Diethyl ethylmalonate

Skin irritation / corrosion

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Diethyl ethylmalonate

EC Number:

205-093-3

EC Name:

Diethyl ethylmalonate

Cas Number:

133-13-1

Molecular formula:

C9H16O4

IUPAC Name:

1,3-diethyl 2-ethylpropanedioate

Test material form:

liquid

Details on test material:

Test item: diethyl ethylmalonateCAS No.: 133-13-1Physical state: colourless liquidStorage: in cool, dry, well-ventilated place

In vitro test system

Test system:

human skin model

Remarks:

three-dimensional human epidermis model

Source species:

human

Vehicle:

unchanged (no vehicle)

Details on test system:

EpiSkinTM Small Model (EpiSkinTMSM), manufactured by EPISKIN SNC Lyon, France, is a three-dimensional human epidermis model. Adult human-derived epidermal keratinocytes are seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis model is obtained after 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum. Its use for skin irritation testing involves topical application of test materials to the surface of the epidermis, and the subsequent assessment of their effects on cell viability.Supplier: SKINETHIC Laboratories

Amount/concentration applied:

A volume of 10 μL test item was applied on the skin surface by using a suitable pipette.

Duration of treatment / exposure:

The plates with the treated epidermis units were incubated for the exposure time of 15 minutes (± 0.5 min) at room temperature. After the incubation time the EpiSkinTMSM units were removed and rinsed thoroughly with PBS 1x solution to remove all of the test material from the epidermal surface. The rest of the PBS was removed from the epidermal surface with suitable pipette tip linked to a vacuum source.

Duration of post-treatment incubation (if applicable):

After rinsing the units were placed into the plate wells with fresh pre-warmed “maintenance medium” (2 mL/well) below them and then incubated for 42 hours (± 1h) at 37°C in an incubator with 5 % CO2.

Number of replicates:

Three replicates were used for the test item and controls respectively.

Test system

Details on study design:

Human Skin Model:EpiSkinTM Small Model (EpiSkinTMSM), manufactured by EPISKIN SNC Lyon, France, is a three-dimensional human epidermis model. Adult human-derived epidermal keratinocytes are seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis model is obtained after 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum (Tinois et al., 1994). Its use for skin irritation testing involves topical application of test materials to the surface of the epidermis, and the subsequent assessment of their effects on cell viability.Justification for selection of the test system: The EPISKIN model has been validated for irritation testing in an international trial.

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Value:

81

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Disks of EPISKIN (three units / chemical) were treated with test item and incubated for 15 minutes at room temperature. Exposure of test material was terminated by rinsing with PBS 1x solution. Epidermis units were then incubated at 37 °C for 42 hours in an incubator with 5% CO2. The viability of each disk was assessed by incubating the tissues for 3 hours with MTT solution at 37°C in 5% CO2 protected from light. The precipitated formazan was then extracted using acidified isopropanol and quantified spectrophotometrically.SDS (5% aq.) and PBS treated epidermis were used as positive and negative controls respectively. For each treated tissue viability was expressed as a percentage relative to negative control.The test item is considered to be irritant to skin, if the mean relative viability after 15 minutes exposure and 42 hours post incubation is less or equal (≤) to 50% of the negative control.In this in vitro skin irritation test using the EPISKIN model, the test item did not show significantly reduced cell viability in comparison to the negative control (mean value: 81 %). All obtained test item viability results were above 50 % when compared to the viability values obtained from the negative control, therefore the test item was considered to be non-irritant to skin.Positive and negative controls showed the expected cell viability values within acceptable limits. The experiment was considered to be valid.

Any other information on results incl. tables

Substance	OpticalDensity(OD)		Viability(%)
Negative Control: 1 x PBS	mean	0.836	100
	standard deviation (SD)		8.44
Positive Control: SDS (5 % aq.)	mean	0.033	4
	standard deviation (SD)		1.83
Test Item: diethyl ethylmalonate (CAS No. 133-13-1)	mean	0.677	81
	standard deviation (SD)		11.75

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The results obtained from this in vitro skin irritation test, using the EPISKIN model, indicated that the test item reveals no skin irritation potential under the utilised testing conditions. The test item diethyl ethylmalonate (CAS No. 133-13-1) is considered to be non-irritant to skin and is therefore not classified (UN GHS No Category).