2-Butene-1,4-diol, 1,4-diacetate, (2Z)-

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2014-10-02 to 2015-01-29

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

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

(2Z)-4-(acetyloxy)but-2-en-1-yl acetate

EC Number:

607-674-0

Cas Number:

25260-60-0

Molecular formula:

C8H12O4

IUPAC Name:

(2Z)-4-(acetyloxy)but-2-en-1-yl acetate

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

Recommended system by the guidelines

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Reconstructed Human Epidermis (RHE Kit)
- Tissue batch number: 14-RHE-024

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 °C
- Temperature of post-treatment incubation: 37 °C

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: rinsing with minimum of 25 mL
- Observable damage in the tissue due to washing: none
- Modifications to validated SOP: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 hours +/- 5 min

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: well
- Barrier function: checked
- Morphology: well
- Contamination: none
- Reproducibility: well

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE : not applicable

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritating to skin if the viability after 42 minutes exposure is less or equal to 50%
- The test substance is considered to be non-irritating to skin if the viability after 42 minutes exposure is greater than 50%

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 16 µL

NEGATIVE CONTROL
- Amount applied: 16 µL

POSITIVE CONTROL
- Amount applied: 16 µL
- Concentration: 5 % aqueous solution

Duration of treatment / exposure:

42 minutes

Duration of post-treatment incubation (if applicable):

42 h

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1

Value:

63.77

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: none
- Direct-MTT reduction: not applicable
- Colour interference with MTT: not applicable

DEMONSTRATION OF TECHNICAL PROFICIENCY: yes

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 obtained after in RHE assay

Dose group	Treatment Interval	Optical density Tissue 1	Optical density Tissue 2	Optical density Tissue 3	Mean optical density	Mean relative viability in %	Standard deviation (SD) in %
Negative Control	42 min	2.445	2.588	2.510	2.515	100.00	2.85
Positive Control	42 min	0.022	0.021	0.021	0.021	0.83	2.77
Test item	42 min	1.571	1.634	1.606	1.604	63.77	1.97

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study the test item is considered to be not irritating to human skin.

Executive summary:

This in vitro study was performed to assess the irritation potential of the test item by means of the Reconstructed Human Epidermis Test according to OECD TG 438. The test consisted of a topical administration of the test item to a human reconstructed skin model followed by a cell viability test. Cell viability was quantitatively measured by dehydrogenase conversion of MTT into a blue formazan salt after extraction from tissues. The percent reduction of cell viability in comparison to untreated negative controls was used to predict the skin irritation potential. Triplicates of the human skin RHE™-model were treated with the test item, the negative or the positive control for 42 minutes (± 1 minute). 16 µL of either the negative control (DPBS-buffer), positive control (5% aqueous solution of sodium dodecyl sulfate) or test item were applied to the tissues.

After treatment with the negative control (DPBS-buffer) the mean OD was 2.515. Treatment with the positive control (5% aqueous solution of sodium dodecyl sulfate) revealed a mean viability value of 0.83%. Therefore, the study fulfilled the validity criteria.

The tissue viability after treatment with the test item was 63.77 % and, thus, higher than 50 %, i.e. according to UN GHS classification the test item is considered as non-irritant to skin.

In conclusion, under the conditions of this study, the test item is not considered to possess an irritant potential to skin.