2.9. Determination of α-Glucosidase Inhibiting Activity

Inhibition of α-glucosidase was determined as reported earlier [22] with slight modification. In brief, 100 μL of test samples dissolved in 10% DMSO (4, 2, 1, and 0.5 mg/mL solution) was incubated with 50 μL ofα-glucosidase from Saccharomyces cerevisiae Type I (Sigma-Aldrich, US) (1.0 U/mL dissolved in 0.1 M phosphate buffer, pH 6.8) for 10 min at 37°C. Afterwards, in reaction mixture, 50 μL substrate (5 mM p-nitrophenyl-α-D-glucopyranoside prepared in the same buffer) was added and release of p-nitrophenol was measured at 405 nm spectrophotometrically after 5 min of incubation. Individual blanks for test samples were prepared to correct background absorbance where substrate was replaced with 50 μL of buffer. Control sample contained 100 μL 10% DMSO instead of test samples. Percentage of enzyme inhibition was calculated using equation AG = (A _control − A _sample)/A _control × 100, where A _control is absorbance of the mixture without test compound (extract) and A _sample represents absorbance of samples containing extracts. As standard reference, acarbose was taken. Applying convenient regression analysis, IC₅₀ (concentration of the test sample necessary to inhibit 50% activity of the enzyme) was obtained.