Preparation of a new fluorogenic substrate of alpha-amylases and a simple alpha-amylase assay by HPLC.
A new substrate of alpha-amylases, O-6-deoxy-6-[(2-pyridyl)amino]-alpha-D-glucopyranosyl-(1 leads to 4)-O-alpha-D-glucopyranosyl-(1 leads to 4)-O-alpha-D-glucopyranosyl-(1 leads to 4)-O-alpha-D-glucopyranosyl-(1 leads to 4)-D-glucopyranose, was prepared using dextrin as a starting material. Compared with other substrates so far reported, the fluorogenic substrate is unique in that it is resistant to exo-alpha-glucosidases due to the blocking group introduced into the non-reducing end glucose residue. The product of alpha-amylase digestion was rapidly separated from the substrate and was detected very sensitively by HPLC and a fluorescence detector. This method for alpha-amylase assay was also applied for determination of alpha-amylase in human serum.
D000681:Amylases / Q000378:metabolism*; D001230:Aspergillus / Q000201:enzymology; D001412:Bacillus subtilis / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000378:metabolism; D009844:Oligosaccharides / Q000032:analysis / Q000138:chemical synthesis*; D012233:Rhizopus / Q000201:enzymology; D013050:Spectrometry, Fluorescence / Q000379:methods; D000516:alpha-Amylases / Q000032:analysis / Q000378:metabolism*
D005456:Fluorescent Dyes; D009844:Oligosaccharides; C021705:maltooligosaccharides; D000681:Amylases; D000516:alpha-Amylases