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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2 NOV 2004 to 24 NOV 2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline Study (OECD TG 429)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report Date:
2005

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
according to German Chemical Law and OECD Principles of GLP
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- strain: CBA/CaOlaHsd
- Source: Harlan Netherlands
- Age at study initiation: 8-12 weeks (beginning of acclimatization)
- Weight at study initiation: mean: 18.9 +/- 2.0 g
- Housing: individually, Makrolon Type I cages
- Diet: pelleted standard diet (Harlan Winkelmann, Borchen), ad libitum
- Water: tap water, ad libitum
- Acclimatization: yes (acclimatization period not given)


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/-3
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12 hrs/12 hrs

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
5, 10, 20 % (w/v)
No. of animals per dose:
4 females
Details on study design:
RANGE FINDING TESTS:
- non GLP
- Compound solubility: 10% (w/v) suspension in acetone:olive oil (4:1 v/v) was the highest technically applicable concentration; higher concentrations could also not be achieved with other vehicles
- Irritation: irritation effects could not be determined due to the intense red colour of the test item. No ear swelling was observed at these concentrations after a single application
- Lymph node proliferation response: no data


MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: LLNA
- Criteria used to consider a positive response:
1: exposure to at least one concentration of the test item resulted in an incorporation of 3H-methyl thymidine at least 3-fold or greater than that recorded in control mice as indicated by the stimulation index
2: data are compatible with a conventional dose response, although allowance must be made for either local toxicity or immunological suppression


TREATMENT PREPARATION AND ADMINISTRATION:
- individual preparation of weight volume dilutions using a magnetic stirrer as homogenizer
- test item preparations were made freshly before each dosing occasion
- the application volume of 25 µl was spread over the entire dorsal surface of each ear lobe once daily for three consecutive days
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
- calculation of mean values and standard deviations for body weight
- statistical evaluation of the dose-response relationship

Results and discussion

Positive control results:
Stimulation indices of 2.0, 3.0 and 4.9 were determined in a separate test with the positive control substance at concentrations of 5%, 10% and 25% (w/v), respectively, in acetone:olive oil. An EC3 value of 9.9% (w/v) was calculated.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: 5%: 2.1 10%: 1.8 20%: 1.6
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Background: 0; 4.7 (duplicate) Control: 2584.5 (2.582.2 without background) 5%: 5397.8 (5395.5 without background) 10%: 4542.4 (4540.1 without background) 20%: 4183.7 (4184.4 without background)

Any other information on results incl. tables

No deaths occurred during the study period.

The body weight of the animals, recorded prior to the 1stapplication and prior to necropsy,

was within the range commonly recorded for animals of this strain and age.

No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period. Due to the colour of the test item, a redness of the ears could not be analysed. However, no signs of ear swelling occurred.  

Calculation of the EC3 value was not performed, because no test concentration produced a SI of 3 or higher.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information
Conclusions:
Under the conditions of this LLNA-assay, the test substance was not sensitising.
Executive summary:

In the study the test item dissolved in acetone:olive oil, 4:1 (v/v) was assessed for its possible contact allergenic potential. For this purpose a local lymph node assay was performed in CBA mice (4 females per group) using test item concentrations of 5, 10 and 20% (w/v). The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. In this study stimulation indices (S.I.) of 2.1, 1.8 and 1.6 were determined with the test item at concentrations of 5, 10 and 20% (w/v) in acetone:olive oil, 4:1 (v/v), respectively. A EC3 value could not be determined. The test item was not a skin sensitiser under the conditions tested. The positive control substance was sensitising (EC value 9.9%), thus confirming the validity of the tests system.

Therefore, the test item has not to be classified as skin sensitiser according to Regulation (EC) No 1272/2008.