Sodium 2-stearoyllactate

Endpoint summary

Description of key information

Skin irritation in vitro (OECD 439, EpiDerm): not irritating

Eye irritation in vitro (OECD 437, BCOP): not irritating

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

12 Sept - 11 Oct 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

adopted in 2019

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Version / remarks:

adopted in 2012

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

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

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: EpiDermTM (EPI-200)

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ (EPI-200-SIT) (MatTek Corporation, 82105 Bratislava, Slovakia)
- Tissue batch number(s): 30830
- Date of initiation of testing: 12 Sept 2019

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 35 min at 37 ± 1.5 °C and 5 ± 0.5% CO2 in Dulbecco's Minimum Essential Medium (DMEM) followed by 25 min in a sterile bench at room temperature
- Temperature of post-treatment incubation: approx. 42 h at 37 ± 1.5 °C and 5 ± 0.5% CO2 in DMEM

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: rinsed with Phosphate Buffered Saline (PBS) several times

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 300 μl/well
- Incubation time: 3 h ± 5 min
- Spectrophotometer: microplate reader (Versamax® Molecular Devices, Software SoftMax Pro Enterprise version 4.7.1)
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: The quality of the EpiDerm™ tissue was assessed by an MTT cell viability test. The determined OD (540 - 570 nm) was 1.946 ± 0.146 (acceptance criteria: 1.0 - 3.0).
- Barrier function: The barrier function was assessed by determination of the exposure time required to reduce tissue viability by 50% (ET-50) upon application of 100 µL of 1% Triton X-100. The ET-50 value was determined to be 5.05 h (acceptance criteria: 4.77-8.72 h).
- Contamination: The cells used to produce the EpiDerm™ tissue were screened for the presence of viruses, bacteria, yeast and other fungi. No contamination was detected.

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
Since the test substance did not directly reduce the MTT solution, an additional functional check was not performed.

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: single experiment

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be irritant or corrosive to skin if the viability after 1 h exposure is ≤ 50%.
- The test substance is considered to be non-irritant to skin if the viability after 1 h exposure is > 50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 25 mg (corresponding to 39.7 mg/cm2) + 25 µL DPBS

Duration of treatment / exposure:

35 min at 37 °C and 25 min at RT

Duration of post-treatment incubation (if applicable):

Approx. 42 h

Number of replicates:

Triplicates for each treatment and control group

Irritation / corrosion parameter:

% tissue viability

Remarks:

mean value of 3 tissues

Run / experiment:

1 h exposure

Value:

100.16

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Direct-MTT reduction: Since the test substance did not directly reduce MTT, an additional test with freeze-killed or viable tissues was not performed.
- Colour interference with MTT: The test substance did not change the colour, when mixed with deionised water and thus passed the colour interference pre-test. Also its intrinsic colour was not intensive.

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: The mean OD of the tissue replicates treated with the negative control was ≥ 0.8 and ≤ 2.8 for every exposure time (values between 1.689 and 1.783).
- Acceptance criteria met for positive control: The mean viability of the tissue replicates treated with the positive control for 1 h was < 15% compared to the negative control (4.32%).
- Acceptance criteria met for variability between replicate measurements: The standard deviations between the three tissue percentage viability values of each group (test item, positive and negative controls) in the main test were below 18.

Table 1: Details on results
Treatment Group	Tissue No.	OD 570 nm			Mean OD of 3 Wells	Mean OD of 3 Wells blank corrected	Mean OD of 3 tissues	Rel. Viability [%] Tissue 1, 2 + 3	Standard Deviation	Mean Rel. Viability [%]
		Well 1	Well 2	Well 3
Blank		0.038	0.039	0.039	0.039
Negative Control	1	1.734	1.713	1.736	1.727	1.689	1.751	96.469	0.054	100.0
	2	1.831	1.818	1.807	1.819	1.780		101.677
	3	1.825	1.783	1.858	1.822	1.783		101.854
Positive Control	1	0.112	0.126	0.114	0.117	0.079	0.076	4.494	0.005	4.32
	2	0.117	0.117	0.117	0.117	0.078		4.469
	3	0.108	0.109	0.109	0.109	0.070		3.989
Test Item	1	1.861	1.795	1.783	1.813	1.774	1.753	101.336	0.082	100.16
	2	1.708	1.702	1.694	1.701	1.663		94.982
	3	1.838	1.886	1.864	1.862	1.824		104.175

