1,4-Benzodioxane-2-carboxylic acid

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:

From 2018-12-17 to 2018-12-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study performed to current OECD guidelines with no significant deviations and run in OECD GLP certified lab.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2016

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1,4-Benzodioxane-2-carboxylic acid

EC Number:

609-276-2

Cas Number:

3663-80-7

Molecular formula:

C9H8O4

IUPAC Name:

1,4-Benzodioxane-2-carboxylic acid

Test material form:

solid

Specific details on test material used for the study:

Purity: 99.7%
Batch No.: 80031745a

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

Recommended test system in international guidelines (OECD and EC).

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm Skin Model
- Tissue batch number(s): 29674 Kit K and L

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: at room temperature
- Temperature of post-treatment incubation (if applicable): 36.8 - 37.6 °C (actual range)

REMOVAL OF TEST MATERIAL AND CONTROLS: washed with phosphate buffered saline to remove residual test item, rinsed tissues were kept in 24 well plates on 300 μL DMEM until 6 tissues (= one application time) were dosed and rinsed.

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 5 mg/mL diluted (1:5)
- Incubation time: 3 hours
- Spectrophotometer: TECAN Infinite® 1200 Pro Plate Reader.
- Wavelength: 570 nm
- Linear OD range of spectrophotometer: 0.8 - 2.8

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: 20 - 100%

NUMBER OF REPLICATE TISSUES: two tissues for 3-minute exposure and two for a 1-hour exposure, 2 tissues for the negative and positive controls for both the 3-minute and 1-hour time point

DECISION CRITERIA
- The test substance is considered to be corrosive to skin if [complete, e.g. the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.]
- The test substance is considered to be non-corrosive to skin if [complete, e.g. the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.]

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 27.0 - 32.3 mg

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 μL Milli-Q water

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 μL 8N KOH

Duration of treatment / exposure:

3 minutes and 1 hour

Number of replicates:

two tissues for 3-minute exposure and two for a 1-hour exposure, 2 tissues for the negative and positive controls for both the 3-minute and 1-hour time point

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Value:

93

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: mean tissue viability at 3-minute application

Results 2

Irritation / corrosion parameter:

% tissue viability

Value:

53

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: mean tissue viability at 1-hour application

Other effects / acceptance of results:

- OTHER EFFECTS:
- Colour interference with MTT: the test item did not interfere with the MTT endpoint.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range.
- Acceptance criteria met for positive control: The positive control had a mean relative tissue viability of 6.6% after the 1-hour exposure.
- Acceptance criteria met for variability between replicate measurements: In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤13%

Applicant's summary and conclusion

Interpretation of results:

other: unclassified

Conclusions:

In conclusion, the test substance is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.

Executive summary:

The objective of this study was to evaluate the test substance for its ability to induce skin corrosion in accordance with OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method).

The possible corrosive potential of the test substance was tested through topical application for 3 minutes and 1 hour. Skin tissue was moistened with 25 μL of Milli-Q water and at least 25 mg of the test substance was applied directly on top of the skin tissue.

The positive control had a mean relative tissue viability of 6.6% after the 1-hour exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the acceptance limits of OECD 431 (lower acceptance limit ≥0.8 and upper acceptance limit ≤2.8) and the laboratory historical control data range. In the range of 20 - 100% viability the Coefficient of Variation between tissue replicates was ≤ 13%, indicating that the test system functioned properly.

The relative mean tissue viability obtained after 3-minute and 1-hour treatments with the test item compared to the negative control tissues was 93% and 53%, respectively. The mean relative tissue viability for the test substance was not below 50% after the 3-minute treatment and not below 15% after the 1-hour treatment.

In conclusion, the test substance is not corrosive in the in vitro skin corrosion test under the experimental conditions described in this report.