Reaction mass of Amines, coco alkyl and β-Alanine, N-(2-carboxyethyl)-, N-coco alkyl derivs. and β-Alanine, N-coco alkyl derivs.

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From August 31, 2001 to April 17, 2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP Guideline study (OECD 404)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

yes

Remarks:

-the temperature and relative humidity recorded in the animal room were sometimes outside of the target ranges specified in the Study plan, -the animal's breeder was CEGAV (instead of Elevage Cunicole de Val de Selle).

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

see above

Principles of method if other than guideline:

These minor deviations were not considered to have compromised the validity or integrity of the study.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Reaction mass of Amines, coco alkyl and ß-Alanine, N-(2-carboxyethyl)-, N-coco alkyl derivs. and ß-Alanine, N-coco alkyl derivs.

EC Number:

915-790-0

Molecular formula:

n/a

IUPAC Name:

Reaction mass of Amines, coco alkyl and ß-Alanine, N-(2-carboxyethyl)-, N-coco alkyl derivs. and ß-Alanine, N-coco alkyl derivs.

Test material form:

liquid

Details on test material:

- Purity: 69% (28% isopropanol – 3% water)
- Batch Number: 01652201
- Description: yellow liquid
- Container: one glass flask
- Date of receipt: 28 June 2001
- Storage conditions: at room temperature and protected from light
- Expiry date/stability: June 2002
- Carbon content: 65.8% (weight/weight).

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Sex: male
- Source: CEGAV, Saint Mars d'Egrenne, France.
- Age at study initiation: no data
- Weight at study initiation: 2.9 ± 0.1 kg.
- Housing: no data
- Diet (e.g. ad libitum): ad libitum, 110 pelleted diet (UAR, Villemoisson, Epinay-sur-Orge, France)
- Water (e.g. ad libitum): Drinking water filtered by a FG Millipore membrane (0.22 micron) was provided ad libitum.
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- temperature: 18 ± 3°C
- relative humidity: 30 to 70%
- light/dark cycle: 12 h/12 h
- ventilation: approximately 12 cycles/hour of filtered, non-recycled air.

IN-LIFE DATES: From: September 11, 2001 To: September 27, 2001

Test system

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 mL

Duration of treatment / exposure:

3 minutes or 4 hours

Observation period:

1 hour, 24, 48 and 72 hours.
Following the OECD and EC guidelines, since there was persistent irritation reactions at 72 hours, the observation period was extended. In the absence of complete reversibility of skin reactions on day 15, the study was ended.

Number of animals:

3 rabbits

Details on study design:

The day before treatment, both flanks of each animal were clipped using electric clippers and the skin of each animal was examined. Only animals with healthy intact skin were used.
Doses of 0.5 mL of the test item were placed on a dry gauze pad, which was then applied to the right flank (application for 4 hours) or the left flank (application for 3 minutes) of the animals.

The test item and the gauze pad were held in contact with the skin by means of an adhesive hypoallergenic aerated semi-occlusive dressing and a restraining bandage.

REMOVAL OF TEST SUBSTANCE
After removal of the dressing, any residual test item was wiped off by means of a dry gauze pad.

SCORING SYSTEM:

Erythema and eschar formation:
. no erythema ........................................................................................................... 0
. very slight erythema (barely perceptible) .............................................................. 1
. well-defined erythema........................................................................................... 2
. moderate to severe erythema................................................................................. 3
. severe erythema (beet redness) to slight eschar formation (injuries in depth) ...... 4

Oedema formation
. no oedema ............................................................................................................. 0
. very slight oedema (barely perceptible) ................................................................ 1
. slight oedema (edges of area well-defined by definite raising)............................. 2
. moderate oedema (raised approximately 1 millimetre)......................................... 3
. severe oedema (raised more than 1 millimetre and extending beyond area
of exposure)........................................................................................................... 4

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

erythema score

Remarks:

(3-min exposure)

Basis:

animal #1

Time point:

other: Mean 24/48/72 hr

Score:

2.3

Max. score:

4

Reversibility:

not fully reversible within: 15 days

Remarks on result:

other: Erythema and dry skin were observed at 15 days.

Results 2

Irritation parameter:

edema score

Remarks:

(3-min exposure)

Basis:

animal #1

Time point:

other: Mean 24/48/72 hr

Score:

0.7

Max. score:

4

Reversibility:

fully reversible within: 6 days

Results 3

Irritation parameter:

erythema score

Remarks:

(4 hr exposure)

Basis:

mean

Remarks:

3 rabbits

Time point:

other: Mean 24/48/72 hr

Score:

3.1

Max. score:

4

Reversibility:

not fully reversible within: 15 days

Remarks on result:

other: Erythema and dry skin were observed at 15 days.

Results 4

Irritation parameter:

edema score

Remarks:

(4 hr exposure)

Basis:

mean

Remarks:

3 rabbits

Time point:

other: Mean 24/48/72 hr

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 7 days

Irritant / corrosive response data:

After a 3-minute exposure (one animal):
A very slight to moderate erythema (grades 1 to 3) was observed, from day 1 up to the end of the observation period (day 15). A slight oedema was noted on days 4 and 5. Dryness of the skin, sometimes together with crusts, was recorded between day 6 and day 15.

After a 4-hour exposure (three animals):
A well-defined to severe erythema (grades 2 to 4) was observed in all animals from day 1; it persisted up to day 5, 9 or up to the end of the observation period (day 15). A slight oedema (grade 2) was noted in all animals, from day 1 (one animal) or day 2 (two animals) up to day 5 (one animal) or 6 (two animals).

