The Ceralpha method can be used for alpha-amylase activity measurements during the hydrolysis of starch at high substrate concentrations (> 40 wt.%). However, the results are affected by the carbohydrates present in the samples. The effect of carbohydrates on the Ceralpha alpha-amylase activity measurements was measured over a broad concentration range. It was found that starch has the largest influence and glucose has the lowest influence on the Ceralpha assay procedure. These results were explained by considering substrate inhibition and substrate competition. A simple kinetic model was used to describe the observed phenomena quantitatively. This model was also used to estimate the Michaelis-Menten constant for a large number of substrates and it requires only a single experiment for each Km determination. (c) 2005 Elsevier Inc. All rights reserved.