Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
29 September 1997- 31 October 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report Date:
1998

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): Gel All DX
- Substance type: white powder
- Physical state: solid
- Lot/batch No.: 4073003
- Storage condition of test material: room temperature

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: David Hall Limited, Burton-on-Trent, Staffodshire, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 311- 374 g
- Housing: animals were housed singly or in pairs in solid-floor polypropylene cages furnished with woodflakes.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19- 22 °C
- Humidity (%): 42- 62 %
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
arachis oil
Concentration / amount:
SIGHTING TESTS
Intradermal induction: 0.1 %, 0.5 %, 1% and 5 % w/v
Topical Induction: 50 %, 25 %, 10 % and 5 % w/w
Topical challange: 25 %, 10 %, 5 % and 2 % w/w

MAIN STUDY
Intradermal induction: 0.5 % w/v
Topical induction: 25 % w/w
Topical challange: 10 % and 5 % w/w
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
arachis oil
Concentration / amount:
SIGHTING TESTS
Intradermal induction: 0.1 %, 0.5 %, 1% and 5 % w/v
Topical Induction: 50 %, 25 %, 10 % and 5 % w/w
Topical challange: 25 %, 10 %, 5 % and 2 % w/w

MAIN STUDY
Intradermal induction: 0.5 % w/v
Topical induction: 25 % w/w
Topical challange: 10 % and 5 % w/w
No. of animals per dose:
Sighting tests:
Intradermal induction: 4/ sex/ dose
Topical Induction: 2/ sex were treated with 4 preparations
Topical challange: 2/ sex were treated with 4 preparations

Main study: 10/ sex/ dose
Details on study design:
RANGE FINDING TESTS:
The concentrations of test material to be used at each stage of the main study were determined by 'sighting tests' in which groups of guinea pigs were treated with various concentrations of test material. The procedures were as follows:

-Selection of Concentration for lntradermal lnduction:
Four concentrations of test material were investigated (0.1 %, 0.5 %, 1 % and 5 % w/v in arachis oil BP). A total of 4 guinea pigs wereused, each guinea pig receiving four 0.1 mL injections of only 1 concentration of test material. The degree of erythema at the injection sites was assessed approximately 24, 48 and 72 h and 7 d after injection according to the Draize scale. The degree of oedema was not evaluated. Any evidence of systemic toxicity was also recorded. The highest concentration that caused only mild to moderate skin irritation, and which was well tolerated systemically, was selected for the intradermal induction stage of the main study.

-Selection of Concentration for Topical lnduction:
Two guinea pigs (intradermally injected with Freund's Complete Adjuvant 17 days earlier) were treated with 4 preparations of the test material (50 %, 25 %, 10 % and 5 % w/w in arachis oil BP). Applications were made to the clipped flanks under occlusive dressings for an exposure period of 48 h. The degree of erythema and oedema was evaluated approximately 1, 24 and 48 h after dressing removal. The highest concentration producing only mild to moderate dermal irritation was selected for the topical induction stage of the main study.

-Selection of Concentration for Topical Challenge:
Four preparations of the test material (25 %, 10 %, 5 % and 2 % w/w in arachis oil BP) were applied to the clipped flanks of 2 guinea pigs under occlusive dressings for an exposure period of 24 h. These guinea pigs did not form part of the main study but had been treated identically to the control animals of the main study, up to Day 14. The degree of erythema and oedema was evaluated approximately 1, 24 and 48 h after dressing removal. The highest non-irritant concentration of the test material and one lower concentration was selected for the topical challenge stage of the main study.

