Alcohols, C6-24 and C6-24-unsatd., distn. residues

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 21 - Nov 12 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study conducted according to GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes

Type of study:

Buehler test

Justification for non-LLNA method:

The test was done before the implementation of the LLNA method.

Test material

Test material information

Constituent 1

Reference substance name:

Alcohols, C6-24 and C6-24-unsatd., distn. residues

EC Number:

310-079-6

EC Name:

Alcohols, C6-24 and C6-24-unsatd., distn. residues

Cas Number:

102242-48-8

Molecular formula:

Not available due to the complexity of the substance

IUPAC Name:

Alcohols, C6-24 and C6-24-unsatd. even numbered, distn. residues

Details on test material:

- Name of test material (as cited in study report): Pernil RU, Espum EGA 213
- Chemical designation: 18:1 fatty alkohol, unsaturated, distillation residue
- Physical state: brown liquid
- Analytical purity: 100%
- Lot/batch No.: Charge of laboratory, no batch number given
- Expiration date: December 31, 1995
- Stability: yes
- Storage: at room temperature

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Strain as cited in study report: Pirbright white, Hsd/Win:DH
- Source: Winkelmann, Borchen, Germany
- Age at study initiation: ca. 7 weeks
- Weight at study initiation: ca. 310 g
- Housing: groups of 2 - 3 animals per cage, Makrolon Type IV cages with standard softwood bedding
- Diet: pelleted Altromin Maintenance Diet 3022 (Fa. Altromin GmbH, Lage, Germany) ad libitum and carrots
- Water: tap water, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 25
- Humidity (%): 45 - 70
- Air changes (per hr): at least 8
- Photoperiod (hrs dark / hrs light): 12/12
- Music was offered during light hours

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

epicutaneous, occlusive

Vehicle:

arachis oil

Remarks:

oleum arachidis DAB 10

Induction 2

Concentration / amount:

Preliminary test series:
6, 12.5, 25, 50, 75, 100%
Main test, induction: 50%
Dose finding for maximal non-irritating concentration suitable for challenge: 20, 30 and 40%
Main test, challenge: 10 and 20%

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

arachis oil

Remarks:

oleum arachidis DAB 10

Challenge 2

Concentration / amount:

Preliminary test series:
6, 12.5, 25, 50, 75, 100%
Main test, induction: 50%
Dose finding for maximal non-irritating concentration suitable for challenge: 20, 30 and 40%
Main test, challenge: 10 and 20%

No. of animals per dose:

Treated group: 20
Control group: 10

Details on study design:

RANGE FINDING
Prior to testing for sensitisation, a preliminary test series was conducted for primary irritation of the test item and determination of the suitable concentration for induction. An amount of 0.5 mL of test solution was applied on the left flank for the concentrations ranging from 6 to 50% and on the right flank of each of 3 animals, for the concentrations ranging from 50 to 100%. Vehicle also was applied on each flank side. The exposure was terminated after 6 hours, by removing the dressing and cleaning the skin from residues. Based on the results of the preliminary test series, 50% was retained as test concentration for induction.

MAIN STUDY
The sensitisation test followed the procedure described by Buehler (Arch. Dermatol. 91: 171-175, 1965).

- INDUCTION
The hair of the flanks each test animal was removed by clipping. An amount of 0.5 mL of a 50% test solution was applied onto the left cranial clipped flank of each of the 20 animals of the treated group; the vehicle was applied similarly onto the flank of each of the 15 control animals. The application site was covered with an occlusive dressing for 6 hours. The application was repeated 3 times, once a week. Following the third induction, skin reaction was assessed after 1 and 24 hours.

- DOSE FINDING FOR CHALLENGE
Five animals of the control group which had already been treated with the vehicle were topically treated with 0.5 mL of 20, 30 and 40% test solution, applied onto the skin of the right flank. Exposure conditions were as already described (occlusive, 6 hours). After 24 and 48 hours following removal of the dressing, the skin was assessed signs of irritation. Based on the results of this test, 10 and 20% were selected as maximally non-irritating concentrations suitable for challenge.

- CHALLENGE EXPOSURE
Challenge was conducted after 14 days following the last induction. Similar as for induction, an amount of 0.5 mL of test material was applied onto an untreated clipped skin area of the flanks under occlusive conditions for 6 hours; both test concentrations, 10 and 20%, were applied bilaterally on the flanks of each animal of the treated group and the control group. After removal of the dressing, the skin was examined for reaction indicating sensitisation after 24, 48 and 72 hours.

EVALUATION OF SKIN REACTION
The skin reaction was scored according to the following grades:
- No reaction -> 0
- Weak erythema and/or edema -> 1
- Moderate erythema and/or edema -> 2
- Strong erythema and/or edema -> 3

EVALUATION CRITERIA
Subtracting the percentage of animals exhibiting dermal effects in the control group from the percentage of positive substance group animals gives the potential of test substance to cause skin sensitisation. A result of at least 15% of animals of the treatment group with dermal effects in a non-adjuvant-test characterizes the test substance as a sensitiser.

