[2-[[2-cyano-3-[4-(diethylamino)phenyl]-1-oxoallyl]oxy]ethyl][3-[[2-cyano-3-[4-(diethylamino)phenyl]-1-oxoallyl]oxy]propyl]dimethylammonium chloride

Endpoint summary

Description of key information

In two skin irritation studies, conducted each with a test item containing 50% dye stuff, no skin irritation was observed and thus it is not expected that the registered dye has any skin irritation properties. The result of the key eye irritation study, similarly conducted with a test item containing 50 %  dye stuff, leads to the conclusion that the registered dye has no eye irritation properties. 

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

GLP compliance:

yes

Species:

rabbit

Strain:

Vienna White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Gaukler, Offenbach/Main
- Weight at study initiation: mean males: 2.54 kg, female 2.57 kg
- Housing: single housing in stainless steel cages with wire mesh walk floors, floor area 40 cm x 51 cm
- Diet: about 130 g/day, Kliba 341, 4 mm; Klingenthalmuehle AG, Kaiseraugst, Switzerland
- Water: about 250 mL tap water/day
- Acclimation period: at least 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated skin sites of the same animal

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 mL
- Concentration: undiluted

Duration of treatment / exposure:

4 hours

Observation period:

72 hours

Number of animals:

3 rabbits (2 males, 1 female)

Details on study design:

TEST SITE
- Area of exposure: 2.5 cm x 2.5 cm on upper third of the back or flanks
- Type of wrap if used: test patches were secured in position with a porous dressing (four layers of absorbent gauze + porous bandage)

REMOVAL OF TEST SUBSTANCE
- Washing: at the end of the exposure period with Lutrol and Lutrol/water (1:1)
- Time after start of exposure: 4 hours

SCORING SYSTEM: according to OECD guideline

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

other: 24, 48, 72h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 48 hours

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

edema score

Basis:

animal #1

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

edema score

Basis:

animal #2

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

edema score

Basis:

animal #3

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritant / corrosive response data:

Treatment with the test substance resulted in erythema (score 1) in 1/3 rabbits at the 24-hour reading, which fully reversed within 48 hours. No erythema was observed in the 2 remaining rabbits. There were no edema in 3/3 rabbits.
For details, see table below.

Irritation scores:

Timepoint	Animal No.	Intact skin		Symptoms
		Erythema	Edema
4 h	1	0*	0	Irritation index could not be read because of staining due to the color of the test substance
	2	0	0
	3	0*	0	Irritation index could not be read because of staining due to the color of the test substance
24 h	1	1	0
	2	0	0
	3	0	0
48 h	1	0	0
	2	0	0
	3	0	0
72 h	1	0	0
	2	0	0
	3	0	0
				Timepoints
Mean	1	0.3	0.0	24, 48, 72h
Mean	2	0.0	0.0	24, 48, 72h
Mean	3	0.0	0.0	24, 48, 72h
Mean	1 - 3	0.1	0.0	24, 48, 72h

* if in 1 – 2 animals assessments referring to erythema cannot be made at a particular reading, the mean rounded to the nearest whole number of the values that can be read is inserted instead.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions (non-GLP).

Qualifier:

according to guideline

Guideline:

other: FED. REG. 38, NO. 187, PARA. 1500.42

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

GLP compliance:

no

Remarks:

pre-GLP

Species:

rabbit

Strain:

Vienna White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Gaukler, Offenbach/Main
- Weight at study initiation: mean males: 2.91 kg, female 2.85 kg
- Housing: single housing, cage area 43 cm x 51 cm
- Diet: approx. 130 g/day, Ovator Solikanin 4 mm; Muskator-Werke, Düsseldorf
- Water: approx. 250 mL/day; monday - friday VE water; saturday & sunday tap water
- Acclimation period: approx. 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 26
- Humidity (%): approx. 30 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated eye of same animal

Amount / concentration applied:

TEST MATERIAL
- Amount applied (volume): 0.1 mL
- Concentration: undiluted

Duration of treatment / exposure:

not specified (single administration, no removal of test material by washing)

Observation period (in vivo):

8 days

Number of animals or in vitro replicates:

6 rabbit (3 males, 3 females)

Details on study design:

SCORING SYSTEM:
according to Draize (APPRAISAL OF THE SAFETY OF CHEMICALS IN FOODS, DRUGS AND COSMETICS. IN: DERMAL TOXICITY PP. 46 - 59, 1959)

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

other: 24, 48, 72h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

iris score

Basis:

mean

Time point:

other: 24, 48, 72h

Score:

0.06

Max. score:

2

Reversibility:

fully reversible within: 48 hours

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

other: 24, 48, 72h

Score:

1.1

Max. score:

3

Reversibility:

fully reversible within: 8 days

Irritation parameter:

chemosis score

Basis:

mean

Time point:

other: 24, 48, 72h

Score:

0.17

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Irritant / corrosive response data:

Conjunctival redness was the main finding after treatment with the test substance seen in all rabbits with the mean score (24, 48, 72 hours) ranging from 0.6 to 1.6. Chemosis was noted in 3/6 rabbits with a mean score of each 0.3. Effects on the iris was noted in one rabbit only (score 0.3). The findings were fully reversible within the observation period with the exeption of corneal opacity (score 1) seen in 1/6 rabbits on day 8.
For details, see table below.

