Registration Dossier

Diss Factsheets

Toxicological information

Skin irritation / corrosion

Currently viewing:

Administrative data

Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
3 June 2020 - 5 June 2020
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2020
Report date:
2020

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
other: EC Guideline No. 440/2008. Part B: Methods for the Determination of Toxicity and other health effects, Guideline B.40 BIS: "In Vitro Skin Corrosion: Human Skin Model Test".
Version / remarks:
31 May 2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)
Version / remarks:
18 June 2019
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
(3aS,3bS,9aR,9bS,11aS)‐11a‐ethyl‐10‐methylidene‐1H,2H,3H,3aH,3bH,4H,5H,7H,8H,9H,9aH,9bH,10H,11H,11aH‐cyclopenta[a]phenanthrene‐1,7‐dione
Cas Number:
54024-17-8
Molecular formula:
C20 H26 O2
IUPAC Name:
(3aS,3bS,9aR,9bS,11aS)‐11a‐ethyl‐10‐methylidene‐1H,2H,3H,3aH,3bH,4H,5H,7H,8H,9H,9aH,9bH,10H,11H,11aH‐cyclopenta[a]phenanthrene‐1,7‐dione
Test material form:
solid: particulate/powder
Details on test material:
Storage Conditions: In refrigerator (2-8°C)
Specific details on test material used for the study:
Physical Description: Off-white powder
Storage conditions: In refrigerator (2-8°C)
Test item handling: No specific handling conditions required

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Details on animal used as source of test system:
EpiDerm™ Reconstructed Human Epidermis (Lot no.: 33045 kit C+D)
- All cells used to produce Epiderm™ are purchased or derived from tissue obtained by MatTek Corporation from acredited institutions.
- Cells are screened for potential biological contaminants (HIV-1, Hepatitis B, Hepatitis C, bacteria, yeast and fungi)
Justification for test system used:
Recommended test system in international guidelines (OECD and EC).
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm Skin Model
- Tissue batch number(s): 33045 kit C+D
- Surface: 0.6 cm²
- The model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: Not specified

REMOVAL OF TEST MATERIAL AND CONTROLS
- After the exposure period, the tissues were washed with phosphate buffered saline

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 5 mg/mL
- Incubation time: 3 hours at 37°C in air containing 5% CO2
- Spectrophotometer: TECAN Infinite® M200 Pro Plate Reader
- Wavelength: 570 nm

NUMBER OF REPLICATE TISSUES: The test was performed on a total of 4 tissues per test item together with a negative control and positive control. Two tissues were used for a 3-minute exposure to the test item and two for a 1-hour exposure.

NUMBER OF INDEPENDENT TEST SEQUENCES: 1

ACCEPTANCE OF RESULTS
The in vitro skin corrosion test is considered acceptable if it meets the following criteria:
a) The absolute mean OD570 of the two tissues of the negative control should reasonably be within the laboratory historical control data range and the acceptance limits of OECD431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8).
b) The mean relative tissue viability following 1-hour exposure to the positive control should be <15 %.
c) In the range 20 - 100% viability, the Coefficient of Variation (CV) between tissue replicates should be ≤ 30%.

DECISION CRITERIA
A test item is considered corrosive in the in vitro skin corrosion test if:
a) The relative mean tissue viability obtained after 3-minute treatment compared to the negative control tissues is decreased below 50%.
b) In addition, a test item considered non-corrosive (viability ≥ 50%) after the 3-minute treatment is considered corrosive if the relative tissue viability after 1-hour treatment with the test item is decreased below 15%.
A test item is considered non corrosive in the in vitro skin corrosion test if:
a) The relative mean tissue viability obtained after the 3-minute treatment compared to the negative control tissues is not decreased below 50%.
b) In addition, the relative tissue viability after the 1-hour treatment is not decreased below 15%.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount applied: 27.0 to 36.0 mg
Duration of treatment / exposure:
3 minutes or 1 hour
Number of replicates:
Two replicates per exposure duration (4 total), two negative control, two positive control.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
1-hour treatment
Value:
96
Vehicle controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
3-minute treatment
Value:
91
Vehicle controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
RESULTS
Skin corrosion is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after the 3-minute and 1-hour treatments with the test item compared to the negative control tissues was 91% and 96% respectively. Because the mean relative tissue viability for the test item was not below 50% after 3 minutes treatment and not below 15% after 1 hour treatment the test item is considered to be not corrosive.

OTHER EFFECTS
- The test item was checked for color interference in aqueous conditions. Addition of the test item to Milli-Q and isopropanol resulted after subtraction of the blank in an OD of -0.001 and 0.004, respectively. Therefore it was concluded that the test item did not induce color interference.
- Because no color change was observed in the presence of MTT it was concluded that the test item did not interact with the MTT endpoint.

ACCEPTANCE OF RESULTS
- The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range.
- The mean relative tissue viability following the 1-hour exposure to the positive control was 6.2%.
- In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤15%, indicating that the test system functioned properly.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The in vitro skin corrosion test was conducted according to OECD 431 guideline and in accordance with GLP principles. In conclusion, EMETAM is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.
Executive summary:

The in vitro skin corrosion test was conducted according to OECD 431 guideline and in accordance with GLP principles. Skin tissue was moistened with 25 μL of Milli-Q water and at least 25 mg of the test item was applied directly on top of the skin tissue.
The positive control had a mean relative tissue viability of 6.2% after the 1-hour exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range. In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤15%, indicating that the test system functioned properly.
Skin corrosion is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after 3-minute and 1-hour treatments with the test item compared to the negative control tissues was 91% and 96%, respectively. Because the mean relative tissue viability for the test item was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment the test item is considered to be not corrosive.
In conclusion, EMETAM is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.