CHECK FOR MORTALITY AND BOY WEIGHT
All animals were observed for mortality and body weight gain was recorded.

Results and discussion

In vivo (non-LLNA)

Results

Results 1

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

induction: 50%, challenge: 20%

No. with + reactions:

16

Total no. in group:

20

Clinical observations:

following first challenge, up to 80% of the animals of the treatment group showed weak to moderate skin reactions.

Remarks on result:

other: see Remark

Remarks:

Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: induction: 50%, challenge: 20%. No with. + reactions: 16.0. Total no. in groups: 20.0. Clinical observations: following first challenge, up to 80% of the animals of the treatment group showed weak to moderate skin reactions..

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

induction: vehicle only, challenge: 20%

No. with + reactions:

7

Total no. in group:

10

Clinical observations:

following first challenge, up to 70% of the animals of the control group showed weak to moderate skin reactions.

Remarks on result:

other: see Remark

Remarks:

Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: induction: vehicle only, challenge: 20%. No with. + reactions: 7.0. Total no. in groups: 10.0. Clinical observations: following first challenge, up to 70% of the animals of the control group showed weak to moderate skin reactions..

Results 3

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

induction: 50%, challenge: 10%

No. with + reactions:

4

Total no. in group:

20

Clinical observations:

following rechallenge, up to 20% of the animals of the treatment group almost showed weak skin reactions (in one case, reaction was assessed as moderate).

Remarks on result:

other: see Remark

Remarks:

Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: induction: 50%, challenge: 10%. No with. + reactions: 4.0. Total no. in groups: 20.0. Clinical observations: following rechallenge, up to 20% of the animals of the treatment group almost showed weak skin reactions (in one case, reaction was assessed as moderate)..

Results 4

Reading:

rechallenge

Hours after challenge:

48

Group:

negative control

Dose level:

induction: vehicle only, challenge: 10%

No. with + reactions:

3

Total no. in group:

10

Clinical observations:

following rechallenge, up to 30% of the animals of the control group showed weak skin reactions.

Remarks on result:

other: see Remark

Remarks:

Reading: rechallenge. . Hours after challenge: 48.0. Group: negative control. Dose level: induction: vehicle only, challenge: 10%. No with. + reactions: 3.0. Total no. in groups: 10.0. Clinical observations: following rechallenge, up to 30% of the animals of the control group showed weak skin reactions..

Any other information on results incl. tables

INDUCTION

After 1 hour following removal of the dressing, skin examination revealed one case of weak skin reaction graded 1 in the control group (1/15 animals) whereas in the treated group 9/20 animals displayed weak signs of skin irritation graded 1 and 2/20 had moderate signs graded 2. After 24 hours, all 15 control animals were free of skin findings whereas in the treated group, 7/20 and 3/20 animals still showed weak and moderate skin reaction, respectively.

CHALLENGE (20% test item)

Skin reactions seen after 24, 48 and 72 hours following removal of the dressing are summarised in the following table:

Overview of skin reaction after challenge with 20% test item in vehicle
Reading Time point	24 hours after challenge				48 hours after challenge				72 hours after challenge
Group	Control (N=10)		Treated (N=20)		Control (N=10)		Treated (N=20)		Control (N=10)		Treated (N=20)
Flank region	left	right	left	right	left	right	left	right	left	right	left	right
No reaction	9	5	20	15	8	3	16	4	10	7	19	15
Weak reaction	1	5	0	5	2	6	4	12	0	3	1	4
Moderate reaction	0	0	0	0	0	1	0	4	0	0	0	1
Strong reaction	0	0	0	0	0	0	0	0	0	0	0	0

RECHALLENGE (10% test item)

Skin reactions seen after 24, 48 and 72 hours following removal of the dressing are summarized in the following table:

Overview of skin reaction after rechallenge with 10% test item in vehicle
Reading Time point	24 hours after challenge				48 hours after challenge				72 hours after challenge
Group	Control (N=10)		Treated (N=20)		Control (N=10)		Treated (N=20)		Control (N=10)		Treated (N=20)
Flank region	left	right	left	right	left	right	left	right	left	right	left	right
No reaction	10	7	20	16	10	7	20	16	10	10	20	19
Weak reaction	0	3	0	4	0	3	0	3	0	0	0	1
Moderate reaction	0	0	0	0	0	0	0	1	0	0	0	0
Strong reaction	0	0	0	0	0	0	0	0	0	0	0	0

CLINICAL PARAMETERS

No mortality occurred. Body weight gain in the treated group was similar to that in the control group.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

DSD: not classified
CLP: not classified