Irritation scores:

Timepoint	Animal No.	Cornea		Iris	Conjunctivae
		Opacity	Area		Redness	Chemosis	Discharge
24 h	1	0	0	0	2	1	0
	2	0	0	1	1	1	1
	3	0	0	0	2	0	1
	4	0	0	0	1	0	0
	5	0	0	0	2	0	0
	6	0	0	0	1	0	0
48 h	1	0	0	0	0	0	1
	2	0	0	0	1	0	0
	3	0	0	0	1	0	1
	4	0	0	0	1	0	0
	5	0	0	0	2	1	1
	6	0	0	0	2	0	0
72 h	1	0	0	0	0	0	0
	2	0	0	0	1	0	0
	3	0	0	0	1	0	1
	4	0	0	0	1	0	1
	5	0	0	0	1	0	1
	6	0	0	0	0	0	0
8 d	1	0	0	0	0	0	0
	2	0	0	0	0	0	0
	3	0	0	0	0	0	0
	4	0	0	0	0	0	0
	5	0	0	0	0	0	0
	6	1	3	0	0	0	0
Mean 24h, 48h, 72h		0	0	0.06	1.1	0.17	0.44

 

Mean (24h,48h,72h)	Cornea Opacity	Iris	Conjunctivae Redness	Conjunctivae Chemosis
Animal 1	0	0	0.6	0.3
Animal 2	0	0.3	1	0.3
Animal 3	0	0	1.3	0
Animal 4	0	0	1	0
Animal 5	0	0	1.6	0.3
Animal 6	0	0	1	0

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

In the key skin irritation study (BASF AG, 1988, according to OECD 404) three rabbits (2 males, 1 female) were treated once with 0.5 mL of a dye preparation via semiocclusive dermal exposure for 4 h. After exposure animals were observed for three days. Skin reactions were observed and scored according to the OECD scoring system. Thereby animals showed an average (24h, 48h, 72h) erythema scores of 0.33, 0.0 and 0.0 (animals 1, 2 and 3), respectively, which were fully reversed within the observation period. The edema score was 0 for all three animals. In conclusion, no relevant skin irritating effects were observed and thus the test material was assessed as not irritating to the skin.

In a supporting skin irritation study (BASF AG, 1982) four rabbits (3 males, 1 female) received 0.5 mL of a dye preparation via semiocclusive dermal exposure for 4 hours followed by an observation period of 8 days. Skin reactions were observed and scored according to Draize scoring system. Erythema score could not be evaluated due to coloration by the test material. No edema was observed 24h, 48h and 72h after treatment. Under the conditions of this supporting study the test material was not irritating to the rabbit skin.

 

Since in the underlying key skin irritation study the dye preparation was shown to be not irritatingn to the skin, even in the presence of a substantial amount of acetic acid, which is classified as skin corrosive 1B (H314) according to Regulation (EC) No 1272/2008 Annex VI Table 3.1, the registered dye, included proportionately with 50%, is not expected to possess skin irritation properties. This assumption is further substantiated by the results obtained in a supporting skin irritation study with another dye preparation containing skin corrosion-classified acetic acid to a lesser extent, and was no skin irritant. In addition, since the dye represents the highest amount in the mixture (50%), it can be concluded that the registered dye does not trigger skin irritation.

Eye irritation

In the key eye irritation study (BASF AG, 1981, according to FED. REG. 38) three male and three female Vienna White rabbits were treated once with 0.1 mL of a dye preparation by single exposure without washing of the eyes. The second eye of each animal was used as control. Eye reactions were observed and scored according to Draize 24h, 48h, 72 h and 8 days after the start of treatment. There were no cornea opacity observed after 24h, 48h and 72h. The mean (24 h, 48 h, 72 h) maxiumum values revealed 0.3 for iris score (one animal), 1.6 for conjunctivae redness (one animal) and 0.3 for conjuntivae chemosis (three animals). The effects were completely reversible after 8 days, except in one animal where mild cornea opacity (score 1) was noted after 8 days. However, as this animals showed no cornea opacity 24 h, 48 h or 72 h after treatment the effect after 8 days was considered negligible. In conclusion, the test item was not irritating to the eyes under the conditions of the study.

In an eye irritation study (BASF AG, 1988, according to OECD 405) one male rabbit was administered with 0.1 mL of a dye preparation applied to one eye of the rabbit while the other eye, remaining untreated, served as control. The test item was not removed by washing. The eye was examined 1, 24, 48, 72 h and on day 8 after treatment. The animal showed a cornea score of 2.5, an iris score of 0, a conjunctivae score of 2 and a chemosis score of 2. 8 days after reading, the irritation index for cornea and iris could not be read due to the color of the test substance and because of severe corneal opacity. The conjunctivae score and the chemosis score were not reversible within 8 days. In conclusion, the test item caused severe eye irritation under the conditions of the study.

Oppositional results were observed in the two in vivo eye irritation studies. The key eye irritation study (BASF AG, 1981) revealed no eye irritation properties when a dye preparation containing 50% dye stuff was used. The equal concentration of dye stuff was used in the second eye irritation study (BASF AG, 1988), however, this dye preparation caused severe eye irritation. This effect was considered to be caused by the high concentration of acetic acid (causes severe eye damage) and butyldiglycol (causes serious eye irritation) in the dye preparation since these substances are classified as skin corrosive cat. 1A (H 314) and eye irritating cat. 2 (H319) according to Regulation (EC) No 1272/2008 (CLP), respectively. Thus, the registered dye has to be regarded as not irritating to the eye. The eye irritation study (BASF AG, 1988) was classified as 'disregarded' for the assessment of the registered substance.

Justification for selection of skin irritation / corrosion endpoint:
most reliable study

Justification for selection of eye irritation endpoint:
most reliable study

Justification for classification or non-classification

Based on the results of the skin and eye irritation studies the test item does not need to be subjected to classification and labelling according to Directive 67/548/EEC (DSD) and Regulation (EC) No 1272/2008 (CLP/GHS).