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Administrative data

skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference Type:
study report

Materials and methods

Test guideline
according to guideline
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
GLP compliance:

Test material

Chemical structure
Reference substance name:
Benzenesulfonic acid, 4-C10-13-alkyl derivs.-, salts with amines, C16-C18 (even numbered) alkyl
EC Number:
Molecular formula:
Benzenesulfonic acid, 4-C10-13-alkyl derivs.-, salts with amines, C16-C18 (even numbered) alkyl
Test material form:
liquid: viscous
Details on test material:
Chemical registery number : 953-378-2
Chemical name : Benzenesulfonic acid, 4-C10-13-alkyl derivs.-, salts with amines, C16-C18 (even numbered) alkyl

Based on the qualitative and quantitative information on the composition, the sample used are representative of the boundary composition shared and agreed by each registrant.

In vitro test system

Test system:
human skin model
Source species:
Cell type:
non-transformed keratinocytes
Details on animal used as source of test system:
Three-dimensional reconstructed human epidermis model consisting of adult human-derived epidermal keratinocytes seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen.
unchanged (no vehicle)
Control samples:
yes, concurrent MTT non-specific colour control
Amount/concentration applied:
10 µL (26.3 µL/cm2) of the test material was applied to the epidermis surface
Duration of treatment / exposure:
exposure period of 15 minutes
Duration of post-treatment incubation (if applicable):
42 Hour post-exposure incubation period

Results and discussion

In vitro

Irritation / corrosion parameter:
% tissue viability
Vehicle controls validity:
Negative controls validity:
Positive controls validity:

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
The test item was classified as non-irritant.
Executive summary:

The purpose of this test was to evaluate the skin irritation potential of the test item using the EPISKIN reconstructed human epidermis model after a treatment period of 15 minutes followed by a post-exposure incubation period of 42 hours. The principle of the assay was based on the measurement of cytotoxicity in reconstructed human epidermal cultures following topical exposure to the test item by means of the colorimetric MTT reduction assay. Cell viability is measured by enzymatic reduction of the yellow MTT tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test item treated tissues relative to the negative controls.

Triplicate tissues were treated with the test item for an exposure period of 15 minutes. At the end of the exposure period each tissue was rinsed before incubating for 42 hours. At the end of the post-exposure incubation period each tissue was taken for MTT-loading. The maintenance medium from beneath each tissue was transferred to pre-labeled micro tubes and stored in a freezer for possible inflammatory mediator determination. After MTT-loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues.
At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 μL samples were transferred to the appropriate wells of a pre-labeled 96-well plate. The optical density was measured at 570 nm.
Data are presented in the form of percentage viability (MTT reduction in the test item treated tissues relative to negative control tissues).

The relative mean viability of the test item treated tissues was 94.6% after the 15-Minute exposure period and 42-Hours post-exposure incubation period.

The quality criteria required for acceptance of results in the test were satisfied.

The test item was classified as non-irritant. The following classification criteria apply: EU CLP Not classified for Irritation.