Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

The Bovine Corneal Opacity and Permeability (BCOP) test method is an in vitro test method that can be used to classify substances as “ocular corrosives / severe irritants” and “non-irritants”. The BCOP is recommended for use as part of a tiered-testing strategy for regulatory classification and labelling within a specific applicability domain. Test substances can be classified as ocular corrosives / severe irritants or non-irritants without further testing in rabbits

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Materials and methods

Test guideline

Test guideline 1

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)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Phosphoric acid, C14-15 branched and linear alkyl esters, potassium salts

EC Number:

812-497-9

Cas Number:

1893414-79-3

Molecular formula:

n/a for an UVCB substance

IUPAC Name:

Phosphoric acid, C14-15 branched and linear alkyl esters, potassium salts

Constituent 2

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

water

Test material form:

solid: crystalline

Specific details on test material used for the study:

Name: Phosphoric acid, C14-15 branched and linear alkyl esters, potassium salts
CAS No.: 1893414-79-3
Physical state: white solid at 20 °C
Batch No.: PU61810016
Re-certification date of batch: 09 March 2018
Purity: 100 % (UVCB, lyophilized solid, water content 0.85 % (w/w))
Stability: stable under test conditions
Storage condition of test material: Room temperature, protected from light

Test animals / tissue source

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

The assay uses isolated corneas obtained as a by-product from animals freshly slaughtered at the abattoir A. Moksel AG, Buchloe, Germany.
On the test day, fresh eyes were collected from the slaughterhouse and were transported in HBSS containing penicillin ( 100 IU/mL) / streptomycin (100 µg/mL) on ice to the laboratories. Immediately after arrival of the eyes, cornea preparation was initiated.
The eyes were carefully examined for defects and any defective eyes were discarded.
The tissue surrounding the eyeball was carefully pulled away and the cornea was excised leaving a 2 to 3 mm rim of sclera. The isolated corneas were stored in a petri dish containing HBSS. Before the corneas were mounted in corneal holders (Duratec GmbH) with the endothelial side against the O-ring of the posterior chamber, they had been visually examined for defects and any defective cornea had been discarded. The anterior chamber was then positioned on top of the cornea and tightened with screws. The chambers of the corneal holder were then filled with RPMI (without phenol red) containing 1% FBS and 2 mM L-glutamine (complete RPMI). The posterior chamber was always filled first. The corneas were incubated for one hour at 32 +- 1 °C.

Test system

Vehicle:

physiological saline

Remarks:

physiological saline 0.9% NaCl

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

The test item was suspended with physiological saline 0.9% NaCl to give a 20% (w/w) concentration.

Duration of treatment / exposure:

750 µL of the test item preparation or the control substance was introduced into the anterior chamber (closed-chamber method). After 4 hours ± 5 minutes incubation at 32 +- 1 °C either the test substance or the control substance was removed and the epithelium washed at least three times with MEM (containing phenol red).

Duration of post- treatment incubation (in vitro):

After the illuminance measurement was performed, the medium was removed from both chambers of the holder. The posterior chamber was refilled with fresh complete RPMI. 1 mL of a 5 mg/mL sodium fluorescein solution was added to the anterior chamber and the corneas were incubated for 90 minutes at 32 +- 1 °C.

Number of animals or in vitro replicates:

3 corneas for the test item
3 corneas as negative controls treated with physiological saline 0.9% NaCl
3 corneas as positive controls treated with imidazole 20% in physiological saline 0.9% NaCl

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Run / experiment:

mean - IVIS

Value:

38.1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

not determinable

Other effects / acceptance of results:

The in vitro irritation score obtained with the positive control fell within the two standard deviations of the current historical mean and therefore this assay is considered to be valid.

Any other information on results incl. tables

The eye irritancy potential of Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts was investigated in the bovine corneal opacity and permeability assay. The test item was suspended with physiological saline 0.9% NaCl (see 10.3) to give a 20% concentration. All 3 corneas treated with Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts showed slightly opacity and fissures in the tissue.

The following mean in vitro irritation score was calculated: 38.10

No prediction can be made regarding the classification of the test substance Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts according to the evaluation criteria. Further testing in another suitable method is required. The in vitro irritation score obtained with the positive control fell within the two standard deviations of the current historical mean and therefore this assay is considered to be valid. The negative control responses should result in opacity and permeability values that are less than the established upper limits for background bovine corneas treated with the respective negative control.

Applicant's summary and conclusion

Interpretation of results:

study cannot be used for classification

Conclusions:

No prediction can be made regarding the classification of the test substance Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts according to the evaluation criteria. Further testing in another suitable method is required.

Executive summary:

The eye irritancy potential of Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts was investigated in the bovine corneal opacity and permeability assay. The test item was suspended with physiological saline 0.9% NaCl (see 10.3) to give a 20% concentration. All 3 corneas treated with Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts showed slightly opacity and fissures in the tissue.

The following mean in vitro irritation score was calculated: 38.10

No prediction can be made regarding the classification of the test substance Phosphoric acid, C14-15-branched and linear alkyl esters, potassium salts according to the evaluation criteria.