Tse P H, Leypoldt J K, Gough D A
Department of Applied Mechanics and Engineering Sciences, Bioengineering Group, University of California, San Diego, La Jolla, California 92093.
Biotechnol Bioeng. 1987 Apr;29(6):696-704. doi: 10.1002/bit.260290606.
Models of membrane systems containing immobilized glucose oxidase and catalase operating together or independently have been developed. A rotated disk electrode apparatus was employed with novel electrochemical operating conditions to experimentally determine mass transfer and intrinsic kinetic parameters of enzyme-containing membranes. The value of a mass transfer parameter that describes internal and external diffusion was first determined under conditions that do not permit the enzyme reactions. In a subsequent experiment with the reaction allowed, kinetic parameters corresponding to the intrinsic maximal velocity and Michaelis constants of the immobilized enzymes were estimated by regression analysis of data based on an appropriate two- or three- parameter model. It was found that immobilization reduced the maximal intrinsic velocity but had no detectable effect on the Michaelis constants. In all but one case- these methods for membrane characterization are nondestructive and can be used repeatedly on a given membrane. These techniques provide the means for quantitative comparisons of immobilization methods and make possible temporal studies of immobilized enzyme inactivation.
已开发出包含固定化葡萄糖氧化酶和过氧化氢酶一起或独立运行的膜系统模型。采用旋转圆盘电极装置和新颖的电化学操作条件，通过实验确定含酶膜的传质和本征动力学参数。首先在不允许酶反应的条件下确定描述内部和外部扩散的传质参数值。在随后允许反应的实验中，通过基于适当的双参数或三参数模型对数据进行回归分析，估算了与固定化酶的本征最大速度和米氏常数相对应的动力学参数。发现固定化降低了本征最大速度，但对米氏常数没有可检测到的影响。除一种情况外，所有这些用于膜表征的方法都是无损的，并且可以在给定的膜上重复使用。这些技术提供了对固定化方法进行定量比较的手段，并使对固定化酶失活的时间研究成为可能。
