disodium 7-(4,6-difluoropyrimidin-2-ylamino)-4-hydroxy-3-(4-methoxy-2-sulfonatophenylazo)naphthalene-2-sulfonate; reaction mass of: disodium 7-(2,4-difluoropyrimidin-6-ylamino)-4-hydroxy-3-(4-methoxy-2-sulfonatophenylazo)naphthalene-2-sulfonate

Endpoint summary

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Delivery of the animals October 23, 1996 - Euthanasia of the animals October 31, 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

adopted July 17, 1992 (reported Paris April 29, 1993)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Version / remarks:

July 31, 1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Remarks:

Chbb: NZW (SPF)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Dr. Karl Thomae GmbH; Birkendorferstrasse 65; D - 88397 Biberach / Riss
- Age at treatment : 15 weeks
- Weight at study initiation:
at start of acclimatization : male 2.5 kg, females 2.5-2.6 kg
at start of treatment : male 2.7 kg, females 2.3-2.6 kg
- Housing: Individually in stainless steel cages with an automatic cleaning system, equipped with feed hoppers, drinking water bowls and wood for gnawing.
- Diet (e.g. ad libitum): Pelleted standard Kliba 341 rabbit maintenance diet (batch 70/96) ad libitum from Kliba Muhlen AG, CH-4303 Kaiseraugst. Results of analysis for contaminants are included in this report.
- Water (e.g. ad libitum): Community tap water from Itingen, ad libitum. Results of bacteriological, chemical and contaminant analyses are included in this report.
- Acclimation period: Four days under test conditions after health examination. Only healthy animals were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3°C
- Humidity (%): between 40-70% (values above 70% during cleaning process possible)
- Air changes (per hr): Air-conditioned with 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light):12 hours artificial fluorescent light/12 hours dark, music during the light period

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

water

Remarks:

bi-distilled water

Controls:

not specified

Amount / concentration applied:

TEST MATERIAL
Amount(s) applied (volume or weight with unit): 0.5g test item/animal (left side only)

VEHICLE
The test article was moistened with bi-distilled water before application

Duration of treatment / exposure:

4 hours

Observation period:

1, 24, 48 and 72 hours after removal of the dressing

Number of animals:

3 young adults: 1 male (#1) and 2 females (#2, #3)

Details on study design:

TEST SITE
- Area of exposure: 0.5 g of the test article was applied to approx. 6 cm2 of the intact skin of the clipped area
- % coverage: It was covered with a 2.5 cm x 2.5 cm patch of surgical gauze
- Type of wrap if used: The gauze was covered with a semi-occlusive dressing. The dressing was wrapped around the abdomen and anchored with an elastic bandage.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): the dressing was removed and the skin was flushed with lukewarm tap water to clean the application site so that any reactions (erythema) were clearly visible at that time
- Time after start of exposure: 4 hours

OBSERVATION TIME POINTS
(indicate if minutes, hours or days): h, 24h, 48h and 72 h after removal of the dressing

SCORING SYSTEM:
- Method of calculation: To classify the irritating potential of the test article, the mean values of erythema I eschar as well as of edema formation were calculated for each individual, using the scores between 24 and 72 hours (EEC Commission Directive 93/21/EEC, April 27, 1993).

Irritation parameter:

erythema score

Basis:

other: Animal : #1 (M), #2 (F) & #3 (F)

Time point:

other: 1h, 24h, 48h & 72h

Score:

0

Max. score:

0

Reversibility:

other: not relevant in absence of effects

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

other: Animal #1 (M), #2 (F) & #3 (F)

Time point:

other: 1h , 24h, 48h & 72h

Score:

0

Max. score:

0

Reversibility:

other: not relevant in absence of effects

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

IRRITATION
SCARLET RN 1165 showed a primary irritation score of 0.00, when applied to healthy intact rabbit skin.
Local signs (mean values from 24 to 72 hours) consisted of grade 0.00 erythema and grade 0.00 edema.

CORROSION
No irreversible alterations of the treated skin were observed nor were corrosive
effects evident on the skin.

Other effects:

VIABILITY/MORTALITY AND CLINICAL SIGNS
No clinical signs of systemic toxicity were observed in the animals during the test and observation period, and no mortality occurred.

COLORATION
Irreversible pale red staining by the test article of the treated skin was observed.

BODY WEIGHTS
The body weight gain of all rabbits was within the normal range of variability.Please refer to results in section "any other information on results incl. tables" hereafter.

