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Description of key information

Skin

In an in vitro skin irritation study (OECD 439) a mean tissue viability of 99.2 % was determined.

In an in vitro skin corrosion study (OECD 431) a mean tissue viability of 102.4 % after 3 min exposure and 107.6 % after 1 h exposure to the test substance was determined.

Eye

In an in vitro eye irritation study (OECD 437) an In Vitro Irritancy score (IVIS) of 0.0 was determined.

In an in vitro Reconstructed Human Cornea like Epithelium (RhCE) eye irritation study a tissue viability of 104.4 % was observed.

Key value for chemical safety assessment

Skin irritation / corrosion

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Two in vitro test batteries were performed in a weight of evidence approach in order to evaluate if the test substance is irritating or corrosive to either skin or eye.

Skin

The objective was to assess the potential for corrosive activity and skin irritation of the test substance. Using the currently available methods a single in vitro assay may not always be sufficient to cover the full range of skin irritating/corrosion potential. Therefore, two in vitro assays were part of this in vitro skin irritation and corrosion test strategy: The Skin Corrosion Test (SCT) according to OECD guideline 431and Skin Irritation Test (SIT) according to OECD guideline 439.

The potential of the test substance to cause dermal corrosion/irritation was assessed by a single topical application of 25 μL bulk volume (about 13 mg) of the undiluted test substance to a reconstructed three dimensional human epidermis model (EpiDerm™).

For the corrosion test two EpiDerm™ tissue samples were incubated with the test substance for 3 minutes and 1 hour, respectively. The irritation test was performed with three EpiDerm™ tissue samples, which were incubated with the test substance for 1 hour followed by a 42-hours post-incubation period.

Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the testsubstance treated epidermal tissues is compared to that of negative control tissues. The quotient of the values indicates the relative tissue viability.

The EpiDerm™ skin corrosion/irritation test showed the following results:

The test substance is not able to reduce MTT directly.

Corrosion test:

The mean viability of the test-substance treated tissues determined after an exposure period of 3 minutes was 102.4 %, and it was 107.6 % after an exposure period of 1 hour.

Irritation test:

The mean viability of the test-substance treated tissues determined after an exposure period of 1 hour with about 42 hours post-incubation was 99.2 %.

Based on the observed results and applying the evaluation criteria, it was concluded, that the test substance does not show a skin irritation potential in the EpiDerm™ in vitro skin irritation and corrosion test strategy under the test conditions chosen.

Eye

The objective of the present study was the determination of a possible eye irritating potential of test substance. Using the currently available methods a single in vitro assay may not always be sufficient to cover the full range of eye irritating potential. Therefore, two in vitro assays were part of this in vitro eye irritation test strategy: The Bovine Corneal Opacity and Permeability Test (BCOP Test) according to OECD guideline 437 and EpiOcular Eye Irritation Test according to OECD guideline 492.

BCOP

The potential of the test substance to cause ocular irritation or serious damage to the eyes was assessed by a single topical application of 750 μL of a 20 % test-substance preparation to the epithelial surface of isolated bovine corneas. Three corneas were treated with the test-substance preparation for an exposure period of 4 hours. In addition to the test substance a negative control (NC; de-ionized water) and a positive control (PC; 20 % imidazole in de-ionized water) were applied to three corneas each. Corneal opacity was measured quantitatively as the amount of light transmitted through the cornea. Permeability was measured quantitatively as the amount of sodium fluorescein dye that passes across the full thickness of the cornea. Both measurements were used to calculate an In Vitro Irritancy Score of the test substance. The following results were obtained in the BCOP Test:

 

Mean Opacity Value

Mean Permeability Value

Mean In Vitro Irritancy Score

Test substance

0.0

0.003

0.0

NC

8.4

0.002

8.4

PC

86.2

2.625

125.6

EpiOcular

The potential of the test substance to cause ocular irritation was assessed by a single topical application of 50 μL bulk volume (about 17 mg) of the undiluted test substance to a reconstructed three dimensional human cornea model (EpiOcular™).

Two EpiOcular™ tissue samples were incubated with the test substance for 6 hours followed by a 18-hours post-incubation period. Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the test substance treated epidermal tissues is compared to that of negative control tissues. The ratio of the values indicates the relative tissue viability.

The EpiOcular™ eye irritation test showed the following results:

The test substance is not able to reduce MTT directly. The mean viability of the test-substance treated tissues was 104.4 %.

Summary of the individual test results of the in vitro eye irritation turnkey testing strategy:

Test method

Test result

Test Evaluation

Evaluation Test Strategy

BCOP Test

The mean IVIS of the

test-substance treated

corneas was 0.0

not identified as corrosive

or severe irritant

non-irritant

EpiOcular

Mean viability of the

test-substance treated

tissues was 104.4 %

non-irritant

Based on the results for BCOP and EpiOcular Test and considering the evaluation criteria, the test substance does not show an eye irritation potential in the in vitro eye irritation test strategy under the test conditions chosen.

Justification for classification or non-classification

Based on the study results of the in vitro test battery and taking into account the provisions laid down in Regulation (EC) No 1272/2008 as amended for the ninth time in Regulation (EU) No 2016/1179, the test substance does not need to be classified as irritating or corrosive to eye or skin.