Evaluation of compound cytotoxicity on human hepatocytes. Human hepatocytes (HC-04 cells) were treated with 1 mM of the compounds, and then analysed for the expression of apoptotic markers [via Annexin V-APC (FL4) and Propidium Iodide (FL2) staining] by flow cytometry. Representative examples of a sub-set of the compounds tested. Recovered cells were then analysed for the expression of both apoptotic markers Annexin-V and PI. At this concentration, none of the compounds induced heightened expression of either apoptotic markers [via Annexin V-APC (FL4) and Propidium Iodide (FL2) staining] by flow cytometry. Note that for compound 24, the fluorescence peaks for Annexin-APC are shifted slightly to the right, indicating that at higher concentrations, some cytotoxicity would likely be observed. However, the compound dosage used here—1 mM—is already much higher than would typically be observed under physiological conditions.