Simple Method for Detecting Amylase Inhibitors in Biological Materials

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Abstract
A simple semiquantitative method for detecting amylase or amylase inhibitors in biological materials is described. The substrate consists of a 0.25% starch-1.5% agar gel slab buffered at pH 6.5 with 0.1 M phosphate-0.01 M sodium chloride. Three millimeters wide cellulose strips saturated with a solution to be examined for inhibitor are placed parallel on the gel slab for 2 hours at 37°. The strips are removed and other 3-mm wide cellulose strips saturated with amylase solutions are placed at right angles across the first strips. The system is incubated for 6 or 18 hours at 37°. After flooding the slab with Lugol (an iodine containing) solution, amylase activity is shown by clear lysis zones on a deep-purple background. Presence of inhibitors is indicated by interruption or narrowing of the lysis zone where the inhibitor-containing and amylase-containing strips crossed. A variation of this method using amylase or amylase-inhibitor mixtures placed into 7-mm wells cut into the starch-agar gel slab is also described. The starch-agar gel slab methods were compared with the Bernfeld method of determining amylase activity.