title: Off-line form of the Michaelis-Menten equation for studying the reaction kinetics in a polymer microchip integrated with enzyme microreactor.
abstract: We firstly transformed the traditional Michaelis-Menten equation into an off-line form which can be used for evaluating the Michaelis-Menten constant after the enzymatic reaction. For experimental estimation of the kinetics of enzymatic reactions, we have developed a facile and effective method by integrating an enzyme microreactor into direct-printing polymer microchips. Strong nonspecific adsorption of proteins was utilized to effectively immobilize enzymes onto the microchannel wall, forming the integrated on-column enzyme microreactor in a microchip. The properties of the integrated enzyme microreactor were evaluated by using the enzymatic reaction of glucose oxidase (GOx) with its substrate glucose as a model system. The reaction product, hydrogen peroxide, was electrochemically (EC) analyzed using a Pt microelectrode. The data for enzyme kinetics using our off-line form of the Michaelis-Menten equation was obtained (K(m) = 2.64 mM), which is much smaller than that reported in solution (K(m) = 6.0 mM). Due to the hydrophobic property and the native mesoscopic structure of the poly(ethylene terephthalate) film, the immobilized enzyme in the microreactor shows good stability and bioactivity under the flowing conditions.
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science, Department of Chemistry, Nanjing University, Nanjing 210093, P. R. China. xhxia@nju.edu.cn
authors.forename: Ai-Lin
authors.initials: AL
authors.lastname: Liu
authors.forename: Ting
authors.initials: T
authors.lastname: Zhou
authors.forename: Feng-Yun
authors.initials: FY
authors.lastname: He
authors.forename: Jing-Juan
authors.initials: JJ
authors.lastname: Xu
authors.forename: Yu
authors.initials: Y
authors.lastname: Lu
authors.forename: Hong-Yuan
authors.initials: HY
authors.lastname: Chen
authors.forename: Xing-Hua
authors.initials: XH
authors.lastname: Xia
mesh_terms: D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005947:Glucose / Q000737:chemistry*; D005949:Glucose Oxidase / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D018625:Microscopy, Atomic Force; D009995:Osmosis; D011093:Polyethylene Terephthalates / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D004800:Enzymes, Immobilized; D011093:Polyethylene Terephthalates; D005949:Glucose Oxidase; D005947:Glucose