Registration Dossier

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 08, 2018 - Septembre 15, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD 439

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Justification for test system used:
The test item is applied topically to a three-dimensional RhE model, comprised of non-transformed, human-derived epidermal keratinocytes, which have been cultured to form a multi-layered, highly differentiated model of the human epidermis.
The RhE test method is based on the premise that irritant chemicals are able to penetrate the stratum corneum and damage the underlying layers of keratinocytes and other skin cells. Cell viability is measured by enzymatic conversion of the vital dye MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, Thiazolyl blue], into a blue formazan salt that is quantitatively measured after extraction from tissues.
Irritant chemicals are identified by their ability to decrease cell viability below defined threshold levels.
Vehicle:
unchanged (no vehicle)
Details on test system:
The test system used for the in vitro skin irritation test was reconstructed human epidermis (SkinEthicTM RHE) as recommended by the OECD 439 guideline. The SkinEthicTM RHE model consists of normal human keratinocytes cultured for 17-days on an insert of 0.5 cm2 polycarbonate filter at the air-liquid interface in a chemically defined growth medium. The cells form a multilayered, highly differentiated and stratified epidermis model of the human epidermis that consists of main basal, supra basal, spinous and granular layers and a functional stratum corneum.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
For the treatment of 11-Methyldodecyl Laurate, negative control tissues and positive control tissues, 16 µL/0.5 cm2 of test item, 16 µL/0.5 cm2 of sterile dulbecco’s phosphate buffered saline (DPBS) and 16 µL/0.5 cm2 of 5% sodium dodecyl sulphate (5% aq.) were applied for 42 minutes at room temperature, respectively. Nylon mesh was used for uniform spreading of liquid materials.
Duration of treatment / exposure:
Tissues were exposed to 11-Methyldodecyl Laurate, negative and positive control for 42 minutes.
Duration of post-treatment incubation (if applicable):
After washing and drying, tissues were incubated in 6-well plate containing 2 mL growth medium at 37± 1°C in 5± 1% CO2 in a humidified incubator for 42 hours.
Number of replicates:
Three replicates were used for the test item, positive control and negative control.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Experiment 1
Value:
95.13
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Experiment 2
Value:
94.28
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Experiment 3
Value:
94.35
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Mean
Value:
94.59
Negative controls validity:
valid
Positive controls validity:
not valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
The negative and positive controls gave acceptable results and the study was accepted as valid.

Any other information on results incl. tables

TABLE1: Data Summary of Percent Viability

Treatment

Tissue Replicate

O.D. at 570 nm

Blank Corrected O.D.

Mean of Corrected O.D.

Mean O.D. of Three Tissues

% Viability/

Tissue

Mean % Viability

S.D. of % Viability

C.V. of % Viability

Corrosivity Class

Negative Control

(Dulbecco’s Phosphate Buffered Saline (DPBS))

1

1.346

1.302

1.296

1.293

100

100

0.01

0.77

NA

1.329

1.285

1.346

1.302

2

1.332

1.288

1.296

1.346

1.302

1.342

1.298

3

1.318

1.274

1.287

1.325

1.281

1.349

1.305

11-Methyldodecyl Laurate

1

1.293

1.250

1.23

1.223

95.13

94.59

0.47

0.50

No Category

1.266

1.223

1.260

1.217

2

1.259

1.216

1.219

94.28

1.265

1.222

1.262

1.219

3

1.257

1.214

1.22

94.35

1.262

1.219

1.269

1.226

Positive control

(Sodium dodecyl sulphate (5% aq.))

1

0.064

0.020

0.019

0.02

1.47

1.52

0.09

5.92

Category 2

0.063

0.019

0.063

0.019

2

0.065

0.021

0.021

1.62

0.064

0.020

0.066

0.022

3

0.062

0.018

0.019

1.47

0.064

0.020

0.064

0.020

Key     : O.D. = Optical Density, S.D. = Standard Deviation, C.V. = Coefficient of Variation, NA = Not Applicable.

Note      :              For Negative control, SD and CV of % viability was calculated using corrected OD at 570 nm; For the test item and positive control SD and CV of %           viability was calculated using % viability/tissue.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The negative and positive controls gave acceptable results and the study was accepted as valid. Based on the results obtained, the test item 11-Methyldodecyl Laurate is classified as non-irritant to skin (UN GHS No Category).
Executive summary:

This study was performed to evaluate the non-irritant and irritant potential of 11-Methyldodecyl Laurate using reconstructed human epidermis (RHE) tissue in accordance with the United Nations Globally Harmonized System of Classification and Labelling of Chemicals (UN GHS).

Tissues were exposed tothenegative control (Dulbecco’s Phosphate Buffered Saline (DPBS)), positive control (sodium dodecyl sulfate,5% aqueous (SDS)) and11-Methyldodecyl Lauratein triplicate for 42 minutes at room temperature.

The mean cell viability in tissues treated with the test item was 94.59% after 42 minutes exposure. A significant reduction in percent cell viability was not observed in treated tissues when compared with that of the concurrent negative control.

The Optical density (OD) values for the negative control replicates were between 1.274 and 1.305, against the guideline requirement of ≥ 0.8 and ≤ 3.0 (≥ 1.2 as per SkinEthic SOP). The OD of the blankwasbetween0.042and0.053which met the guideline requirement of OD< 0.1.The positive control showed a 1.52% cell viability, against the acceptance criteria of <40% for the SkinEthic RHE model, when compared with that of the concurrent negative control. Variation between tissue replicates (i.e., CV% value) was 0.50% for the test item group, 5.92% for positive control and 0.77% for negative control against the guideline requirement of ≤ 18%, which demonstrate the efficiency of the test system, SkinEthicTMRHE model. All criteria for a valid study were met as describe in the study plan.