5-hydroxyiminobarbituric acid

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-07-25 to 2018-09-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Deviations:

no

GLP compliance:

no

Remarks:

The study has been performed in House according to current guidelines by trained technicians. The study is commonly performed at the lab and leaves up to the validty criteria.Thus data from the study can be considered reliable.

Test material

Test material information

Constituent 1

Reference substance name:

5-hydroxyiminobarbituric acid

EC Number:

201-741-4

EC Name:

5-hydroxyiminobarbituric acid

Cas Number:

87-39-8

Molecular formula:

C4H3N3O4

IUPAC Name:

5-(hydroxyimino)-1,3-diazinane-2,4,6-trione

Test material form:

solid: particulate/powder

Details on test material:

Lot/batch No.: 201713807

Test animals / tissue source

Species:

human

Details on test animals or tissues and environmental conditions:

- Justification of the test method and considerations regarding applicability : The SkinEthic Human Corneal Epithelial Model (SkinEthicTM HCE) in order to assess eye irritation potential of violuric acid. The test method has been chosen in order to investigate whether or not the test material should be classified. Based on prvious QSAR assessment, which did not indicate yirritation potential, this method was chosen.
- Description of the cell system used
The assay is a cytotoxicity test performed with SkinEthic Human Corneal Epithelium kit. The model consists of fully differentiated epithelium of normal human keratinocytes in a three-dimensional structure. The cells are grown on a microporous polycarbonate filter, which mimics the structure of the outer layer of the cornea of the eye. Batch No 18-HCE-029 was used for the present assay.
The absence of mycoplasma, bacteria and fungus has been verified.

Test system

Vehicle:

physiological saline

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 mg

Duration of treatment / exposure:

Exposure: 4h +/- 5 min

Duration of post- treatment incubation (in vitro):

18h +/- 30 min

Number of animals or in vitro replicates:

2 replicates

Details on study design:

- RhCE tissue construct used, including batch number : Immortalized Human Corneal Epithelim Cells batch no. 18-HCE-029
- Doses of test chemical and control substances used : 30 mg violuric acid, 30 µL Methyl acetate, 30 µl PBS
- Duration and temperature of exposure, post-exposure immersion and post-exposure incubation periods (where applicable) : Treatment period: 4 hours ± 5min (37°C, 5% CO2, ≥ 95% humidity) , post exposure immersion 30 minutes ± 2min at room temperature; post-incubation period: 18 hours ± 30min incubation at 37°C, 5% CO2
- Justification for the use of a different negative control than ultrapure H2O (if applicable) : PBS was used to premoisten the tissue before application of violuric acid
- Number of tissue replicates used per test chemical and controls (positive control, negative control, NSMTT, NSCliving and NSCkilled, if applicable) 2
- Wavelength and band pass (if applicable) used for quantifying MTT formazan: 570 nm
- Description of the method used to quantify MTT formazan
- Description of evaluation criteria used including the justification for the selection of the cut-off point for the prediction model
- Acceptable variability between tissue replicates for positive and negative controls : Viability of PC expressed as % of neg control <=30%
- Acceptable variability between tissue replicates for the test chemical: Difference in viability between 2 replica is <=20%

Results and discussion

In vitro

Results

Irritation parameter:

other: percent cell viability compared to control tissue, which is set to 100%

Value:

0.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Interpretation of results:

study cannot be used for classification

Conclusions:

Violuric acid was found be irritating to the eye and should be classified. However, the present system cannot distinguish between UN GHS Category 1 from Category 2.

Executive summary:

The in vitro eye irritation potential of violuric acid was investigated with SkinEthic Human Corneal Epithelium model. The test was performed in accordance with OECD 492.

The irritation potential was assessed indirectly by assessing the uptake of MTT in living cells, which results in conversion to MTT in a blue dye, which can be measured spectophotometrically and by visual assessment of the tissue after exposure of 4h followed by immersion period of 30 min and post treatment period of 18h at 37°C and 5% CO2. Viability of cells exposed to violuric acid was 0.6 % compared to control cells and thus indicates eye irritation potential of the test substance.

According to the test system violuric acid should be classified as irritant, but the test system cannot distinguish between category 1 and 2 classification.

In conclusion, violuric acid was found to be irritaing in the present test system