3.5. DPPH Radical-Scavenging Assay

The DPPH radical-scavenging assay analyzes antioxidant behavior based on an electron-transfer reaction. The change in color from purple to yellow is observed as DPPH reacts with any antioxidant compounds present. The reaction mixture consisted of a solution of antioxidant compounds (100 μL) and an equal volume of DPPH solution (0.1 mM) in ethanol, and was incubated for 30 min at room temperature in the dark. Ascorbic acid was used as the positive control. DPPH reaction was monitored by measuring the absorbance (Abs.) at 517 nm by using a microplate reader (PowerWave XS; Bio-Tek Instruments, Winooski, VT, USA). The radical-scavenging activity is presented as an IC₅₀ value, which is the concentration of the antioxidant compounds required to inhibit 50% of the free radicals, and was calculated using the following equation: Scavenging activity (%) = [(Control Abs. − Sample Abs.)/Control Abs.] × 100.