Action pattern of human pancreatic alpha-amylase on maltoheptaose, a substrate for determining alpha-amylase in serum

Journal of Chromatography
E O HaegeleM Grassl

Abstract

An enzymatic assay for the determination of alpha-amylase in serum was developed which employed a soluble substrate, maltoheptaose, and a coupled enzymatic indicator reaction consisting of alpha-glucosidase and the hexokinase-glucose-6-phosphate dehydrogenase system. We used high-performance liquid chromatography (HPLC) to establish the action pattern of maltoheptaose under the test conditions: (A) the action pattern of alpha-amylase, (B) that of the combined action of alpha-amylase and alpha-glucosidase. Conductive to this effect was: the availability of pure maltoheptaose and human pancreatic alpha-amylase; the development of an adequate procedure for sample pretreatment (partition chromatography on a mixed-bed ion exchange) and of an HPLC system for separation of substrate and reaction products without interference from by products of the assay (partition chromatography on a cation-exchange column with acetonitrile-water); and the use of a new, very sensitive refractometric detector revealing sugar amounts as low as 40 ng. We derived the following stoichiometric equations: (see formula index).

Citations

Sep 23, 1994·Journal of Chromatography. B, Biomedical Applications·G W WellingS Welling-Wester
Oct 24, 2002·European Journal of Biochemistry·Gyöngyi GyémántLili Kandra
Jul 1, 1983·Journal of Clinical Chemistry and Clinical Biochemistry. Zeitschrift Für Klinische Chemie Und Klinische Biochemie·K Lorentz
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Jan 1, 1985·Digestive Diseases and Sciences·C NiederauJ Erckenbrecht
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Dec 29, 2000·Carbohydrate Research·L Kandra, G Gyémánt
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