-[(4-amino-5-methoxy-2-methylphenyl)sulfonyl]ethyl hydrogen sulfate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experiment start date - 08 July 2003; Experiment end date - 04 August 2003; Study completion date - 03 September 2003.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

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

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): FAT 40'812/A
- Purity: Approx. 75%
- Lot/batch No.: WP 3/03
- Expiration date: 23 April 2010
- Stability: Stable under storage conditions
- Storage conditions: At room temperature (range of 20 ± 3 °C), away from direct sunlight

Specific details on test material used for the study:

Identity: FAT 40812/A
Batch: WP 8/03
Purity: approx. 75 %
Appearance: Solid, dark red-brownish powder
Expiration date: 23 April 2010
Storage: At room temperature at about 20 °C

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Elevage Scientifique des Dombes, F-01400 Chatillon sur Chalaronne / France
- Age at treatment: 10 weeks (male), 11 weeks (females)
- Bodyweight at treatment: 2227 - 2574 g
- Housing: Individually in stainless steel cages equipped with feed hoppers and drinking water bowls. Wood blocks (RCC Ltd., Füllinsdorf) and haysticks 4646 (batch no. 0301, Provimi Kliba AG) were provided for gnawing.
- Diet: Pelleted standard Provimi Kliba 3418 rabbit maintenance diet ad libitum (batch no. 28/03) provided by Provimi Kliba AG, CH-4303 Kaiseraugst.
- Water: Community tap water from Füllinsdorf, ad libitum.
- Acclimation period: Under laboratory conditions after health examination. Only animals without any visual signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17 - 23
- Humidity (%): 30 - 70
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12/12

Room temperature and humidity were monitored continuously and values outside of these ranges may have occasionally occurred, usually following room cleaning. These transient variations are considered not to have any influence on the study and, therefore, these data are not reported but are retained at RCC.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

0.1 g

Duration of treatment / exposure:

21 days

Observation period (in vivo):

up to 21 days

Number of animals or in vitro replicates:

1 male, 2 females

Details on study design:

TREATMENT:
The eyes of the animals were examined one day prior to test item administration. Animals with overt signs of ocular injury or irritation which may have interfered with the interpretation of the results were not used in the test. On the day of treatment, 0.1 g of FAT 40'812/A was placed in the conjunctival sac of the left eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about one second to prevent loss of test item. The right eye remained untreated and served as the reference control. The treated eyes were not rinsed after instillation.

OBSERVATIONS:
Viability/Mortality: Daily from acclimatization of the animals to the termination of test.
Clinical signs: Daily from acclimatization of the animals to the termination of test.
Body weights: At start of acclimatization, on the day of application and at termination of observation.

SCORING SYSTEM:
The ocular reaction was assessed according to the numerical scoring system listed in the EEC Commission Directive 92/96/EEC, July 31, 1992 at approximately 1, 24, 48 and 72 hours, as well as 7, 10, 14, 17, and 21 days after instillation. When present, corrosion and/or staining of conjunctivae, sclerae and cornea by the test item were recorded and reported.

TOOL USED TO ASSESS SCORE:
Eye examinations were made with a Varta Cliptrix diagnostic-lamp (Roth AG, CH-4153 Reinach/Switzerland).


SCORING SYSTEM: Scale for scoring Ocular Lesions.
1.Cornea
(A) Opacity-degree of density (area most dense taken for reading)
- No opacity: 0
- Scattered or diffuse area, details of iris clearly visible: 1
- Easily discernible translucent areas, details of iris slightly obscured: 2
- Opalescent areas, no details of iris visible, size of pupil barely discernible: 3
- Opaque, iris invisible: 4

(B) Area of cornea involved
- One quarter (or less) but not zero: 1
- Greater than one quarter but less than half: 2
- Greater than half, but less than three quarters: 3
- Greater than three quarters, up to whole area: 4

2.Iris
(A) Values
- Normal: 0
- Folds above normal, congestion, swelling, circumcorneal injection (any or all of these or combination of any thereof) iris still reacting
to light (sluggish reaction is positive): 1
- No reaction to light, haemorrhage, gross destruction (any or all of these): 2
Ax5 Total maximum=10

3.Conjunctivae
(A) Redness (refers to palpebral and bulbar conjunctivae excluding cornea and iris)
- Vessels normal: 0
- Vessels definitely injected above normal: 1
- More diffuse, deeper crimson red, individual vessels not easily discernible: 2
- Diffuse beefy red: 3

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

(C) Discharge'
- No discharge: 0
- Any amount different from normal (does not include small amounts observed" in inner canthus of normal animals): 1
- Discharge with moistening of the lids and hairs just adjacent to lids: 2
- Discharge with moistening of the lids and hairs, and considerable area around the eye: 3

TOOL USED TO ASSESS SCORE: Slit lamp

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Results 2

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Results 3

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0.67

Max. score:

3

Reversibility:

fully reversible within: 7 days

Remarks on result:

no indication of irritation

Results 4

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0.22

Max. score:

4

Reversibility:

fully reversible within: 48 hours

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

- No abnormal findings were observed in the cornea or iris of any animal at any of the measurement intervals. Slight reddening of the conjunctivae was observed in all animals at the 1- and 24-hour reading. Slight reddening was still noted in two animals at the 48-hour examination and persisted in one animal up to 72-hours after treatment. Moderate swelling of the conjunctivae (oedema) was noted in all animals at the 1-hour reading. Slight swelling of the conjunctivae was still evident in two animals 24 hours after treatment. Slight reddening of the sclerea was present in all animals at the 1-hour reading and persisted in one animal up to 24-hours after treatment. Slight to moderate discharge was noted in all animals at the 1-hour examination. One animal screamed immediately after application of the test item for a few seconds.
- No corrosion of the cornea was observed at any of the reading times.

Other effects:

- Slight red staining of the treated eyes produced by the test item was observed in all animals one hour after treatment and persisted in all animals up to the end of the observation period. Red remnants of the test item were observed in the eye or conjunctival sac of all animals one hour after treatment.
- No clinical signs of systemic toxicity were observed in the animals during the study and no mortality occurred. One animal screamed after the application.
- The body weights of all rabbits were considered to be within the normal range of variability.

Applicant's summary and conclusion

Interpretation of results:

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

Conclusions:

Red staining of the treated eyes produced by the test item observed in all animals persisted up to the 21 days after treatment.

Executive summary:

In a GLP-compliant eye irritation study, performed according to OECD guideline 405, test substance (0.1 g) was placed in the conjunctional sac of the left eye of 3 New Zealand White rabbits. Scoring of irritation effects was performed approximately 1, 24, 48 and 72 hours, as well as 7, 10, 14, 17, and 21 days after test item instillation. The instillation of the test substance into the eye resulted in mild, early-onset and transient ocular changes, such as reddening of the conjunctivae and sclerea, discharge and chemosis. These effects were reversible and were no longer evident 7 days after treatment. The mean 24/48/72h corneal opacity, iris scores were zero, the conjuctiva and chemosis score were recorded to be 0,67 and 0.22, respectively. Red staining of the treated eyes produced by the test item was observed in all animals at the 1-hour examination and persisted up to the observation 21 days after treatment. No abnormal findings were observed in the iris of any animal at any of the examinations, no corrosion was observed at any of the measuring intervals and no clinical signs were observed.