Briefly, for the cytotoxicity assay, the human prostate cancer cell line PC-3 and the colon adenocarcinoma cancer cell line HT-29 (both from ATCC, Manassas, VA, USA) were used. The cell handling and assay techniques were in accordance with the method described by Khan et al. [34 (link)]. The extract was tested at the concentrations of 0.05 and 50 μg/mL. Anti-proliferative and cytotoxic effects of the extract were investigated by performing colorimetric MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and CV (crystal violet)-based cell viability assays (Sigma-Aldrich, Taufkirchen, Germany) after 48 h treatment time, respectively. The absorbance was measured with an automated microplate reader at 540 nm with a reference wavelength of 670 nm. Digitonin (125 μM) was used as positive control, which was set for data normalization to 0% cell viability. The results are presented as a percentage of control values obtained from untreated cultures.
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