Effects of macroporous resin size on Candida antarctica lipase B adsorption, fraction of active molecules, and catalytic activity for polyester synthesis
Effects of macroporous resin size on Candida antarctica lipase B adsorption, fraction of active molecules, and catalytic activity for polyester synthesis
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DOI：
10.1021/la062258u
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发表时间：
2007-01-30
期刊：
影响因子：
3.9
通讯作者：
Gross, Richard A.
中科院分区：
文献类型：
--
作者：
Chen, Bo;Miller, Elizabeth M.;Gross, Richard A.
采用平均孔径（250 A）和表面积（500 m（2）/g）相同但粒径不同（35 ~ 560 ~ 710 μ m）的甲基丙烯酸甲酯树脂，研究固定化树脂粒径如何影响Candida anastritica脂肪酶B（CALB）的负载量、活性部位分数和聚酯合成的催化性能。CALB在较小的珠上吸附更快。对于直径为35和560-710 μ m的珠粒，分别在小于30 s和48 h内发生饱和。通过Scatchard分析的吸附等温线数据的线性化表明，对于35 μ m树脂：（i）CALB在饱和时的负载量远低于形成单层并完全覆盖载体表面所需的负载量，和（ii）CALB对该树脂表面具有高亲和力。红外显微光谱表明，CALB形成的蛋白质装载前线的树脂粒径为560-710和120 μ m。相比之下，CALB似乎均匀地分布在整个35 μ m树脂。用对硝基苯基正己基磷酸酯（MNPHP）滴定，活性CALB分子在树脂上的吸附率小于50%，且不受粒径的影响。随着酶负载量从0.9%（w/w）增加到5.7%（w/w），35 μ m载体上的活性CALB分子的分数从30%增加到43%，导致ε-己内酯（ε-CL）开环聚合活性增加。在约5%w/w CALB负载下，通过将固定化载体直径从560-710减小到120、75和35 μ m，ε-CL %转化为聚酯的转化率增加（在80分钟时分别为20到36、42和61%）。对于1，8-辛二醇和己二酸之间的缩合聚合，观察到类似的趋势。
Methyl methacrylate resins with identical average pore diameter (250 A) and surface area (500 m(2)/g) but with varied particle size (35 to 560-710 mu m) were employed to study how immobilization resin particle size influences Candida antarctica Lipase B (CALB) loading, fraction of active sites, and catalytic properties for polyester synthesis. CALB adsorbed more rapidly on smaller beads. Saturation occurred in less than 30 s and 48 h for beads with diameters 35 and 560-710 mu m, respectively. Linearization of adsorption isotherm data by the Scatchard analysis showed for the 35 mu m resin that: (i) CALB loading at saturation was well below that required to form a monolayer and fully cover the support surface and (ii) CALB has a high affinity for this resin surface. Infrared microspectroscopy showed that CALB forms protein loading fronts for resins with particle sizes 560-710 and 120 mu m. In contrast, CALB appears evenly distributed throughout 35 mu m resins. By titration with p-nitrophenyl n-hexyl phosphate (MNPHP), the fraction of active CALB molecules adsorbed onto resins was < 50% which was not influenced by particle size. The fraction of active CALB molecules on the 35 mu m support increased from 30 to 43% as enzyme loading was increased from 0.9 to 5.7% (w/w) leading to increased activity for epsilon-caprolactone (epsilon-CL) ring-opening polymerization. At about 5% w/w CALB loading, by decreasing the immobilization support diameter from 560-710 to 120, 75, and 35 mu m, conversion of epsilon-CL % to polyester increased (20 to 36, 42, and 61%, respectively, at 80 min). Similar trends were observed for condensation polymerizations between 1,8-octanediol and adipic acid.