DPPH Radical Scavenging Activity Assay

The 2,2-diphenyl-1-picrylhydrazyl (DPPH; Thermo Fisher Scientific, Ward Hill, MA, USA) free radical scavenging activity was measured using a previously reported method [23 (link)]. Briefly, a DPPH solution (0.2 mM, 100 μL) was mixed with a sample (100 μL) on a 96-well plate and reacted for 30 min in the shade. The absorbance was measured at 517 nm using a micro reader (Epoch, Biotek Instruments, Inc., Winooski, VT, USA). Ascorbic acid (100 μg/mL) (Sigma-Aldrich, Co., St. Louis, MO, USA) was used as the positive control group. The difference in DPPH radical scavenging activity between the negative control and treated sample was calculated with the following formula: %EC = (A control − A sample) * 100/(A control); A sample, absorbance of the sample; A control, absorbance of untreated sample.