4-Cyclopentadecen-1-one, (4Z)-

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From December 22, 1997 to January 23, 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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 406 (Skin Sensitisation)

Deviations:

yes

Remarks:

details of environmental conditions not reported

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

At the time of study completion (1998), the LLNA OECD test method was not adopted.

Test material

Test material information

Constituent 1

Reference substance name:

(4Z)-cyclopentadec-4-en-1-one

EC Number:

630-473-4

Cas Number:

14595-54-1

Molecular formula:

C15H26O

IUPAC Name:

(4Z)-cyclopentadec-4-en-1-one

Test material form:

solid

Details on test material:

- Physical state: Colourless cristallized solid
- Storage condition of test material: Room temperature, away from the light

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles RIVER, 76410 Saint-Aubin-Les-Elbeuf, France.
- Weight at study initiation: 315-374 g
- Housing: Animals were housed individually in polypropylene cage (310 x 465 x 190).
- Diet: Complete pelleted diet UAR 106 (91360 - Epinay sur Orge, France), ad libitum
- Acclimation period: 5 days

IN-LIFE DATES: From December 22, 1997 to January 23, 1998

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

other: mineral oil

Concentration / amount:

40%

Day(s)/duration:

Day1

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

other: acetone

Concentration / amount:

80%

Day(s)/duration:

Day 8

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

other: acetone

Concentration / amount:

2.5 and 5%

Day(s)/duration:

Day 21

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

5 and 10 animals for control and treatment group, respectively

Details on study design:

RANGE FINDING TESTS:
- Determination of the MIC (Maximal concentration slightly to moderately irritating) by intradermal route (MICID): One week before the main test, two guinea-pigs received intradermal injections of test material at 10, 20, 40 and 80 % w/v in mineral oil and evaluated for skin reactions at 24 h after injections. The concentration selected for the main study (intradermal induction) was 40 % w/v in mineral oil.
- Determination of the MIC by topical route (MICT): Between Day -8 and Day 4, two guinea pigs were applied topically with test material at 10, 20, 40 and 80 % in acetone and acetone only under occlusive patch for 48 ± 1 h and cutaneous irritation reactions were evaluated after the patch removal. The concentration selected for the main study (topical induction) was 80 % in acetone.
As irritation reactions were still observed with the lowest concentration (10 %), in order to determine the concentrations to be used for the maximal non irritant concentration (MNIC) determination, a second test was performed in two other animals with lower concentrations (1.25, 2.5, 5 and 10 % in acetone).
-Determination of the MNIC by topical route: One week before the main test, at Day -8, two guinea pigs received the intradermal injection of 0.1 mL of the 3 following preparations on both sides of the spinal column, on the clipped dorsal area: Freund's Complete Adjuvant (FCA) diluted at 50 % with distilled water (v/v); vehicle : mineral oil; a 50/50 mixture of the FCA diluted at 50 % with distilled water (v/v) and mineral oil. After a 21 day resting period, the animals were applied topically with test material at 2.5 and 5% in acetone (determined in w/v from the results obtained during the evaluation of the MICT) under occlusive patches for 24 ± 1 h. Skin reactions were evaluated 24, 48 and 72 h after patch removal. The concentrations selected for the main study (challenge phase) were 2.5 and 5 % in acetone (MNIC/2 and MNIC).

MAIN STUDY
A. INDUCTION EXPOSURE: INTRADERMAL
- No. of exposures: One
- Test groups: Intradermally injected with 3 injections (0.1 mL each) of FCA and distilled water in the ratio 1:1 (v/v); a 40 % w/v formulation of the test material in mineral oil; a 40 % w/v formulation of the test material in a 1:1 (v/v) preparation of FCA and distilled water, on Day 1.
- Control group: Intradermally injected with 3 injections (0.1 mL each) of FCA and distilled water in the ratio 1:1 (v/v); vehicle (mineral oil); a mixture 1:1 (v/v) of FCA plus distilled water and mineral oil, on Day 0.
- Site: On both sides of the spinal column, at the scapular level
- The injections 1 and 2 were performed close to each other, as nearer as possible to the head. The injection 3 was performed towards the caudal part of the experimental area.

