2.7.1. Inhibition Assays for α-glucosidase Activity

The α-glucosidase inhibition was determined by the 96-well microtiter plate method based on the calorimetric assay as previously described by Vinholes et al. [23]. α-glucosidase enzyme solution (2U mL−1, Sigma) was prepared in 100 mM phosphate buffer (pH 7.0). Ethyl acetate extracts were used in concentrations ranging from 10–100 μg mL<sup>−</sup>1. 2 mM of *para*-nitrophenyl-α-D-glucopyranoside (Sigma) was prepared in 50 mM phosphate buffer (pH 7.0). 50 μL of the partially purified fraction was pre-incubated with an equal volume of yeast enzyme at 37 ◦C for 5 min, followed by the addition of 30 μL of pNPG and further incubation for 30 min. After incubation, 100 μL of stopping reagent (0.1 M Na2CO3) was added to cease the reaction. Color produced was quantified by UV spectrophotometer (Shimadzu, Kyoto, Japan) at 405 nm. Each experiment was performed in triplicate. Acarbose (Sigma) was used as a positive control, whereas purified fraction was replaced by phosphate buffer in control. Reaction mixture without enzyme was taken as blank. The percentage inhibition (%) was determined by the formula:

$$\text{percentage inhibition } (\%) = \frac{\text{absorbance of control} - \text{absorbance of sample}}{\text{absorbance of control}} \times 100\%$$
