2.7.3. ABTS Radical Scavenging Assay

ABTS assay was done using the modified methods from Re et al. and Kenny et al. [41,42]. ABTS solution (7 mM) was mixed with potassium persulfate (2.45 mM) and incubated in the dark for 16 h to form ABTS radical cations (ABTS^•+). This solution was diluted with ethanol until reaching an absorbance of 0.70 ± 0.05 at 734 nm. A total of 2.0 mL of this solution was mixed with 0.5 mL of test sample. The mixture was vortexed and incubated for 6 min in a dark condition. Test sample mixed with ethanol was used as the blank, and diluted ABTS^•+ solution without mixing the test sample was used as the control. The absorbance was measured with the same UV–vis spectrophotometer described above at 734 nm. The ABTS scavenging percentage of each sample was estimated according to Equation (3). IC₅₀ values were calculated using the dose–response model.

where A_sample: test sample with ABTS^•+ solution absorbance; A_blank: blank absorbance; and A_control: control absorbance. The result of the testing sample was expressed as ABTS IC₅₀ value (μg/mL).