MAIN STUDY
- Test group: 1 group of 10 animals
- Control group: 1 group of 5 animals

A. INDUCTION EXPOSURE
- No. of exposures: 2; intradermal and topical
- Exposure period: 48 h (topical induction)
- Site: 40 mm x 20 mm on the shoulder region

Induction of test animals
- Shortly before treatment on Day 0 the hair was removed from an area approximately 40 mm x 60 mm on the shoulder region of each animal with veterinary clippers. A row of 3 injections (0.1 mL each) was made on each side of the mid-line. The injections were:
a) Freund's Complete Adjuvant plus distilled water in the ratio 1:1
b) a 0.5 % w/v formulation of the test material in arachis oil BP
c) a 0.5 % w/v formulation of the test material in a 1:1 preparation of Freund's Complete Adjuvant plus distilled water.
Approximately 24 and 48 hours after intradermal injection the degree of erythema at the test material injection sites (ie. Injection site b) was evaluated.
-One week later (Day 7), the same area on the shoulder region used previously for intradermal injections was clipped again and treated with a topical application of the test material formulation. A filter paper patch loaded with the test material formulation (25 % w/w in arachis oil BP) as a thick, even layer was applied to the prepared skin and held in place with a strip of surgical adhesive tape covered with an overlapping length of aluminium foil. The patch and foil were further secured with a strip of elastic adhesive wound in a double layer around the torso of each animal. This occlusive dressing was kept in place for 48 h.
- The degree of erythema and oedema was quantified 1 and 24 h following removal of the patches. Any other reactions were also recorded.

Induction of the control animals
-lntradermal injections were administered using an identical procedure to that used for the test animals, except that the injections were:
a) Freund's Complete Adjuvant plus distilled water in the ratio 1:1
b) arachis oil BP
C) a 50 % w/v formulation of arachis oil BP in Freund's Complete Adjuvant/distilled water 1:1
Approximately 24 and 48 hours after intradermal injection the degree of erythema at the vehicle injection sites was evaluated.
- The topical applications followed the same procedure as for the test animals except that the vehicle alone was applied to the filter paper. Skin reactions were quantified as for the test animals.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 1
- Exposure period: 24 h
- Site: 20 mm x 20 mm
- Concentrations: 5 % and 10 % w/w in arachis oil BP
- Evaluation (hr after challenge): 24 and 48 h after challange dressing removal, the degree of erythema and oedema was quantified.
- Shortly before treatment on Day 21, an area of approximately 50 mm x 70 mm on both flanks of each animal, was clipped free of hair with veterinary clippers. A square filter paper patch loaded with a thick, even layer of test material at the maximum non-irritant concentration (10 % w/w in arachis oil BP) was applied to the shorn right flank of each animal and was held in place with a strip of surgical adhesive tape. To ensure that the maximum non-irritant concentration was used at challenge, the test material at a concentration of 5 % w/w in arachis oil BP was similarly applied to a skin site on the left shorn flank. The patches were occluded with an overlapping length of aluminium foil and secured with a strip of elastic adhesive bandage wound in a double layer around the torso of each animal.

After 24 h, the dressing was carefully cut using blunt-tipped scissors, removed and discarded. The challenge sites were swabbed with cotton wool soaked in diethyl ether to remove residual material. The position of the treatment sites was identified by using a black indelible marker-pen. Prior to the 24-h observation the flanks were clipped using veterinary clippers to remove regrown hair.
Positive control substance(s):
no

Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
10 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 10 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None noted..
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
5 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
10 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 10 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
5 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 5 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None noted..
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
10 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 10 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None noted..
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
5 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 5 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
10 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 10 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
5 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
None noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 5 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: None noted..

Any other information on results incl. tables

lntradermal and Topical Sighting Tests

Intradermal sighting test- summary of results. Vehicle: arachis oil BP

Animal identification

Time of observation

Concentration of test material (% w/v)

Grade of erythema at injection sites

Evidence of systemic toxicity

A

24 h

1

3

None

48 h

3-4N*

None

72 h

3-4N*

None

7 d

0

None

B

24 h

5

3-4N

None

48 h

4NE

None

72 h

4E

None

7 d

4E

None

C

24 h

0.1

1-2

None

48 h

2

None

72 h

2

None

7 d

1

None

D

24 h

0.5

1-2

None

48 h

2-3

None

72 h

2

None

7 d

1

None

E= eschar; N= green necrosis; N*= possible necrosis

Topical sighting test for induction application (48-h exposure)- individual skin reactions. Vehicle: arachis oil BP