Evolution of body weights in grams:

Animal#	Sex	First Day of Acclimatization	Day of Treatment	Last Day of Observation
1	Male	2540g	2675g	2748g
2	female	2469g	2349g	2577g
3	female	2360g	2551g	2664g

Interpretation of results:

GHS criteria not met

Conclusions:

Based upon the referred classification criteria (EEC Commission Directive 93/21/EEC of April 27, 1993), with the exception of the irreversible pale red staining which persisted in all animals up to termination of the test, SCARLET RN 1165 is considered to be "not irritating" to rabbit skin.

Executive summary:

The primary skin irritation potential of the test article was investigated by topical application of 0.5 g onto 6 cm2 intact dorsal skin of each of three young adult New Zealand rabbits. The scoring of skin reactions was performed l, 24, 48 and 72 hours after removal of the dressing. The scores of each animal at the fol lowing reading times (24, 48, 72 hours) were used in calculating the respective mean values for each type of lesion.

The primary irritation score was calculated by totalling the individual cumulative scores at 24, 48 and 72 hours and then dividing by the number of figures. Under the conditions of this experiment, SCARLET RN 1165 was found to cause a primary irritation score of 0.00 (max. 8.0) when applied to healthy intact rabbit skin.

Local signs (mean values from 24 to 72 hours) consisted of grade 0.00 erythema and grade 0.00 edema.

The test article caused irreversible pale red staining of the treated skin.

No corrosive effects were noted on the treated skin of any animal at any measuring interval.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Delivery of the animals : October, 30, 1996 - Euthanasia of the animals : November 25, 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted February 24, 1987

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

July 31, 1992

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

PROTOCOL DEVIATION: the test article classification was omitted at the request of the sponsor.

Species:

rabbit

Strain:

New Zealand White

Remarks:

Chbb:NZW (SPF)

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Dr. Karl Thomae GmbH; Birkendorferstrasse 65; D - 88397 Biberach / Riss
- Age at treatment: 15 weeks
- Weight at study initiation:
body weight at start of acclimatization: male 2.5 kg, females 2.8 kg
body weight at start of treatment: male 2.5 kg, females 2.8-2.9 kg
- Housing: Individually in stainless steel cages with an automatic cleaning system, equipped with feed hoppers, drinking water bowls and wood for gnawing.
- Diet (e.g. ad libitum): Pelleted standard Kliba 341 rabbit maintenance diet (batch 70/96 and 71/96) ad libitum from Kliba Muhlen AG, CH-4303 Kaiseraugst. Results of analysis for contaminants are included in this report.
- Water (e.g. ad libitum): Community tap water from Itingen, ad libitum. Results of bacteriological, chemical and contaminant analyses are included in this report.
- Acclimation period: Four days under laboratory conditions after health examination. Only healthy animals were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3 °C
- Humidity (%): between 40-70 % (values above 70% during cleaning process possible)
- Air changes (per hr): Air-conditioned with 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light/12 hours dark, music during the light period

Vehicle:

unchanged (no vehicle)

Remarks:

the test article was applied as delivered by the sponsor

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1g/animal (into one eye of each animal)
- Concentration (if solution): undiluted

Duration of treatment / exposure:

21 days as the treated eyes were not rinsed after instillation days

Observation period (in vivo):

Scoring of irritation effects was performed approximatively 1, 24, 48 and 72 hours as well as 7, 14 and 21 days after test article application

Number of animals or in vitro replicates:

3: 1 male (#16) and 2 females (#17 and #18)

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): none
- Time after start of exposure: not relevant

SCORING SYSTEM: see page 14 and 15 of the study report

TOOL USED TO ASSESS SCORE: Eye examinations were made with a Varta Cliptrix diagnostic-lamp (A. Riegger, Basel I Switzerland).

Irritation parameter:

cornea opacity score

Basis:

animal: #16, #17 & #18

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Not relevant as scores remain nil

Irritation parameter:

iris score

Basis:

animal: #16, #17, #18

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Not relevant as scores remain nil

Irritation parameter:

conjunctivae score

Basis:

animal: #16, #17 & #18

Time point:

24 h

Score:

1

Max. score:

1

Reversibility:

fully reversible within: 48h

Irritation parameter:

conjunctivae score

Basis:

animal: #16, #17 & #18

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible within: 48h

Irritation parameter:

other: Conjunctivae chemosis

Basis:

animal: #16 & #18

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Not relevant as scores remain nil

Irritation parameter:

chemosis score

Basis:

animal: #17

Time point:

24 h

Score:

1

Max. score:

