Sodium 2-stearoyllactate

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 Sept - 11 Oct 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

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:

adopted in 2019

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:

adopted in 2012

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hessisches Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 2-stearoyllactate

EC Number:

246-929-7

EC Name:

Sodium 2-stearoyllactate

Cas Number:

25383-99-7

Molecular formula:

This reference substance is a UVCB of the NCS type. It is a complex mixture of compounds and therefore molecular formula, molecular weight, SMILES, InChI and structural formula cannot be given.

IUPAC Name:

sodium 2-hydroxy-2-methyl-3-oxoicosanoate

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: EpiDermTM (EPI-200)

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ (EPI-200-SIT) (MatTek Corporation, 82105 Bratislava, Slovakia)
- Tissue batch number(s): 30830
- Date of initiation of testing: 12 Sept 2019

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 35 min at 37 ± 1.5 °C and 5 ± 0.5% CO2 in Dulbecco's Minimum Essential Medium (DMEM) followed by 25 min in a sterile bench at room temperature
- Temperature of post-treatment incubation: approx. 42 h at 37 ± 1.5 °C and 5 ± 0.5% CO2 in DMEM

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: rinsed with Phosphate Buffered Saline (PBS) several times

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 300 μl/well
- Incubation time: 3 h ± 5 min
- Spectrophotometer: microplate reader (Versamax® Molecular Devices, Software SoftMax Pro Enterprise version 4.7.1)
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: The quality of the EpiDerm™ tissue was assessed by an MTT cell viability test. The determined OD (540 - 570 nm) was 1.946 ± 0.146 (acceptance criteria: 1.0 - 3.0).
- Barrier function: The barrier function was assessed by determination of the exposure time required to reduce tissue viability by 50% (ET-50) upon application of 100 µL of 1% Triton X-100. The ET-50 value was determined to be 5.05 h (acceptance criteria: 4.77-8.72 h).
- Contamination: The cells used to produce the EpiDerm™ tissue were screened for the presence of viruses, bacteria, yeast and other fungi. No contamination was detected.

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
Since the test substance did not directly reduce the MTT solution, an additional functional check was not performed.

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

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be irritant or corrosive to skin if the viability after 1 h exposure is ≤ 50%.
- The test substance is considered to be non-irritant to skin if the viability after 1 h exposure is > 50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 25 mg (corresponding to 39.7 mg/cm2) + 25 µL DPBS

Duration of treatment / exposure:

35 min at 37 °C and 25 min at RT

Duration of post-treatment incubation (if applicable):

Approx. 42 h

Number of replicates:

Triplicates for each treatment and control group

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean value of 3 tissues

Run / experiment:

1 h exposure

Value:

100.16

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Direct-MTT reduction: Since the test substance did not directly reduce MTT, an additional test with freeze-killed or viable tissues was not performed.
- Colour interference with MTT: The test substance did not change the colour, when mixed with deionised water and thus passed the colour interference pre-test. Also its intrinsic colour was not intensive.

DEMONSTRATION OF TECHNICAL PROFICIENCY:
The technical proficiency of the test facility was demonstrated. Appropriate data are provided in 'Any other information on results incl. tables' below.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD of the tissue replicates treated with the negative control was ≥ 0.8 and ≤ 2.8 for every exposure time (values between 1.689 and 1.783).
- Acceptance criteria met for positive control: The mean viability of the tissue replicates treated with the positive control for 1 h was < 15% compared to the negative control (4.32%).
- Acceptance criteria met for variability between replicate measurements: The standard deviations between the three tissue percentage viability values of each group (test item, positive and negative controls) in the main test were below 18.

Any other information on results incl. tables

Table 1: Details on results
Treatment Group	Tissue No.	OD 570 nm			Mean OD of 3 Wells	Mean OD of 3 Wells blank corrected	Mean OD of 3 tissues	Rel. Viability [%] Tissue 1, 2 + 3	Standard Deviation	Mean Rel. Viability [%]
		Well 1	Well 2	Well 3
Blank		0.038	0.039	0.039	0.039
Negative Control	1	1.734	1.713	1.736	1.727	1.689	1.751	96.469	0.054	100.0
	2	1.831	1.818	1.807	1.819	1.780		101.677
	3	1.825	1.783	1.858	1.822	1.783		101.854
Positive Control	1	0.112	0.126	0.114	0.117	0.079	0.076	4.494	0.005	4.32
	2	0.117	0.117	0.117	0.117	0.078		4.469
	3	0.108	0.109	0.109	0.109	0.070		3.989
Test Item	1	1.861	1.795	1.783	1.813	1.774	1.753	101.336	0.082	100.16
	2	1.708	1.702	1.694	1.701	1.663		94.982
	3	1.838	1.886	1.864	1.862	1.824		104.175

 

Table 2: Historical control data
Positive Control; OD at 570 nm after exposure to 5% SDS solution in deionized water (MatTek)		Negative Control OD at 570 nm DPBS (MatTek)
Tissue Viability [%]	3.89	Mean OD	1.69
Standard Deviation	1.01 % points	Standard Deviation	0.19
Range of Viabilities	2.24 % - 6.19 %	Range of OD*	1.28 - 2
Mean OD	0.07	* should be 0.8 - 2.8 (OECD 439) or 1.0 - 2.5 (MatTek)
Standard Deviation	0.02
Range of OD	0.03 - 0.11
Data of 50 sets of controls shared between 195 studies performed from August 2015 until May 2019 (p.p.: percentage points)

 

Table 3: Technical proficiency
Proficiency Substance	Viability [%]	Category
Naphtalene acetic acid	101.7	No Cat.
Isopropanol	85.5	No Cat.
Methyl stearate	91.1	No Cat.
Heptyl butyrate	109.0	No Cat.
Hexyl salicylate	98.1	No Cat.
Cyclamen aldehyde	24.1	Cat. 2
1-Bromohexane	16.3	Cat. 2
Potassium hydroxide (5% aq.)	40.1	Cat. 2
1-Methyl-3-phenyl-1-piperazine	21.6	Cat. 2
Heptanal	20.6	Cat. 2
In vitro Skin Irritation Assay: OECD 439
Proficiency Data (March 2014, non-GLP) performed at ICCR-Roßdorf GmbH Study
Director: Dipl.-Ing. Andreas Heppenheimer

Applicant's summary and conclusion

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008.

Conclusions:

Under the conditions of the conducted test, the test substance did not possess irritating properties towards reconstructed human epidermis tissue in the EpiDerm™ model.