Trisodium [3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-2-hydroxy-5-nitrobenzenesulphonato(3-)][2-hydroxy-3-[(2-hydroxy-1-naphthyl)azo]-5-nitrobenzenesulphonato(3-)]chromate(3-)

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted according to internationally accepted testing guideline.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

Trisodium [3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-2-hydroxy-5-nitrobenzenesulphonato(3-)][2-hydroxy-3-[(2-hydroxy-1-naphthyl)azo]-5-nitrobenzenesulphonato(3-)]chromate(3-)

EC Number:

274-230-7

EC Name:

Trisodium [3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-2-hydroxy-5-nitrobenzenesulphonato(3-)][2-hydroxy-3-[(2-hydroxy-1-naphthyl)azo]-5-nitrobenzenesulphonato(3-)]chromate(3-)

Cas Number:

69943-66-4

Molecular formula:

C32H18CrN8O14S2Na3

IUPAC Name:

Trisodium [3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H pyrazol-4-yl)azo]-2-hydroxy-5-nitrobenzenesulphonato(3-)][2hydroxy-3-[(2-hydroxy-1-naphthyl)azo]-5-nitrobenzenesulphonato(3-)]chromate(3-)

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Spolkem breeder of Hrace Králové.
- Weight at study initiation: above 2.5 kg.
- Housing: individually housed, in commercially metal cages (supplied by s.p. Velaz Prague), without litter and climatic control.
- Diet: standard granular commercial food for rabbits, KO-16 provided by s.p. Velaz Prague.
- Water: drinking water, ad libitum.
Cleaning and disinfection of premises menagerie were made at dates determined, according to the standard operating procedures and compliance regime measures.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 3 °C.
- Humidity: 50 ± 15 %.
- Photoperiod: 12 hrs dark / 12 hrs light, by fluorescent lamp.

Test system

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1 g

Observation period (in vivo):

72 hours (observations were made at 1, 24, 48 and 72 hours after installation of test material).

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: water.
- Time after start of exposure: 24 hours. Areas with the loss of corneal tissue are intensely coloured and the progressive rinsing of the dye in the damaged areas require more time than the place intact.

TOOL USED TO ASSESS SCORE: 2 % sodium salt, fluorescein. Reactions after 48 hours were evaluated without fluorescein. Reactions after 72 hours were evaluated with 2 % aqueous fluorescein solution.

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

IRITIS:
Normal: 0
Folds above normal, congestion, swelling, circum cornea 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
A x 5; Maximum possible score = 10

CONJUNCTIVAE:
A Redness (refers to lids 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

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

> 1

Max. score:

4

Results 2

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

>= 1

Max. score:

4

Results 3

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

> 2

Max. score:

4

Results 4

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

> 2

Max. score:

4

Irritant / corrosive response data:

Primary irritation score: 63.3
Degree of irritation: 4

The following effects were recorded at 1, 24, 48 and 72 hours: visual examination was found massive purulent discharge from the corner of eye, blepharospasm, chemosis coniunctivae, diffuse injection of the lids and bulbar conjunctiva, cornea in quadrant rich milky turbid-leucoma with a total loss of transparency, iris obscured with loss of reaction to light.

Any other information on results incl. tables

Reactions observed

N. animal cage	Assessment	Cornea		Iritis	Conjunctival
		Reaction	Area		Congestion	Swelling	Discharge
1	1	1	4	2	2	3	3
	24	2	4	1	2	3	3
	48	2	4	1	2	2	3
	72	2	4	2	2	2	3
	Mean 24, 48 and 72 hrs	2.00		1.33	2.00	2.33
2	1	1	4	1	2	3	3
	24	2	4	1	2	3	3
	48	2	4	1	2	2	3
	72	1	4	1	2	2	3
	Mean 24, 48 and 72 hrs	1.67		1.00	2.00	2.33
3	1	1	4	2	2	3	3
	24	2	4	1	2	3	3
	48	2	4	1	2	1	3
	72	2	4	1	1	2	2
	Mean 24, 48 and 72 hrs	2.00		1.00	1.67	2.00

Applicant's summary and conclusion

Interpretation of results:

other: Eye Irrit. 2 (H319), according to the CLP Regulation (EC 1272/2008)

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

Eye Irrit. 2, H319.

Executive summary:

The skin irritation potential of the substance was determined by the skin irritation test, according to the OECD guideline 404.

0.5 g of test material were applied to intact and scarified skin of rabbits and removed after 24 hours. Observations were made for 72 hours.

The substance resulted strong irritant. Primary irritation score was stated at 63.3 and the degree of irritation as 4.

The criteria for the evaluation of the eye irritation potential followed in the test are different from those reported into the OECD guideline. The scoring system used for the cornea opacity and degree of density, for the iris and conjunctivae redness and chemosis is the same of that reported into the OECD, but the timing for the removal of the substance are very different (24 hours in the current study and 1 hour in the OECD). Nevertheless, because the raw tables are included into the study report, it is possible to re-evaluate the results according to the CLP Regulation (EC 1272/2008).

The mean values from gradings at 24, 48 and 72 hours was:

- higher than 1 in all the animals tested for corneal opacity. Nevertheless in none was equal/higher than 3.

- equal than 1 in two rabbits and 1.33 in the last one for iritis

- higher than 2 in two rabbit, for conjunctival redness.

- higher than 2 in two animals, for conjunctival chemosis

Information are not available about the reversibility of the effects recorded. A relevant aspect, that may be determinant in the assessment, is that the solid substance tested was removed after 24 hours after application, instead of 1 hour as required by the OECD guideline 405 updated version 2012. Thus, it can not be excluded that the irritation potential determined by the powder form may contribut to the reactions observed.

In conclusion, it is considered appropriate a classification as eye irritant, category 2 (H319), according to the CLP Regulation (EC 1272/2008).