Purification and characterization of alpha-amylase inhibitor from the seeds of underutilized legume, Mucuna pruriens

An alpha-amylase inhibitor (EC 3.2.1.1) was purified by buffer extraction, ammonium sulfate fractionation, CM-cellulose, and sephadex G-75 chromatography from the soaked seeds of Mucuna pruriens. The molecular weight determined by gel permeation chromatography on Sephadex G-100 and SDS-PAGE, both in the presence and absence of 2-mercaptoethanol, was found to be 27.24 kDa and 25.6 kDa, respectively. The purified Mucuna pruriens amylase inhibitor showed a specific inhibitor activity of 61.18, fold purity of 36.68, and the yield obtained was 14.01%. The purified amylase inhibitor was found to be heat-stable and retained 80.50% activity at 65 degrees C. Inhibitor was found to have pH optima of 6.9. Hundred percent zone of inhibition was observed when added on the plated organisms of purified inhibitor. Purified amylase inhibitor was found to inhibit the activity of human salivary alpha-amylase. Inhibitory activity of alpha-amylase inhibitor against mammalian amylases could suggest its potential in treatment of diabetes and cure of nutritional problems, which results in obesity.