3,5-xylidine

Endpoint summary

Description of key information

Skin irritation: OECD 404; New Zealand white rabbits; 4h exposure semi occlusive; non irritating

Eye irritation: OECD 405; New Zealand white rabbits; 24h exposure; irritating

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:

02 JAN 1990 - 09 JAN 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Principles of method if other than guideline:

Slight modifications from the guidelines (e.g: more accurate description and scoring of corneal defects, additional examination of aqueous humour) this does not affect the validity of the study.

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Interfauna U.K. Ltd. Wyton, Huntingdon, England
- Age at study initiation: adult
- Weight at study initiation: 3.1 - 3.2 kg
- Housing: the rabbits were individually housed in stainless steel cages with flat rod bases or polyethylene cages with perforated bases, under standardized conventional conditions. Excrement trays beneath the cages contained low dust (wood) bedding (type S 8/15, Ssniff spezialdiaten GmbH, Soest). Bedding was regularly spot checked for contaminants at the instance of the Department of Laboratory Animal Services and changed at least twice weekly.
All the animals in this study were kept in one room. For capacity reasons, rabbits treated with other test substances were also housed in the same room. Adequate separation, clear cage and individual marks, as well as appropriate organization of working procedures ensured that the test animals were not mixed up.
- Diet (e.g. ad libitum): Standard diet Ssniff K 4, approx. 100 - 120g per animal/day; once per day in the morning. The nutritive composition and contaminant content of the standard diet were routinely spot checked and analysed at the instance of the department of laboratory animal services.
- Water (e.g. ad libitum): Tap water; ad libitum. The tap water was of drink water quality (Drink water ordinance of May 22, 1986, Federal Law Gazette (Bundesgesetzblatt) part I, page 760). The drink water was regularly examined at the instance of the Quality Assurance, Bayer AG. The results of the feed and water analyses provided no evidence of interference with the study objective. Drink water was supplied either in polycarbonate bottle containing approx. 750mL or from automatic watering.
- Acclimation period: Prior to the initiation of the treatment the animals were kept for at least 14 days in the quarantine station of the Dpartment of the Laboratory Animal Services and monitored for diseases. During this period pooled faeces specimens were examined for Coccidia oocysts.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3 °C
- Humidity (%): approx 50 %
- Air changes (per hr): approx 10 times per hour
- Photoperiod (hrs dark / hrs light): 12 hours, artificial illumination from 6 to 18hrs CET
The humidity and air temperature were continuously recorded using a calibrated thermohygrograph (type 252, Lambrecht Co. Gottingen). Occasional excursions from these conditions, e.g., due to cleaning of the room, had no detectable influence on the outcome of the study.

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 500 μg of the test substance was applied to a hypoallergenic Hansmed patch (Beiersdorf no. 2342 PV3).

NEGATIVE CONTROL: A further patch was moistened with water

Duration of treatment / exposure:

4 h

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: dorsolateral area of the trunk (6 x 6 cm) of each of three rabbits.
- Type of wrap if used: Each animal was loosely held in place with a semi occlusive dressing (Fixomull - stretch klebevlies, Beiersdorf no. 2293) for the duration of the exposure period. Thus, access by the animal to the patch and resultant ingestion/ inhalation of the test substance was prevented.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The exposed skin areas were carefully washed with water without altering the existing response, or the integrity of the epidermis
- Time after start of exposure: 4 hours

OBSERVATION TIME POINTS: 1h, 24h, 48h, 72h and 7 days

SCORING SYSTEM:
- Method of calculation: DRAIZE

Irritation parameter:

erythema score

Basis:

animal:

Time point:

other: 1 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

other: 1 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Basis:

mean

Time point:

7 d

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

7 d

Score:

0

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

The substance was found to be not irritating on semi occlusive application on the skin of white rabbits for 4 h.

Evaluation of results

For each animal the DRAIZE scores recorded approx. 24, 48 and 72 hours after application were added. The total of these three values was divided by three to give the irritation index. This index was separately calculated for erythema/eschar formation and for edema formation. Where three animals were used, interpretation was based on the individual indices obtained from the two most sensitive animals. Where more than three animals were used, the mean irritation index was calculated by averaging the total scores of all rabbits tested. If delayed reactions occurred, or where no irritation indices could be calculated (e.g. due to coloration by the test substance) other interpretation criteria were applied.

