3-methoxyoestra-1,3,5(10)-trien-17-one

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

12 Oct 2020 - 15 Oct 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

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:

2020

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:

2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

3-methoxyoestra-1,3,5(10)-trien-17-one

EC Number:

216-613-3

EC Name:

3-methoxyoestra-1,3,5(10)-trien-17-one

Cas Number:

1624-62-0

Molecular formula:

C19 H24 O2

IUPAC Name:

3-methoxyoestra-1,3,5(10)-trien-17-one

Specific details on test material used for the study:

Batch number: UN103MA
Purity: 94.98 %
Expiry date: 23 Jul 2021
Storage: room temperature

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: no data available

Source strain:

not specified

Justification for test system used:

according to guideline

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: reconstructed human epidermis model epiCS® (Cat.-No.CS-1001, CellSystems, Troisdorf, Germany)
- Tissue batch number(s): 100-AJ0414-1
- Date of initiation of testing: 12 Oct 2020

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: RT (room temperature)
- Temperature of post-treatment incubation (if applicable): Incubator temperature: 37 ± 2° C (CO2 gas concentration: 5 %; Humidity: 95%)

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1mg/ml
- Incubation time: 3 hours
- Spectrophotometer: EL808, Bio-Tek
- Wavelength: 570 nm

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Killed control and Color control were not required.

NUMBER OF TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: the optical density of the isopropanol-extracts of 3 insert was determined by duplicate per insert = 6 OD values.

PREDICTION MODEL / DECISION CRITERIA:

- The mean optical density (OD) values obtained with the test item were used to calculate the percentage of viability relative to the negative control, which is set at 100 %.
- according to UN GHS (Category 2 or Category 1) if the mean percent tissue viability after exposure and post treatment incubation is less than or equal (≤ ) to 50 %.


Acceptance criteria:
The following acceptance criteria determined the validity of an assay:
- mean OD negative control >/=0.8 and - mean relative viability of the positive control is - If the mean viability of the 3 replicates > 20% the coefficient of variation (CV) should not exceed 0.3.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 mg (plus 30 µl 0.9% NaCl to moisten and ensure good contact with the epidermis surface)

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µl
- Concentration (if solution): 0.9% NaCl in water

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 µl
- Concentration (if solution): 5% SDS in physiological saline

Duration of treatment / exposure:

20 minutes

Duration of post-treatment incubation (if applicable):

42 h

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Value:

105.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

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

- DEMONSTRATION OF TECHNICAL PROFICIENCY:
Reliability of the test was previously confirmed by interlaboratory validation

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

Conclusions:

A study was performed for the assessment of the skin irritancy of the test item with reconstructed human epidermis (RhE). The experiment was carried out using the commercially available test method epiCS®. The study was conducted in accordance with OECD TG 439 and EU Test Method B.46. The mean value of cell viability was recorded to be 105.6%. The test item was thus shown to be not irritating to reconstructed human skin in vitro.

Executive summary:

A study was performed for the assessment of the skin irritancy of Estron methylether with reconstructed human epidermis (RhE). The experiment was carried out using the commercially available test method epiCS®. The study was conducted in accordance with OECD TG 439 and EU Test Method B.46.

The test item was applied undiluted topically to the RhE tissue construct in triplicates and incubated for 20 minutes, followed by a 42 hours post-treatment incubation period.
Cell viability was measured in a photometer by the amount of MTT (methylthiazole tetrazolium) reduction. The optical density value obtained for the test item was used to calculate the percentage of viability relative to the negative control, which is set at 100% The results of the concurrent negative control (NC, 0.9 % NaCl) and positive control (PC, 5 % SDS) demonstrated the viability (NC) and sensitivity (PC) of the test model.
The mean value of cell viability was recorded to be 105.6%. The test item was thus shown to be not irritating to reconstructed human skin in vitro.