Disodium 4,4'-bis[[6-anilino-4-[(2-hydroxyethyl)methylamino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Internationally accepted guideline, well documented and scientifically acceptable.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1975

Report date:

1975

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: "Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics" (1959) of the US Association of Food and Drug Officials (AFDO).

GLP compliance:

no

Remarks:

Pre GLP.

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 4,4'-bis[[6-anilino-4-[(2-hydroxyethyl)methylamino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate

EC Number:

237-600-9

EC Name:

Disodium 4,4'-bis[[6-anilino-4-[(2-hydroxyethyl)methylamino]-1,3,5-triazin-2-yl]amino]stilbene-2,2'-disulphonate

Cas Number:

13863-31-5

Molecular formula:

C38H40N12Na2O8S2

IUPAC Name:

disodium 2,2'-ethene-1,2-diylbis[5-({4-anilino-6-[(2-hydroxyethyl)(methyl)amino]-1,3,5-triazin-2-yl}amino)benzenesulfonate]

Test animals / tissue source

Species:

rabbit

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Test animals: rabbit Russian.
- Weight at study initiation: 1.5 to 2 kg.
- Housing: individually housed in metal cages.
- Diet: ad libitum, standard rabbit food NAFAG, Gossau SG.
- Water: ad libitum.
- Acclimation period: for a minimum of 8 days.
- Other: only rabbith with normal ophthalmic findings were used for the test.

ENVIRONMENTAL CONDITIONS
- Temperature: 18 ± 1 °C
- Humidity: 55 ± 5 %
- Photoperiod: 14 hrs light cycle day.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1 ml.

Duration of treatment / exposure:

In 3 rabbits the treated eye was flushed with 10 ml of lukewarm water, 30 seconds after the treatment.

Observation period (in vivo):

7 days

Number of animals or in vitro replicates:

3 males and 3 females.

Details on study design:

APPLICATION
The test material was inserted in the conjunctival sac of the left eye of the rabbits and the lids were gently closed for one seconds. The right eye was not treated and served as an untreated control.

REMOVAL OF TEST SUBSTANCE
- Washing: 3 rabbits eyes wasehd with 10 ml lukewarm water.
- Time after start of exposure: 30 seconds.

TOOL USED TO ASSESS SCORE: hand-slit lamp; the irritation was appraised on day 1, 2, 3, 4 and 7 and was scored for each individual rabbit.
The primary irritation index represents the sum of the mean values for cornea, iris and conjunctivae.

SCORING SYSTEM

CORNEA:
A Opacity and degree of density (most dense area scored) .
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
A x B x 5; Maximum possible score = 80

IRIS:
A Values
Normal: 0
Folds above normal, congestion, swelling, circumcornea 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, hemorrhage, gross destruction (any or all of these): 2
A x 5; Maximum possible score = 10

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
(A + B + C) x 2; Maximum possible score = 20

The total maximum score was thus 80 for the cornea, 10 for the iris, 20 for the conjunctivae and 110 for the entire eye. The severity of overall eye irritation was rated as:
0 - 10: minimal
11 - 25: slight
26 - 56: moderate
57 - 84: marked
above 84: extreme

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

4

Reversibility:

fully reversible

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

4

Reversibility:

fully reversible

Results 3

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

4

Reversibility:

fully reversible

Results 4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

2

Reversibility:

fully reversible

Results 5

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

2

Reversibility:

fully reversible

Results 6

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

ca. 0

Max. score:

2

Reversibility:

fully reversible

Results 7

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

other: see "any other information on results incl. tables" section

Results 8

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

not measured/tested

Irritant / corrosive response data:

In the present experiments the irritation index was established as 2.8 for the cornea, 0 for the iris and 2.7 for the conjunctivae.
The substance caused therefore a minimal overall eye irritation to rabbits.

Any other information on results incl. tables

Animal	Eye	24 hrs	48 hrs	72 hrs	4 days	7 days	Mean 24, 48, 72 hrs
Cornea
1 F	rinsed	0	0	0	0	0	0
2 F	rinsed	0	0	0	0	0	0
3 F	rinsed	0	0	0	0	0	0
Iritis
1 F	rinsed	0	0	0	0	0	0
2 F	rinsed	0	0	0	0	0	0
3 F	rinsed	0	0	0	0	0	0
Conjunctival reactions
1 F	rinsed	6*	0	0	0	0	*not evaluable
2 F	rinsed	0	0	0	0	0	0
3 F	rinsed	0	0	0	0	0	0

* Effect reported only in one rabbit; results are not re-evaluable according to the current regulation and, in any case, it would be not sufficient to classify, according to the CLP criteria (which require at least in 2 of 3 tested animals).

Animal	Eye	24 hrs	48 hrs	72 hrs	4 days	7 days	Mean 24, 48, 72 hrs
Cornea
4 M	Unrinsed	15	10	5	0	0	not evaluable
5 M	Unrinsed	10	10	5	0	0	not evaluable
6 M	Unrinsed	15	10	5	0	0	not evaluable
Iritis
4 M	Unrinsed	0	0	0	0	0	0
5 M	Unrinsed	0	0	0	0	0	0
6 M	Unrinsed	0	0	0	0	0	0
Conjunctival reactions
4 M	Unrinsed	10	8	0	0	0	not evaluable
5 M	Unrinsed	15	10	5	0	0	not evaluable
6 M	Unrinsed	12	10	8	0	0	not evaluable

Applicant's summary and conclusion

Interpretation of results:

not irritating

Remarks:

Migrated information according to the CLP Regulation Criteria used for interpretation of results: EU

Conclusions:

Not irritating.

Executive summary:

Method

The eye irritation potential was assessed following the procedure described in the "Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics" (1959) of the US Association of Food and Drug Officials (AFDO).

 

Results

The irritation index was established as 2.8 for the cornea, 0 for the iris and 2.7 for the conjunctivae; minimum irritating.

Discussion and conclusion

The scoring system used for the corea opacity and degree of density, for the iris and conjunctivae redness and chemosis is the same of that reported into the OECD, but the calculation of results for the evaluation of the eye irritation potential followed in the test is different from that reported into the OECD guideline. The raw tables are not included into the study report, thus the scores related to the reactions of the unrinsed eye can not be re-evaluated according to the CLP Regulation (EC 1272/2008).

Despite also the timing for the removal of the substance is very different between AFDO and OECD (30 seconds in this experiment and 1 hours in the OECD), the results clearly indicate that the mean values from gradings at 24, 48 and 72 hours for rinsed eyes were lower than 1 for corneal opacity, lower than 1 for irtitis, lower than 2 for both conjunctival redness and oedema, in all animals.

Therefore, the substance can be classified as not irritating.