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Toxicological information

Skin irritation / corrosion

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Administrative data

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

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): C17 Methacrylate
- Physical state: liquid
- Analytical purity: > 99%
- Lot/batch No.: S732030105

Test animals

Species:
other: in vitro
Strain:
other: in vitro

Test system

Vehicle:
unchanged (no vehicle)
Details on study design:
METHODE
TEST SYSTEM
The EpiDerm TM model consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multi layered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multilayered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in viva. The EpiDerm TM tissues (surface 0.6 cm2)
are cultured on specially prepared cell culture inserts (MILLICELLs®, 10 mm 0) and commercially available as kits (EpiDerm TM 200), containing 24 tissues on shipping agarose.
Skin model: Epi-200
Supplier: MatTek In Vitro Life Science Laboratories, Bratislava, Slovakia

TEST PROCEDURE

- Irritation test: Three tissues were treated with the test substance, the PC and NC, respectively. Thirty microliter (30 μL) of the undiluted liquid test substance (after heating at ca. 30 °C) was applied using a pipette.
Control tissues were concurrently applied with 30 μL of sterile PBS (negative control, NC) or with 30 μL of 5% SDS (positive control, PC). A nylon mesh was placed carefully onto the tissue surface afterwards. The tissues were kept under the laminar flow hood at room temperature for 25 minutes overall and for 35 minutes in the incubator. The tissues were washed with sterile PBS to remove residual test material 1 hour after start of application. Rinsed tissues were blotted on sterile absorbent paper and transferred into new 6-well plates, pre-filled with 0.9 mL fresh medium. When all tissues were rinsed, the surface of each tissue was carefully dried with a sterile cotton swab. Subsequently, the tissues were incubated in the incubator at 37°C for 24 ± 2 hours. After 24 ± 2 hours the tissues were transferred into new 6-well plates pre-filled with 0.9 mL of fresh medium and placed into the incubator for additional 18 ± 2 hours post-incubation period. After the post-incubation period, the assay medium was replaced by 0.3 mL MTT solution and the tissues were incubated in the incubator for 3 hours. After incubation, the tissues were washed with PBS to stop the MTT-incubation. The formazan that was metabolically produced by the tissues was extracted by incubation of the tissues in isopropanol. The optical density at a wavelength of 570 nm (OD570) of the extracts was determined spectrophotometrically. Blank values were established of 6 microtiter wells filled with isopropanol for each microtiter plate.

Results and discussion

Any other information on results incl. tables

Individual and mean OD570 values, individual and mean viability values and standard deviations

Test substance

 

tissue 1

tissue 2

tissue 3

mean

SD

NC

mean OD570

2.078

2.215

2.189

2.160

 

viability

[% of NC]

96.2

102.5

101.3

100

3.37

14/0094-1

mean OD570

2.157

2.118

2.013

2.096

 

viability

[% of NC]

99.8

98.0

93.2

97

3.43

PC

mean OD570

0.076

0.072

0.074

0.074

 

viability

[% of NC]

3.5

3.3

3.4

3

0.09

The mean viability of the test-substance treated tissues determined after an exposure period of 1 hour with about 42 hours post-incubation was 97%.

Irritation test

Mean tissue viability

(% of negative control)

Prediction

50

Irritant

>50

Non-irritant

Applicant's summary and conclusion