Evaluation of alpha-amylase assays with 4-nitrophenyl-alpha-oligosaccharides as substrates

Measurements of alpha-amylase with 4-nitrophenyl glucosides offer the following advantages over methods that rely on the formation of NADH: a short lag phase, no apparent interference by metabolites and enzymes of the sample and extremely stable substrates with low blank values. The intrinsic sensitivity of nitrophenol formation was equal to that of hydrolysis of maltotetraose, but was less than that of glucose-producing methods using oligosaccharides. In contrast to starch, the chromogenic substrates are more rapidly hydrolysed by salivary than by pancreatic amylase. Disadvantages of these substrates are: higher turnover by animal than by human amylases, and a marked susceptibility of the chromophore to small changes of pH and protein concentration. Some analytical qualities such as specificity, accuracy, precision, stability of the substrate and linear range are described in detail and compared with those of other methods.