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Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study consistent with OECD guidelines

Data source

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

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
At the time of testing, the LLNA method (OECD 429) was not yet available as OECD adopted guideline.

Test material

Constituent 1
Chemical structure
Reference substance name:
1,4-Benzenediamine, N,N'-mixed Ph and tolyl derivs.
EC Number:
273-227-8
EC Name:
1,4-Benzenediamine, N,N'-mixed Ph and tolyl derivs.
Cas Number:
68953-84-4
Molecular formula:
C18-20H16-20N2
IUPAC Name:
Reaction mass of N-phenyl,N'-o-tolyl-phenylene diamine, N,N'-diphenyl-p-phenylene diamine and N,N'-di-o-tolyl-phenylene diamine

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
Animals were obtained from Harlan Sprague Dawley, Michigan. They were young adults of both sexes with study initiation body weights of 350 - 500 g. The test animals were housed individually, and fed Purina Guinea Pig chow ad libitum. Drinking water also available ad libitum. The animals were quarantined for minimum of 5 days prior to study initiation. Animal rooms were 65 - 75 degrees F, and humidity 36-63% with 12 hr light/12 hr dark photocycle. Ventilation was 10-12 room air changes per hr.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
other: Induction - acetone/propylene glycol (0.5% v/v)
Concentration / amount:
Induction - 5% test substance in acetone/propylene glycol soln
Challenge - Either 100% of test substance or a 25% solutionn of test substance in acetone/mineral oil.
Challengeopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
other: Induction - acetone/propylene glycol (0.5% v/v)
Concentration / amount:
Induction - 5% test substance in acetone/propylene glycol soln
Challenge - Either 100% of test substance or a 25% solutionn of test substance in acetone/mineral oil.
No. of animals per dose:
20 animals were used in each of the two challenge test groups (25% & 100% DAPD)
Details on study design:
Range finding tests showed that application of up to and including 100% test chemical in a Hilltop chamber did not cause significant irritation, and thus was acceptable for use in induction & challenge phases. For intradermal assessments, injected soln concentration up to & including 5% was also without significant signs of irritation.

In the main study, induction exposures to test groups on Day 0 consisted of intradermal injections (at 2 opposing dorsal skin sites) of Freund's adjuvant, 5% test chemical in acetone/propylene glycol, and test chemical in Freund's adjuvant. Topical exposures were performed on Day 7 when sodium lauryl sulfate was applied to defat skin sites prior to application of 100% test chemical (0.35 g) 24 hr later. This chemical exposure was 48 hr in duration. Both induction exposures were done on single occasions. 20 guinea pigs (10 of each sex) were in this test group.

Control groups were established with 10 animals (5M/5F). These guinea pigs were treated with the Freund's adjuvant and solvents as employed in the test group above.

The challenge exposures were performed on Day 21. The chemical test group was exposed to 25 or 100% test chemical by applying it to the right side of each animal under a semi-occlusive patch or in a chamber, respectively. The chemical was kept in dermal contact for 24 hrs at which time, the site was wiped clean. These dermal sites were then scored 24 & 48 hrs later for signs of irritation (allergic responses).

A rechallenge was performed on Day 28 of the study. Dosing and skin scoring was as that described above for the initial challenge exposure.


Challenge controls:
Challenge controls were given identical test chemical exposures on Days 21 and 28 as described above for test groups.
Positive control substance(s):
yes
Remarks:
Dichloronitrobenzene

Results and discussion

Positive control results:
DNCB results conducted in 6 guinea pigs on Day 21 showed significant skin irritating responses. Irritation scores were 2.5/4.0 & 2.5/4.0 for 24 & 48 hr readings. Controls (4 animals) had irritation scores of 0.5 & 0.3/4.0 for 24 & 48 hrs, respectively. These data indicated assay responsiveness to exposures to allergenic chemicals.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
25%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no signs
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 25%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: no signs.
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
25%
No. with + reactions:
5
Total no. in group:
20
Clinical observations:
5 with edema
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 25%. No with. + reactions: 5.0. Total no. in groups: 20.0. Clinical observations: 5 with edema.
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
25%
No. with + reactions:
3
Total no. in group:
20
Clinical observations:
1 with edema, 2 with desquamation
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 25%. No with. + reactions: 3.0. Total no. in groups: 20.0. Clinical observations: 1 with edema, 2 with desquamation.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no signs
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: no signs.
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
100%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no signs
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no signs.
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
100%
No. with + reactions:
5
Total no. in group:
20
Clinical observations:
5 with desquamation
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 5.0. Total no. in groups: 20.0. Clinical observations: 5 with desquamation.
Reading:
rechallenge
Hours after challenge:
24
Group:
negative control
Dose level:
25%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no signs
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: negative control. Dose level: 25%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: no signs.
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
25%
No. with + reactions:
15
Total no. in group:
20
Clinical observations:
1 desquamation, 15 edema, 6 blanching
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 25%. No with. + reactions: 15.0. Total no. in groups: 20.0. Clinical observations: 1 desquamation, 15 edema, 6 blanching.
Reading:
rechallenge
Hours after challenge:
48
Group:
test chemical
Dose level:
25%
No. with + reactions:
11
Total no. in group:
20
Clinical observations:
11 desquamation, 10 edema, 6 blanching
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 25%. No with. + reactions: 11.0. Total no. in groups: 20.0. Clinical observations: 11 desquamation, 10 edema, 6 blanching.
Reading:
rechallenge
Hours after challenge:
24
Group:
negative control
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no signs
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: negative control. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: no signs.
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
100%
No. with + reactions:
15
Total no. in group:
20
Clinical observations:
8 desquamation, 15 edema, 13 blanching
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 100%. No with. + reactions: 15.0. Total no. in groups: 20.0. Clinical observations: 8 desquamation, 15 edema, 13 blanching.
Reading:
rechallenge
Hours after challenge:
48
Group:
test chemical
Dose level:
100%
No. with + reactions:
15
Total no. in group:
20
Clinical observations:
14 desquamation, 14 edema, 9 blanching
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 15.0. Total no. in groups: 20.0. Clinical observations: 14 desquamation, 14 edema, 9 blanching.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information
Conclusions:
The results in guinea pigs show the chemical to have sensitization activity in dermal tests in guinea pigs.
Executive summary:

Test material was subjected to a guinea pig maximization sensitization assay according to OECD and GLP guidelines. The induction phase included both intradermal and epidermal exposures to test chemical plus adjuvant.

The dermal response following 48h after the first challenge to 25% or 100% of chemical resulted in clinical effects (edema or desquamation) for 15% or 25% of test animals, respectively.

48 Hours after the second challenge 55% of the animals exposed to 25% of test chemical showed signs of desquamation, edema or blanching. In the test group exposed to 100% of test chemical, 75% of the animals showed signs of a sensitization reaction after 48 hours.