科研之友
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摘要
Oxidases catalyze the oxidation of a variety of substrates with the concomitant reduction of molecular oxygen as a final electron acceptor. UV-visible spectrophotometry is a simple and high-throughput method commonly used to measure oxidase activities. However, drawbacks such as light scattering exist especially concerning the activity assessment of enzymes immobilized on supports. Monitoring of the universal cosubstrate O-2 circumvents these drawbacks. This study aimed at developing a methodology that allows activity measurement of many types of oxidases based on O-2 consumption applicable to various open systems. Dissolved oxygen in the reaction medium was monitored by an O-2 sensor and the reaction rate was deduced from the O-2 mass balance equation correcting for atmospheric diffusion. Common activity units (mu(product) min(-1) or U/L) could be subsequently derived using calibration curves. The sensitivity of the method toward temperature, atmospheric pressure, and ionic strength variations was evaluated, and made it possible to define operating windows for the simplification of the proposed methodology.
- 出版日期2015-11
