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Eye irritation

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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Sep. 6-18, 2019, experimental phase: Sep. 9-12, 2019
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

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 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:
2019
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
Silicic acid, aluminum magnesium sodium salt
EC Number:
234-919-5
EC Name:
Silicic acid, aluminum magnesium sodium salt
Cas Number:
12040-43-6
Molecular formula:
Na(0.017-1.739)Mg(0.008-0.823)Al(0.002-0.2473)SiO(2.018-3.837)
IUPAC Name:
Aluminium (3+) magnesium (2+) sodium oxosilanediolate
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
Name: Silicic acid, magnesium aluminium sodium salt
Batch no.: 1901274010
Appearance: white fluffy powder
Composition: Sodium; Magnesium; Aluminium; Silica
Purity: >95%
Homogeneity: homogeneous

Test system

Controls:
yes, concurrent positive control
yes, concurrent negative control
Duration of treatment / exposure:
6 h
Duration of post- treatment incubation (in vitro):
18 h
Number of animals or in vitro replicates:
2
Details on study design:
- Negative Control: Sterile demineralised water, prepared by LAUS GmbH using an ion exchanger and membrane filtration through sterile filters, batch no.: T20190717.
- Positive Control: Methyl acetate (C3H6O2, CAS No. 79-20-9), procured from MatTek, batch no.: 021319ISC.
- Test System: Commercially available EpiOcular(TM) kit.
The EpiOcular tissue consists of normal, human-derived keratinocytes which have been cultured to form a stratified squamous epithelium similar to that found in the human cornea. It consists of highly organized basal cells. These cells are not transformed or transfected with genes to induce an extended life span. The EpiOcular tissues are cultured in specially prepared cell culture inserts with a porous membrane through which nutrients can pass to the cells. The tissue surface is 0.6 cm².
- MTT solution: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (=MTT), which can be reduced to a blue formazan, prepared by LAUS GmbH.
A MTT stock solution of 5 mg/mL in DPBS buffer was prepared and stored in aliquots of 2 mL at – 20 ± 5 °C. 2 mL of the stock solution were thawed and diluted with 8 mL of assay medium (resulting in 1 mg/mL). This MTT-solution with the concentration of 1 mg/mL was used in the test.

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Run / experiment:
relative tissue viability
Value:
ca. 86.3 - ca. 91
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation

Any other information on results incl. tables

Measured Values

As blank, the optical density of isopropanol was measured in eight wells of the 96-well-plate. The measured values and their mean are given in the following table:

Table 9.1-a : Absorbance Values Blank Isopropanol (OD at 570 nm)

Replicate              1              2               3               4             5             6             7             8              Mean

Absorbance       0.032       0.033       0.033       0.034       0.034       0.034       0.033       0.035       0.034

The absorbance values of negative control, test item and positive control are given in the following table:

Table 9.1-b: Absorbance Values Negative Control, Positive Control and Test Item (OD at 570 nm)

Designation        Measurement        Negative Control        Positive Control       Silicic acid, magnesium aluminium                                                                                                                                                   sodium salt

Tissue 1                    1                            1.878                     0.791                                          1.671

2                            1.889                     0.772                                          1.658

Tissue 2                    1                            1.771                     0.866                                          1.586

2                            1.761                     0.859                                          1.574

From the measured absorbances, the mean of each tissue was calculated, subtracting the mean absorbance of isopropanol as given in table 9.1-a (= corrected values).

Table 9.1-c: Mean Absorbance Negative Control, Positive Control and Test Item

Designation                     Negative Control        Positive Control        Silicic acid, magnesium aluminium sodium salt

Mean – blank (Tissue 1)       1.850                            0.748                                          1.631

Mean – blank (Tissue 2)       1.732                            0.829                                          1.546

Comparison of Tissue Viability

For the test item and the positive control, the following percentage values of tissue viability were calculated in comparison to the negative control:

Table 9.2-a: % Viability Positive Control and Test Item

Designation                     Positive Control               Silicic acid, magnesium aluminium sodium salt

% Viability (Tissue 1)              41.7%                                                 91.0%

% Viability (Tissue 2)              46.3%                                                 86.3%

% Viability Mean                     44.0%                                                88.7%

Eye hazard potential was assessed using the criteria given in the following table:

Table 9.3-a: Assessment of Eye Hazard Potential

% Viability               Assessment               UN GHS classification

> 60 %                    Non eye irritant              No Category

≤ 60 %                   At least eye irritant         No prediction can be made (category 1 or 2)

Validity criteria and results are stated in the following table:

Table 9.4 -a:

Validity                                           CriterionDemanded                                           Found    

Mean OD of negative control              > 0.8 and < 2.8                                                 1.8

% mean relative viability of

positive control                                   < 50% of negative control                                 44.0%

Variation within replicates                   < 20%                                                              6.6% (negative control)

4.5% (positive control)

4.7% (test item)

Values for negative control and for positive control were within the range of historical data of the test facility.

Therefore, the experiment was considered valid.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of the test, Silicic acid, aluminum magnesium sodium salt (CAS 12040-43-6, SMAS) is considered non-eye irritant in the EpiOcular(TM) Eye Irritation Test.
Executive summary:

The test item Silicic acid, aluminum magnesium sodium salt (CAS 12040-43-6, SMAS) was applied to a three-dimensional human cornea tissue model in duplicate for an exposure time of 6 hours.