F4 fraction has no activity on mitochondrial membrane depolarization. A. K562 human cells were treated with F4 fraction (31.2 μg/ml), or positive control S2 fraction (15.6 μg/ml) or ethanol (0.2%) for 4, 8 and 12 h. All cells were stained with JC-1 (Sigma) dye and analyzed by flow cytometry (FACScalibur CellQuest software program) (Becton Dickinson). Bars represent cell percentage (%) ± SEM of depolarized (filled bars) or non-depolarized (hatched bars) cells, representing two independent experiments. ***p < 0.001 versus control (ethanol 0.2%; Unpaired Student's t-test). B. A375 cells treated with ethanol (0.2%) (left panel) or F4 fraction (31.2 μg/ml) (right panel) for 24 h were permeabilized, stained with DAPI and analyzed under fluorescence microscope (Olympus). Results show photos representing four independent experiments.