 

Table 2: Historical control data
Positive Control; OD at 570 nm after exposure to 5% SDS solution in deionized water (MatTek)		Negative Control OD at 570 nm DPBS (MatTek)
Tissue Viability [%]	3.89	Mean OD	1.69
Standard Deviation	1.01 % points	Standard Deviation	0.19
Range of Viabilities	2.24 % - 6.19 %	Range of OD*	1.28 - 2
Mean OD	0.07	* should be 0.8 - 2.8 (OECD 439) or 1.0 - 2.5 (MatTek)
Standard Deviation	0.02
Range of OD	0.03 - 0.11
Data of 50 sets of controls shared between 195 studies performed from August 2015 until May 2019 (p.p.: percentage points)

 

Table 3: Technical proficiency
Proficiency Substance	Viability [%]	Category
Naphtalene acetic acid	101.7	No Cat.
Isopropanol	85.5	No Cat.
Methyl stearate	91.1	No Cat.
Heptyl butyrate	109.0	No Cat.
Hexyl salicylate	98.1	No Cat.
Cyclamen aldehyde	24.1	Cat. 2
1-Bromohexane	16.3	Cat. 2
Potassium hydroxide (5% aq.)	40.1	Cat. 2
1-Methyl-3-phenyl-1-piperazine	21.6	Cat. 2
Heptanal	20.6	Cat. 2
In vitro Skin Irritation Assay: OECD 439
Proficiency Data (March 2014, non-GLP) performed at ICCR-Roßdorf GmbH Study
Director: Dipl.-Ing. Andreas Heppenheimer

Interpretation of results:

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

Conclusions:

Under the conditions of the conducted test, the test substance did not possess irritating properties towards reconstructed human epidermis tissue in the EpiDerm™ model.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

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

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

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

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.

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.

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.

 

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.

 

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.

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

12 Sept - 10 Oct 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Version / remarks:

adopted in 2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

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

Species:

human

Strain:

other: EpiOcular™

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 50 mg (corresponding to 83.3 mg/cm2 of the solid test item)

NEGATIVE CONTROL SUBSTANCE
- Amount(s) applied: 50 µL

POSITIVE CONTROL SUBSTANCE
- Amount(s) applied: 50 µL
- Concentration: ≥ 98%

Duration of treatment / exposure:

6 h at 37 ± 1.5 °C and 5 ± 0.5% CO2 in Dulbecco's Minimum Essential Medium (DMEM)

Duration of post- treatment incubation (in vitro):

25 min at RT and approx. 18 h at 37 ± 1.5 °C and 5 ± 0.5% CO2 in DMEM

Number of animals or in vitro replicates:

Duplicates for each treatment and control group

Details on study design:

- Details of the test procedure used : The EpiOcular™ Eye Irritation Test (EIT) consists of a topical exposure of the neat test item to a human reconstructed cornea model followed by a cell viability test. Cell viability is measured by dehydrogenase conversion of MTT [(3-4,5-dimethyl thiazole 2-yl) 2,5-diphenyl-tetrazoliumbromide], present in cell mitochondria, into a blue formazan salt that is quantitatively measured after extraction from tissues. The percent reduction of cell viability in comparison of untreated negative controls is used to predict the eye irritation potential.

- RhCE tissue construct used, including batch number : Standard EpiOcular™ Eye Irritation Test (EIT) kit purchased from MatTek Corporation (82105 Bratislava, Slovakia), Lot No.: 30629

- Indication of controls used for direct MTT-reducers and/or colouring test chemicals: The ability of the test substance to directly reduce MTT and to form a blue/purple reaction product was assessed in a pre-experiment. Since the MTT solution colour did not turn blue/purple, the test substance is not presumed to have reduced the MTT. An additional test with freeze-killed tissues did not have to be performed.

