Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
15 June 1995 to 21 July 1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, performed to valid guidelines and the study was conducted under GLP conditions.

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
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): M-377
- Physical state: solid
- Appearance: garnet powder
- Storage condition of test material: room temperature and protected from light

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: approximately 3 months
- Weight at study initiation (mean (standard deviation)): 338 (30) g males; 312 (27) g females
- Housing: individually in polycarbonate cages
- Diet: diet provided ad libitum
- Water: filtered drinking water (0.22 µm), ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2 °C
- Humidity (%): 30 - 70 %
- Air changes (per hr): 12 cycles per hour of filtered, non-recycled air.
- Photoperiod (hrs dark / hrs light): 12 hours dark / 12 hours light


IN-LIFE DATES: From 15 June 1995 to 21 July 1995

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
other: isotonic saline solution (0.9 % NaCl)
Concentration / amount:
Induction
- intradermal: 10 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)
- topical: 10 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)

First challenge:
- topical: 5 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)

Second challenge:
- topical: 1 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: isotonic saline solution (0.9 % NaCl)
Concentration / amount:
Induction
- intradermal: 10 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)
- topical: 10 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)

First challenge:
- topical: 5 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)

Second challenge:
- topical: 1 % (w/w) in sterile isotonic saline solution (0.9 % NaCl)
No. of animals per dose:
5 males and 5 females (control group)
10 males and 10 females (treatment group)
Details on study design:
RANGE FINDING TESTS
A preliminary study was conducted to inform on test concentrations to be used in the main study. The maximal practicable concentration by both the intradermal, and cutaneous route, was 10 % and this was selected for the main study.

MAIN STUDY
On day 1, an area of 4 cm x 2 cm in the dorsal region between the shoulders was clipped free of fur. Three injections of 0.1 mL were made in this region as follows:

Injection Site Treated group Control group
Anterior 1: FCA diluted at 450 % (v/v) with 0.9% NaCl 1: FCA diluted at 450 % (v/v) with 0.9% NaCl
Middle 2: test material at 10 % (w/w) in vehicle 2: vehicle
Posterior 3: mixture of 50/50 (w/v) of 1 and 2 3: mixture of 50/50 (w/v) of 1 and 2

On day 7, the same region received a topical application of 0.5 mL of sodium laurylsulphate in vaseline (10 % w/w) in order to induce local irritation.

On day 8, the same test site was treated by topical application of 0.5 mL test material at the chosen concentration (treatment group) or the vehicle (control group) and was covered by an occlusive dressing, for 48 hours. On removal of the dressing, any residual test material was removed using a moistened gauze pad.

After a rest period of 12 days, all animals of the treatment and control groups were challenged by a topical application of the test material (0.5 mL) on the right flank. The left flank served as the control and received the vehicle (0.5 mL) only. Test material and vehicle were maintained under an occlusive dressing for 24 hours. On removal of the dressing, any residual test material was removed using a moistened gauze pad. Skin reactions were evaluated approximately 24 and 48 hours after the challenge application according to the scale outlined in the field "Any other information on materials and methods incl. tables", and any other lesions were recorded.

After a rest period of 10 days, a second challenge application was performed under the same experimental conditions. Where the test material and the vehicle were applied to the left and right flanks, respectively. Skin reactions were evaluated approximately 24 and 48 hours after the challenge application and any other skin lesions wee

During the study the animals were observed for clinical signs and mortality twice a day. Individual bodyweights were recorded on study days 1, 8, 15, 25 and on the last day of the study.

At the end of the study, animals were killed. Macroscopic examination of the main organs was performed on the animal found dead during the study. No necropsy on the surviving animals was performed and no skin samples were taken from the challenge application sites.
Challenge controls:
During the challenge applications the left flank served as a control and was treated topically with the vehicle only (0.5 mL).
Positive control substance(s):
yes
Remarks:
2,4-dinitro-1-chlorobenzene

Results and discussion

Positive control results:
A positive response was observed in 95 % of animals treated with the positive control (1% w/w) during a concurrent study.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
48
Group:
negative control
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: negative control. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

Clinical examinations

- No clinical signs were observed during the study. In the treated group, one animal was found dead on day 33. Such spontaneous mortality is often observed in guinea-pigs and it was not thought to be related to treatment.

- The body weight gain of the treated animals was normal when compared to the controls.

Scoring cutaneous reactions

- On day 10, after topical application of the induction period, signs of irritation were observed at the test site in the control group. In the treatment group it was not possible to detect signs of dermal irritation due to the test site colouration.

- No cutaneous reactions were observed after both challenge applications. A red colouration of the test site which could mask erythema grade 1 or 2 was noted in all animals after the first challenge application.

- After the second challenge application, a slight colouration which could mask a very slight erythema (grade 1) was observed in 3/10 and 8/19 animals of the control and treated groups, respectively. This was noted at the 24 hour reading only.

Necropsy

- Macroscopic examination of the animal found dead during the study revealed no abnormalities.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of the study, no cutaneous reactions, which could be attributable to the sensitisation potential of the test material, were observed. The study is considered to be reliable, relevant and adequate for risk assessment and classification and labelling purposes.
Executive summary:

The skin sensitisation potential of the test material was determined in accordance with standardised guidelines OECD 406 and EU Method B.6 using the Guinea-pig Maximisation Test. The test material was dosed as a 10 % w/w preparation in isotonic saline solution in an intradermal and topical induction phase. Challenge applications were performed topically with a 5 % w/w application made during the first challenge and a 1 % w/w application made during the second challenge. The positive control was shown to have the capacity to cause skin sensitisation confirming the validity of the protocol used for this study. Under the conditions of the study, no cutaneous reactions, which could be attributable to the sensitisation potential of the test material, were observed.