[No public or meaningful name is available]

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

15 May 2012 - 17 May 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

At present there are no validated or OECD adopted in vitro tests for eye irritation. However, the test used here, SkinEthic™ HCE (SkinEthic, France), was conducted in accordance with GLP standards. The study was also well documented and the method is scientifically acceptable.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

The test is based on the SkinEthic HCE model (SkinEthic Laboratories, Nice, France). Triplicate tissues were treated with the test item for an exposure period of 10 minutes. At the end of the exposure period each tissue was taken for MTT assay and optical density at 540nm to measure cell viability were determined. The individual and mean OD540 values and tissue viabilities for the test item, negative control item, and positive control item were calculated.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Renewable hydrocarbons (diesel type fraction)

EC Number:

700-916-7

Molecular formula:

UVCB substance, not available. View remarks field.

IUPAC Name:

Renewable hydrocarbons (diesel type fraction)

Details on test material:

- Name of test material (as cited in study report): Diesel
- Substance type: UVCB substance
- Physical state: liquid
- Analytical purity: 100%; UVCB substance
- Lot/batch No.: 11-09149-001
- Expiration date of the lot/batch: 01 January 2013
- Storage condition of test material: room temperature in the dark

Test animals / tissue source

Species:

other: Human corneal epithelium

Test system

Vehicle:

unchanged (no vehicle)

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 30 microliters

Duration of treatment / exposure:

10 minutes

Observation period (in vivo):

See details on study details

Number of animals or in vitro replicates:

Not used. In vitro test.

Details on study design:

Triplicate tissues were treated with 30 microliters of the test item for 10 minutes. Triplicate tissues were treated with positive and negative control solutions. At the end of the exposure period each tissue was rinsed with PBS without Ca/Mg. Following rinsing, the tissues (two per group) were transferred to a pre-labelled 24-well plate containing 300 microliters of a 0.5mg/ml MTT solution. The plate was placed into an incubator for three hours. At the end of the incubation period the tissue inserts were rinsed twice with PBS and transferred to a new pre-labelled 24-well plate containing 0.75 ml of isopropanol in each well. An extra 0.75 ml of isopropanol was added onto each tissue and the plate was seled to prevent isopropanol evaporation. The plate was allowed to stand overnight at room temperature to extract the formazan crystals out of the tissue. At the end of the extraction period, the solution in each tissue insert were homogenized and measured for optical density at 540nm.

Results and discussion

In vitro

Results

Irritation parameter:

mean percent tissue viability 

Run / experiment:

Mean

Value:

106.7

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

Relative tissue viability (% negative control) < 60 is classified as an irritant.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

Criteria used for interpretation of results: expert judgment

Conclusions:

The eye irritation potential of the test item was evaluated using SkinEthic HCE reconstructed human corneal epithelium model. Based on the study result the the test item is considered to be a non-irritant.

Executive summary:

The study was regarded reliable without restrictions, since the study is carried out in compliance with OECD principles of Good Laboratory Paractice (GLP) and the documentation is sufficient for assessment. However, at present the method is not validated in vitro tests for assessment of eye irritation.

Triplicate tissues were treated with the test item for an exposure period of 10 minutes. At the end of the exposure period each tissue was taken for MTT-loading. After MTT loading tissue inserts were placed into isopropanol for extraction of formazan crystals out of the MTT-loaded tissues. At the end of the extraction period each well was mixed and samples were measured for optical density at 540nm. The individual and mean OD540 values and tissue viabilities for the test item, negative control item, and positive control item were calculated. The relative mean viability of the test item treated tissues was 106.7 % after a 10 min exposure period. Based on this data test item is considered to be a non-irritant. The obtained viability (%) for the test item (higher than 100 %) is due to the mean viability of the negative control tissues is set 100%.

This study is used as key study in the hazard assessment.