4.4. The Radical Scavenging Assay (The DPPH Assay)

For the radical scavenging assay, the 1,1-diphenyl-2-picrylhidrazine (DPPH, ≥98%, Sigma Aldrich) radical was used [60] and ascorbic acid (≥99%, Sigma Aldrich) was the reference compound. For the DPPH assay, 250 μL of extract diluted in ethanol (p.a., Kemika) was mixed with 500 μL of deionized water and prepared DPPH reagent in methanol (72 μg/mL, p.a., Kemika). The reaction mixture was kept in the dark for 30 min, after which the absorbance of the solution was measured at 517 nm in a 96-well plate. Appropriate blanks (ethanol) and standards (ascorbic acid solutions in ethanol) were run simultaneously. Results were expressed as milligram ascorbic acid equivalents per gram of sample (mg AAE/g sample).