Continuous-flow determination of an alpha-amylase inactivator in fermentation samples using a ferricyanide reagent

A continuous-flow system has been developed in which pancreatic alpha-amylase is incubated with soluble starch at 37 degrees C. Reducing sugars being delivered at the "steady-state" hydrolysis of starch are dialyzed into a solution of alkaline ferricyanide. Ferricyanide is then reduced at 95 degrees C. The decrease in the absorbance of ferricyanide solution is recorded as "enzyme baseline." When samples containing alpha-amylase inactivator are introduced into the system, hydrolysis of starch is reduced according to the concentration of enzyme inactivator. The amount of inactivator in the samples is deduced from a standard curve of six standard concentrations using the Technicon Autoanalyzer II system manager in connection with a calculator for automated interpolation. This method allows to determine 40 samples/h with high sensitivity and precision (mean = 1.53 mg/liter; C.V. = 1.4%).