Coupling reaction products of 3,3’-dichlorobiphenyl-4,4’-bis(diazonium) dichloride with N-(2,4-dialkylphenyl)-3-oxobutanamide and potassium 4-[(3-oxobutanoyl)amino]substituted benzene and their basic aluminium and earth alkali metal salts

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 December 2007-14 January 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted according to OECD and EC guidelines and according to GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material (as cited in study report): S-500
- Physical state: yellow powder
- Lot/batch No.: MF1456-2345
- Expiration date of the lot/batch: 18 August 2011
- Storage condition of test material: At room temperature in the dark
- Stability under storage conditions: Stable

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River France, L’Arbresle Cedex, France
- Age at study initiation: approx. 9 weeks old
- Weight at study initiation: 19-23g
- Housing: Individual housing in labeled Macrolon cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.8 – 23.1
- Humidity (%): 42 - 69
- Air changes (per hr): approximately 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

0, 10, 25 and 50%

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: Homogeneity was obtained to visually acceptable levels.
- Irritation: No irritation was observed in any of the animals examined.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: local lymph node assay
- Criteria used to consider a positive response: DPM values are presented for each animal and for each dose group. A Stimulation Index (SI) is calculated for each group. The SI is the ratio of the DPM/group compared to DPM/vehicle control group. If the results indicate a SI >= 3, the test substance may be regarded as a skin sensitizer, based on the test guideline and recommendations done by ICCVAM.


TREATMENT PREPARATION AND ADMINISTRATION: Three groups of five experimental animals were treated with test substance concentrations of 10%, 25% or 50% on three consecutive days, by open application on the ears (25 µL/ear). Five vehicle control animals were similarly treated, but with vehicle alone (Propylene glycol ).
Three days after the last exposure, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes were excised.
After precipitating the DNA of the lymph node cells, radioactivity measurements were performed.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:

The SI values calculated for the substance concentrations 5, 10 and 25% were 1.0, 2.0 and 5.7 respectively. An EC3 value of 14.1% was calculated using linear interpolation.
The calculated EC3 value was found to be in the acceptable range of 2 and 20%. The results of the 6 monthly HCA reliability checks of the recent years were 10.3, 9.5, 13.1 and 15.6%.
Based on the results, it was concluded that the Local Lymph Node Assay as performed at NOTOX is an appropriate model for testing for contact hypersensitivity.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: The SI values calculated for the substance concentrations 10, 25 and 50% were 0.6, 1.1 and 0.7 respectively.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Mean DPM/animal values for the experimental groups treated with test substance concentrations 10, 25 and 50% were 173, 330 and 209 respectively. The mean DPM/animal value for the vehicle control group was 310.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

Since there was no indication that the test substance could elicit an SI >= 3 when tested up to 50%, it was established that the EC3 value (if any) exceeds 50%. Therfore, the test substance S-500 does not have to be classified and has no obligatory labeling requirement for sensitization by skin contact.