Determination of α-glucosidase inhibition

The α-glucosidase inhibition by the samples was assessed according to the method described by Zeng et al. (12) with slight modifications. Briefly, the reaction mixture contained 25 μL of 10 mM p-nitrophenyl-α-d-glucopyranoside (PNPG; Sigma-Aldrich) and 25 μL of the sample preincubated at 37 °C for 10 min. The reaction was initiated by the addition of 50 μL α-glucosidase solution (0.16 U/mL; Sigma-Aldrich) diluted with 0.1 M phosphate buffer, and incubated at 37 °C for 30 min. The reaction was terminated by adding 100 μL of 0.1 M Na₂CO₃ (Sigma-Aldrich). The enzymatic activity was quantified based on the measurements of the absorbance of the samples at 405 nm on a Multiskan MK3 plate reader (Thermo Fisher Scientific). Each test sample was analyzed in technical triplicate, and the absorbance values were corrected against sample blanks in which α-glucosidase was replaced with phosphate buffer. The positive control (α-glucosidase activity with no inhibitor) and negative control (no α-glucosidase activity) were prepared by using phosphate buffer instead of the sample or instead of the sample and the α-glucosidase solution, respectively. The α-glucosidase inhibition rate was calculated as follows: