Volokitina Mariia V, Nikitina Anna V, Tennikova Tatiana B, Korzhikova-Vlakh Evgenia G
Russian Academy of Sciences, Institute of Macromolecular Compounds, St. Petersburg, Russia.
Institute of Chemistry, Saint-Petersburg State University, St. Petersburg, Russia.
Electrophoresis. 2017 Nov;38(22-23):2931-2939. doi: 10.1002/elps.201700210. Epub 2017 Sep 13.
Macroporous monolithic columns with different mean pore size (from 360 to 2020 nm) and appropriate flow-through properties were synthesized using free radical in situ copolymerization of glycidyl methacrylate, 2-hydroxyethyl methacrylate and ethylene dimethacrylate. In order to predict the composition of porogen mixture to generate the pores in the interested size interval, the Hildebrand theory was used. Ribonuclease A and its specific low- and macromolecular substrates cytidine-2',3'-cyclic monophosphate sodium salt and RNA were applied as model system. The effect of mean pore size of macroporous monoliths used for enzyme immobilization on molecular recognition and biocatalytic characteristics was examined. The monitoring of RNA degradation was performed using anion-exchange HPLC on monolithic CIM DEAE analytical column. The high efficiency of heterogeneous biocatalysts obtained comparatively to the catalytic reaction of RNA degradation in solution was demonstrated. Additionally, the series of six monolithic immobilized enzyme reactors with different amount of biocatalyst was prepared and studied regarding to the biocatalytic properties at recirculation mode at two experimental variants, e.g. (i) fixed range of concentrations of circulated substrate solutions, and (ii) fixed range of substrate/enzyme molar ratios.
采用甲基丙烯酸缩水甘油酯、甲基丙烯酸2-羟乙酯和二甲基丙烯酸乙烯酯的自由基原位共聚反应，合成了具有不同平均孔径（360至2020纳米）且具有适当流通性能的大孔整体柱。为了预测能在感兴趣的尺寸区间内产生孔隙的致孔剂混合物的组成，使用了希尔德布兰德理论。将核糖核酸酶A及其特定的低分子和高分子底物2',3'-环磷酸胞苷钠盐和RNA用作模型体系。研究了用于酶固定化的大孔整体柱的平均孔径对分子识别和生物催化特性的影响。在整体式CIM DEAE分析柱上，采用阴离子交换高效液相色谱法监测RNA的降解情况。结果表明，与溶液中RNA降解的催化反应相比，所制备的多相生物催化剂具有较高的催化效率。此外，制备了一系列六种含有不同生物催化剂量的整体式固定化酶反应器，并在两种实验变体的循环模式下研究了其生物催化性能，例如：（i）循环底物溶液浓度的固定范围，以及（ii）底物/酶摩尔比的固定范围。
