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EC number: 946-959-7
CAS number: -
Table No. 1: OD570 values obtained at the MTT test, their averages, standard deviations (%) and relative viabilities.
OD570 (tissue 1,2,3)
99% MA (PC)
The test substance, Acid Black 227, was assayed for the in vitro eye irritation in human epidermal model EpiOcularTM. The test was performed according to the OECD Test Guideline No. 492: Reconstructed human Cornea-like Epithelium (RhCE) test method for identifying chemicals not requiring classification and labelling for eye irritation or serious eye damage. Details of the procedure are given in Protocol: EpiOcular™ Eye Irritation Test (OCL-200-EIT) for the prediction of acute ocular irritation of chemicals (MatTek 06/29/2015).
After pre-incubation and wetting of tissues, 50 mg of the test substance was placed directly atop to the tissue and it was spread on the entire tissue surface. Length of exposition was 6 hours at 37±1°C in humidified CO2 incubator (5±1% CO2). Three tissues were used for the test substance and every control. Two tissues more were used as colorant control to correction of possible colour interference, which was undergo the entire testing procedure excepting of incubation with MTT medium.
After removal of the test substance, tissues were post-soaked in medium for approximately 25 minutes and post-incubated for about 18 hours at culture conditions. Three hours incubation with MTT and two hours extraction period with shaking followed then. Optical density (OD570) of isopropyl alcohol extracts was measured on a plate reader. Relative cell viability was calculated for each tissue as % of the mean viability of the negative control tissues.
Test for colour interference revealed direct reduction caused with the test substance. “Viability” of tissues treated by the test substance was corrected.
Test for direct reduction was performed as a part of another study. Direct reduction was not discernible. With regard to the result of MTT, test assay in frozen tissues was not performed.
Under the above-described experimental design average viability of treated tissues was -2.7 % (after correction) i.e. viability was ≤ 60 %.
The effect of the test substance was positive in EpiOcularTM model (tissues were damaged).
According to the classification criteria given in chapter 4.5., the test substance, Acid Black 227, is identified as substance potentially requiring classification and labelling according to UN GHS (Category 2 or Category 1). Further testing with other test methods will be required.
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