Reaction product of 2,3-epoxypropyl neodecanoate and Benzenesulfonic acid, C10-13-sec-alkyl derivatives

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

21-25 July 2017

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

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:

26 July 2013

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Reference substance 001

Cas Number:

None

Molecular formula:

C10-13H21-27-C6H4-SO2-O-CH2-C(H)OH-CH2-CO2-C9H19

Test material form:

liquid

Specific details on test material used for the study:

Identification: Reaction product of 2,3-epoxypropyl neodecanoate and benzenesulfonic acid, C10-13-sec-alkyl derivatives
Appearance: Brown liquid
Batch: P718261998
Storage: At room temperature
Stable under storage conditions until: 01 June 2018 (expiry date)

Test animals / tissue source

Species:

other: Bovine eyes were used as soon as possible after slaughter.

Details on test animals or tissues and environmental conditions:

Bovine eyes from young cattle were obtained from the slaughterhouse (Vitelco, 's Hertogenbosch, The Netherlands), where the eyes were excised by a slaughterhouse employee as soon as possible after slaughter. Eyes were collected and transported in physiological saline in a suitable container under cooled conditions. 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. 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 (Life Technologies)). 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 32C. The corneas were incubated for a minimum of 1 hour at 32C. 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. The opacity of each cornea was read against a cMEM filled chamber, and the initial opacity reading thus determined wasrecorded. Corneas that had an initial opacity reading higher than 7 were not used. Three corneas were selected at random for each treatment group.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

750 µL

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

120 minutes

Number of animals or in vitro replicates:

Three

Details on study design:

The medium from the anterior compartment was removed and 750 µL of either the negative control, positive control or test item was introduced onto the epithelium of the cornea. The holders were slightly rotated, with the corneas maintained in a horizontal position, to ensure uniform distribution of the control or the test item over the entire cornea. Corneas were incubated in a horizontal position for 10 minutes at 32°C. After the incubation the solutions were removed and the epithelium was washed with MEM with phenol red (Earle’s Minimum Essential Medium, Life Technologies) and thereafter with cMEM. Possible pH effects of the test item on the corneas were recorded. The medium in the posterior compartment was removed and both compartments were refilled with fresh cMEM. Subsequently the corneas were incubated for 120 minutes at 32C. After the completion of the incubation period opacity determination was performed. Each cornea was inspected visually for dissimilar opacity patterns. The opacity of a cornea was measured by the diminution of light passing through the cornea. The change in opacity for each individual cornea (including the negative control) was calculated by subtracting the initial opacity reading from the final post-treatment reading. The corrected opacity for each treated cornea with the test item or positive control was calculated by subtracting the average change in opacity of the negative control corneas from the change in opacity of each test item or positive control treated cornea. The mean opacity value of each treatment group was calculated by averaging the corrected opacity values of the treated corneas for each treatment group.

Following the final opacity measurement, permeability of the cornea to Na-fluorescein was evaluated. The medium of both compartments (anterior compartment first) was removed. The posterior compartment was refilled with fresh cMEM. The anterior compartment was filled with 1 mL of 4 mg Na-fluorescein/mL cMEM solution. The holders were slightly rotated, with the corneas maintained in a horizontal position, to ensure uniform distribution of the sodium-fluorescein solution over the entire cornea. Corneas were incubated in a horizontal position for 90 minutes at 32°C.

After the incubation period, the medium in the posterior compartment of each holder was removed and placed into a sampling tube labelled according to holder number. 360 μL of the medium from each sampling tube was transferred to a 96-well plate. The optical density at 490 nm (OD490) of each sampling tube was measured in triplicate using a microplate reader. Any OD490 that was 1.500 or higher was diluted to bring the OD490 into the acceptable range (linearity up to OD490 of 1.500 was verified before the start of the experiment). OD490 values of less than 1.500 were used in the permeability calculation. The mean OD490 for each treatment was calculated using cMEM corrected OD490 values. If a dilution has been performed, the OD490 of each reading of the positive control and the test item was corrected for the mean negative control OD490 before the dilution factor was applied to the reading.

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Run / experiment:

Experiment

Value:

26

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

Mean IVIS -1.6

Positive controls validity:

valid

Remarks:

Mean IVIS 26

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

The 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. The mean in vitro irritancy score of the positive control (Ethanol) was 61 and within two standard deviations of the current historical positive control mean. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.

Any other information on results incl. tables

 Summary of results

Treatment	Mean opacity	Mean permeability	Mean IVIS
PBS	-1.5	-0.00	-1.6
Ethanol	22	2.585	61
Test material	6.7	1.317	26

(IVIS) = mean opacity value + (15 x mean OD490 value)

Applicant's summary and conclusion

Interpretation of results:

study cannot be used for classification

Conclusions:

Since [reaction product of 2,3-epoxypropyl neodecanoate and benzenesulfonic acid, C10-13-sec-alkyl derivatives] induced an IVIS > 3 ≤ 55, no prediction on the CLP classification can be made.

Executive summary:

The objective of this study was to evaluate the eye irritation potential of the submission substance [reaction product of 2,3-epoxypropyl neodecanoate and benzenesulfonic acid, C10-13-sec-alkyl derivatives], as measured by its ability to induce opacity and increase permeability in an isolated bovine cornea (BCOP test). The eye damage of the test item was tested through topical application for 10 minutes. The test item was applied (750 μL, undiluted) directly onto of the corneas. Reaction product of 2,3-epoxypropyl neodecanoate and benzenesulfonic acid, C10-13-secalkyl derivatives induced ocular irritation through both endpoints, resulting in a mean in vitro irritancy score (IVIS) of 26 after 10 minutes of treatment. In conclusion, since reaction product of 2,3-epoxypropyl neodecanoate and benzenesulfonic acid, C10-13-sec-alkyl derivatives induced an IVIS > 3 ≤ 55, no prediction on the CLP classification can be made.