2.3. Cell Viability Assay

Cell lines were seeded into 96-well plates at 5 × 10³ (HepG2) and 3 × 10³ (Hep3B and HH4) cells/well 24 h before treatment. Increasing concentrations of the monomer trans-resveratrol, the dimers ampelopsin A and trans-ε-viniferin and the tetramers isohopeaphenol, hopeaphenol, R2-viniferin and R-viniferin were then added and cells were incubated for 24, 48, and 72 h. Stilbenes were dissolved in dimethyl sulfoxide (DMSO), Sigma-Aldrich, St Louis, MO, USA) at a final concentration of 0.01%. The same amount of DMSO was added to control cells. After treatment, the cell viability was determined using the crystal violet assay [18]. The absorbance was recorded at 590 nm in a Synergy HT microplate reader (BioTek, Winooski, VT, USA). Cell viability was calculated as the percentage of viable cells treated with stilbene versus untreated control cells using the following equation: Cell viability (%) = [OD (Treatment) − OD (Blank)]/[OD (Control) − OD (Blank)] × 100. The cytotoxic effect of stilbenes was determined by calculating IC₅₀ values using non-linear regression analysis (GraphPad Prism 6, San Diego, CA, USA).