Total Phenolic and Flavonoid Contents.
Total phenolic (TPC) and total flavonoid (TFC) content of authenticated HMF, MMF, and EMF standards was determined using a model EL 340 UV/vis microplate reader (Biotek, Inc., Winooski, VT). A modified Folin− Ciocalteu assay was employed to determine the TPC value measured at 765 nm against reagent blank.25 TPC was expressed as milligrams of gallic acid equivalents per gram of standard (mg GAE/g) determined using the standard calibration curve of gallic acid with a linear range of 0.0156−0.5 mg/mL (r = 0.999). Total flavonoid content (TFC) was determined using a modified 96-well microplate method.26 Absorbance was measured at 510 nm and TFC value was expressed as milligrams of catechin equivalents per gram of standard (mg CAE/g) using a calibration curve of (+)-catechin with linear range of 0.0156−0.5 mg/mL (r = 0.999)