Evaluating Cytotoxic Effects of Organic Extracts

Evaluation of the effect of each crude organic extract and fractions A-H, as well as subfractions C1-C6 and C3–1–C3–12 on cell viability was performed using the standard XTT assay and an established protocol [44 (link)]. In brief, HCT116 cells were seeded in 96-well plates (10⁴ cell/well) and 24 h later were treated for a period of 24 h with two doses from the crude extract and fractions A-H; 200 and 400 μg/mL, and with 4 doses for each subfraction; 15, 25, 50, and 100 μg/mL. Medium and DMSO were added to control wells. For sub-fraction C3–5, the XTT assay was additionally conducted using 30 μg/mL for 24 h. Following treatment, cell viability was determined by the XTT assay (Biological Industries, Beit Haemek, Israel) according to the manufacturer’s instructions using a plate reader (version, BioTek, Winooski, VT, USA). Experiments were repeated 3 times. Data were presented as the average proliferation percentage of the respective control.