Off-line form of the Michaelis-Menten equation for studying the reaction kinetics in a polymer microchip integrated with enzyme microreactor.
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.
Liu
Ai-Lin
AL
Key Laboratory of Analytical Chemistry for Life Science, Department of Chemistry, Nanjing University, Nanjing 210093, P. R. China. xhxia@nju.edu.cn
Zhou
Ting
T
He
Feng-Yun
FY
Xu
Jing-Juan
JJ
Lu
Yu
Y
Chen
Hong-Yuan
HY
Xia
Xing-Hua
XH
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
D004800:Enzymes, Immobilized; D011093:Polyethylene Terephthalates; D005949:Glucose Oxidase; D005947:Glucose