A 200 l reaction mixture containing 1% (w/v) soluble starch and
20 l of the enzyme sample in 20 mM Tris–HCl buffer (pH 7.5) was
incubated at 50 ◦C for 10 min and the amount of reducing sugars
released was measured using 2,5-dinitrosalicylic acid (Analytical
grade) reagent. One unit (U) of enzyme activity was defined as the
amount of enzyme that liberated 1 mol of reducing sugar as glucose
equivalents in 1 min under the assay conditions. Protein was
quantified by Bradford’s method using bovine serum albumin as
the standard protein [18].