Interpretation of results:

GHS criteria not met

Remarks:

EU GHS criteria not met

Conclusions:

All three animals did not show any signs of irritation such as erythema or edema on the skin of white rabbits.

Executive summary:

Skin irritation test was done according to OECD Guideline 404 under GLP compliance. 3 of New Zealand white rabbits were exposed to the test item for 4 hours semi occlusive. Observation was done 1, 24, 48, 72hr and 7days after the exposure. All three animals did not show any signs of irritation such as erythema or edema on the skin.

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:

02 JAN 1990 - 09 JAN 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Principles of method if other than guideline:

Slight modifications from the guidelines (e.g: more accurate description and scoring of corneal defects, additional examination of aqueous humour) this does not affect the validity of the study.

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Interfauna U.K. Ltd. Wyton, Huntingdon, England
- Age at study initiation: adult
- Weight at study initiation: 3.4 - 3.7 kg
- Housing: the rabbits were individually housed in stainless steel cages with flat rod bases or polyethylene cages with perforated bases, under standardized conventional conditions. Excrement trays beneath the cages contained low dust (wood) bedding (type S 8/15, Ssniff spezialdiaten GmbH, Soest). Bedding was regularly spot checked for contaminants at the instance of the Department of Laboratory Animal Services and changed at least twice weekly.
All the animals in this study were kept in one room. For capacity reasons, rabbits treated with other test substances were also housed in the same room. Adequate separation, clear cage and individual marks, as well as appropriate organization of working procedures ensured that the test animals were not mixed up.
- Diet (e.g. ad libitum): Standard diet Ssniff K 4, approx. 100 - 120g per animal/day; once per day in the morning. The nutritive composition and contaminant content of the standard diet were routinely spot checked and analysed at the instance of the department of laboratory animal services.
- Water (e.g. ad libitum): Tap water; ad libitum. The tap water was of drink water quality (Drink water ordinance of May 22, 1986, Federal Law Gazette (Bundesgesetzblatt) part I, page 760). The drink water was regularly examined at the instance of the Quality Assurance, Bayer AG. The results of the feed and water analyses provided no evidence of interference with the study objective. Drink water was supplied either in polycarbonate bottle containing approx. 750mL or from automatic watering.
- Acclimation period: Prior to the initiation of the treatment the animals were kept for at least 14 days in the quarantine station of the Dpartment of the Laboratory Animal Services and monitored for diseases. During this period pooled faeces specimens were examined for Coccidia oocysts.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3 °C
- Humidity (%): approx 50 %
- Air changes (per hr): approx 10 times per hour
- Photoperiod (hrs dark / hrs light): 12 hours, artificial illumination from 6 to 18hrs CET
The humidity and air temperature were continuously recorded using a calibrated thermohygrograph (type 252, Lambrecht Co. Gottingen). Occasional excursions from these conditions, e.g., due to cleaning of the room, had no detectable influence on the outcome of the study.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): After gently pulling the lower lid away from the eyeball a volume of 100 μL test substance was instilled into the conjunctival sac of one eye of each of three rabbits. The other eye remained untreated and served as control.

Duration of treatment / exposure:

24 hours

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The treated eye was rinsed with saline.
- Time after start of exposure: 24 hours

SCORING SYSTEM & TOOL USED TO ASSESS SCORE:
Eye irritation was scored and recorded after administration given 1, 24, 48, 72h and 7 days. The signs of cornea (opacity and area affected), iris (hyperaemia, reaction to light), conjunctivae - i.e. conjunctiva of bulbs, lids, and nictitation membrane - (erythema, chemosis), and discharge were recorded as described by DRAIZE, and the aqueous humour (opacity) as described by MCDONALD and SHADDUCK. In addition, any serious lesions or toxic effects other than ocular ones were recorded. The examinations of cornea, iris and aqueous humour were facilitated using optical instruments (e.g. hand slit lamp). To define epithelial damage, one drop of a 1! fluorescein solution was applied to the cornea surface 24 hours after administration of the test substance; where positive effects were recorded this was repeated at the later observation times. The eye was then rinsed with saline to remove excess and non-absorbed fluorescein. Evaluation was performed by means of ultraviolet illumination (area) in a darkened room and diffuse white illumination (intensity), according to MCDONAD and SHADDUCK.

