5-bromo-5-nitro-1,3-dioxane

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: 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:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted 28 July 2015

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

5-bromo-5-nitro-1,3-dioxane

EC Number:

250-001-7

EC Name:

5-bromo-5-nitro-1,3-dioxane

Cas Number:

30007-47-7

Molecular formula:

C4H6BrNO4

IUPAC Name:

5-bromo-5-nitro-1,3-dioxane

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: A14-Q0201

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- Stability under test conditions: yes

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The solid test substance was ground with pestle and mortar before application

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from multiple donors

Vehicle:

water

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDermTM 200 kit; 24 EPI-200 tissues (reconstructed epidermis): surface 0.6 cm²
cultured in Millicells, diameter: 1 cm
- Tissue batch number(s): EPI-200, EPI-212,EPI-218
- Date of initiation of testing: 04 August 2016

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

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: once with PBS
- Observable damage in the tissue due to washing: no
- Modifications to validated SOP: no

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT: 1.0 mg / mL MTT diluent
- Incubation time: 3 hrs
- Spectrophotometer: SunriseTM Absorbance Reader
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: 1.850 ± 0.053
- Barrier function: 6.18 hrs
- Contamination: no
- Reproducibility: yes

NUMBER OF REPLICATE TISSUES: 2

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- killed tissues
- Procedure used to prepare the killed tissues (if applicable): freezing
- N. of replicates : 2

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than 50% and the viability after 1 hour exposure is greater than 15%.]

Acceptance criteria for tissue variability:
For every treatment 2 tissues are treated in parallel. In the range of 20% and 100% viability, variability between the
tissues is considered to be acceptable if the coefficient of variation (CV) of %-viability is ≤ 30%.

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 μL de-ionized water was applied first; thereafter, a bulk volume of ca. 25 μL of the solid
ground test material was applied with a sharp spoon and homogeneously distributed with the water

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 μL of de-ionized water

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

Duration of treatment / exposure:

3 min or 1 hour

Number of replicates:

2

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean

Run / experiment:

3 min incubation

Value:

16.2

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean

Run / experiment:

1 hour incubation

Value:

3.8

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: no
- Direct-MTT reduction: no

DEMONSTRATION OF TECHNICAL PROFICIENCY: yes

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes

The CV of %-viability of the 3-minute exposure of the test-substance treated tissues, which was 37.4%, is out of the acceptance range, which was ≤ 30%. Since all other quality criteria of the study were met and both viability values are well below the cut off for skin corrosion, this deviation is not considered to adversely affect the result of this study.

Applicant's summary and conclusion

Interpretation of results:

Category 1A (corrosive) based on GHS criteria