Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Equivalent to OECD 406. GLP study. The sensitivity and reliability of the experimental technique was not assessed by use of substances which are known to have mild-to-moderate skin sensitisation properties. However, it is considered that this deviation does not affect the outcome of the study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1988

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: Magnusson and Kligman (1969)
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
(However, it is considered that this deviation does not affect the outcome of the study)
GLP compliance:
yes
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Physical state: Pale yellow powder
- Analytical purity: 99.04%
- Lot/batch No.: 5/87; Indent 9200/9622
- Storage condition of test material: Following its arrival in Compound Control this test substance was stored in the dark at ambient temperature.
- Stability: Infra-red spectra of the test substance were taken. There were no significant differences between the spectra and so, on this basis, the test substance was judged to have been stable for the duration of this study.

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Porcellus Ltd., U.K.
- Age at study initiation: Not documented.
- Weight at study initiation: males: 558-667 g; females: 501-565 g
- Housing: Animals were housed in groups of five animals (two and three per cage). Hanging, stainless-steel cages with all-mesh floors and tops and half-mesh fronts were used. Each cage measured 54 cm x 31 cm x 36 cm.
- Diet (e.g. ad libitum): SG1 with vitamin C supplement, Grain Harvesters Ltd., Kent, ad libitum.
- Water (e.g. ad libitum): Filtered from public supply, ad libitum.
- Acclimation period: At least two weeks.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23 ºC
- Humidity (%): Not documented.
- Air changes (per hr): Not documented.
- Photoperiod (hrs dark / hrs light): Lighting was automatically switched on for the day (06.00 to 18.00 hours GMT) and off for the night (18.00 to 06.00 hours GMT).


Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
corn oil
Concentration / amount:
Induction - Intradermal injections (0.5% m/v in corn oil):
Test animals:
Anterior sites: 0.1 ml of Freunds Complete Adjuvant (FCA)
Middle sites: 0.1 ml of test material in corn oil
Posterior sites: 0.1 ml of test material in 50:50 FCA/corn oil

Control animals:
Anterior sites: 0.1 ml of FCA
Middle sites: 0.1 ml of corn oil
Posterior sites: 0.1 ml of 50:50 FCA/corn oil

Induction - topical:
0.45 ml of the diluted test material (50% m/m petroleum jelly).

Challenge:
0.15 ml of the diluted test material (50% m/m in petroleum jelly).

Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
corn oil
Concentration / amount:
Induction - Intradermal injections (0.5% m/v in corn oil):
Test animals:
Anterior sites: 0.1 ml of Freunds Complete Adjuvant (FCA)
Middle sites: 0.1 ml of test material in corn oil
Posterior sites: 0.1 ml of test material in 50:50 FCA/corn oil

Control animals:
Anterior sites: 0.1 ml of FCA
Middle sites: 0.1 ml of corn oil
Posterior sites: 0.1 ml of 50:50 FCA/corn oil

Induction - topical:
0.45 ml of the diluted test material (50% m/m petroleum jelly).

Challenge:
0.15 ml of the diluted test material (50% m/m in petroleum jelly).

No. of animals per dose:
Ten animals per sex.
Control group: five animals per sex.
Details on study design:
RANGE FINDING TESTS:
Two male and two female guinea pigs were closely shorn in the shoulder region using electric clippers followed by an electric razor. 0.1 ml doses of several dilutions (0.05, 0.1, 0.5 and 1.0% m/v in corn oil) of the test material were injected intradermally on each side of the mid-line. The animals were examined to determine the maximum concentration that could be used without causing untoward toxicity.

The flanks of each animal in further groups of two male and two female guinea pigs was closely shorn. 0.3 ml doses of several dilutions (25% and 50 m/m in petroleum jelly) of the test material were absorbed onto 16 cm2 Whatman No. 3 filter paper patches. The patches were placed on the flanks and held in place with a “Sleek” adhesive tape patch, then covered with an 8 cm “Poroplast” elastic adhesive bandage for 24 hours. After removal of the patches and bandages the dermal test sites were examined for signs of irritation.

MAIN STUDY
A. INDUCTION EXPOSURE
Pairs of intradermal injections were administered to the animals as described under the heading "concentration". One week after induction by intradermal injection, the same area of skin was shaven using electric clippers. A 16 cm2 patch of Whatman No. 3 filter paper was coated with 0.45 ml of the test material and placed over the sites of intradermal injections. The patches were covered with a “Sleek” dressing. The dressing was then securely covered with an 8 cm “Poroplast” elastic adhesive bandage for 48 hours.


B. CHALLENGE EXPOSURE
Challenge was carried out two weeks after the topical induction. Hair was removed from a 3 cm x 3 cm area of one flank by clipping then shaving. A 4cm2 patch of Whatman No. 3 filter paper coated with 0.15 ml of the test material was placed on the shaven area, covered with a 3 cm square of adhesive tape, held in place by an 8 cm “Poroplast” elastic adhesive bandage. After 24 hours the patches were removed and the challenges sites examined for any response, 24 and 48 hours after its removal.
Challenge controls:
Controls were also treated with the test material.
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
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. No with. + reactions: 0.0. Total no. in groups: 20.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. No with. + reactions: 0.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

Table 7.4.1: Summary of the Sensitisation Response after Challenge

Sensitisation Response

Hours

24

48

Test animals

0/20

0/20

Control animals

0/10

0/10

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In the guinea-pig maximization test of Magnusson and Kligman, none of the twenty test animals showed positive responses at 24 or 48 hours after removal of the challenge patches. The test material was therefore not a skin sensitizer in guinea pigs.
Executive summary:

A range-finding test was performed to determine the appropriate concentrations that could be used for both intradermal and topical induction as well as topical challenge. The first induction phase involved six intradermal injections. These doses were comprised of pairs of injections of the test substance in corn oil, the test substance combined with Complete Freund´s Adjuvant and corn oil and the Adjuvant alone. A control group (ten animals) was maintained under the same environmental conditions and received injections of corn oil, corn oil combined with complete Freund´s Adjuvant and Adjuvant alone. The second phase of induction was conducted by topical application for a period of 48 hours. Two weeks after the topical induction, a challenge dose of the test material was applied for 24 hours. Twenty-four and 48 hours after challenge patch removal, the animals were scored for a sensitisation response. None of the twenty test animals showed any positive response at either 24 or 48 hours after removal of the challenge patches.