[N,N,N',N',N'',N''-hexaethyl-29H,31H-phthalocyaninetrimethylaminato(2-)-N29,N30,N31,N32]copper

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and OECD guideline compliant studies with well characterized test material.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Experimental Toxicology and Ecology, BASF SE, Germany

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

[N,N,N',N',N'',N''-hexaethyl-29H,31H-phthalocyaninetrimethylaminato(2-)-N29,N30,N31,N32]copper

EC Number:

249-125-4

EC Name:

[N,N,N',N',N'',N''-hexaethyl-29H,31H-phthalocyaninetrimethylaminato(2-)-N29,N30,N31,N32]copper

Cas Number:

28654-73-1

Molecular formula:

C47 H49 Cu N11

IUPAC Name:

Copper,[N,N,N',N',N'',N''-hexaethyl-29H,31H-phthalocyanine-C,C,C-trimethanaminato(2-)-κN29,κN30,κN31,κN32]-

Details on test material:

- Analytical purity: The test substance has been characterized by analyses.
- Stability: The stability under storage conditions over the study period was guaranteed by the sponsor. The stability of the test substance in the vehicle for the maximum application period was confirmed indirectly by analysis of the homogeneity / correctness of the concentration.
- Homogeneity: The test substance is homogeneous by visual inspection. Additionally, homogeneity / correctness of the concentrations were confirmed by analysis.

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sulzfeld, Germany
- Age at study initiation: 8 - 12 weeks
- Weight at study initiation: 18.3 g – 23.8 g
- Housing: single housing in Makrolon cage, type II
- Diet: Kliba-Labordiaet (Maus / Ratte Haltung "GLP"), Provimi Kliba SA, Kaiseraugst, Basel, Switzerland, ad libitum
- Water: Tap water ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24°C
- Humidity (%): 30 - 70%
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

5, 10, and 25%

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: A solubility experiment was performed according to the recommendations given by OECD 429. The highest test-substance concentration which can be technically used was a 25% test-substance preparation. The test-substance preparations at different concentrations were formulated in propylene glycol.
- Irritation: To determine the highest test-substance concentration that does not induce local signs of skin irritation and/or systemic toxicity, a pre-test (experimental conduct in accordance with GLP but without a GLP status) was performed. Two mice were treated with a 25% test-substance concentration on three consecutive days. In the pre-test clinical signs were recorded after each application as well as on day 5. Signs of local irritation were recorded on day 1, 2 and 5. Furthermore, the ears were punched after sacrifice at the apical area using a biopsy punch (Ø 0.8 cm) and were immediately pooled per animal and weighed using an analytical balance. Additionally the weight of the pooled lymph nodes from both sides was determined for each animal. At the tested concentration of 25% the animals showed some increases in ear weights and lymph node weights. Moreover compound residues on the application area and blue discolored ear skin were noted during the whole observation period. Signs of systemic toxicity were not observed in the pre-test.
Therefore, the following dose levels were selected for the main study: 5%, 10% and 25%.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Murine Local Lymph Node Assay
- Criteria used to consider a positive response: increase in 3H-thymidine incorporation by a factor of >/= 3, cell-count stimulation index (SI >/= 1.5), lymphnode-weight stimulation index and ear weight stimulation index together with dose-response.

TREATMENT PREPARATION AND ADMINISTRATION: The test-substance preparation was produced on a weight per weight basis shortly before the application by stirring with a high speed homogenizer (Ultra-Turrax) and/or a magnetic stirrer. The homogeneity of the test-substance preparation during application was provided by stirring with a magnetic stirrer. Propylene glycol was used as the vehicle because good homogeneity of the preparations was achieved. 25 μL per ear were applied on 3 consecutive days (day 0 – day 2) to the same application site.

Positive control substance(s):

other: A positive control is not included in the study. A separate study with a positive control (Alpha-Hexylcinnamaldehyde, techn. 85%) is carried out in the laboratory twice a year.

Statistics:

Mean values and standard deviations of the measured parameters were calculated for the test and control groups from the individual values. The stimulation indices of 3H-thymidine incorporation, cell count, lymph node weight and ear weight measurements were calculated as the ratio of the test group mean values for these parameters divided by those of the vehicle control group. Further statistical analyses were conducted: 3H-thymidine incorporation, cell count, lymph node weight and ear weight (WILCOXON - Test).

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: vehicle: 1.00 5%: 1.579 (statistically significant for the value p ≤ 0.05) 10%: 2.55 (statistically significant for the value p ≤ 0.01) 25%: 3.44 (statistically significant for the value p ≤ 0.01)

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: dpm / lymph node pair: vehicle: 238 5% 385.1 10%: 626.4 25%: 845.7

Any other information on results incl. tables

Cell Count Stimulation Index

	Mean [Counts / lymph node pair]	Stimulation Index
vehicle	8,098,800	1.00
5%	8,752,800	1.04
10%	10,173,600	0.99
25%	11,318,400	1.34#

Ln-Weight Stimulation Index

	Mean [mg]	Stimulation Index
vehicle	5.2	1.00
5%	5.4	1.08#
10%	6.0	1.26##
25%	6.5	1.40##

Ear-Weight Stimualtion Index

	Mean [mg]	Stimulation Index
vehicle	28.5	1.00
5%	31.9	1.08
10%	31.8	1.17#
25%	34.0	1.26##

# = statistically significant for the value p ≤ 0.05

## = statistically significant for the value p ≤ 0.01

No signs of systemic toxicity were noticed in all animals during general observation. Blue stained ear skin was observed at all test substance concentrations during the whole observation period.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information GHS Cat 1B Criteria used for interpretation of results: EU