Polychloro copper phthalocyanine

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

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)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Polychloro copper phthalocyanine

EC Number:

215-524-7

EC Name:

Polychloro copper phthalocyanine

Cas Number:

1328-53-6

Molecular formula:

C32HxClyCuN8

IUPAC Name:

[1,2,3,4,8,9,10,11,15,16,17,18,22,23,25-pentadecachloro-5,26-dihydro-29H,31H-phthalocyaninato(2-)-kappa~2~N~29~,N~31~]copper

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): Pigment Green 7, C.I. 74260

Specific details on test material used for the study:

Green solid
Batch DEBF 015527
Stable under storage conditions
Expiry data 31-MAR-2006
Purity: >95%

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands
- Age at study initiation: 8 - 12 weeks
- Weight at study initiation: 16 g - 24 g (ordered)
- Housing: individually in Makrolon type-2 cages with standard softwood bedding
- Diet: Pelleted Standard Kliba 3433, batch no. 78/03 mouse maintainance diet (Provimi Kliba AG, Kaiseraugst, Switzerland), ad libitum
- Water: tap water, ad libitum
- Acclimation period: under test conditions after haelth examination; only animals without any visible signs of illness were used for the study

ENVIRONMENTAL CONDITIONS
- Temperature: 22 +- 3 °C
- Humidity: 30 - 70 %
- Air changes per hr: 10 - 15
- Photoperiod: 12 hrs dark / 12 hrs light

Study design: in vivo (LLNA)

Vehicle:

dimethyl sulphoxide

Concentration:

5, 10 and 25%

No. of animals per dose:

4 animals per dose

Details on study design:

RANGE FINDING TESTS:
To determine the highest non-irritant and technically applicable test item concentration, a non-GLP test was conducted in 2 mice with concentrations of 5, 10 and 25 %. The top dose is the highest technically achievable concentration.

MAIN STUDY:
Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear lobe (left and right) with different test item concentrations of 5 %, 10 % and 25 % (w/v) in dimethylsulfoxide (DMSO). The application volume, 25 µI, was spread over the entire dorsal sutiace (0 - 8 mm) of each ear lobe once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals). A hair dryer was passed briefly over the ear's surface to prevent the loss of any of the test item applied. Five days after the first topical application, all mice were administered with 250 µI of
79.6 µCi/ml 3HTdR (equal to 19.9 µCi 3HTdR) by intravenous injection via a tailv ein.
Approximately five hours after treatment with 3HTdR all mice were euthanized with dry ice (C02).
The draining lymph nodes were rapidly excised and pooled for each experimental group (8 nodes per group). Single cell suspensions (phosphate buffered saline) of pooled lymph node cells were prepared by gentle mechanical disaggregation through stainless steel gauze (200 µm mesh size). After washing two times with phosphate buffered saline (approx. 10 ml) the lymph node cells were resuspended in 5 % trichloroacetic acid (approx. 3 ml) and incubated at approximately +4 °C for at least 18 hours for precipitation of macromolecules. The precipitates were then resuspended in 5 % trichloroacetic acid (1 ml) and transferred to glass scintillation vials with 10 ml of 'Ultima Gold' scintillation liquid and thoroughly mixed.
The level of 3HTdR incorporation was then measured on a ß-scintillation counter. Similarly, background 3 HTdR levels were also measured in two 1ml-aliquots of 5 % trichloroacetic acid. The ß-scintillation counter expresses 3HTdR incorporation as the number of radioactive disintegrations per minute (DPM).

Positive control substance(s):

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

Statistics:

A test item was considered as sensitizer if the following criteria were fulfilled:
- Exposure to at least one concentration resulted in an incorporation of 3HTdR of at least 3-fold or greater than in controls as indicated by S.I.
- Data were compatible with a conventional dose-response, although allowance must be for either local toxicity or immunological suppression.

For statistical analysis, the mean values and standard deviations were calculated in the body weight tables.

Results and discussion

Positive control results:

Results with the latest postive control study are attached to the report (RCC Study Number 852080),

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

1.4

Test group / Remarks:

5%

Results 2

Parameter:

SI

Value:

1.3

Test group / Remarks:

10%

Results 3

Parameter:

SI

Value:

1.3

Test group / Remarks:

25%

Cellular proliferation data / Observations:

No deaths occured during the study period.
No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period.
The body weight of the aninals was within the normal range recorded for animals of this strain and age.

Any other information on results incl. tables

Test item concentration %( w/v)		Measurement dpm	Calculation			Result
			dpm - BG a)	number of lymph nodes	dpm per lymph node b)	S.I.
--	BG1	10	--	--	--	--
--	BG II	3	--	--	--	--
--	CG 1	5238	5231	8	654	--
5	TG 2	7418	7411	8	926	1.4
10	TG 3	6791	6784	8	848	1.3
25	TG 4	6816	6809	8	851	1.3

BG=Background (1 ml 5 % trichloroacetic acid) in duplicate

CG = Control Group

TG = Test Group

S.I. = Stimulation Index

a)      = The mean value was taken from the figures BG I and BG II

b)      = Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled

The positive control, alpha-hexylcinnamaldehyde, was tested in concentrations of 5.0, 10.0 and 25.0 % and produced S.I.s of 1.5, 2.3 and 8.4, respectively.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met