On Day 7: After 6 days resting period, animals were clipped at the injection site level. The experimental area of the control and treated group animals were coated with 0.5 mL of a sodium lauryl sulphate at 10 % in petrolatum to produce a local irritation.

B. INDUCTION EXPOSURE: TOPICAL
- No. of exposures: One, on Day 8
- Exposure period: 48 ± 1 h
- Test groups: 0.5 mL of the test material at 80 % in acetone was applied to the filter paper patch of 8 cm2 (2x4 cm), put on a piece of gauze of 8 cm2 (2x4 cm) put on a square Blenderm occlusive film of 25 cm2. The occlusive patch was held in place with an adhesive Micropore film put around the animal trunk during 48 ± 1 h exposure.
- Control group: 0.5 mL of the vehicle (acetone COOPER batch 122470) was applied according to the method described above and let in contact with the skin during 48 ± 1 h exposure.
- Site: Same intradermally injected area
- Frequency of applications: Single application

C. CHALLENGE EXPOSURE: TOPICAL
- No. of exposures: One
- Day of challenge: Day 21
-After a 11-day rest period, the guinea-pigs from both groups were clipped one last time at the dorso-lumbar region, the skin surface chosen corresponding to a zone which had not previously been in contact with the test product.
- Exposure period: 24 ± 1 h
- Test groups and control groups: 0.2 mL of the test material at 2.5 and 5 % in acetone were applied respectively on 2 patches made with a square of filter paper of about 2 cm2 which receives the product, put on a square of gauze of 4 cm2, put in the middle of an occlusive Blenderm film of 25 cm2. The patches were loosely held on the right flank of each animal by a Micropore film. As the vehicle used was not distilled water, 0.2 mL of acetone was put on a patch and applied to the same flank of each animal as the test material.
- Evaluation (h after removal of challenge patch): 24, 48 and 72 h

OTHER:
- Body weight of each animal was recorded at the start and end of the study.

Challenge controls:

See "Details on study design"

Positive control substance(s):

yes

Remarks:

2-Mercapto Benzothiazole

Results and discussion

Positive control results:

The sensitivity and the validity of the experimental technique adopted were confirmed by the study EVIC-CEBA Td 718 of October 09, 1997. The product 2-Mercapto Benzothiazole appeared highly to very highly sensitizing for the guinea-pig (80 % to 90 % of reactive animals at the 24, 48 and 72 h reading - class 4) when it was used at challenge diluted at 5 % and 2 % with petrolatum.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 3

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 4

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

2.5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 5

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

2.5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 6

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

2.5 % in acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 7

Reading:

1st reading

Hours after challenge:

24

Group:

other: Solvent control

Dose level:

Acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 8

Reading:

2nd reading

Hours after challenge:

48

Group:

other: Solvent control

Dose level:

Acetone

No. with + reactions:

1

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 9

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

other: Solvent control

Dose level:

Acetone

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 10

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

2.5 or 5 % in acetone or acetone only

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 11

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

2.5 or 5 % in acetone or acetone only

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 12

Reading:

other: 3rd reading

Hours after challenge:

72

Group:

negative control

Dose level:

2.5 or 5 % in acetone or acetone only

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 13

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

5 to 2%

No. with + reactions:

8

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Remarks:

80 to 90% of reactive animals; exact number not reported

Results 14

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

5 to 2%

No. with + reactions:

8

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Remarks:

80 to 90% of reactive animals; exact number not reported

Results 15

Reading:

other: 3rd readling

Hours after challenge:

72

Group:

positive control

Dose level:

5 to 2%

No. with + reactions:

8

Total no. in group:

10

Remarks on result:

positive indication of skin sensitisation

Remarks:

80 to 90% of reactive animals; exact number not reported

Any other information on results incl. tables

Determination of the sensitizing potential (main test):

In the control group, no local irritation reaction was observed 24, 48 and 72 h after patch removal.

