Calcium bis[4-[[3-[[2-hydroxy-3-[[(4-methoxyphenyl)amino]carbonyl]-1-naphthyl]azo]-4-methylbenzoyl]amino]benzenesulphonate]

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

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:

2006

Report date:

2006

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

Test material information

Constituent 1

Reference substance name:

Calcium bis[4-[[3-[[2-hydroxy-3-[[(4-methoxyphenyl)amino]carbonyl]-1-naphthyl]azo]-4-methylbenzoyl]amino]benzenesulphonate]

EC Number:

256-050-0

EC Name:

Calcium bis[4-[[3-[[2-hydroxy-3-[[(4-methoxyphenyl)amino]carbonyl]-1-naphthyl]azo]-4-methylbenzoyl]amino]benzenesulphonate]

Cas Number:

43035-18-3

Molecular formula:

C32H26N4O7S.1/2Ca

IUPAC Name:

calcium bis(4-{[3-({2-hydroxy-3-[(4-methoxyphenyl)carbamoyl]-1-naphthyl}diazenyl)-4-methylbenzoyl]amino}benzenesulfonate)

Test material form:

solid: nanoform, no surface treatment

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands, Harlan, The Netherlands
- Age at study initiation: 7-8 weeks
- Weight at study initiation: 19.0 g (17.9g - 21.0 g)
- Housing: individually, Makrolon Type I with mesh top
- Diet: pelleted standard diet, ad libitum
- Water: tap water, ad libitum
- Acclimation: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30- 72
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

0 %, 5 %, 10 %, 20 %

No. of animals per dose:

4 females per dose group,
2 females in the pre-test

Details on study design:

RANGE FINDING TESTS:
- non GLP
- Compound solubility: 20 % (w/v) was the highest concentration at which the test item could be suspended (in acetone:olive oil (4+1)) 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 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
- 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:

- calculation of mean values and standard deviations for body weight .

Results and discussion

Positive control results:

Stimulation indices of 2.04, 2.88 and 3.56 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 12.6 % (w/v) was calculated.

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Remarks on result:

other: Stimulation indices were all below 3. The following SI were calculated: 5 % test item: 1.49 10 % test item: 1.59 20 % test item: 1.84

Results 2

Key result

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

There was a dose dependent increase in the dpm, which were measured for the pooled lymph nodes of each treatment group (8 lymph nodes per dose group): Background I: 18.56 dpm Background II: 19.58 dpm Control group: 7331.05 dpm 5% test item: 10911.90 dpm 10% test item: 11617.40 dpm 20% test item: 13440.40 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. Due to the intense red colour of the test item local irritation reactions such as ear redness could not be examined. 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:

Criteria used for interpretation of results: EU

Conclusions:

The test substance was not sensitising in this LLNA in concentrations up to 20% (w/v) in acetone:olive oil (4+1), the highest technically achievable concentration.

Executive summary:

In the study the test item suspended in acetone:olive oil (4 +1) was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed according to OECD TG 429 using test item concentrations of 5, 10 and 20 % (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 1.49, 1.59 and 1.84 were determined with the test item at concentrations of 5, 10 and 20 % (w/v) in acetone:olive oil, 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.