2-[-N-(2-cyanoethyl)-4-[(p-nitrophenyl)azo]anilino]ethyl benzoate

Endpoint summary

Description of key information

Not skin or eye irritating

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 August 1998 to 21 August 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Species: New Zealand albino rabbit
Origin: Charles River Deutschland GmbH
Number of animals: 3
Body weight at start of the study: 2.8-3.0 kg
Animal maintenance: in fully air-conditioned rooms in separate cages arranged in a battery
Room temperature: 20±3°C
Relative humidity: 50 ± 20 %
Lighting time: 12 hours daily
Acclimatisation: 1 week under study conditions
Food: ssniff® K-H (V2333), ad libitum and hay (approx. 15 g daily)
Water: water from automatic water dispensers, ad libitum
Animal identification: numbered ear tags

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

other: sesame oil

Controls:

no

Amount / concentration applied:

Dose levels: 500 mg per patch
Each animal was treated with 0.5 g test substance pasted with 1.0 ml sesame oil (Oleum sesami DAB 10).

Duration of treatment / exposure:

Exposure 4 hours

Observation period:

72 hours after removal of patch

Number of animals:

3

Details on study design:

About 24 hours before the start of the study the hair in the dorsal region of the body of 3 rabbits was removed with an electric clipper over an area of about 25 cm2. Only animals with intact skin were used.

Each animal was treated with 0.5 g test substance pasted with 1.0 ml sesame oil (Oleum sesami DAB 10). The substance was administered over the whole surface of a 2.5 x 2.5 cm cellulose patch on a piece of surgical plaster (specially produced by Beiersdorf Aktiengesellschaft). The plaster was fixed to the prepared skin area and then covered with a semi-occlusive bandage.

The exposure period was 4 hours. After the exposure period all remnants of the test substance were carefully removed from the skin with warm tap water.

Examinations of the skin took place after 30 - 60 minutes as well as 24, 48 and 72 hours after removal of the patches.

Erythema, eschar formation and oedema were evaluated numerically according to the score of DRAIZE. All other changes of the skin were recorded.

Irritation parameter:

erythema score

Basis:

mean

Remarks:

all animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible within: 3 days

Remarks on result:

no indication of irritation

Remarks:

skin discoloured orange

Irritation parameter:

edema score

Basis:

mean

Remarks:

all animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible within: 3 days

Remarks on result:

no indication of irritation

Remarks:

skin discoloured orange

Irritant / corrosive response data:

The reversibility results detailed correspond to minor skin discolouration. No other effects were noted. Three days after removal of the patch the irritations were reversible

Other effects:

From one hour up to one day after removal of the patch the skin surface of the animals was discoloured orange.

Generalised Results by Animal – Animals Against Parameters – Multiple Times

Activity : Skin Irritation Results

Study: 98.0574 – Skin Irritation Test in Rabbits

Group sex	Animal Number	Day Number	Time Slot	Bodyweight g	Treated Flank	Duration Treatment	Erythema	Edema
1f	313	1		2840	Left	4 hours
			30-60 min. after decontamination				0	0
		2	24 hours after decontamination				0	0
		3	48 hours after decontamination				0	0
		4	72 hours after decontamination				0	0
	314	1		2800	Left	4 hours
			30-60 min. after decontamination				0	0
		2	24 hours after decontamination				0	0
		3	48 hours after decontamination				0	0
		4	72 hours after decontamination				0	0
	315	1		2930	Left	4 hours
			30-60 min. after decontamination				0	0
		2	24 hours after decontamination				0	0
		3	48 hours after decontamination				0	0
		4	72 hours after decontamination				0	0

Nominal Dose: Group 1 – 0.5g (moistened)

 

Clinical Observations – Clinical Signs by Animal

Study: 98.0574 – Skin Irritation Test in Rabbits

Day numbers relative to Start Date
Group Sex	Animal Number	Clinical Sign	1	2	3	4
			30-60 min a. decom.	24 hours a. decom.	48 hours a. decom.	72 hours a. decom
1f	313	No Abnormalities Detected	-	-	X	X
		Skin Discoloured- Small Area	O	O	-	-
	314	No Abnormalities Detected	-	-	X	X
		Skin Discoloured – Small Area	O	O	-	-
	315	No Abnormalities Detected	-	-	X	X
		Skin Discoloured – Small Area	O	O	-	-

X = Present   O = orange

Nominal Dose: Group 1 – 0.5g (moistened)

Interpretation of results:

GHS criteria not met

Conclusions:

Based on the results of this study the test substance is not irritating to skin.

