Direct spectrophotometric determination of alpha-amylase activity in salive, with p-nitrophenyl alpha-maltoside as substrate.
We describe a simple, direct kinetic method for determination of salivary alpha-amylase (1,4, alpha-D-glucan 4-glucanohydrolase, EC 3.2.1.1). The assay makes use of a well-defined substrate, p-nitrophenyl alpha-maltoside, which is hydrolyzed by alpha-amylase to a chromogenic product, p-nitrophenol. Activity is determined by directly monitoring the increase in absorbance of the reaction mixture. Amylase activity can be defined in international (IUB) units of micromoles of product/min per liter of saliva. For 22 healthy subjects, the mean +/- SD of amylase activity in mixed saliva was 2.77 +/- 1.12 U/liter. Activity and instrumental response were linearly related over the entire range tested (0.224 to 11.90 U/liter). The within-run precision (CV) over this range was better than 3% for all but the lowest activities. Values obtained with this assay correlate well with those obtained with a modified Nelson-Somogyi saccharogenic method (r = 0.979). The precision and simplicity of this assay suggest that it is the method choice for determining amylase activity in human saliva.
D000681:Amylases / Q000032:analysis*; D006801:Humans; D007700:Kinetics; D009596:Nitrophenols; D012463:Saliva / Q000201:enzymology*; D013053:Spectrophotometry / Q000379:methods; D000516:alpha-Amylases / Q000032:analysis* / Q000378:metabolism
D009596:Nitrophenols; D000681:Amylases; D000516:alpha-Amylases