Fatty acids, C16-18, reaction products with ethanolamine

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 23 March 2020 to 17 June 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Fatty acids, C16-18, reaction products with ethanolamine

EC Number:

287-643-2

EC Name:

Fatty acids, C16-18, reaction products with ethanolamine

Cas Number:

85566-32-1

Molecular formula:

Not applicable

IUPAC Name:

Fatty acids, C16-18, reaction products with ethanolamine

Test material form:

solid: particulate/powder

In vitro test system

Test system:

human skin model

Cell type:

non-transformed keratinocytes

Cell source:

other: Human-derived epidermal keratinocytes

Justification for test system used:

The EpiDermTM skin model and assay for skin corrosion testing is endorsed by OECD TG 431.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: MatTek Corporation EpidermTM reconstructed tissue model: EPI-200
- Tissue batch number(s): 30862
- Delivery date: 28 April 2020

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37°C, 5% CO2, Relative Humidity: ≥95%

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- Incubation time: 60 minutes
- Spectrophotometer: BMG LabTech FluoStar Optima
- Wavelength: 570 nm
- Filter: no reference filter

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: Pass
- Barrier function: Pass
- Morphology: Pass
- Contamination: Pass

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE: Prior to the assay, the test item was checked for interference (water colouration or MTT interference) and found not to interfere.

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION:
3 tissues per condition

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

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL

Duration of treatment / exposure:

3 and 60 minutes

Number of replicates:

3

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minutes

Value:

94.679

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minutes

Value:

110.868

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: None reported
- Direct-MTT reduction: No direct reduction
- Colour interference with MTT: No colour interference

DEMONSTRATION OF TECHNICAL PROFICIENCY:

- Acceptance criteria met for negative control: The mean OD570 of the negative control tissues must be ≥0.8. Results for 3 minutes = 1.781 and 1 hour = 1.942.
- Acceptance criteria met for positive control: The mean of the positive control relative percentage viability, after 1 hour exposure must be < 15% of the mean of the negative control. The result was a pass at 8.780.
- Acceptance criteria met for variability between replicate measurements: should not exceed 0.3 (30%), result = pass.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The skin corrosion potential of Fatty acids, C16-18, reaction products with ethanolamine was evaluated. The tissue viability of the test item treated tissues were assessed and compared to a negative control. The percentage viability obtained after 3 minutes was 110.868% and after 60 minutes was 94.679%. The substance did not meet the criteria for classification as a corrosive in accordance with Regulation (EC) No.1272/2008.

Executive summary:

The skin corrosion potential of Fatty acids, C16-18, reaction products with ethanolamine was evaluated using OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method). The test was GLP compliant. Triplicate EpiDermTM skin model tissues were treated with a single topical application of 25 mg of Fatty acids, C16-18, reaction products with ethanolamine. Additional triplicate tissues were treated with 50 µl of Sterile Water (Negative control) and 50µl of Potassium Hydroxide (Positive control). All tissues were exposed for 30 minutes and 60 minutes at 37°C, 5% CO2, ≥95% Relative Humidity, prior to the MTT endpoint. Corrosion is directly related to cytotoxicity in the EpiDerm™ tissue. Cytotoxicity is demonstrated by the reduction of MTT (3-[4,5dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to formazan by viable cells in the test item treated tissues compared to the corresponding negative control. The results are then used to make a prediction of the corrosivity potential of the test item. All controls were valid and demonstrated the reliability of the test system. Mean tissue viability (as a percentage of the negative control), was 110.868% and 94.679% after 3 and 60 minutes of exposure, respectively. Therefore, Fatty acids, C16-18, reaction products with ethanolamine does not meet the criteria for classification according to CLP Regulation (EC) No. 1272/2008.