Executive summary:

The present study was conducted in compliance with EEC-Guideline B.4."Acute Toxicity Skin Irritation" of the Directive 92/69/EEC: Commission Directive of July 31,1992 adapting to technical progress for the seventeenth time Council Directive 67/548/EEC on the approximation of the laws, regulations and administrative provisions relating to the classification, packaging and labeling of dangerous substances and OECD Guidelines for Testing of Chemicals, 404 „Acute Dermal Irritation / Corrosion", updated guideline, adopted: July 17,1992

From one hour up to one day after removal of the patch the skin surface of the animals was discolored orange.

Three days after removal of the patch the irritations were reversible

Based on the individual scores for erythema and oedema after 24, 48 and 72 hours, the following mean values were calculated as zero. No effects other than skin staining were noted.

The substance is not considered to be an irritant to the skin.

Reference 2

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1978

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

GLP compliance:

yes

Species:

rabbit

Strain:

not specified

Type of coverage:

occlusive

Preparation of test site:

shaved

Vehicle:

water

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Concentration (if solution): 50% (w/w)

Duration of treatment / exposure:

One exposure at 0 h

Observation period:

min. 8 d, parameters observed at 24 and 72 h

Number of animals:

6

Irritation parameter:

edema score

Basis:

mean

Time point:

other: 24 and 72 h

Score:

0

Max. score:

4

Reversibility:

other: not irritating

Irritation parameter:

erythema score

Basis:

mean

Time point:

24 h

Remarks on result:

not determinable

Remarks:

The substance stained the skin so that erythema could not be determined properly

Irritation parameter:

erythema score

Basis:

animal: 2, 3

Time point:

72 h

Score:

0

Max. score:

4

Reversibility:

other: no irritation observed

Remarks on result:

not determinable because of methodological limitations

Remarks:

average score based on two animals only since staining of skin was to strong to derive an appropriate erythema score

Irritant / corrosive response data:

Scarified skin showed a stronger edema score (1 out of 4, average of 24 and 72 h) but this method is not useful for classification according to CLP criteria. All irritation symptoms on scarified skin were fully reversible within 8 d.

Interpretation of results:

GHS criteria not met

Conclusions:

Based on a skin irritation test with rabbits, the test substance is not irritating to skin. The average edema score was determined to be 0 (out of 4) on intact skin after 24 and 72 h.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 August 1998 to 11 September 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Species: New Zealand albino rabbit
Origin: Charles River Deutschland GmbH
Number of animals: 3
Body weight at start of the study: 3.1 -3.8 kg
Animal maintenance: in fully air-conditioned rooms in separate cages arranged in a battery
Room temperature: 20 ± 3eC
Relative humidity: 50 ± 20 %
Lighting time: 12 hours daily
Acclimatisation: 1 week under study conditions
Food: ssniff K-H (V2333), ad libitum and hay (approx. 15 g daily)
Water: water from automatic water dispensers, ad libitum
Animal identification: numbered ear tags

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

Dose levels: 100 mg per eye, applied neat.

Duration of treatment / exposure:

24 hours duration

Observation period (in vivo):

Initially 72 hours extended to 7 days

Number of animals or in vitro replicates:

3

Details on study design:

About 24 hours before the start of the study the test eyes of all animals were examined under UV light for corneal lesions after instillation of one drop of a 0.01 % fluorescein-sodium solution. Only animals without ocular abnormalities were used for the study.

100 mg T-9601 was administered once to the conjunctival sac of the left eye of three rabbits. The untreated eyes served in each case as a control.

24 hours after administration and at all other designated examination times at which the treated eyes still showed discharge or at which a corneal examination with fluorescein sodium solution took place, the treated eyes were washed out thoroughly with isotonic saline at approx. 37 °C.

