Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 28 to August 25, 2010
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Although the study is performed according to current OECD test guidelines and GLP principles, the result of the positive and negative control show limited validity, where the historical data confirm the (sensitising/not sensitising) results of this study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
yes
Remarks:
see below
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): E-C104
- Physical state: Blue powder, solid
- Lot/batch No.: MB-1
- Expiration date of the lot/batch: July 6, 2015
- Storage condition of test material: Room temperature, shielded from light, tight container

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/JNCrlj
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc.
- Age at study initiation: 8 weeks
- Weight at study initiation: 19.9 to 23.6 g
- Housing: Polycarbonate cages (92Wx205Dx127H mm) with after radio-isotope administration (220Wx325Dx130H mm) stainless steel mesh flooring, (stainless) steel racks, one animal per cage and after administration of radioisotope 6 animals per cage.
- Diet (ad libitum): Pellet diet (MF, Oriental Yeast Co., Ltd.), analysed: contaminants were confirmed to be within acceptable limits established by the test facility.
- Water (ad libitum): Tap water filtered through a 5 microm filter and disinfected by UV irradiation, analysed twice a year: contaminants were confirmed to be within acceptable limits establised by the test facility.
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.2-23.7
- Humidity (%): 47.7-67.5
- Air changes (per hr): 6 to 30
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Remarks:
=AOO
Concentration:
0, 5, 15, 50 w/v%.
No. of animals per dose:
6
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: The dosing suspension was stirred with a magnetic stirrer throughout the application procedure.
- Irritation:25 microl of the maximum feasible dose, 50 w/v%, was applied to two mice once a day for two days. This revealed no abnormalities in either animal, hence the maximum concentration was set at 50 w/v%.
- Lymph node proliferation response: no data.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: LLNA
- Criteria used to consider a positive response: Stimulation Index (SI) value of 3 or greater was judged to be positive.

TREATMENT PREPARATION AND ADMINISTRATION:
Each animal was applied 25 microliter to one side of both auricular using a micropipette. This was performed once daily for three consecutive days. The radioisotope was admnistered (250 microliter/body) by injection five days after the initial auricular application.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Numerical data in the test substance groups were analysed by a multiple comparison test. Homogeneity of the variance among the groups was first tested by Bartlett's test. When variance was homogenous, which was, the one way analysis of variance was applied. The numerical data in the positive control group against the vehicle group was examined for homogeneity of the variance by F test and Welch test.

Results and discussion

Positive control results:
HCA group had a radioactivity mean of 3979.8 dpm (range 1364-6472 dpm), SI value was 2.85. Althought SI<3, this mean dpm value was normal compared to historical data with mean of 2525 and range of 507.2 to 7148.0 dpm. See also overall remarks.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: SI of 5 w/v% froup is 0.77, of 15% group is 0.44 and of 50% group is 0.53.
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Mean radioactivity values of AOO group is 1396.8 dpm (range 980 to 2104 dpm), of 5 w/v% group is 1076.8 dpm (range 755 to 1597 dpm), of 15% group is 616.0 dpm (range 420 to 735 dpm), of 50% group is 746.5 dpm (range 323 to 1090 dpm).

Any other information on results incl. tables

No abnormal clinical signs or lymph node weight or body weight were observed in any of the animals throughout the study period.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
E-C104 has no sensitising potential under the conditions of this LLNA study (OECD 429).
Executive summary:

The sensitisation potential of E-C104 was assessed in 30 female mice using the LLNA study performed according to OECD 429. E-C104 was suspended in a mixture of acetone/olive oil (4:1) to make three concentrations of test substance: 5, 15 and 50 w/v%. HCA 25 v/v% was used as positive control and the vehicle was used as negative control. The SI values were 0.77, 0.44 and 0.53 in the 5, 15 and 50 w/v% test substance groups, respectively. The SI value of HCA was 2.85, was slightly less than the criterion (SI value>3). However, the negative control showed very high dpm values compared to the historical control data, resulting in a lower SI value for the positive control. In addition, the test groups would still have an SI<3 and no dose related increase is observed. Therefore, the study is considered valid. No abnormalities in body weight, lymph node weight or clinical signs were observed in any of the animals. From the results, it is concluded that E-C104 has no sensitisation potential under the conditions of this study.