3.5. Alpha-Glucosidase Inhibition Assay

The alpha-glucosidase (0.2 U/mL) and substrate (5.0 mM p-nitrophenyl-α-D-glucopyranoside) were dissolveed in 100 mM of pH 6.9 sodium phosphate buffer [27]. The inhibitor (50 µL) was preincubated with alpha-glucosidase at 37 °C for 20 min, and then the substrate (40 µL) was added to the reaction mixture. The enzymatic reaction was carried out at 37 °C for 20 min and stopped by adding 0.2 M of Na₂CO₃ (130 μL). Enzymatic activity was quantified by measuring absorbance at 405 nm. All samples were analyzed in triplicate at five different concentrations around the IC₅₀ values, and the mean values were retained. The inhibition percentage (%) was calculated by the following equation:

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