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

72 h

Score:

1

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

48 h

Score:

0

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24 h

Score:

1

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Irritation parameter:

iris score

Basis:

animal #3

Time point:

72 h

Score:

1

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

area

Basis:

animal #3

Time point:

24/48/72 h

Score:

4

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

72 h

Score:

0

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

48 h

Score:

0

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24 h

Score:

1

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

area

Basis:

animal #2

Time point:

72 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

area

Basis:

animal #2

Time point:

48 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

area

Basis:

animal #2

Time point:

24 h

Score:

4

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #2

Time point:

72 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #2

Time point:

48 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #2

Time point:

24 h

Score:

1

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

72 h

Score:

1

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

48 h

Score:

0

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24 h

Score:

1

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

72 h

Score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48 h

Score:

1

Irritation parameter:

iris score

Basis:

animal #1

Time point:

72 h

Score:

1

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48 h

Score:

0

Irritation parameter:

cornea opacity score

Remarks:

area

Basis:

animal #1

Time point:

24/48/72 h

Score:

4

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Irritant / corrosive response data:

The application of 0,1 ml of 3,5 -xylidine caused only slight corneal opacity and redding. The symptoms were reversible within 7 days.

animal no. (body weight)	tissue	signs	DRAIZE grade							irritation index
			1h	24h	48h	72h	7d	14d	21d
L27 (3.4 kg)	cornea	o	1	1	1	1	0	-	-	1.0
		a	4	4	4	4	0	-	-
	fluorescein	i	-	1	1	1	0	-	-
		a	-	3c	3c	3c	0	-	-
	iris		0	0	0	1	0	-	-	0.3
	conjunctivae	r	1	1	1	2	0	-	-	1.3
		s	2	1	0	1	0	-	-	0.7
	aqueous humour		0	0	0	0	0	-	-
	discharge		2	2	1	1	0	-	-
L43 (3.6 kg)	cornea	o	1	1	0	0	0	-	-	0.3
		a	4	4	0	0	0	-	-
	fluorescein	i	-	1	1	1	0	-	-
		a	-	2c	2c	2c	0	-	-
	iris		0	0	0	0	0	-	-	0.0
	conjunctivae	r	1	1	1	1	0	-	-	1.0
		s	2	1	0	0	0	-	-	0.3
	aqueous humour		0	0	0	0	0	-	-
	discharge		3	1	0	0	0	-	-
L13 (3.7 kg)	cornea	o	1	1	1	1	0	-	-	1.0
		a	4	4	4	4	0	-	-
	fluorescein	i	-	1	1	1	0	-	-
		a	-	4c	4c	4c	0	-	-
	iris		0	0	0	1	0	-	-	0.3
	conjunctivae	r	1	1	1	1	0	-	-	1.0
		s	2	1	0	1	0	-	-	0.7
	aqueous humour		0	0	0	0	0	-	-
	discharge		2	2	1	2	0	-	-

o=opacity, a=area, i=intensity

r=redness, s=swelling

-=not examined

c=confluent diffuse areas

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Remarks:

Based on EU GHS criteria

Conclusions:

Exposure of the test substance to the eye caused slight reactions of the mucous membranes and the cornea which were more pronounced immediately following administration. Signs proved to be fully reversible within 7days. As there were 2 of 3 tested animals showed a positive response of corneal opacity ≥ 1, test substance can be concluded as Category 2 according to guidanc to regulation (EC) No 1272/2008.

Executive summary:

Eye irritation test was done according to OECD Guideline 405 under GLP compliance. 3 or New Zealand white rabbits' were exposed to the test item (only one eye each, the other one was served as control) for 24 hours. Observation was done 1, 24, 48 and 72 hours and 7days after the exposure. Exposure of the test substance to the eye caused slight reactions of the mucous membranes and the cornea which were more pronounced immediately following administration. Signs proved to be fully reversible within 7days. As there were 2 of 3 tested animals showed a positive response of corneal opacity ≥ 1, test substance can be concluded as Category 2 according to guidanc to regulation (EC) No 1272/2008.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on eye irritation test, 2 of 3 tested animals showed a positive response of corneal opacity ≥ 1, the test substance is concluded as irritant to the eye, Category 2 (H319) according to Regulation (EC) No 1272/2008. No potential for skin irritation was observed.