Mixture of reaction products of substituted copper phthalocyanine, hydrogenated resin acids, alkaline earth metal chlorides, and copper phthalocyanine

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 June 2008 - 30 June 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

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

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2009

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(2002)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(2004)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

(2003)

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

B508

IUPAC Name:

B508

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): B508
- Physical state: powder

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. 11 weeks old
- Weight at study initiation: 21-25 g
- Housing: Individual housing in labeled Macrolon cages (MI type; height 12.5 cm) containing sterilized sawdust as bedding material. Paper was supplied as cage-enrichment. The paper was removed on Day 1 prior to dosing and was supplied again after scoring of the ears on Day 3.
- Diet (e.g. ad libitum): Free access to pelleted rodent diet
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: 7-21 days before the start of treatment under laboratory conditions.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.4 – 23.4
- Humidity (%): 39 - 78
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

Preliminary irritation study: 25% and 50%
Main test: 10%, 25%, 50%

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
As no irritation was observed, the 50% dilution was used for the main study.

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.
The EC3 value (the estimated test substance concentration that will give a SI = 3) was determined, using linear interpolation.

TREATMENT PREPARATION AND ADMINISTRATION:
Test substance preparation: The test substance formulations (w/w) were prepared within 4 hours prior to each treatment. Homogeneity was obtained to visually acceptable levels.
Rationale for vehicle: The vehicle was selected based on trial formulations performed at the testing lab and on test substance data supplied by the sponsor.

Induction - Days 1, 2 and 3; Excision of nodes - Day 6; Tissue processing for radioacitivity - Day 6; Radioactivity measurements - Day 7; Performed according to test guidelines.

Observations:
Mortality/Viability: Twice daily.
Body weights: On Day 1 (pre-treatment) and Day 6.
Toxicity: At least once daily.
Irritation: On Day 3 (3-4 hours after treatment), the skin reactions were assessed according to the numerical scoring system. Furthermore, a description of all other (local) effects was recorded according to guidelines.

Positive control substance(s):

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

Statistics:

Not performed.

Results and discussion

Positive control results:

The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Local Lymph Node Assay as performed at the testing laboratory is an appropriate model for testing for contact hypersensitivity. See attached document 'Reliability check'.

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 2.7, 5.5 and 4.3 respectively. These results indicate that the test substance could elicit an SI ≥ 3. An EC3 value of 11.6% was calculated.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

Median DPM/animal values are reported due to possible outlier responses for individual animals within groups. Median DPM/animal values for the experimental groups treated with test substance concentrations 10, 25 and 50% were 797, 1674 and 1299 respectively. The median DPM/animal value for the vehicle control group was 300.

Any other information on results incl. tables

Other results - main study:

Animal 11 (treated at 25%) showed one extremely enlarged node. Since the size of this node was outside the range of this study and occurred in the mid-dose group, the extremely enlarged node of animal 11 was considered not to be related to treatment. Therefore, the results of animal 11 were rejected and not used for interpretation.

No oedema was observed in any of the animals examined. No erythema was noted in the control animals. Blue staining by the test substance prevented scoring for erythema in all animals treated with test substance. Alopecia was noted behind the ears and on the head of all animals at 50%, which was considered not to have a toxicologically significant effect on the activity of the nodes.

The majority of nodes were considered normal in size, except for one or both nodes of one animal at 10%, two animals at 25% and two animals at 50% which were considered enlarged in size.

One animal at 25% showed a cloudy eye which was considered not to have a toxicologically significant effect on the activity of the nodes. No macroscopic abnormalities of the surrounding area were noted in all other animals.

Body weights and body weight gain of experimental animals remained in the same range as controls over the study period.

No mortality occurred and no symptoms of systemic toxicity were observed in the animals of the main study.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information

Conclusions:

In an LLNA skin sensitisation study, performed according to OECD 429 guideline and GLP principles, the substance was considered to be a skin sensitiser, as the SI appeared to be ≥ 3 when tested up to 50%.

Executive summary:

An LLNA test was performed according to OECD 429 guideline and GLP principles. The majority of nodes were considered normal in size, except for one or both nodes of one animal at 10%, two animals at 25% and two animals at 50% which were considered enlarged in size. Median DPM/animal values were reported due to possible outlier responses for individual animals within groups. Median DPM/animal values for the experimental groups treated with test substance concentrations 10, 25 and 50% were 797, 1674 and 1299 respectively. The median DPM/animal value for the vehicle control group was 300. The SI values calculated for the substance concentrations 10, 25 and 50% were 2.7, 5.5 and 4.3 respectively. An EC3 value of 11.6% was calculated.

Based on these results, the test substance should be classified as a skin sensitizer (category 1B) according to Regulation EC 1272/2008 .