Registration Dossier

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1995
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
92/69/EWG, B.6 (Guinea Pig maximization test (GPMT))
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Valid in vivo data from a guinea pig study is available.

Test material

Reference
Name:
Unnamed
Type:
Constituent

In vivo test system

Test animals

Species:
guinea pig
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: D. Hall, Newchurch, Staffordshire, England.
- Age at study initiation: Approximately four to five weeks of age.
- Weight at study initiation: 292 to 339 g.
- Housing: in groups of five in suspended metal cages with wire mesh floors
- Diet: ad libitum (A vitamin C enriched guinea-pig diet FD2; hay was given weekly) diet was not analysed for nutrients, possible contaminants or microorganisms.
- Water: ad libitum (analysed weekly for contamination).
- Acclimation period: For five days prior to allocation to the main study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.
- Humidity (%): 30 - 70.
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

An additional six animals, from the same supplier, were used for the preliminary investigations.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
coconut oil
Concentration / amount:
1 % v/v in Alembicol D (A product of coconut oil, supplied by Alembic Products, Saltney, Chester, England).
Route:
epicutaneous, occlusive
Vehicle:
coconut oil
Concentration / amount:
100 %
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
coconut oil
Concentration / amount:
100 % and 50 % v/v in Alembicol D.
No. of animals per dose:
Number of animals in test group: 10
Number of animals in negative control group: 5
Details on study design:
RANGE FINDING TESTS:
Animals for these investigations were pre-treated with an intradermal injection of Freund's complete adjuvant, 50:50 with water for irrigation (Ph. Eur), approximately two weeks prior to the start of the preliminary investigations.

MAIN STUDY
A. INDUCTION EXPOSURE
Intradermal Injection
- Site/Area: Back (scapular region)/2 x 4 cm
- No. of injections/animal: 6 injections in pairs each.
- Test substance group (cranial; 2 injections side by side): Freund's complete adjuvant (FCA) blended (1:1, v/v) with water (Ph.Eur.).
- Test substance group (middle; 2 injections side by side): The test article (1%) in vehicle.
- Test substance group (caudal; 2 injections side by side): 50% Freund's complete adjuvant blended (1:1, v/v) with water + 1 % test article in vehicle.
- Control group (cranial; 2 injections side by side): Freund's complete adjuvant (FCA) blended (1:1, v/v) with water (Ph.Eur.).
- Control group (middle; 2 injections side by side): Vehicle.
- Control group (caudal; 2 injections side by side): 50% Freund's complete adjuvant blended (1:1, v/v) with water + vehicle
- Volume per injection: 0.1 ml
- Evaluation (hr after injection): from 24 hour till day 7
- Exposure period: Single exposure

Epicutaneous induction exposure
- Time schedule: 7 days after intradermal injection
- Site: same as intradermal injections (site was pretreated 24 hours before with 0.5 ml per site of 10% w/w sodium lauryl sulphate in petrolatum)
- Frequency of applications: Once.
- Area of application: 2 x 4 cm (on Whatman No. 3)
- Concentrations: undiluted test article (test group) and water (control group)
- Volume applied: 0.4ml/animal
- Type of coverage: occlusive (impemeable plastic adhesive tape (50 ram width "Blenderm") + adhesive bandage (50 mm width "Elastoplast")
- Duration: 48 hours
- Evaluation (hr after challenge): directly after removal of patch

B. CHALLENGE EXPOSURE (all animals)
- Frequency of exposure: once
- Time schedule: 2 weeks after termination of induction exposures
- Site: left flank
- Area of application: 2 x 2 cm (Whatman No. 3 paper)
- Concentrations: undiluted (anterior flank) and 50% (v/v) in vehicle
- Volume applied: 0.2 ml/animal
- Type of coverage: occlusive (impemeable plastic adhesive tape (50 ram width "Blenderm") + adhesive bandage (50 mm width "Elastoplast")
- Duration: 24 hours
- Evaluation (hr after challenge): 24 and 48 hours after termination of exposure

SCORING SYSTEM: OECD Draize system
Erythema and eschar formation:
No erythema 0
Slight erythema 1
Well-defined erythema 2
Moderate erythema 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth) 4

Oedema formation:
No oedema 0
Slight oedema 1
Well-defined (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 the area of exposure) 4

OTHER
All animals were observed daily for signs of ill health or toxicity. The body weight of each guinea-pig on the main study was recorded on Day 1 (day of intradermal injections) and on the last day observations were made of dermal responses to the challenge applications.
Positive control substance(s):
yes
Remarks:
The sensitivity of the guinea-pig strain used is checked periodically at HRC with hexyl cinnamic aldehyde, a known sensitiser

Results and discussion

Positive control results:
See Table 1 for summary of positive control data

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Refer to Table 2
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Refer to Table 2
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Refer to Table 2
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Refer to Table 2
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
Refer to Table 2
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
Refer to Table 2
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
Refer to Table 2
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
Refer to Table 2

Any other information on results incl. tables

Table 1. Summary of positive control data using hexyl cinnamic aldehyde

M&K

R&D

No.

Number of animals

Dates of study

Dates of receipt of HCA

Dose levels % (aqueous dilutions)

Results

Induction

Challenge

Test

Control

Start

Finish

Intradermal

Topical

Positive

Inconclusive

Negative

1

10

5

08.12.92

09.01.93

29.10.92

10

As

Supplied

As supplied

and 50

10/10

0/10

0/10

IE

10

10

20.07.93

16.08.93

11.06.93

10

As

Supplied

As supplied

and 50

9/10

0/10

1/10

2A

10

10

15.03.94

11.04.94

11.06.93

10

As

Supplied

As supplied

and 50

10/10

0/10

0/10

2D

10

10

06.09.94

01.10.94

12.08.94

10

As

Supplied

As supplied

and 50

9/10

0/10

1/10

2G*

10

10

28.09.94

23.10.94

12.08.94

10

As

Supplied

As supplied

and 50

10/10

0/10

0/10

2H

10

10

05.10.94

30.10.94

12.08.94

10

As

Supplied

As supplied

and 50

10/10

0/10

0/10

3A

10

10

24.01.95

18.02.95

24.10.94

10

As

Supplied

As supplied

and 50

8/10

1/10

1/10

Animals supplied by D. Hall Newchurch.

* Animals supplied by Interfauna (UK) Ltd.

Hexyl cinnamic aldehyde obtained from Aldrich Chemicals Co.

Table 2. Dermal reactions observed after each induction

Site

Intradermal injection

Topical application

Test animals

Control animals

Test animals

Control animals

1

Necrosis

Necrosis

Slight erythema

Slight erythema

2

Slight irritation

Slight irritation

3

Necrosis

Necrosis

 

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met