Sodium 2-stearoyllactate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 Dec 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

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)

Version / remarks:

adopted in 2017

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted in 2010

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hessisches Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 2-stearoyllactate

EC Number:

246-929-7

EC Name:

Sodium 2-stearoyllactate

Cas Number:

25383-99-7

Molecular formula:

This reference substance is a UVCB of the NCS type. It is a complex mixture of compounds and therefore molecular formula, molecular weight, SMILES, InChI and structural formula cannot be given.

IUPAC Name:

sodium 2-hydroxy-2-methyl-3-oxoicosanoate

Test animals / tissue source

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: AB Schlachthof GmbH & Co. KG, 63739 Aschaffenburg, Germany
- Characteristics of donor animals: 14 months old
- Storage, temperature and transport conditions of ocular tissue (e.g. transport time, transport media and temperature, and other conditions): Hank’s Buffered Salt Solution (HBSS) containing 1% (v/v) penicillin/streptomycin (100 units/mL penicillin and 100 μg/mL streptomycin), cooled in the slaughter-house and during transport.
- Time interval prior to initiating testing: The eyes were transported on the morning of slaughter of the donor animals to the laboratory. The corneae were isolated on the same day after delivery of the eyes.
- indication of any existing defects or lesions in ocular tissue samples: The eyes were carefully examined macroscopically for defects. Only eyes without defects (e.g. vascularization, pigmentation, opacity and scratches) were used in the assay.

Test system

Vehicle:

physiological saline

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.75 mL
- Concentration: 20% suspension (w/v) in saline using sonication for 10 min

Duration of treatment / exposure:

240 min at 32 ± 1 °C

Number of animals or in vitro replicates:

Triplicates for each treatment and control group

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
The cornea was carefully removed from the eye using scalpel and rounded scissors. A rim of about 2 mm of tissue (sclera) was left for stability and handling of the isolated cornea. Each cornea was mounted in a specially designed cornea holder.

QUALITY CHECK OF THE ISOLATED CORNEAS
At the end of the equilibration period, the basal opacity was determined (t0). Each cornea with a value of the basal opacity > 7 was discarded.

TREATMENT METHOD: closed chamber
The endothelial side of the cornea was positioned against the O-ring of the posterior part of the holder. The cornea was gently flattened over the O-ring but stretching was avoided. The anterior part of the holder was positioned on top of the cornea and fixed in place with screws. Both compartments of the holder were filled with incubation medium. The posterior compartment was filled first to return the cornea to its natural convex position. Care was taken to assure that no air bubbles were present within the compartments.

REMOVAL OF TEST SUBSTANCE
After exposure, the test substance was rinsed off from the corneae with Eagle’s minimum essential medium (EMEM) containing phenol red for at least three times or more until phenol red was still discoloured (yellow or purple), or the test item was still visible. Since the test item proved difficult to be removed by the rinsing method, the front cover of the holder was opened and the cornea was carefully washed using a gentle stream of incubation medium. Once the medium was free of the test item the corneae were given a final rinse with complete Minimum Essential Medium (cMEM) without phenol red.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Corneal opacity was determined by the amount of light transmission through the cornea via an opacitometer (OP_KiT opacitometer (Electro Design, 63-Riom, France)).
- Corneal permeability: Passage of sodium fluorescein dye measured with the aid of a microplate reader (Versamax® Molecular Devices, Software SoftMax Pro Enterprise version 4.7.1) (OD490)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA:
Test substance with an IVIS > 55 was regarded as serious eye damage and labelled Category 1 according to CLP/EPS/GHS.
Test substance with an IVIS ≤ 3 was regarded as non-irritant and labelled in no category.
For test substance with an IVIS in the range > 3 ≤ 55 no prediction can be made.

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Run / experiment:

240 min exposure

Value:

2.27

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

DEMONSTRATION OF TECHNICAL PROFICIENCY:
The technical proficiency of the test facility was demonstrated. Appropriate data are provided in 'Any other information on results incl. tables' below.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes, the negative control resulted in opacity and permeability values that were less than the established upper limits for background opacity and permeability values for bovine corneae treated with the respective negative control.
- Acceptance criteria met for positive control: yes, the positive control resulted in an IVIS which was within two standard deviations of the current historical mean.

Any other information on results incl. tables

 

Table 1: Details on results

Test Group	Opacity value = Difference (t240-t0) of Opacity		Permeability at 490 nm (OD490)		IVIS	Mean IVIS	Standard Deviation in vitro score	Proposed in vitro Irritancy Score
		Mean		Mean
Negative Control	0	0.00	0.067	0.062	1.01	0.94	0.06	No Category
	0		0.059		0.89
	0		0.061		0.92
Positive Control	95.00*		0.161*		97.41	106.87	8.61	Category 1
	111.00*		0.217*		114.25
	107.00*		0.130*		108.95
Test Substance	1.00*		0.010*		1.15	2.27	0.97	No Category
	2.00*		0.056*		2.84
	2.00*		0.055*		2.82

* corrected values

 

Table 2: Historical control data

	Positive Control	Negative Control
Mean IVIS	113.39	1.29
Standard Deviation of IVIS	12.73	0.32
Range of IVIS	87.39 - 146.55	0.77 - 2.85
95 % Control limits of IVISpos	87.92-138.85
Mean Opacity t240min	110.20	0.25
Standard Deviation of Opacity t240min	18.98	0.32
Range of Opacity t240min	54.00 - 187.00	0.00 - 1.33
Mean Permeability	0.21	0.07
Standard Deviation of Permeability	0.29	0.01
Range of Permeability	-0.02 - 1.96	0.05 - 0.10

Values of 83 studies with solid test items sharing 46 sets of controls, performed between January 2016 and July 2019.

 

Applicant's summary and conclusion

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008.

Conclusions:

Under conditions of the Bovine Corneal Opacity and Permeability Test (BCOP) the test substance was not irritating to the eye. Application of the test substance to bovine corneas resulted in a calculated mean IVIS of 2.27.