Cytotoxicity against HepG2 cell line

HepG2 cells (ATCC HB-8065) were grown in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% of Fetal calf serum (FCS), 100 U/mL penicillin, 2 mM L-glutamine, 100 μg/mL streptomycin and 1 mM Na-pyruvate and were incubated in a 5% humified CO₂ atmosphere at 37 °C. The confluent cell layer was harvested using 0.5 mM trypsin/EDTA. MTT (3–4, 5-dimethylthiazol-2-yl) − 2, 5-diphenyltetrazolium bromide), a tetrazolium dye was used to access the cytotoxic potential of different leaf extracts in vitro. In this assay,, MTT become reduced into its insoluble purple product formazan which is measured spectrophotometrically. In a 96-well plate, pre-seeded HepG2 cells (> 90% viability; 1 × 10⁴ cells/well or 10,000 cells per well) were treated with 200 μg/mL of test samples for 24 h. Later, 10 μL of MTT dye (5 mg/mL) was added per well, followed by incubation of 3 h. Insoluble formazan was then dissolved by adding 10% acidified sodium dodecyl sulfate (SDS). Cells were then incubated overnight. Plates were analyzed at 570 nm using a microplate reader (Platos R 496, AMP). Non-treated HepG2 cells (NTC) were included as control. DMSO was used as a negative control for plant extracts. Optical Density of treated samples and NTC was measured at 570 nm. Percent (%) viability was calculated relative to the NTC sample using the following formula: