2.7. MTT Assay

Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay following the published procedure [40]. Briefly, cells were seeded at 10⁵/cm² density in 96-well plates. Twenty-four hours later, the medium was changed and supplemented with the specified concentrations of metabolite (from 0.5 to 10 mM in DMSO) for 24 and 48 h. MTT solution 1:10 (stock solution 5 mg/mL) was added to each well and the absorbance was measured in dual-wavelength mode (570 nm and 630 nm). The percentage of cell viability was calculated as follows: mean (A570–A630) and compared to cells supplemented with DMSO alone. Values shown in the plot are mean ± SD of sixfold determinations. Mean and the standard deviation was calculated on biological triplicates using GraphPad Prism8 software (GraphPad, San Diego, CA, USA).