In the treated group (previously induced by intradermal and epidermal routes on another site of the skin), the presence of slight local irritation reaction in 1 animal for the three reading times after application of the tested substance at the concentrations 2.5 and 5 % in acetone. The same animal presented a slight erythematous reaction after application of the vehicle only for the 24 and 48 h readings.

 

Table 7.4.1/1: Main study – Challenge phase results

 

Group / Animal No.	5 % with acetone			2.5 % with acetone			Acetone
	24 h	48 h	72 h	24 h	48 h	72 h	24 h	48 h	72 h
Group 1: Control group
8983	0	0	0	0	0	0	0	0	0
8984	0	0	0	0	0	0	0	0	0
8985	0	0	0	0	0	0	0	0	0
8986	0	0	0	0	0	0	0	0	0
8987	0	0	0	0	0	0	0	0	0
Reactive animals (%)	0	0	0	0	0	0	0	0	0
Group 2: Treated group
8988	1	2	1	1	1	1	1	1	0
8989	0	0	0	0	0	0	0	0	0
8990	0	0	0	0	0	0	0	0	0
8991	0	0	0	0	0	0	0	0	0
8992	0	0	0	0	0	0	0	0	0
8993	0	0	0	0	0	0	0	0	0
8994	0	0	0	0	0	0	0	0	0
8995	0	0	0	0	0	0	0	0	0
8996	0	0	0	0	0	0	0	0	0
8997	0	0	0	0	0	0	0	0	0
Reactive animals (%)	10	10	10	10	10	10	10	10	0

 

General state - Mortality

The general state of the guinea-pigs was not modified throughout the test period.

The weight growth of all the animals was satisfactory. The mean weight gain calculated for the treated animals during the test period was not significantly different from the control animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the test conditions, the test item is not classified as sensitiser according to the criteria of Annex VI to Regulation (EC) No. 1272/2008 (CLP) and to the GHS.

Executive summary:

In a Magnusson & Kligman maximisation study (GPMT) performed according to OECD Guideline 406 and in compliance with GLP, 10 female Hartley guinea pigs were induced with three pairs of intradermal injections (0.1 mL each) of FCA and distilled water in the ratio 1:1 (v/v); a 40 % w/v formulation of the test material in mineral oil; a 40 % w/v formulation of the test material in a 1:1 (v/v) preparation of FCA and distilled water, on Day 1 on both sides of the spinal column, at the scapular level. Whereas, control group of 10 females was intradermally induced with 0.1 mL of FCA and distilled water in the ratio 1:1 (v/v); vehicle (mineral oil); a mixture 1:1 (v/v) of FCA plus distilled water and mineral oil. On Day 8, the experimental area of the control and treated group animals were coated with 0.5 mL of a sodium lauryl sulphate at 10 % in petrolatum to produce a local irritation. On Day 8, the same area was topically induced with 80 % test material in acetone via occluded filter paper patch for 48 ± 1 h in test group, whereas control group was applied with vehicle alone. On Day 21, a challenge filter paper patch loaded with test material (2.5 and 5 % in acetone) was applied to the right flank of all animals, and acetone was applied to the same flank of each animal as the test material. The test concentrations for the main study were determined from a sighting study.

In the control group, no local irritation reaction was observed after 24, 48 and 72 h after patch removal. In the treated group, the presence of slight local irritation reaction in 1 animal for the three reading times after application of the tested substance at the concentrations 2.5 and 5 % in acetone. The same animal presented a slight erythematous reaction after application of the vehicle only for the 24 and 48 h readings. The sensitivity and validity of the experiment was confirmed with known sensitiser, 2-Mercapto benzothiazole.

 

Under the test conditions, the test material is not classified according to the annex VI of the Regulation (EC) No. 1272/2008 (CLP) and to the GHS.

This study is considered as acceptable and satisfies the requirement for sensitisation endpoint.