Benzyl propionate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-12-11 and 2016-03-22

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2015

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Benzyl propionate

EC Number:

204-559-3

EC Name:

Benzyl propionate

Cas Number:

122-63-4

Molecular formula:

C10H12O2

IUPAC Name:

benzyl propanoate

In vitro test system

Test system:

human skin model

Source species:

other: reconstituted human epidermis

Cell type:

non-transformed keratinocytes

Justification for test system used:

The potential of chemical induced skin irritation is usually determined in vivo in the Draize rabbit skin irritation test as described in OECD guideline 404. Because systemic reactions play a minor role in modulating local skin toxicity potential of chemicals, skin irritation potential may be predicted by in vitro systems, provided they are sufficiently complex to mimic human skin barrier and cell reactivity. In an international prevalidation study performed by ECVAM, the in vitro skin irritation test using the human skin model EpiDerm™ and EpiSkin™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for skin irritancy potential.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Epi-200 SIT kit
- Tissue batch number(s): 23308
- Delivery date: 19 February 2016
- Date of initiation of testing: On day of receipt the pre-incubation phase of the EpiDerm™ tissues started.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 35 min at 37°C and 25 min at room temperature
- Temperature of post-treatment incubation: 37°C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: rinsed with DPBS at least 15 times; submerged in DPBS at least three times; afterwards once again rinsed with sterile DPBS from the inside and outside
- Observable damage in the tissue due to washing: no data
- Modifications to validated SOP: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: Versamax® Molecular Devices, Softmax Pro, version 4.7.1
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD = 1.673 +/- 0.048
- Barrier function: 8.20 hours
- Contamination: sterile

NUMBER OF REPLICATE TISSUES: 3

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

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin category 2 if the viability is less than or equal to 50%.
- The test substance is considered to be non-irritant to skin if the viability is greater than 50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 30 µL (47 µL/cm2)

NEGATIVE CONTROL
- Amount applied: 30 µL

POSITIVE CONTROL
- Amount applied: 30 µL
- Concentration: 5%

Duration of treatment / exposure:

60 minutes

Duration of post-treatment incubation (if applicable):

41 hours and 37 minutes

Number of replicates:

3 triplicate tissues (3 wells per tissue)

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 1

Value:

106.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 2

Value:

91.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 3

Value:

98.9

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:

It can be stated that in this study and under the experimental conditions reported, the test item is not irritant to skin.

Executive summary:

The irritation potential of the test substance was investigated by using the in vitro Human Skin Model Test (OECD 439, EU B.46). Each three tissues of the human skin model EpiDerm were treated with 30 µL of the test item, the negative (Dulbecco's Phosphate Buffered Saline) or the positive control (5% sodium lauryl sulfate) for 60 minutes in total. Cell viability was determined using the MTT assay. The absorbance value obtained for the positive control was 0.081 and this result corresponds to 4.3% viability when compared to the results obtained from the negative controls. After treatment with the test item the mean relative absorbance value decreased irrelevantly to 99.9% compared to the relative absorbance value of the negative control. This value is not below the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential. Thus, the test substance is not considered to be irritant to skin under the test conditions chosen.