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The enzyme, alpha-glucosidase (1 U/mL), and 20 μL of different concentrations of each extract fraction (100, 200, 300, 400, and 500 mg/mL) were added to a test tube. In each working test tube, a reaction mixture contained 0.1 mL of alpha-glucosidase solution was mixed with 0.2 mL from each extract dilution and 0.5 mL of phosphate buffer (100 mM, pH = 6. 8). The samples were incubated at nearly 37 °C for 15 min. After this incubation period, 0.2 mL of 5 mM PNPG (the substrate used for this experiment) was added to the reaction mixture, and the samples were again incubated at 37 °C for 20 min. The reaction was terminated by adding 0.1 M sodium carbonate (Na2CO3). The absorbance at the 405 nm wavelength was recorded for all samples. Acarbose was used as a positive control at the same concentrations as the plant extracts [25]. The results were expressed as percentage inhibition according to the following equation:
AB is the absorbance without enzyme inhibitor; AE is the absorbance in the presence of E. alata sample.
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