The eyes were examined 1, 24, 48 and 72 hours after administration of the test substance. At 24 and 72 hours as well as after 7, 14 days, the eyes were also examined for corneal lesions under UV light after instillation of one drop of a 0.01 % fluorescein-sodium solution. Lesions in cornea, iris or conjunctivae were graded numerically. All other changes or toxic effects were recorded. Since effects were still present in the eyes after 72 hours, further examinations were carried out after 7 days.

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

all animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

mean

Remarks:

all animals

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Reversibility:

fully reversible within: 7 days

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.67

Max. score:

3

Reversibility:

fully reversible within: 48 hours

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

3

Reversibility:

fully reversible within: 48 hours

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

animal #2 and animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

7 days after application the irritations observed were all reversible.

Other effects:

From one hour up to 3 days after application the conjunctiva of the animal showed definitely injected blood vessels up to deeper crimson red colours and slight swellings up to swelling with partial eversion of lids. The irritations were attended by clear coloured eye discharge.

Generalised Results By Animal – Animals Against Parameters – Multiple Times

Activity : - Eye Irritation Results

Eye Irritation Test in Rabbits

Group Sex	Animal Number	Day Number	Time Slot	Bodyweight g	Treated Eye	Control Eye normal	Duration Treatment	Initial Pain Reaction	Eye Irrigated	Fl.ceine staining	Opacity Grade	Opacity Area	Iris	Redness	Chemosis	Discharge
1f	57	1	Treatment	3480	Left	Yes	24 hour	None
			1 hour after treatment			Yes			No		0	0	0	1	2	2
		2	24 hours after treatment			Yes			Yes	0	0	0	0	2	2	1
		3	48 hours after treatment			Yes			No		0	0	0	2	1	0
		4	72 hours after treatment			Yes			Yes	0	0	0	0	1	0	0
		8	7 days after treatment			Yes			Yes	0	0	0	0	0	0	0
	607	1	Treatment	3180	Left	Yes	24 hour	None
			1 hour after treatment			Yes			No		0	0	0	2	2	2
		2	24 hours after treatment			Yes			Yes	0	0	0	0	2	0	0
		3	48 hours after treatment			Yes			No		0	0	0	0	0	0
		4	72 hours after treatment			Yes			Yes	0	0	0	0	0	0	0
	681	1	Treatment	3770	Left	Yes	24 hour	None
			1 hour after treatment			Yes			No		0	0	0	2	1	1
		2	24 hours after treatment			Yes			Yes	0	0	0	0	1	0	0
		3	48 hours after treatment			Yes			No		0	0	0	0	0	0
		4	72 hours after treatment			Yes			Yes	0	0	0	0	0	0	0

Nominal Dose: Group 1 – 0.1g

 

Clinical Observation – Clinical Signs by Animal

Eye Irritation Test in Rabbits

Day numbers relative to Start Date
Group Sex	Animal Number	Clinical Signs	1	2	3	4	8
			1 hour p. appl.	24 hours p. appl.	48 hours p. appl.	72 hours p. appl.	7 days p. appl.
1f	57	No Abnormalities Detected	X	-	X	X	X
		Eye discharge serous	M	S	-	-	-
	607	No Abnormalities Detected	-	X	X	X	-
		Eye discharge serious	M	-	-	-	-
	681	No Abnormalities Detected	-	X	X	X	-
		Eye discharge serious	S	-	-	-	-

X = Present  S = slight  M = Moderate

Nominal Dose: Group 1 – 0.1g

Interpretation of results:

GHS criteria not met

Conclusions:

Testing for primary eye irritation in the rabbit showed, that the substance is not irritating to eye.

Executive summary:

Testing for primary irritation in the rabbit eye provides information on ocular lesions after a single administration of the test substance to the conjunctival sac of the eye and serves as a basis for classification and labelling.

 

The present study was conducted in compliance with EEC-Guideline B.5."Acute Toxicity Eye Irritation" of the Directive 92/69/EEC: Commission Directive of July 31,1992 adapting to technical progress for the seventeenth time Council Directive 67/548/EEC on the approximation of the laws, regulations and administrative provisions relating to the classification, packag­ing and labelling of dangerous substances and OECD Guidelines for Testing of Chemicals, 405 „Acute Eye Irritation / Corrosion", updated guideline, adopted: February 24, 1987.This study was conducted in compliance with the Principles of Good Laboratory Practice (GLP).

