Mice were treated with Guttiferone F (i.p. 20 mg/kg/day)

Human breast adenocarcinoma (MDAMB-231) , and human normal hepatic cells (HL-7702) were used for activity evaluation. Cells were cultured in RPMI 1640, DMEM, or DMEM/F12 medium, containing 10% fetal bovine serum. The cell lines were maintained at 37 °C in a humidified environment containing 5% CO2. To determine the effects of the compounds on cell viability, the cell number was quantified using a standard colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cells were plated in a 96-well plate (5 × 10^3 cells/ well) and allowed to attach overnight. Cells were treated with 5, 10, 20, and 40 μM of each compound (Guttiferone F) in culture medium for 72 h. Then, new culture medium was added with 20 μL of MTT (5 mg/mL stock in PBS) per well and incubated for 4 h at 37 °C. Finally, the culture medium was discarded, and 150 μL of DMSO was added per well to dissolve the purple formazan crystals. Absorbance of the solution was measured using a microplate reader spectrophotometer (Bio-Rad Laboratories, Inc., Hercules, CA, USA), at a wavelength of 490 nm