N-(2-ethylhexyl)-1-[[3-methyl-4-[(3-methylphenyl)azo]phenyl]azo]naphthalen-2-amine

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Nov 1997 - Mar 1998

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

1987

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

N-(2-ethylhexyl)-1-[[3-methyl-4-[(3-methylphenyl)azo]phenyl]azo]naphthalen-2-amine

EC Number:

260-125-3

EC Name:

N-(2-ethylhexyl)-1-[[3-methyl-4-[(3-methylphenyl)azo]phenyl]azo]naphthalen-2-amine

Cas Number:

56358-10-2

Molecular formula:

C32H37N5

IUPAC Name:

N-(2-ethylhexyl)-1-[[3-methyl-4-[(3-methylphenyl)azo]phenyl]azo]naphthalen-2-amine

Test material form:

solid

Details on test material:

Batch number ZD 00883/044. Manufactured Oct 1996
91.3% purity. Impurity is residual solvent Shellsol AB

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Dr K. Thomae GMBH, BIBERACH
- Age at study initiation: Young adult
- Weight at study initiation: 3.77kg - 3.96kg
Animal 1: 3.77kg (Male)
Animal 2: 3.96kg (Male)
Animal 3: 3.92kg (Male)
Animal 4: 3.78kg (Male)
- Housing: 1 animal per cage. Stainless steel wire mesh. Floor area 3000cm2. No bedding in the cages, sawdust in waste trays.
- Diet: Kliba-Labordiaet, Klingentalmuehle AG, Switzerland. About 130g per animal per day.
- Water: About 250mL tap water per animal per day
- Acclimation period: At least 1 week

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

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other:

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.1ml
The material was first heated to 90C then cooled to body temperature before application.

Duration of treatment / exposure:

24 hours (24 hours after application the substance was washed out with tap water)

Observation period (in vivo):

1 hour
24 hour (the 24 hour reading was taken after the substance had been washed out.
48 hour
72 hour
8 days (Animals 2 and 3 only)
15 days (Animals 2 and 3 only)
21 days (Animals 2 and 3 only)
28 days (Animals 2 and 3 only)

Number of animals or in vitro replicates:

4 animals in total
1 animal in an intitial step.
As no further findings were noted, a further 2 confirmatory animals. Some symptoms were observed in one of these animals and it was unclear if they were due to the test substance.
Therefore, to clarify the results, a 4th animal was then tested.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes
- Time after start of exposure: 24 hours

Initially 3 animals had the test substance applied. As the irritant potential of the substance was unknown, 1 animal was tested as a first step. As no severe findings were noted, another 2 rabbits were tested. To clarify the results, a further animal was tested.
Animal 1 initially application date 03-Nov-97
Animals 2 and 3 application date: 10-Nov-97
The effects were reversible in 2 animals, In one animal, opacity grade 1 over an area of less than a quarter was observed from 24 hours to 28 days. As no effects on the cornea were observed in the other 2 animals, and the grade or size of the observed corneal opacity did not change during the observation period, it was assumed that it was not related to the test substance. Therefore another animal was tested and the animal with corneal opacity was excluded from the evaluation of eye irritancy properites of the test substance.
Animal 4 application date: 12-Jan 1998.

The findings which were noted in animal 4 were reversible within 48 hours after application, thus the observation period for this animal was terminated after 72 hours.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Results 2

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Results 3

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0.5

Max. score:

2

Results 4

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0.2

Max. score:

1

Remarks on result:

no indication of irritation

Any other information on results incl. tables

The mean scores were calculated based on the readings at 24 hour, 48 hours and 72 hours. The full individual readings are tabulated below. Animal 2 was excluded from the mean calculations as it was determined that the effects seen were not due to the test substance.

Tabulation of irritant response data for each individual animal at each observation time as well as calculation of the means.

 

Time point of reading	Animal	Cornea		IRIS	CONJUCTIVA			SYMPTOMS (see key below)
		OP	AR		RED	SW	DI
1 hour	01	-	-	-	-	1	2	A
	02	-	-	-	-	1	2	A
	03	-	-	-	-	1	2	A
	04	-	-	-	-	1	1	A, B
24 hours	01	0	0	0	1	1	1	B
	02	1	1	0	2	1	1	B
	03	0	0	0	2	0	1	B
	04	0	0	0	0	1	1	B
48 hours	01	0	0	0	0	0	0	B
	02	1	1	0	2	1	0	B
	03	0	0	0	1	0	0	B
	04	0	0	0	0	0	0	B
72 hours	01	0	0	0	0	0	0	SD, B
	02	1	1	0	1	0	0	B
	03	0	0	0	1	0	0	B
	04	0	0	0	0	0	0	SD, B
8 days	02	1	1	0	0	0	0	B
	03	0	0	0	0	0	0	B
15 days	02	1	1	0	0	0	0	B
	03	0	0	0	0	0	0	B
21 days	02	1	1	0	0	0	0	B
	03	0	0	0	0	0	0	B
28 days	02	1	1	0	0	0	0	B
	03	0	0	0	0	0	0	B
Mean (24h, 48h 72h results)	01	0.0		0.0	0.3	0.3
	02	1.0		0.0	1.7	0.7		Animal excluded from evaluation
	03	0.0		0.0	1.3	0.0
	04	0.0		0.0	0.0	0.3
Overall Mean		0.3		0.0	0.8	0.3
Overall mean excluding animal 2		0.0		0.0	0.5	0.2

 

Symptoms Key

A: Index for corneal opacity, area, iris and conjunctivae redness could not be read because of staining due to the colour of the test substance. 

B: Fur of the head reddish discoloured.

SD: Study discontinued because the animal was free of symptoms.

 

 

 

 

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

Not irritating

Conclusions:

The test substance is not classified as irritating to eyes according to EU CLP criteria.

Executive summary:

The test material was assessed for eye irritation/corrosion according to the Method B.5 Acute Toxicity: Eye Irritation/Corrosion. Council Regulation (EC) No.440/2008, and OECD Test guideline 405.

The potential of the test substance to cause damage to the conjuntiva, iris or cornea was assessed in 4 white New Zealand rabbits, subjected to a singular ocular application of 0.1ml of the test susbstance. The eyes were examined at 1 hour, 24 hours, 46 hours and 72 hours. For 2 of the animals examinations were also carried out at 8, 15, 21 and 28 days. One animal was subsequently excluded from the study due to eye effects not attributable to the test substance. The avergage score (24 to 74 hours) for irritation was calculated to be 0.0 for corneal opacity, 0.0 for Iris, 0.5 for Conjunctival redness and 0.2 for Chemosis.

The individual animal mean scores for the 24, 48 and 72 hr time-point indicated that this material would not be classified under EU regulations.