 

From one hour up to 3 days after application the conjunctiva of the animal showed definitely injected blood vessels up to deeper crimson red colours and slight swellings up to swelling with partial eversion of lids. The irritations were attended by clear coloured eye discharge.

7 days after application the irritations were reversible.

 

Based on the individual scores after 24, 48 and 72 hours, the following mean values were calculated:

 

Opacity of cornea	All animals	0.00	Iris	All animals	0.00
	Animal 57	0.00		Animal 57	0.00
	Animal 607	0.00		Animal 607	0.00
	Animal 681	0.00		Animal 681	0.00
Redness of conjunctiva	All animals	0.89	Chemosis of conjunctiva	All animals	0.33
	Animal 57	1.67		Animal 67	1.00
	Animal 607	0.67		Animal 607	0.00
	Animal 681	0.33		Animal 681	0.00

 

Testing for primary eye irritation in the rabbit showed, that the substance is not irritating to eye.

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1978

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

GLP compliance:

yes

Species:

rabbit

Strain:

not specified

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Duration of treatment / exposure:

one treatment at 0 h, observation at 24, 48 and 72 h

Observation period (in vivo):

min. 8 d

Number of animals or in vitro replicates:

6

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

9.4

Max. score:

80

Reversibility:

fully reversible within: 8 d

Remarks on result:

probability of weak irritation

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

10

Reversibility:

other: not irritating

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

5.3

Max. score:

20

Reversibility:

fully reversible within: 8 d

Remarks on result:

probability of weak irritation

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

24/48/72 h

Score:

14.78

Max. score:

110

Reversibility:

fully reversible within: 8 d

Remarks on result:

probability of weak irritation

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

6 rabbits

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 8 d

Remarks on result:

no indication of irritation

Remarks:

EU GHS

Irritation parameter:

iris score

Basis:

mean

Remarks:

6 rabbits

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Remarks:

EU GHS

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

6 rabbits

Time point:

24/48/72 h

Score:

1.11

Max. score:

3

Reversibility:

fully reversible within: 8 d

Remarks on result:

no indication of irritation

Remarks:

EU GHS

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

6 rabbits

Time point:

24/48/72 h

Score:

0.56

Max. score:

4

Reversibility:

fully reversible within: 8 d

Remarks on result:

no indication of irritation

Remarks:

EU GHS

Interpretation of results:

GHS criteria not met

Conclusions:

Due to the low overall eye irritation score of 14.7 (mean of 24, 48 and 72 h) in rabbits, the classification criteria according to CLP were not met. The test substance showed only very weak signs of irritation (cornea opacity, conjunctivae) which were fully reversible within 8 d.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation / Corrosion.

A single study was evaluated on this endpoint. In these the substance was tested for primary dermal irritation and corrosiveness. No inflammatory signs (eschar or oedema formation) became apparent within the observation period and signs of toxicity were not observed. Orange coloured staining was observed; however this was fully reversible within 3 days. This evidence indicates no hazard potential to the skin and the test substance may therefore be regarded as 'not irritating to the skin’. It is not a dermal corrosive. No risk phrase or classification is required.

 

Eye irritation.

 A single study was evaluated on this endpoint. In these the substance was tested for acute irritation and was found not to be an irritant to the rabbit eye. Effects were noted in all animals assessed; however these were fully reversible within 24 hours. No classification is applicable.

 

Respiratory irritation

Respiratory irritation was not assessed; however no effects on the animals were noted in any associated studies.

 

Justification for classification or non-classification

The above studies have all been ranked reliability 1 or 2 according to the Klimisch et al system. This ranking was deemed appropriate because the studies were conducted to GLP an in compliance with agreed protocols. Sufficient dose ranges and numbers are detailed; hence it is appropriate for use based on reliability and animal welfare grounds.

The above results triggered no classification under the Dangerous Substance Directive (67/548/EEC) and the CLP Regulation (EC No 1272/2008). No classification for irritation effects is therefore required.