Determination of alpha-amylase using 4-O-beta-D-galactopyranosylmaltotetraose (Gal-G4) as a substrate.
A new substrate, 4-O-beta-D-galactopyranosylmaltotetraose (Gal-G4) is applied for the determination of alpha-amylase in serum and urine in a coupled assay with alpha-glucosidase (EC 3.2.1.20), glucokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) as auxiliary enzymes. Gal-G4 having a 4-position of the non-reducing-end glucose residue modified by a beta-galactopyranose group is resistant for degradation by alpha-glucosidase as auxiliary enzyme. Moreover, this substrate is hydrolyzed at just one position by alpha-amylase in serum and urine. More than 99% of the products generated from Gal-G4 by alpha-amylase are identified 4-O-beta-D-galactopyranosylmaltose (Gal-G2), maltose, respectively. Glucose and maltose do not interfere the value of alpha-amylase activity at least up to 0.056 mmol/l (1 g/dl) glucose and 0.027 mmol/l (1 g/dl) maltose, respectively. We are now carrying out this work under the authority of The Enzyme committee of Japanese Society of Clinical Chemistry (JSCC) as a standard method for determination of alpha-amylase in clinical chemistry.
Kikuchi
T
T
Tsuruga Institute of Biotechnology, Toyobo Co., Ltd., Fukui, Japan.
Kimata
S
S
Majima
K
K
Asano
S
S
D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007202:Indicators and Reagents; D007700:Kinetics; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D012031:Refractometry; D013379:Substrate Specificity; D000516:alpha-Amylases / Q000097:blood / Q000378:metabolism* / Q000652:urine
C113459:4-O-galactopyranosylmaltotetraose; D007202:Indicators and Reagents; D009844:Oligosaccharides; D000516:alpha-Amylases