Perylene-3,4:9,10-tetracarboxydiimide

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:

1999

Report date:

1999

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Perylene-3,4:9,10-tetracarboxydiimide

EC Number:

201-344-6

EC Name:

Perylene-3,4:9,10-tetracarboxydiimide

Cas Number:

81-33-4

Molecular formula:

C24H10N2O4

IUPAC Name:

7,18-diazaheptacyclo[14.6.2.2²,⁵.0³,¹².0⁴,⁹.0¹³,²³.0²⁰,²⁴]hexacosa-1(23),2,4,9,11,13,15,20(24),21,25-decaene-6,8,17,19-tetrone

Details on test material:

Test materials used in this dossier are all considered to fall under the definition of nano-materials according to the European Commission Recommendation 2011/696/EU as the synthesis and manufacturing of this pigment always yields particulate material with a fine particle size distribution.

Specific details on test material used for the study:

- Physical state: solid/powder
- Analytical purity: 98.9%

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan UK, Blackthorne, Bicester, Oxon, UK
- Age at study initiation: young adults
- Housing: cages suitable for animals of this strain and weight range (max. of 4 mice per cage)
- Diet (e.g. ad libitum): R&M No.1 (Special Diet Services Limited, Witham, Essex, UK), ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: The animals were housed under the experimental conditions for at least 5 days, prior to the start of the study .

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3
- Humidity (%): 30-70
- Air changes (per hr): A minimum of 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

the test substance was applied as 3%, 10% or 30% w/v preparations in propylene glycol

No. of animals per dose:

4

Details on study design:

TREATMENT PREPARATION AND ADMINISTRATION:
Groups of four male mice were used for this study. Approximately 25μ1 of a 3%, 10% or 30% w/v preparation of the test substance in propylene glycol was applied, using a variable volume micro-pipette, to the dorsal surface of each ear. A vehicle control group was similarly treated using propylene glycol alone. The procedure was repeated daily for 3 consecutive days. A concurrent naïve control group was not treated with the test substance or the vehicle .
Three days after the third application, all the animals were injected, via the tail vein, with approximately 250 μl of phosphate buffered saline (PBS) containing approximately 20 μCi of a 2.OCi/mmol specific activity 3H-methyl thymidine . Approximately 5 hours later, the animals were humanely killed by inhalation of halothane vapour followed by cervical dislocation. The draining auricular lymph nodes were removed from each animal and, together with the nodes from the other animals in the group, were placed in a container of PBS .
A single cell suspension was prepared by mechanical disaggregation of lymph nodes through a 200-mesh stainless steel gauze . The cell suspensions were then washed three times by centrifugation with approximately 10 ml of PBS. Approximately 3 ml of 5% w/v trichloroacetic acid (TCA) was added and after overnight precipitation at 4°C, the samples were pelleted by centrifugation and the supernatant was discarded. The cells were then resuspended in approximately 1 ml of TCA .
The lymph node suspensions were transferred to scintillation vials and 10 mL of scintillant (Optiphase) was added prior to ß-scintillation counting using a Packard Tri-Carb 2500TR Liquid Scintillation Counter.

CLINICAL OBSERVATIONS:
Animals were checked at least once daily for signs of systemic toxicity.

CRITERIA USED TO CONSIDER A POSITIVE RESPONSE:
The criterion for a positive response is that one or more concentrations of the test substance should elicit a 3-fold or greater increase in isotope incorporation relative to the vehicle control group.

Positive control substance(s):

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

Results and discussion

Positive control results:

The application of hexylcinnamaldehyde at concentrations of 1%, 3% and 10% w/v in acetone resulted in a greater than 3-fold increase in isotope incorporation at the 10% w/v concentration. Therefore, hexylcinnamaldehyde was shown to be a skin sensitiser, confirming the validity of the protocol used for the study.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

1.16

Test group / Remarks:

3%

Results 2

Parameter:

SI

Value:

1.61

Test group / Remarks:

10%

Results 3

Parameter:

SI

Value:

1.31

Test group / Remarks:

30%

Results 4

Parameter:

other: disintegrations per minute (DPM)

Value:

>= 2 322 - <= 3 216

Remarks on result:

other: naïve control: 2301 0 (vehicle only): 1989 3% w/v: 2322 10% w/v: 3216 30% w/v: 2613

Any other information on results incl. tables

The application of the test substance at concentrations of 3%, 10% and 30% w/v in propylene glycol resulted in an increase in isotope incorporation which was less than 3-fold at all three concentrations. Consequently, the test substance is designated as unlikely to be a moderate or strong sensitiser under the conditions of the test.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions chosen, the test substance did not show any skin sensitizing potential and thus, was concluded not to be a skin sensitizer.

Executive summary:

In a GLP-compliant LLNA study conducted according to OECD guideline 429, groups of four male CBA/Ca mice per dose level were treated with a 3%, 10% or 30% w/v preparation in propylene glycol or with the vehicle alone. Each test animal was applied with 25 µL of the test substance preparation to the dorsum of both ears for three consecutive days. Three days after the last application the mice were sacrificed and the auricular lymph nodes were removed. Lymph node response was evaluated by measuring the proliferative capacity of the prepared lymph node cells. The application of the test substance at concentrations of 3%, 10% and 30% w/v in propylene glycol resulted in an increase in isotope incorporation which was less than 3-fold at all three concentrations (SI indices were: 1.16 (3%); 1.61 (10%); 1.31 (30%)). Therefore, the test substance is considered to be not sensitizing.