Methyl 2,3-dibromopropionate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-12-19

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 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Version / remarks:

2017

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Methyl 2,3-dibromopropionate

EC Number:

217-044-3

EC Name:

Methyl 2,3-dibromopropionate

Cas Number:

1729-67-5

Molecular formula:

C4H6Br2O2

IUPAC Name:

methyl 2,3-dibromopropanoate

Test material form:

liquid

Specific details on test material used for the study:

Purity: 98%
Batch No.: 800327680

Test animals / tissue source

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: Bovine eyes from young cattle were obtained from the slaughterhouse, where the eyes were excised by a slaughterhouse employee as soon as possible after slaughter.
- Storage, temperature and transport conditions of ocular tissue:
Storage: The isolated corneas were stored in a petri dish with cMEM (Earle's Minimum Essential Medium containing 1% (v/v) L-glutamine and 1% (v/v) Foetal Bovine Serum).
Transport: Eyes were collected and transported in physiological saline in a suitable container under cooled conditions.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 μL
- Concentration (if solution): 98%

Duration of treatment / exposure:

10 ± 1 minutes

Number of animals or in vitro replicates:

Three corneas for each treatment group

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
- Preparation of Corneas: The isolated corneas were mounted in a corneal holder (one cornea per holder) with the endothelial side against the O-ring of the posterior half of the holder. The anterior half of the holder was positioned on top of the cornea and tightened with screws. The compartments of the corneal holder were filled with cMEM of 32 ± 1°C. The corneas were incubated for the minimum of 1 hour at 32 ± 1°C.
- Cornea Selection: After the incubation period, the medium was removed from both compartments and replaced with fresh cMEM. Opacity determinations were performed on each of the corneas using an opacitometer. Corneas that had an initial opacity reading higher than 7 were not used.

QUALITY CHECK OF THE ISOLATED CORNEAS: The eyes were checked for unacceptable defects, such as opacity, scratches, pigmentation and neovascularization by removing them from the physiological saline and holding them in the light. Those exhibiting defects were discarded.

NUMBER OF REPLICATES : Three corneas for each treatment group

NEGATIVE CONTROL USED : physiological saline

POSITIVE CONTROL USED : Ethanol

APPLICATION DOSE AND EXPOSURE TIME : 750 μL of either the negative control, positive control (Ethanol) or test item; 10 ± 1 minutes

REMOVAL OF TEST SUBSTANCE: After the incubation the control or test item was washed with MEM with phenol red.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: measured by the diminution of light passing through the cornea
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader (OD490)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: the decision criteria as indicated in the TG was used.

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Value:

>= -0.7 - <= 1.1

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
The corneas were clear after the 10 minutes of treatment with test item. No pH effect of the test item was observed on the rinsing medium.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean negative control responses for opacity and permeability were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas.
- Acceptance criteria met for positive control: The mean in vitro irritancy score of the positive control (Ethanol) was 47 and was within two standard deviations of the current historical positive control mean.

Applicant's summary and conclusion

Interpretation of results:

other: unclassfied

Conclusions:

In conclusion, since the test substance induced an IVIS ≤ 3, no classification is required for eye irritation or serious eye damage.

Executive summary:

The study was performed to evaluate the eye hazard potential of the test substance in accordance with OECD Guideline 437: Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage.

The test item was applied as it is (750 μL) directly on top of the corneas for 10 minutes. After exposure the cornea was thoroughly rinsed to remove the test item and incubated for 2 hours with fresh medium followed by opacity measurement and the permeability of the corneas was determined after a 90 minutes incubation period with sodium fluorescein.

The test conditions were adequate and that the test system functioned properly.

The test substance did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of -0.1 after 10 minutes of treatment.

In conclusion, since the test substance induced an IVIS ≤ 3, no classification is required for eye irritation or serious eye damage.