Chen Bo, Miller M Elizabeth, Gross Richard A
NSF I/UCRC for Biocatalysis and Bioprocessing of Macromolecules, Polytechnic University, 6 Metrotech Center, Brooklyn, New York 11201, USA.
Langmuir. 2007 May 22;23(11):6467-74. doi: 10.1021/la063515y. Epub 2007 Apr 24.
Polystyrene resins with varied particle sizes (35 to 350-600 microm) and pore diameters (300-1000 A) were employed to study the effects of immobilization resin particle size and pore diameter on Candida antarctica Lipase B (CALB) loading, distribution within resins, fraction of active sites, and catalytic properties for polyester synthesis. CALB adsorbed rapidly (saturation time </= 4 min) for particle sizes </= 120 microm (pore size = 300 A). Infrared microspectroscopy showed that CALB forms protein loading fronts regardless of resin particle size at similar enzyme loadings ( approximately 8%). From the IR images, the fractions of total surface area available to the enzyme are 21, 33, 35, 37, and 88% for particle sizes 350-600, 120, 75, 35 microm (pore size 300 A), and 35 microm (pore size 1000 A), respectively. Titration with methyl p-nitrophenyl n-hexylphosphate (MNPHP) showed that the fraction of active CALB molecules adsorbed onto resins was approximately 60%. The fraction of active CALB molecules was invariable as a function of resin particle and pore size. At approximately 8% (w/w) CALB loading, by increasing the immobilization support pore diameter from 300 to 1000 A, the turnover frequency (TOF) of epsilon-caprolactone (epsilon-CL) to polyester increased from 12.4 to 28.2 s-1. However, the epsilon-CL conversion rate was not influenced by changes in resin particle size. Similar trends were observed for condensation polymerizations between 1,8-octanediol and adipic acid. The results herein are compared to those obtained with a similar series of methyl methacrylate resins, where variations in particle size largely affected CALB distribution within resins and catalyst activity for polyester synthesis.
使用具有不同粒径（35至350 - 600微米）和孔径（300 - 1000埃）的聚苯乙烯树脂，研究固定化树脂的粒径和孔径对南极假丝酵母脂肪酶B（CALB）负载量、在树脂中的分布、活性位点比例以及聚酯合成催化性能的影响。对于粒径≤120微米（孔径 = 300埃）的树脂，CALB吸附迅速（饱和时间≤4分钟）。红外显微光谱显示，在相似的酶负载量（约8%）下，无论树脂粒径如何，CALB都会形成蛋白质负载前沿。从红外图像可知，对于粒径为350 - 600、120、75、35微米（孔径300埃）以及35微米（孔径1000埃）的树脂，酶可利用的总表面积比例分别为21%、33%、35%、37%和88%。用对硝基苯基正己基磷酸甲酯（MNPHP）滴定表明，吸附到树脂上的活性CALB分子比例约为60%。活性CALB分子比例不随树脂粒径和孔径变化。在CALB负载量约为8%（w/w）时，将固定化载体孔径从300埃增加到1000埃，ε-己内酯（ε-CL）转化为聚酯的周转频率（TOF）从12.4增加到28.2 s⁻¹。然而，ε-CL转化率不受树脂粒径变化的影响。对于1,8 - 辛二醇与己二酸之间的缩聚反应，也观察到了类似趋势。本文的结果与使用类似系列甲基丙烯酸甲酯树脂得到的结果进行了比较，在甲基丙烯酸甲酯树脂中，粒径变化在很大程度上影响了CALB在树脂中的分布以及聚酯合成的催化剂活性。
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