Reaction mass of 5-[(Disubstituted-dihydrocarbopolycyclyl)diazenyl]-6-hydroxy-4-methyl-2-substituted-1-propyl-1,2-dihydropyridine-3-carbonitrile and 1-Butyl-5-[(disubstituted-dihydrocarbopolycyclyl)diazenyl]-6-hydroxy-4-methyl-2-substituted-1,2-dihydropyridine-3-carbonitrile

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:

2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

OECD Guideline for the Testing of Chemicals, Guideline 429: Skin Sensitization: Local
Lymph Node Assay (adopted 24 April 2002).

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Commission Directive 2004/73/EC, B.42: Skin Sensitization: Local Lymph Node Assay, 29
April 2004.

Deviations:

no

Principles of method if other than guideline:

Guidelines followed

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

solid

Details on test material:

Batch: P2/05 UL
Appearance: light orange crystalline powder
Expiration date: 31.05.2011
Storage: at room temperature

Specific details on test material used for the study:

- Identity: FAT 41039/A
- Description: Light orange crystalline powder
- Batch number: P2/05 UL
- Purity / Formulation: content of organic pigments > 99 %; main components: 97 %
- Stability of test item: Stable under storage conditions.
- Expiry date: 31-MAY-2011
- Stability of test item dilution: Stable in PEG 300 for at least 7 days at room temperature.
- Storage conditions: At room temperature (range of 20 ± 5 °C), light protected.
- Safety precautions: Routine hygienic procedures were used to ensure the health and safety of the personnel.

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaHsdRcc(SPF)

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd, Laboratory Animal Service, CH-4414 Füllinsdorf / Switzerland
- Females- nulliparous and non-pregnant: yes
- Age at study initiation: 8-12 weeks (beginning of acclimatization)
- Weight at study initiation: 16 g - 24 g (ordered)
- Number of animals for the pre-test (non-GLP): 2 females
- Number of animals for the main study: 16 females
- Number of animals per group: 4 females (nulliparous and non-pregnant)
- Number of test groups: 3
- Number of negative control group: 1
- Identification: Each cage by unique cage card.
- Housing: Individual in Makrolon type-2 cages with standard softwood bedding ("Lignocel", Schill AG, CH-4132 Muttenz).
- Diet: Pelleted standard Kliba 3433, batch no. 25/05 mouse maintenance diet (Provimi Kliba AG, CH-4303 Kaiseraugst) available ad libitum.
- Water: Community tap water from Itingen, available ad libitum.
- Acclimation period: 7 d
- Indication of any skin lesions: no

ENVIRONMENTAL CONDITIONS
- Temperature: 22 + 3 °C,
- Humidity: 30 - 70 %
- Air changes: 10-15 per h
- Photoperiod: 12 h dark / 12 h light
- IN-LIFE DATES: From: 03-AUG-2005; To: 17-AUG-2005

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Remarks:

Polyethylene glycol 300

No. of animals per dose:

4

Results and discussion

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

1.8

Test group / Remarks:

Group 2 (0.25% concentration)

Results 2

Key result

Parameter:

SI

Value:

1.4

Test group / Remarks:

Group 3 (0.5% concentration)

Results 3

Key result

Parameter:

SI

Value:

1.4

Test group / Remarks:

Group 4 (1% concentration)

Any other information on results incl. tables

STIMULATION INDEX

STIMULATION INDICES of 1.8, 1.4 and 1.4 were determined with the test item at concentrations of 0.25 %, 0.5 % and 1 % (w/v), respectively, in polyethylene glycol 300. No dose-response relationship was observed. Calculation of the EC 3 value was not performed because no test concentrations produced a STIMULATION INDEX (S.I.) of 3 or higher.

VIABILITY / MORTALITY

No deaths occurred during the study period.

CLINICAL SIGNS

No clinical signs of local toxicity at the ears of the animals and no systemic findings were observed during the study period.

BODY WEIGHTS

The body weight of the animals, recorded prior to the first application and prior to necropsy, was within the range commonly recorded for animals of the strain and age.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

FAT 41039/A was found to be a non-sensitizer when tested up to the highest applicable concentration of 1 % (w/v) in polyethylene glycol 300.

Executive summary:

In order to study a possible contact allergenic potential of FAT 41039/A, three groups each of four female mice were treated daily with the test item at concentrations of 0.25, 0.5 and 1 % (w/v) in polyethylene glycol 300 by topical application to the dorsum of each ear lobe (left and right) for three consecutive days. 1 % (w/v) was the highest technically applicable concentration in the vehicle. A control group of four mice was treated with the vehicle polyethylene glycol 300 only. Five days after the first topical application the mice were injected intravenously into a tail vein with radio-labelled thymidine (3H-methyl thymidine). Approximately five hours after intravenous injection, the mice were sacrificed, the draining auricular lymph nodes excised and pooled per group. Single cell suspensions oflymph node cells were prepared from pooled lymph nodes which were subsequently washedand incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3H-methyl thymidine measured in a ßscintillation counter. All treated animals survived the scheduled study period. No clinical signs were observed.

A test item is regarded as a sensitizer in the LLNA if the exposure to one or more test concentrations resulted in 3-fold or greater increase in incorporation of 3HTdR compared with concurrent controls, as indicated by the STIMULATION INDEX (S.I.)- In this study STIMULATION INDICES of 1.8, 1.4 and 1.4 were determined with the test item at concentrations of 0.25 %, 0.5 % and 1 % (w/v), respectively, in polyethylene glycol 300. FAT 41039/A was therefore found to be a non-sensitizer when tested up to the highest applicable concentration of 1 % (w/v) in polyethylene glycol 300.