Sodium methyl sulphate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Nov 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

28 July 2015

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Sodium methyl sulphate

EC Number:

208-142-7

EC Name:

Sodium methyl sulphate

Cas Number:

512-42-5

Molecular formula:

CH4O4S.Na

IUPAC Name:

sodium methyl sulphate

Test material form:

solid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Lot/batch No. of test material: KIRSCHAZ2-00182
- Content: 99.1 g/100 g
- pH-value: Ca. 6 (undiluted test substance, moistened with de-ionized water; determined in the lab prior to start of the GLP study)

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- Stability under test conditions: The stability under storage conditions over the study period was guaranteed by the sponsor.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The solid test substance was ground with pestle and mortar before application and applied minimally moistened with PBS.

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Origin: MatTek In Vitro Life Science Laboratories, Bratislava, Slovakia. Tissue model: EPI-200.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ 200 kit, EPI-200
- Tissue batch number: 23377

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 25 minutes at room temperature and for 35 minutes in the incubator at 37°C
- Temperature of post-treatment incubation (if applicable): 37°C

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1.0 mg / mL MTT diluent
- Incubation time: 3 hours
- Spectrophotometer: SunriseTM Absorbance Reader
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD570 of the negative control between 0.8 and 2.8 and of the positive control (5% SDS) ≤ 20%
- Barrier function: Lower acceptance limit: ET50 = 4.0 hours. Upper acceptance limit: ET50 = 8.7 hours.

NUMBER OF REPLICATE TISSUES: 3

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin if the mean tissue viability after exposure is less than 45%.
- The test substance is considered to be borderline irritant (inconclusive) to skin if the mean tissue viability after exposure is 45-55%.
- The test substance is considered to be non-irritant to skin if the viability after exposure is greater than or equal to 55%.
- Justification for the selection of the cut-off point(s) if different than recommended in TG 431 and 439: The “borderline“ evaluation (50 ± 5%) was statistically determined by using historic data of the test facility and hence considers the variance of the test method. This evaluation is confirming the borderline range provided in OECD Guideline 439.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 μL sterile PBS was applied first. Thereafter, a bulk volume of ca. 25 μL (about 15 mg) of the solid ground test material was applied with a sharp spoon and homogeneously distributed together with the fluid.

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL
- Concentration (if solution): 5% (w/v)

Duration of treatment / exposure:

25 minutes at room temperature and for 35 minutes in the incubator at 37°C

Duration of post-treatment incubation (if applicable):

42 hours

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Run 1, mean of 3 replicate tissues

Value:

85.1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

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

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

Any other information on results incl. tables

Table 1: Results of the Skin Irritation Test. Individual and mean OD₅₇₀values, individual and mean viability values, standard deviations and coefficient of variation.

Testsubstance identification		tissue 1	tissue 2	tissue 3	mean	SD	CV [%]
NC	mean OD570	1.802	1.693	1.695	1.730
	viability [% of NC]	104.2	97.9	98.0	100.0	3.6	3.6
16/0352-1	mean OD570	1.381	1.459	1.576	1.472
	viability [% of NC]	79.8	84.3	91.1	85.1	5.7	6.7
PC	mean OD570	0.041	0.055	0.051	0.049
	viability [% of NC]	2.4	3.2	2.9	2.8	0.4	14.3

Applicant's summary and conclusion

Executive summary:

The objective was to assess the skin corrosion and irritation potential of the test substance. Using the currently available methods a single in vitro assay is not sufficient to cover the full range of skin irritating/corrosion potential including transport classification. Therefore, three in vitro assays were proposed for this in vitro skin irritation and corrosion test strategy including transport classification: The Skin Corrosion Test (SCT), Skin Irritation Test (SIT) and In vitro Membrane Barrier Test (Corrositex®). However, in the current case the results derived with SIT alone were sufficient for a final assessment. Therefore further testing in SCT and Corrositex® was waived.

Skin Irritation Test (SIT):

The potential of the test substance to cause dermal corrosion/irritation was assessed by a single topical application of ca. 25 μL bulk volume (about 15 mg) of the undiluted test substance to a reconstructed three-dimensional human epidermis model (EpiDerm™). The irritation test was performed with three EpiDerm™ tissues, which were incubated with the test substance for 1 hour followed by a 42-hours post-incubation period. Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the test-substance treated epidermal tissues is compared to that of negative control tissues. The quotient of the values indicates the relative tissue viability.

The following results were obtained in the EpiDerm™ skin irritation test (SIT):

The test substance was not able to reduce MTT directly. The mean viability of the test substance treated tissues determined after an exposure period of 1 hour with about 42 hours post-incubation was 85.1%.

Based on the observed results and applying the evaluation criteria, it was concluded that the test substance does not show a skin irritation potential in the EpiDerm™ in vitro skin irritation and corrosion test strategy including transport classification under the test conditions chosen.