Registration Dossier

Administrative data

Description of key information

Based on the results of the in vitro tests, the test substance is considered to be not irritating to skin and eyes.

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

In vitro skin irritation study:

An in vitro study was conducted to determine the skin irritation potential of the test substance according to OECD Guideline 439 and EU Method B.46 (Reconstructed Human Epidermis Test Method), in compliance with GLP. Human three dimensional epidermal models (triplicates) were exposed to 25µL test substance for 15 min. After a 42 hour post-incubation period, a determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity was expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment. Skin irritation was expressed as the remaining cell viability after exposure to the test substance. The positive control (25 µL 0.5% SDS) had a mean cell viability of 5.3% after 15 ± 0.5 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control (25 µL PBS) tissues was within the laboratory historical control data range. The relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with the test substance compared to the negative control tissues was 99%. The standard deviation value of the percentage viability of three tissues treated identically was less than 16%, indicating that the test system functioned properly. Th experiment was considered valid. Since the mean relative tissue viability for the test substance was above 50% after 15 ± 0.5 minutes treatment, the test substance was considered to be non-irritant to human skin under the study conditions (Eurlings, 2016).

In vitro eye irritation studies:

1. An in vitro study was conducted to determine the eye irritation potential of the test substance according to OECD guideline 437 (bovine corneal opacity and permeability (BCOP) test), in compliance with GLP. Bovine corneas were exposed to the test substance for a period of 10 minutes (followed by a post-incubation of 2h with fresh medium). Opacity and permeability were measured and an in vitro irritancy score was then established. Physiological saline and ethanol were used as negative and positive controls, respectively. The corneas treated with the test substance showed opacity values ranging from 0.0 to 4.5 and permeability values ranging from 0.036 to 0.172. The corneas were slightly translucent after the treatment. No pH effect of the test substance was observed on the rinsing medium. The in vitro irritancy scores ranged from 0.6 to 5.2 after 10 minutes of treatment. The negative and positive control values were within the expected range; hence the experiment was considered valid. Under the study conditions, the test substance induced ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 3.3 after 10 minutes of treatment. Since the test substance induced an IVIS > 3 ≤ 55, no prediction on the classification can be made (Eurlings, 2016).

Another in vitro study was conducted to determine the eye irritation potential of the test substance according to OECD Guideline 492 (EpiOcular™ Cornea Epithelial Model), in compliance with GLP. The EpiOcular tissue construct is a nonkeratinized epithelium prepared from normal human keratinocytes. It models the cornea epithelium with progressively stratified, but not cornified cells. The test consisted of an application of the test substance (50 µL) to the surface of the cornea epithelial construct for 30 minutes. After exposure, the cornea epithelial construct was thoroughly rinsed to remove the test substance and transferred to fresh medium for an immersion incubation. After transfer to fresh medium for a 2 hours incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) at the end of the treatment (mean absorption measurement - optical density reading at 570 nm).  Eye hazard potential is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 30 ± 2 minutes treatment with the test substance compared to the negative control tissues was 85%. Since the mean relative tissue viability for the test substance was above 60%, the test substance was considered to be non-irritant. The positive control had a mean cell viability of 27% after 30 ± 2 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was 1.5, which is within the acceptability range from > 0.8 to < 2.5. The standard deviation value of the percentage viability of two tissues treated identically was less than 11%, indicating that the test system functioned properly. It was concluded that this test was valid. Under the study conditions, the test substance was considered to be non irritant to human eye (Verbaan, 2017).

Justification for classification or non-classification

Based on the results of the in vitro skin and eye irritation tests, the test substance does not meet the criteria for classification for this endpoint according to CLP (Regulation 1272/2008/EC) criteria.