Alpha-glucosidase inhibition assay in an enzyme-immobilized amino-microplate.
Alpha-glucosidase (AGH) from the small intestine of rat was immobilized onto a glutaraldehyde (GA) activated NH(2)-96 well microplate to establish a convenient and rapid AGH inhibition assay system. After AGH immobilization, remaining GA groups were blocked by beta-alanine to induce a negative charge on the surface of the well. The AGH-plate showed an enzyme activity of 444 nU/well under an assayed condition at 37 degrees C for 2 h using 0.3 mM 4-methylumbelliferyl-alpha-D-glucopyranoside as a fluorogenic substrate. Inhibitory powers of voglibose and acarbose as therapeutic AGH inhibitors were successfully evaluated to have IC(50) values of 13 and 114 nM, respectively.
Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, Higashi, Fukuoka 812-8581, Japan. tmatsui@agr.kyushu-u.ac.jp
Toshiro
T
Matsui
Mayu
M
Shimada
Nozomi
N
Saito
Kiyoshi
K
Matsumoto
D020909:Acarbose / Q000494:pharmacology; D000818:Animals; D055162:Biocatalysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D005976:Glutaral / Q000737:chemistry*; D065089:Glycoside Hydrolase Inhibitors; D020128:Inhibitory Concentration 50; D007294:Inositol / Q000031:analogs & derivatives / Q000494:pharmacology; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D000520:alpha-Glucosidases / Q000737:chemistry* / Q000378:metabolism; D015091:beta-Alanine / Q000494:pharmacology
D004791:Enzyme Inhibitors; D004800:Enzymes, Immobilized; D065089:Glycoside Hydrolase Inhibitors; D015091:beta-Alanine; D007294:Inositol; D000520:alpha-Glucosidases; C102817:voglibose; D005976:Glutaral; D020909:Acarbose