title: Development of a fast and efficient CE enzyme assay for the characterization and inhibition studies of α-glucosidase inhibitors.
abstract: The inhibition of the α-glucosidase enzyme plays an important role in the treatment of diabetes mellitus. We have established a highly sensitive, fast, and convenient CE method for the characterization of the enzyme and inhibition studies of α-glucosidase inhibitors. The separation conditions were optimized; the pH value and concentration of the borate-based separation buffer were optimized in order to achieve baseline separation of p-nitrophenyl-α-d-glucopyranoside and p-nitrophenolate. The optimized method using 25 mM tetraborate buffer, pH 9.5, was evaluated in terms of repeatability, LOD, LOQ, and linearity. The LOD and LOQ were 0.32 and 1.32 μM for p-nitrophenyl-α-D-glucopyranoside and 0.83 and 3.42 μM for p-nitrophenolate, respectively. The value of the Michaelis-Menten constant (K(m)) determined for the enzyme is 0.61 mM, which is in good agreement with the reported data. The RSDs (n = 6) for the migration time was 0.67 and 1.83% for substrate and product, respectively. In the newly established CE method, the separation of the reaction analytes was completed in <4 min. The developed CE method is rapid and simple for measuring enzyme kinetics and for assaying inhibitors.
authors.affiliation: Department of Pharmaceutical Sciences, COMSATS Institute of Information Technology, Abbottabad, Pakistan.
authors.forename: Shoaib
authors.initials: S
authors.lastname: Iqbal
authors.lastname: Nisar ur Rehman
authors.forename: Ulrich
authors.initials: U
authors.lastname: Kortz
authors.forename: Jamshed
authors.initials: J
authors.lastname: Iqbal
mesh_terms: D004305:Dose-Response Relationship, Drug; D019075:Electrophoresis, Capillary; D057075:Enzyme Assays / Q000379:methods*; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000737:chemistry / Q000494:pharmacology*; D065089:Glycoside Hydrolase Inhibitors; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D015394:Molecular Structure; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013329:Structure-Activity Relationship; D013997:Time Factors; D000520:alpha-Glucosidases / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D065089:Glycoside Hydrolase Inhibitors; D000520:alpha-Glucosidases
keywords: Capillary electrophoresis; Enzyme kinetics; Inhibition assay; α-Glucosidase