Biphenyl-4-yl-(9,9-dimethyl-9H-fluoren-2-yl)-[2-(9,9-diphenyl-9H-fluoren-4-yl)-phenyl]-amine

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

25 January 2017 - 25 January 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

July 29, 2016

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EU Method B.40.bis. In vitro skin corrosion: human skin model test

Version / remarks:

31 May 2008

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: INVITTOX Protocol SkinEthic™ Skin Corrosivity Test

Version / remarks:

April 2012

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

N-{[1,1'-biphenyl]-4-yl}-N-[2-(9,9-diphenyl-9H-fluoren-4-yl)phenyl]-9,9-dimethyl-9H-fluoren-2-amine

EC Number:

811-852-5

Cas Number:

1792219-00-1

Molecular formula:

C58H43N

IUPAC Name:

N-{[1,1'-biphenyl]-4-yl}-N-[2-(9,9-diphenyl-9H-fluoren-4-yl)phenyl]-9,9-dimethyl-9H-fluoren-2-amine

Test material form:

solid: particulate/powder

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Episkin/SkinEthic Laboratories, Lyon, France

Source strain:

not specified

Justification for test system used:

The reconstructed human epidermis model in vitro method is an accepted in vitro method to replace animal testing. The human skin RHE™ model closely mimics the biochemical and physiological properties of the upper parts of the human skin, i.e the epidermis.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: SkinEthic ™ RHE-model RHE/S/ 17
- Tissue batch number(s): 17-RHE-009
- Date of initiation of testing: 25 January 2017

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: 20 mL DPBS

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1mg/mL
- Incubation time: 3 hours (+/- 15 minutes)
- Spectrophotometer:ELx800, BioTek Instruments GmbH,
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD 1.4 (Acceptance criteria: OD > 0.7)
- Barrier function: 5.5 h (Acceptance criteria: 4 h <= ET50 <= 10 h
- Morphology: 5.5 cell layers, absent of significant histological abnormalities, satisfactory (acceptance criteria: number of cell layers >= 4; absence of significant histological abnormalities; well differentiated epidermis consisting of basal, spinous, granular layers and a stratum corneum)

NUMBER OF REPLICATE TISSUES: 2

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues
- N. of replicates: one

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

yes, concurrent MTT non-specific colour control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 20 mg (+/- 3 mg)

NEGATIVE CONTROL (deionised water)
- Amount applied: 40 ± 3 μL

POSITIVE CONTROL
- Amount applied: 40 ± 3 μL

Duration of treatment / exposure:

3 minutes and 1 hour

Number of replicates:

2

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 1/ 3 min

Value:

99.54

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 1/ 1 hours

Value:

92.52

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 2/ 3 min

Value:

112.98

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 4

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 2/ 1 hour

Value:

98.48

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: no
- Colour interference with MTT: no


ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Acceptance criteria met for variability between replicate measurements: Yes

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the present study, the test item is not considered to possess a corrosive potential to skin.

Executive summary:

The objective of the present study was to investigate the potential of the test item to induce skin corrosion in an in vitro human skin model.

The test item was applied topically to a human reconstructed skin model followed by determination of the cell viability. Cell viability was determined by enzymatic conversion of vital dye MTT into a blue formazan salt and measurement of the formazan salt after extraction from tissues. The percent reduction of cell viability in comparison to untreated negative controls was used to predict the skin corrosion potential. Duplicates of the human skin RHE-model were treated with the test item or the negative control for 3 minutes and additional 1 hour. Duplicates with the positive control were only treated for 1 hour. 40 ± 3 μL of either the negative control (deionised water) or the positive control (potassium hydroxide, 8N) were applied to the tissues. Before application of 20 ± 3 mg of the solid test item, 20 ± 2 μL of deionised water was spread to the epidermis surface to improve the contact between the test item and the epidermis. After treatment with the negative control (deionized water) the mean OD per tissue replicate was 1.696 and 1.913 after 3 minutes exposure and 1.935 and 2. 145 after 1 hour exposure (study acceptance criterion: >= 0.8 and <= 3.0). Treatment with the positive control (Potassium hydroxide, 8N) revealed a mean viability of 0.48% after 1 hour (study acceptance criterion: <15%). Therefore, the study fulfilled the validity criteria. Following treatment with the test item, the tissue viability was >= 50% after 3 minutes exposure (mean viability: 106.26%) and >= 15% after 1 hour exposure (mean viability: 95.50%), i.e. according to OECD 431 the test item is not considered as corrosive to skin.