4-ethoxy-2,3-difluoro-4'-propyl-1,1'-biphenyl

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2011-01-11 to 2011-03-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Version / remarks:

2009

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

2010

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

4-ethoxy-2,3-difluoro-4'-propyl-1,1'-biphenyl

EC Number:

638-734-4

Cas Number:

157248-24-3

Molecular formula:

C₁₇H₁₈F₂O

IUPAC Name:

4-ethoxy-2,3-difluoro-4'-propyl-1,1'-biphenyl

Test material form:

solid

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: not specified

Justification for test system used:

The test system used is a established standard model for in vitro skin irritation testing.

Vehicle:

unchanged (no vehicle)

Remarks:

The test item was not dissolved in a vehicle. However, the tissue surface was wetted with 10 µL deionised water before application.

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: The applied human in vitro skin model RHE was produced by SkinEthic Laboratories (Lyon, France).
- Tissue batch number: 11022A0209
No further details were specified.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation: 37 °C

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: 1, using at least 25 mL PBS
- Observable damage in the tissue due to washing: not specified
- Modifications to validated SOP: no

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1.0 mg/L
- Incubation time: 3 h +/- 5 min
- Spectrophotometer: plate spectrophotometer, not further specified
- Wavelength: 570 nm
No further details were specified.

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
No details were specified.

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
The pretest for MTT-reducing capacity of the test item showed that the test item did not reduce MTT. Therefore, no controls were used.

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

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be irritant to skin if the mean tissue viability is equal or less than 50 %.
- The test substance is considered to be non-irritant to skin if the mean tissue viability is more than 50 %.

Control samples:

yes, concurrent negative control

yes, concurrent vehicle

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 16 mg

NEGATIVE CONTROL
- Amount applied: 16 µL

POSITIVE CONTROL
- Amount applied: 16 µL
- Concentration: 5 % in deionised water

Duration of treatment / exposure:

42 +/- 1 min

Duration of post-treatment incubation (if applicable):

42 +/- 1 h

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

mean value

Value:

126.44

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: Not specified
- Direct-MTT reduction: MTT was not reduced by the test item.
- Colour interference with MTT: No change in color observed.

DEMONSTRATION OF TECHNICAL PROFICIENCY: Not specified

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes. The optical density of negative control (mean value: 1.822 +/- 0.06 %) was >= 1.2 and standard deviation was <= 18 %.
- Acceptance criteria met for positive control: Yes. The relative cell viability of the positive control (mean value: 1.51, standard deviation 0.21 %) was < 40 % and standard deviation was <= 18 %.
- Acceptance criteria met for variability between replicate measurements: Yes. The standard deviation of 6.88 was <= 18 %.

Any other information on results incl. tables

Group	Time / [min]	Mean OD	Mean Relative viability / [%]
Negative Control	42	1.822	100
Positive Control	42	0.028	1.51
Test Material	42	2.303	126.44

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item showed no skin irritation potential.

Executive summary:

The objective of the present study was to investigate potential of the test material to induce skin irritation in an in vitro human skin model. The test conducted according to OECD 439 consisted of a topical exposure of the test item to a human reconstructed skin model followed by a cell viability test. Cell viability was quantitatively measured by dehydrogenase conversion of MTT into a blue formazan salt after extraction from tissues. The percent reduction of cell viability in comparison to untreated negative controls was used to predict the skin irritation potential. Triplicates of the human skin RHE-model were treated with the test material, the negative or the positive control for 42 minutes (± 1 minute). 16 µL of either the negative control (PBS-buffer) or the positive control (5 % aqueous solution of sodium dodecyl sulfate) were applied to the tissues. Before adding the solid test item, 10 µL of deionised water was spread to the epidermis surface to improve the contact between the test item and the epidermis. Afterwards, 16 mg of the test item were applied to the tissues. After treatment with the negative control the mean OD was 1.822 and thus >1.2. Treatment with the positive control revealed a mean viability value of 1.51 % (study acceptance criteria: < 40 %). Therefore, the study fulfilled the validity criteria. The mean tissue viability after treatment with the test material was 126.44 %, clearly exceeding 50 %. Therefore, the test material is considered as non-irritant to the skin in this in vitro assay.