Other effects:

Other cutaneous reactions, including dryness of the skin and crusts, were recorded in all animals during the study. A superficial and rapidly reversible necrosis was observed in 2/3 animals on day 4 or 5. The necrosis observed corresponds to a lesion of the superficial layers of the skin (epidermis) occurring 72 hours for animal 601 and at day 5 for the 602 one. These lesions heal quickly but with the presence of a crust for the animal 602 until day 10. It is not an eschar (complete necrosis of the skin, including the dermis).
A yellow coloration of the skin was noted in 2/3 animals.

Any other information on results incl. tables

Table 1: 3-minute exposure – Individual cutaneous examinations and mean values of the scores recorded for the first animal (24, 48 and 72 hours).

 

Rabbit number	Dermal irritation	Scores				Mean irritation score (1)
		1h – D1	24h – D2	48h – D3	72h – D4
862	Erythema	2	2	2	3	2.3
	Oedema	0	0	0	2	0.7
	Other	*	*	*	*

(1)  mean scores on days 2, 3 and 4

h = hour; D = day; * = none

 

Rabbit number	Dermal irritation	Scores
		D5	D6	D7	D8	D9	D10	D11	D12	D13	D14	D15
862	Erythema	3	3	3	3	2	2	1	1	1	1	1
	Oedema	2	0	0	0	0	0	0	0	0	0	0
	Other	*	S	S/A	S/A	S/A	S	S	S	S	S	S

 

 

 

Table 2: 4-hour exposure – Individual cutaneous examinations and mean values of the scores recorded for each animal (24, 48 and 72 hours).

 

Rabbit number	Dermal irritation	Scores				Mean irritation score (1)
		1h – D1	24h – D2	48h – D3	72h – D4
862	Erythema	2	3	3	3	3.0
	Oedema	0	2	2	2	2.0
	Other	*	*	C	C
601	Erythema	2	3	3	4	3.3
	Oedema	0	2	2	2	2.0
	Other	*	*	*	N
602	Erythema	2	3	3	3	2.3
	Oedema	2	2	2	2	2.0
	Other	*	C	C	C

 

Rabbit number	Dermal irritation	Scores
		D5	D6	D7	D8	D9	D10	D11	D12	D13	D14	D15
862	Erythema	3	LS	LS	LS	3	3	2	2	2	2	2
	Oedema	2	0	0	0	0	0	0	0	0	0	0
	Other	C	C/S	C/S	C/S	S	S	S	S	S	S	S
601	Erythema	4	3	2	2	1	0	0	0	0	-	-
	Oedema	2	2	0	0	0	0	0	0	0	-	-
	Other	N	S	S	S	S	S	S	S	*	-	-
602	Erythema	4	LA	LA	LA	LS	LS	0	0	0	0	0
	Oedema	2	2	0	0	0	0	0	0	0	0	0
	Other	N	A	A	A	A	*	S	S	S	S	S

 

(1)  mean scores on days 2, 3 and 4

h = hour; D = day; * = none

S = dryness of the skin

C = yellow coloration of the skin

N = necrosis

A = crust

LS = scoring masked by dryness of the skin

LA: scoring masked by the crust

- : cutaneous examination not performed

Applicant's summary and conclusion

Interpretation of results:

Category 1C (corrosive)

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The substance should be considered as severely irritant when applied topically to rabbits. In a conservative approach, a classification corrosive cat. 1 C is proposed.

Executive summary:

The potential of the test item to induce skin irritation was evaluated in rabbits according to OECD (No. 404, 17th July 1992) and EC (92/69/EEC, B.4, 31st July 1992) guidelines.The study was conducted in compliance with the principles of Good Laboratory Practice Regulations.

In the first instance, the test item was applied for periods of 3 minutes and 4 hours to a single male New Zealand White rabbit. Since the test item was not corrosive on this first animal, it was then applied for 4 hours to two other animals.

A single dose of 0.5 mL of the undiluted test item was applied to the closely-clipped skin of one flank.

The test item was held in contact with the skin by means of a semi-occlusive dressing.

Cutaneous reactions were observed approximately 1 hour, 24, 48 and 72 hours after removal of the dressing and then daily until the end of the observation period. The mean values of the scores for erythema and oedema were calculated for each animal.

After a 3-minute exposure (one animal):

A very slight to moderate erythema was observed, from day 1 up to the end of the observation period (day 15). A slight oedema was noted on days 4 and 5. Dryness of the skin, sometimes together with crusts, was recorded between day 6 and day 15.

After a 4-hour exposure (three animals):

A well-defined to severe erythema was observed in all animals from day 1; it persisted up to day 5, 9 or up to the end of the observation period (day 15).

A slight oedema was noted in all animals, from day 1 (one animal) or day 2 (two animals) up to day 5 (one animal) or 6 (two animals).

Other cutaneous reactions, including dryness of the skin and crusts, were recorded in all animals during the study. A superficial and rapidly reversible necrosis was observed in 2/3 animals on day 4 or 5.

A yellow coloration of the skin was noted in 2/3 animals.

Mean scores over 24, 48 and 72 hours for each animal were 3.0, 3.3 and 3.0 for erythema and 2.0, 2.0 and 2.0 for oedema.

Under these experimental conditions, the test item should be considered as severely irritant when applied topically to rabbits. However by a conservative approach a classification as corrosive cat. 1C. is proposed.