2-ethylhexyl acrylate

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

12 Dec 2000 - 13 Dec 2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study conducted in compliance with GLP regulations

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-ethylhexyl acrylate

EC Number:

203-080-7

EC Name:

2-ethylhexyl acrylate

Cas Number:

103-11-7

Molecular formula:

C11H20O2

IUPAC Name:

2-ethylhexyl acrylate

Details on test material:

- Test substance: 2-ethylhexyl acrylate
- Chemical name: 2-propenoic acid, 2-ethylhexyl ester
- Test substance No.: 00/0903-1
- Batch/Lot No.: Tank 102
- Analytical Purity: 99.7 area-% (GC)
(according to BASF AG, Analytical report)
- Physical state: liquid
- Stability under test conditions: stable
- Storage condition of test material: temperature, protection from light

In vitro test system

Test system:

human skin model

Vehicle:

unchanged (no vehicle)

Details on test system:

The potential of the test substance to cause dermal corrosion was assessed by a single topical application of 50 uL of the test substance to a reconstructed three dimensional human epidermis model (EpiDermTM). Duplicates of the EpiDermTM tissue were incubated with the test subtance for 3 minutes and 1 hour, followed by a colorimetric determination of the possibly induced cytotoxic effect. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt. The formazan production of the test substance treated epidermal tissues is compared to the negative control tissues, calculated as relative tissue viability.

TEST SYSTEM
- The EpiDerm System consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis. It consits of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in vivo. The EpiDermTM tissues (surface 0.6 cm2) are cultured on specially prepared cell culture inserts (MILLICELLs®, 10 mm QS) and shipped world-wide as kits (EpiDermTM 200), containing 24 tissues on shipping agarose.
- Tissue model: Epi-200
- Origin: MatTek Corporation, Ashland MA, USA

TEST PROCEDURE
- The test was performed on a total of 4 tissues per test material, negative control and positive control. Out of these 4 tissues 2 tissues are used for the 3 minutes application and 2 tissues for the 1 hour application.
- After 3 minutes (room temperature) or after 1 hour (incubator) tissue contact with the test material, the tissues were washed with buffer to remove residual test material. Rinsed tissues were kept in 24-well plates (holding plates) at room temperature on assay medium until all tissues per application time were dosed and rinsed. The assay medium was then replaced by MTT solution and tissues were incubated for 3 hours. After incubation tissues were washed with buffer and formazan was extracted with Isopropanol over night at room temperature. The optical density at a wavelength of 570 nm (OD570) of extracted formazan was determined spectrophotometrically and cell viability was calculated for each tissue as % of the mean of the negative control tissues. Skin corrosivity potential of the test material was classified according to remaining cell viability obtained after test material treatment with either of the 2 exposure times.

ASSAY ACCEPTANCE CRITERIA:
- Negative control (NC ): mean OD570 of the NC per exposure time > = 0.8
- Positive control (PC ): mean relative tissue viability of the 3 min positive control is <= 30 %
- Tissue variability: The inter tissue variability is considered to be acceptable if the difference of the OD570 values of
the two tissues is <= 0.3
- Killed controls (KC ): OD570 of the killed control tissues treated as negative control should be <= 0.35

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

50 µL

Duration of treatment / exposure:

3 min and 1h

Duration of post-treatment incubation (if applicable):

none

Number of replicates:

2 per exposure time

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3-min exposure

Value:

99

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: not corrosive

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1-h exposure

Value:

104

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: not corrosive

Any other information on results incl. tables

Exposure: 3 min

Test article	Mean OD570	Viability [% of NC]
NC	1.621	100
TS	1.600	99
PC	0.237	15

Exposure: 1 h

Test article	Mean OD570	Viability [% of NC]
NC	1.593	100
TS	1.663	104
PC	0.182	11

NC = Negative control

TS = Test substance

PC = Positive control

Applicant's summary and conclusion

Interpretation of results:

other: not corrosive

Conclusions:

Based on the observed results it was concluded, that the test substance does not have a corrosive potential in the EpiDerm skin corrosivity test under the test conditions chosen.

Executive summary:

In order to support the decision if the local lesions caused after skin contact are to be classified as corrosion or as severe irritation, BASF AG carried out an alternative to the Draize skin test which was developed in order to differentiate between irritation and corrosion (EU Guideline B.40). The so called EpiDermTM Skin Corrosivity Test was performed using 2-ethylhexyl acrylate, purity 99.7%. The potential of 2-EHA to cause dermal corrosion was assessed by a single topical application of 50 µl of the test substance to a reconstructed three dimensional human epidermal model (EpiDermTM). Dublicates of the EpiDermTM tissue were incubated with 2-EHA for 3 minutes and 1 hour, followed by a colorimetric determination of the possibly induced cytotoxic effects. Viability of the test substance treated tissues determined after an exposure period of 3 minutes was 99% and for the exposure period of 1 hour 104% (2-EHA is not able to directly reduce MTT, the indicator used for detection of cytotoxicity). It is demonstrated that 2-EHA reacts like the negative control, while a caustic compound used as positive control proved that this system is able to detect caustic chemicals. Based on the observed results and applying the evaluation criteria of the test, 2-EHA does not have a corrosive potential in this test under the conditions chosen.