- Wavelength used for quantifying MTT formazan: 570 nm

- Description of the method used to quantify MTT formazan : The absorbance (OD570) was measured with a plate reader (Versamax® Molecular Devices, Software Softmax Pro, version 4.7.1).

- Description of evaluation criteria used including the justification for the selection of the cut-off point for the prediction model : The test substance is considered to be not irritating to the eye if the tissue viability after 30 min exposure, 12 min post-exposure immersion and 120 min post-exposure incubation is > 60% relative to the negative control treated tissue.

- Reference to historical data of the RhCE tissue construct : Historical control data are summarised in 'Any other information on results incl. tables' below.

- Demonstration of proficiency in performing the test method before routine use by testing of the proficiency chemicals : The technical proficiency of the test facility was demonstrated and certified by the manufacturer of the test system (MatTek Corporation (Bratislava, Slovakia).

- Acceptable variability between tissue replicates for positive and negative controls : The results are acceptable if the negative control OD is > 0.8 and < 2.5 and if the mean relative viability of the positive control is below 50% of the negative control viability.

- Acceptable variability between tissue replicates for the test chemical: The results are acceptable if the difference of viability between the two relating tissues of a single test item is < 20% in the same run (for positive and negative control tissues and tissues of test items).

Irritation parameter:

other: % tissue viability

Run / experiment:

6 h exposure

Value:

52.02

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 and certified by the manufacturer of the test system (MatTek Corporation (Bratislava, Slovakia).

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD of the tissue replicates treated with the negative control was ≥ 0.8 and ≤ 2.8 for every exposure time (values between 1.939 and 2.045).
- Acceptance criteria met for positive control: The mean viability of the tissue replicates treated with the positive control was < 50% compared to the negative control (31.07%).
- Acceptance criteria met for variability between replicate measurements: The difference of relative viability between the two relating tissues was < 20%. (values between 0.22% and 10.11%) in the same run (for test item tissues, positive and negative control tissues).

 

Table 1: Details on results

Test Group	Tissue No.	Well 1 [OD570]	Well 2 [OD570]	Mean [OD570] (Well 1 and well 2)	Mean [OD570] blank corr. (Well 1 and well2)	Mean [OD570] of T1 and T2	Tissue viabil. [%]	Viabil. of T1 and T2 [%]	Diff. of viabil. between T1 and T2 [p.p.]
Blank		0.037	0.037	0.037
Negative Control	1	2.136	2.029	2.082	2.045	1.992	100.0	102.7	5.35
	2	2.024	1.928	1.976	1.939			97.3
Positive Control	1	0.659	0.649	0.654	0.617	0.619	31.07	31.0	0.22
	2	0.665	0.652	0.658	0.621			31.2
Test Item	1	0.986	0.960	0.973	0.936	1.036	52.02	47.0	10.11
	2	1.183	1.165	1.174	1.137			57.1

 

Table 2: Historical control data

Positive Control		Negative Control [OD570]
Mean Viability	27.04%	Mean Absorption	1.54
Standard Deviation	10.39 p.p.	Standard Deviation	0.248
Range of Viabilities	6.73% - 42.54%	Range of Absorbance	1.02 – 2.09
Mean Absorption	0.463
Standard Deviation	0.169
Range of Absorbance	0.078 – 0.776

 

Data of 51 studies performed from July 2015 until March 2019.

