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Toxicological information

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
Study period:
1984
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: MAGNUSSON B. - KLIGMAN A.M. The identification of contact allergens by animal assay "The guinea pig maximization test". J. Invest. Derm. 1969,52,3,268-276
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
An in vitro or in chemico skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study (different from LLNA test) is available.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder

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: C.E.R.J. (53940 Saint Berthevin, France)
-Weight at study initiation:
test group: 374 g for the males and 360 g for the females
control group: 369 g for the males and 356 g for the females.
-Housing: the animals were housed individually in polypropylene cages (0,43 x 0,23 x 0,17 m). Each cage was equipped with a food container and a water bottle. Wooden saw dust chemical analysis was carried out periodically by the Laboratoire Municipal de Rouen (76000 Rouen, France).
-Diet: the animals were fed ad libitum during the study, with a certified pelleted diet Rabbit diet S.Q.C. (Special Diets Services Ltd., Nitham, Essex, England).
-Water: the animals had free access to bottles containing tap water filtered with Millipore filters (0,22 micron) and supplemented with vitamin C. Bacteriological and chemical water analysis and detection for major contaminants were performed regularly by the Laboratoire Municipal de Rouen (76000 Rouen, France).
-Acclimation period: 7 days.

ENVIRONMENTAL CONDITIONS
-Temperature (°C): 18 ± 3 °C
-Humidity (%): 50 ± 20 %
-Air changes (per hr): The animals were maintained in an air conditioned room. The incoming, non-recycled air, was filtered with an absolute filter.
-Photoperiod (hrs dark / hrs light): the light/dark cycle was 12 hours per day.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
water
Concentration / amount:
2.5 %
Day(s)/duration:
day 0
Route:
epicutaneous, semiocclusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
0.5 g
Day(s)/duration:
day 8 for 48 hours
Challenge
No.:
#1
Route:
epicutaneous, semiocclusive
Vehicle:
water
Concentration / amount:
0.5 g
Day(s)/duration:
day 25 for 24 hours
No. of animals per dose:
10 per sex per dose
Details on study design:
RANGE FINDING TESTS: yes

MAIN STUDY
-INDUCTION EXPOSURE
- No. of exposures: 3 x 2 intradermal injections 0.1 ml each
-Exposure period: day 0
-Test groups: 20 animals (10 ♂ - 10 ♀)
-Control group: 10 animals (5 ♂ - 5 ♀)
-Site: on the scapula on a surface area of 4 x 2 cm (one injection of each solution per flank).
-Concentrations:
Test group: 2.5 %
Control group: vehicle only
-Solutions for each injection site
Test group
1. Freund's complete adjuvant at 50 % in saline (NaCl 9 x 0/00)
2. Test sample at the concentration of 2.5 % in distilled water.
3. A 50/50 (v/v) mixture of Freund's complete adjuvant diluted at 50 % in saline (NaCl 9 x 0/00) and test sample at the concentration previously determined in a preliminary study.

Control group
1. Freund's complete adjuvant at 50 % in saline (NaCl 9 x 0/00)
2. Distilled water.
3. A 50/50 (v/v) mixture of Freund's complete adjuvant diluted at 50 % in saline (NaCl 9 x 0/00) and distilled water.

-CHALLENGE EXPOSURE
-No. of exposures: one
-Day(s) of challenge: day 8
- Test groups: 20 animals (10 ♂ - 10 ♀)
- Control group: 10 animals (5 ♂ - 5 ♀)
-Site: scapular region
-Concentrations:
Test group: 0.5 g
Control group: 0.5 ml of the vehicle
-Evaluation (hr after challenge): 48 h
-Other: On Day 7; 0.5 ml of sodium lauryl sulfate 10 % in codex vaseline were applied on the scapular region to provoke a local irritation.

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 25
- Test groups: 20 animals (10 ♂ - 10 ♀)
- Control group: 10 animals (5 ♂ - 5 ♀)
- Site: flanks
- Concentrations:
Test group: 0.5 g undiluted test sample
Control group: 0.5 ml of the vehicle (distilled water)
- Evaluation (hr after challenge): 24 h
-Other: occlusive patch tests were applied on 4 cm². The patches were kept in place by an hypo-allergenic tape. The right flank was used as negative control.
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:
2.5 % in distilled water (intradermal induction)
No. with + reactions:
0
Total no. in group:
19
Clinical observations:
no change in behaviour nor clinical signs due to treatment were observed during the study
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
2.5 % in distilled water (intradermal induction)
No. with + reactions:
0
Total no. in group:
19
Clinical observations:
no change in behaviour nor clinical signs due to treatment were observed during the study
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no change in behaviour nor clinical signs due to treatment were observed during the study
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
no change in behaviour nor clinical signs due to treatment were observed during the study
Group:
positive control
Remarks on result:
not measured/tested

Any other information on results incl. tables

-Mortality

One male of the treated group was found dead on Day 22.

-Macroscopic and microscopic observation:

Individual macroscopic and microscopic observations are given thereafter.

1. Macroscopic observations

No local reaction was observed on the right flank 24 and 48 h after patch removal. Due to the staining of the skin by the test sample, the scoring of the cutaneous reaction was not recorded of the left flank. No oedematous reaction was observed 24 h and 48 h after patch removal.

2. Microscopic observation:

The slight differences in cutaneous reactions noted between animals from groups I and II were considered to be of no toxicological importance. Occasionnal foci of inflammatory cells in the upper dermis were more frequently observed in animals from group II (more often in the right flank than the left one).

One animal showed an epidermic vesicle (n° 314, group II, left flank). Two others showed lninimal spongiosis (n° 326, group II, right flank n° 329, group II, left flank).

Treated areas were coloured by the test sample. In general, the microscopic observations showed that only ea few keratinized flakes were coloured by the test sample. In two cases (n° 305 and n° 317, group I), they appeared on the control flank due to a technical artefact. In two cases (n° 318, group I - n° 325 group II), they did not appear on any flank.

Applicant's summary and conclusion

Interpretation of results:
other: not classified according to the CLP Regulation (EC 1272/2008)
Conclusions:
Under the experimental conditions, the test item did not provoke any cutaneous sensitization in the guinea pig.
Executive summary:

The sensitizing potential of substance was tested after intradermal injections and cutaneous applications in ten male and ten female Albino Guinea Pigs, to the method established by B. Magnusson and A. Kligman *. The intradermal induction was performed at the concentration of 2.5 % in distilled water. In the epicutaneous induction and in the challenge patch with undiluted test item were used. The behaviour of the animals was not influenced by the treatment. Due to the staining of the skin, no local reaction was noted 24 h and 48 h after removal of the challenge patch test.

Skin samples of both flanks were taken and fixed in formalin 10 %. Histological examination of the skin samples did not reveal evidence of a sensitization reaction.

In conclusion, under the experimental conditions, the test sample did not provoke any cutaneous sensitization in the guinea pig.

 

*MAGNUSSON B. - KLIGMAN A.M. The identification of contact allergens by animal assay "The guinea pig maximization test". J. Invest. Derm. 1969,52,3,268-276