Animal identification

Concentration of test material (% w/w)

Skin reaction (hours after removal of patches)

1

24

48

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

E

50*

2

2

-

2

0

-

1

0

-

25

2

2

-

2

0

-

1

0

D

10

1

0

-

0

0

-

0

0

-

5

1

0

-

0

0

-

0

0

-

F

50*

3

2

-

2

1

-

1

0

-

25

2

2

-

2

1

-

1

0

D

10

1

0

-

0

0

-

0

0

-

5

1

1

-

0

0

-

0

0

-

Er= erythema; Oe= oedema; -= no other reactions noted; D= desquamation; *= maximum attainable concentration suitable for topical application

Topical sighting test for challenge application (24-h exposure)- individual skin reactions. Vehicle: arachis oil BP

Animal identification

Concentration of test material (% w/w)

Skin reaction (hours after removal of patches)

1

24

48

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

G

25*

2

2

-

1

0

-

0

0

-

10

2

2

-

0

0

-

0

0

-

5

2

1

-

0

0

-

0

0

-

2

2

1

-

0

0

-

0

0

-

H

25*

2

2

-

1

1

-

0

0

-

10

2

1

-

0

0

-

0

0

-

5

1

1

-

0

0

-

0

0

-

2

1

1

-

0

0

-

0

0

-

Er= erythema; Oe= oedema; -= no other reactions noted; *= maximum attainable concentration suitable for topical application

Based on these results, the following concentrations were selected for the main study:

Intradermal induction: 0.5 % w/v in arachis oil BP

Topical induction: 25 % w/w in arachis oil BP

Topical challenge: 10 % and 5 % w/w in arachis oil BP

Main study

-Skin Reactions Observed After lntradermal lnduction:

Well-defined erythema was noted at the intradermal induction sites of all test group animals at the 24 and 48-hour observations. Very slight erythema was noted at the intradermal induction sites of all control group animals at the 24-h observation and in 4 control group animals at the 48-h observation.

-Skin Reactions Observed After Topical lnduction:

Very slight to well-defined erythema was noted at the induction sites of all test group animals at the 1-h observation with very slight erythema at the induction sites of 9 test group animals at the 24-h observation. Bleeding from the intradermal induction sites was noted in 5 test group animals at the 1-h observation. Residual test material was noted in all test group animals. Bleeding from the intradermal induction sites was noted in 1 control group animal at the 1-h observation. No signs of erythema or oedema were noted at the treatment sites of control group animals at the 1 and 24-h observations.

-Skin Reactions Observed After Topical Challenge:

No skin reactions were noted at the challenge sites of the test or control group animals at the 24 or 48-h observations.

-Bodyweight:

Bodyweight gains of guinea pigs in the test group, between Day 0 and Day 24, were comparable to those observed in the control group animals over the same period.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material produced a 0 % (0110) sensitisation rate and was classified as a non-sensitiser to guinea pig skin.
Executive summary:

A study was performed to assess the contact sensitisation potential of the test material in the albino guinea pig. The study was performed in compliance with the OECD Guidelines for Testing of Chemicals No. 406 "Skin Sensitisation" and Method B6 of Commission Directive 92/69/EEC.

Ten test and five control animals were used for the main study.

Based on the results of sighting tests, the concentrations of test material for the induction and challenge phases were selected as follows:

- lntradermal lnduction: 0.5 % w/v in arachis oil BP

- Topical lnduction: 25 % w/w in arachis oil BP

- Topical Challenge: 10 % and 5 % w/w in arachis oil BP

The test material produced a 0 % (0/10) sensitisation rate and was classified as a non-sensitiser to guinea pig skin.