 

Interpretation of results:

other: no prediction possible

Conclusions:

After treatment with the test substance a mean relative viability value of 52.02% was measured compared to the mean value of the negative control. This value is below the threshold for irritancy of ≤ 60%. Therefore, no prediction of the eye irritation potency can be made for the test substance in this study and under the experimental conditions reported. Furthermore, RhCE test methods show a high percentage of false positive results. In conclusion, additional data are required to conclude on the hazard assessment and classification & labelling of the test substance.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation / corrosion

The skin irritation potential of esterification product of C16-C18 even numbered, saturated fatty acids and lactic acid, sodium salt (sodium stearoyl lactylate, SSL, CAS No. 25383-99-7, EC No. 246-929-7) was assessed in the in vitro Human Skin Model Test (EpiDerm™) according to OECD guideline 439 under GLP conditions (Muchalla, 2019, EpiDerm). The test item did not prove to be a MTT reducer and it did not change colour when mixed with deionised water or isopropanol. Three tissues of the human skin model EpiDerm™ were treated with the test item, the negative control Dulbecco's Phosphate Buffered Saline (DPBS) and the positive control sodium dodecyl sulphate (5% SDS) for 60 min, respectively. After treatment with the negative control the absorbance values were well within the required range of the acceptability criterion thus assuring the quality of the tissues. Treatment with the positive control induced a sufficient decrease in the relative absorbance as compared to the negative control thus assuring the validity of the test system. After treatment with the test item, the mean relative viability value was 100.16% compared to the relative absorbance value of the negative control. This value is above the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential.

Eye irritation / serious eye damage

An in vitro study was performed to assess the eye irritation potential of SSL by means of the Human Cornea Model Test (EpiOcular) according to OECD guideline 492 under GLP conditions (Sokolowski, 2019, EpiOcular). The test item did not reduce MTT and its intrinsic color was not intensive and the optical density (OD) of the test item in deionised water or isopropanol at 570 nm after blank correction was < 0.08. Each 50 mg of the test item or 50 μL of the negative control (deionised water) and of the positive control (methyl acetate), were applied to each duplicate tissue for 6 h. The mean OD of the tissue replicates treated with the negative control was > 0.8 and < 2.5, thus showing the quality of the tissues. Treatment with the positive control induced a decrease below 50% viability compared with the negative control value in the relative absorbance, thus ensuring the validity of the test system. All other quality and validity criteria were also met. After treatment with the test item a mean relative viability value of 52.02% was measured compared to the mean value of the negative control. This value is below the threshold for irritancy of ≤ 60%. Therefore, no prediction of the eye irritation potency can be made for the test substance in this study and under the experimental conditions reported. Furthermore, test methods based on reconstructed human epithelium (RhCE) show a high percentage of false positive results. In conclusion, additional data are required to conclude on the hazard assessment and classification and labelling of the test substance.

A second in vitro study was performed to assess the corneal irritation and damage potential of SSL using the Bovine Corneal Opacity and Permeability test method (BCOP) according to OECD guideline 437 and observing GLP conditions (Sokolowski, 2020, BCOP). After a first opacity measurement of the fresh bovine corneae, the 20% (w/v) suspension in saline of the test item as well as the positive and the negative controls were each applied to different corneae and incubated for 240 min at 32 ± 1 °C. After the incubation phase the test item, the positive controls (10% (w/v) benzalkonium chloride in saline) and the negative controls (physiological saline) were each rinsed from the corneae and opacity was measured again. After the opacity measurements, permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein after incubation for 90 min at 32 ± 1 °C. With the negative control, neither an increase of opacity nor permeability of the corneae was observed. The positive control showed clear opacity and distinctive permeability of the corneae. Relative to the negative control, the test item did not cause a relevant increase of the corneal opacity or permeability. The calculated mean in vitro irritancy score (IVIS) was 2.27. According to OECD guideline 437 the test item does not need to be classified for eye irritation or serious eye damage.

Justification for classification or non-classification

The available data on skin and eye irritation of esterification product of C16-C18 even numbered, saturated fatty acids and lactic acid, sodium salt (sodium stearoyl lactylate, SSL, CAS No. 25383-99-7, EC No. 246-929-7) do not meet the criteria for classification according to Regulation (EC) No. 1272/2008 (CLP) and are therefore conclusive but not sufficient for classification.