1

Reversibility:

fully reversible within: 48h

Irritation parameter:

chemosis score

Basis:

animal: #17

Time point:

48 h

Score:

0

Max. score:

0

Reversibility:

fully reversible within: 48h

Irritation parameter:

other: Staining of conjunctivae and sclera

Basis:

animal: #16, #17 & #18

Time point:

other: 24/48/72h & 7/14/21d

Reversibility:

not reversible

Remarks on result:

positive indication of irritation

Irritant / corrosive response data:

IRRITATION
SCARLET RN 1165 showed a primary irritation score of 0.44, when applied to the conjunctival sac of the rabbit eye.
Slight to moderate swelling and slight swelling of the conjunctivae was seen in all animals. These findings were reversible in all animals after 48 hours.

CORROSION
No corrosion of the cornea was observed at any of the reading times.

Other effects:

VIABILITY/MORTALITY AND CLINICAL SIGNS
No clinical signs of systemic toxicity were observed in the animals during the test and observation period, and no mortality occurred.

COLORATION
Irreversible light red staining of the conjunctivae and sclera by the test article was observed, although reversible light red staining of the iris and cornea of all animals was also seen.

BODY WEIGHTS
The body weight of the animals was within the normal range of variability (for results, please refer to the point "Any other information on results incl. tables" hereafter).

EYE IRRITATION SCORES - INDIVIDUAL VALUES

Animal	Sex	Cornea opacity	Iris	Conjunctivae		Cumul. score	Mean cumul. score
				Redn.	Chem.
1 hour
16	M	0	0	-	2*	2	1.33
17	F	0	0	-	1	1
18	F	0	0	-	1	1
24 hours
16	M	0	0	1	0	1	1.33
17	F	0	0	1	1	2
18	F	0	0	1	0	1
48 hours
16	M	0	0	0	0	0	0.00
17	F	0	0	0	0	0
18	F	0	0	0	0	0
72 hours
16	M	0	0	0	0	0	0.00
17	F	0	0	0	0	0
18	F	0	0	0	0	0
7 days
16	M	0	0	0	0	0	0.00
17	F	0	0	0	0	0
18	F	0	0	0	0	0
14 days
16	M	0	0	0	0	0	0.00
17	F	0	0	0	0	0
18	F	0	0	0	0	0
21 days
16	M	0	0	0	0	0	0.00
17	F	0	0	0	0	0
18	F	0	0	0	0	0

BODY WEIGHTS

Animal #	Sex	First Day of Acclimatization	Day of Treatment	Last Day of Observation
16	male	2456	2507	3161
17	female	2802	1761	3670
18	female	2813	2945	3769

Interpretation of results:

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

Conclusions:

Based on criteria defined in Regulation (EC) No 1272/2008, SCARLET RN 1165 is classified as Serious eye damage (category 1). Eye irritation test shows persistence of red light staining at the end of the 21 days recovery period which is according to the CLP guideline a reason for Serious eye damage Cat 1 (discoloration of the cornea by a dye substance).

Executive summary:

The primary irritation potential of the test article was investigated by instillation of 0.1 g into one eye of each of three young adult New Zealand rabbits. The treated eyes were not rinsed after application. Scoring of irritation effects was performed approximately 1, 24, 48 and 72 hours as well as 7, 14 and 21 days after test article application.

The scores of each animal at the following reading times (24, 48 and 72 hours) were used in calculating the respective mean values for each type of lesion.

The primary irritation score was calculated by totalling the individual cumulative scores at 24, 48 and 72 hours and then dividing the resulting total by the number of figures. The primary irritation score was 0.44 (max. 13).

Swelling and reddening of the conjunctivae was seen in all animals. These findings were reversible after 48 hours.

Irreversible light red staining of the conjunctivae and sclera by the test article was observed, although reversible light .red staining of the iris and cornea of all animals was also seen.

No corrosion was observed at any of the measuring intervals.

The mean values of the scores for each type of lesion, calculated for each animal separately, are the following:

Animal No.	Mean values 24-72 hours
	Cornea Opacity	Iris	Conjunctivae
			Redness	Chemosis
16, male	0.00	0.00	0.33	0.00
17, female	0.00	0.00	0.33	0.33
18, female	0.00	0.00	0.33	0.00

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irreversible damage)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on criteria defined in Regulation (EC) No 1272/2008, SCARLET RN 1165 is classified as Serious eye damage (category 1). Eye irritation test shows persistence of red light staining at the end of the 21 days recovery period which is according to the CLP guideline a reason for Serious eye damage Cat 1 (discoloration of the cornea by a dye substance)