Kinetics of glucose oxidase immobilized in p(HEMA)-hydrogel microspheres in a packed-bed bioreactor
Kinetics of glucose oxidase immobilized in p(HEMA)-hydrogel microspheres in a packed-bed bioreactor
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DOI：
10.1016/s1381-1177(02)00061-9
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发表时间：
2002-09-13
影响因子：
--
通讯作者：
Guiseppi-Elie, A
中科院分区：
文献类型：
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作者：
Brahim, S;Narinesingh, D;Guiseppi-Elie, A
通过物理包埋和共价连接将葡萄糖氧化酶（GO（x））固定到通过反相悬浮聚合合成的交联聚（甲基丙烯酸羟乙酯-共-甲基丙烯酸二甲氨基乙酯，（p（HEMA-DMEMA））水凝胶微球（直径20-150μm）上。通过物理包埋实现了每克水凝胶7-8 mg GO（x）的负载能力，与<采用共价技术将每克凝胶添加 1.8 mg GO(x)，将含有物理包埋酶的微球装入生物反应器中，并在各种流动条件下研究固定化酶的动力学，当外推至接近无扩散条件时，结果分别为 13.2 mM 和 2.7 x 10(-3) mol min(-1)。对 K-m(app) 和 C-max 的 pH 依赖性的研究表明，咪唑鎓和巯基基团可能参与固定化 GO(x) 的温度依赖性。对 C 和 C-max 的温度依赖性的研究证实，在 >35°C 的温度下，葡萄糖氧化的活化能较低，这表明水凝胶内扩散限制的影响。保留。
Glucose oxidase (GO(x)) was immobilized via both physical entrapment and covalent linkage to crosslinked poly(hydroxyethyl methacrylate-co-dimethylaminoethyl methacrylate, (p(HEMA-DMEMA)) hydrogel microspheres (20-150 mum in diameter) that were synthesized by inverse suspension polymerization. Loading capacities of 7-8 mg GO(x) per gram of hydrogel were achieved with physical entrapment, compared to < 1.8 mg GO(x) per gram of gel with the covalent technique. The microspheres containing physically entrapped enzyme were packed into bioreactors and the kinetics of the immobilized enzyme investigated under various flow conditions. Flow rate dependence of K-m(app) and C-max, when extrapolated to near diffusion-free conditions, resulted in values of 13.2 mM and 2.7 x 10(-3) mol min(-1), respectively, for the immobilized enzyme. Studies of pH-dependence of K-m(app) and C-max suggest that the imidazolium and sulphydryl groups may be involved at the active site of the immobilized GO(x). Studies of the temperature dependence of C and C-max confirm lower activation energies for the oxidation of glucose at temperatures >35degreesC, suggesting the influence of diffusional limitations within the hydrogel. (C) 2002 Elsevier Science B.V. All rights reserved.