Interference of thiol-compounds with dextrinizing activity assay of α-amylase by starch-iodine colour reaction: Modification of the method to eliminate this interference

Abstract

Interference of Luria broth and other bacteriological media with the starch–iodine colour assay of the dextrinizing activity of α-amylase (Fuwa's method) was observed, complete bleaching occurring with 0.4ml of the Luria broth. The interference was found to be due to the thiol groups present in the medium which compete with starch for iodine. Among the various metal salts tested for counteracting the interference, ZnSO₄ was found to be the best which reverted the colour to about 73–85% of that of the blank. A combination of hydrogen peroxide (10 μl of 30% solution) and CuSO₄ · 5H₂O (50 μl of 0.1 M solution) completely protected the starch–iodine reaction in the presence of even 0.5 ml of Luria broth and a modified assay was developed based on this finding. The colour intensity, however, was almost double than that obtained for the same amount of starch and iodine in the absence of these protective agents. Nevertheless, with different concentrations of starch as well as with varying amounts of enzymes, the modified method showed perfect linearity and could be effectively used for estimation of dextrinizing activity of α-amylase in the presence of thiol groups.