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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 14 MAY 2003 to 28 MAY 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (OECD 429 and GLP).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report date:
2003

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Constituent 1
Chemical structure
Reference substance name:
4-[(2,5-dichlorophenyl)azo]-3-hydroxy-N-phenylnaphthalene-2-carboxamide
EC Number:
227-930-1
EC Name:
4-[(2,5-dichlorophenyl)azo]-3-hydroxy-N-phenylnaphthalene-2-carboxamide
Cas Number:
6041-94-7
Molecular formula:
C23H15Cl2N3O2
IUPAC Name:
4-[(2,5-dichlorophenyl)diazenyl]-3-hydroxy-N-phenyl-2-naphthamide
Test material form:
solid: nanoform, no surface treatment
Details on test material:
Name of test material (as cited in study report): Pigment Composition P.R.2 Trial AK 503
Analytical purity: 100 % Permanent Red FRR

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Netherlands, Horst, The Netherlands
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 17.4 - 20.9 g
- Housing: in groups of four
- Diet: pelleted standard Kliba 3433, batch no. 84/02 mouse maintenance diet, ad libitum
- Water: tap water, ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +-3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
0, 2.5, 5 and 10 % (w/v)
No. of animals per dose:
4 females
Details on study design:
RANGE FINDING TESTS:
- non GLP
- Compound solubility: 10 % (w/v) was the highest technically achievable concentration in acetone:olive oil (4:1 v/v) so that an applicable test item formulation was obtained. Higher concentrations could also not be achieved with other vehicles.
- Irritation: No irritation effects were observed after single application of test item concentrations of 1%, 2.5%, 5% and 10%
- 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
- 25 µl were 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:
The mean values and standard deviations for body weights were calculated.

Results and discussion

Positive control results:
Stimulation indices of 2.5, 3.7 and 9.7 were determined with the positive control substance at concentrations of 5 %, 10 % and 25 % (w/v), respectively, in acetone:olive oil (4+1). An EC3 value of 7.08 % (w/v) was calculated.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Value:
ca. 2.1
Test group / Remarks:
Concentration 10%
Parameter:
SI
Value:
ca. 2.7
Test group / Remarks:
Concentration 5%
Parameter:
SI
Value:
ca. 1.8
Test group / Remarks:
Concentration 2.5%
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: The DPM were measured for the pooled lymph nodes of each treatment group (8 lymph nodes per dose group): Background I: 3 dpm Background II:1 dpm Control group: 5699 dpm 2.5 % test item: 10425 dpm 5 % test item: 15632 dpm 10 % test item: 11745 dpm

Any other information on results incl. tables

The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. The body weight of the animals was within the normal range.

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 Criteria used for interpretation of results: EU
Conclusions:
The test substance was not sensitising in this LLNA in concentrations up to 10 % (w/v) in acetone-olive oil, the highest achievable concentration.
Executive summary:

In this guideline study (according to OECD 429) the test item suspended 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 using test item concentrations of 2.5 %, 5 %, and 10 % (w/v).

The animals (4 female mice/dose group) 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 1.8, 2.7 and 2.1 were determined with the test item at concentrations of 2.5 %, 5 % and 10 % (w/v) in acetone:olive oil (4:1, v/v), respectively. The results obtained with the positive control confirmed the validity of the test.

The test item was not a skin sensitiser in this assay.

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