α-Amylase Inhibition Assay: DNSA Method

The 3,5-dinitrosalicylic acid (DNSA) method was used to determine α-amylase inhibition activity [19 (link)] in which 200 μL each of extracts (20–360 μg/mL) and 3 unit/mL α-amylase (Sigma-Aldrich, USA) were mixed and incubated for 15 min at 37°C. An additional 5 min of incubation at 37°C was carried out after the addition of 200 μL of 1% starch solution. Then, a 200 μL of DNSA solution was added to it and the whole reaction mixture was heated for 10 min at 95°C. The final volume was adjusted to 5 mL with the addition of distilled water. The absorbance was measured at λ = 540 nm in a UV-visible spectrophotometer. The results were compared with the activities of acarbose and 1% dimethyl sulfoxide (DMSO) was used as a positive control.