横向交变电场电色谱的传递理论和应用研究
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摘要
本论文对三腔室横向交变电场电色谱的传热、传质理论以及该电色谱的应用进行了系统的研究。
首先构建了横向电场电色谱测温系统,测定了电色谱冷却和施加电场的动态升温过程,研究了电流强度和流动相离子强度对柱内温度分布的影响。建立了横向电场电色谱过程的传热模型,模拟从非稳态操作到稳态操作的温度分布。利用该模型可同时获得柱内电压分布、有效电压和功耗等参数,分析电色谱放大的温度效应,以及各种条件下的温度分布行为。
制备了不同配基修饰密度的离子交换介质,研究了电色谱的蛋白质动态吸附行为。结果表明,由于高配基密度介质的表面电荷密度较高,孔内电渗流对促进传质的作用更明显,故蛋白质动态吸附容量随电场强度增加较多。
建立横向交变电场离子交换电色谱过程的理论模型,利用上述蛋白质动态吸附实验结果,验证了模型的可靠性。系统分析了电色谱过程的传质和色谱行为。理论分析发现,电渗引起的孔内对流受蛋白质不可逆吸附的影响,孔内对流速度对吸附饱和度呈负指数下降趋势;在电场作用下,吸附剂内蛋白质浓度分布呈现出沿电场方向的偏移,在操作的初始时刻电场作用效果有利于提高动态吸附容量。
利用填充阴离子交换介质Q Sepharose Fast Flow的横向电场色谱,研究了卵清蛋白质的分离纯化,获得了高纯度卵铁传递蛋白和卵清白蛋白。与普通离子交换色谱相比,横向电场电色谱的分离能力提高一倍以上,证明了该横向电场电色谱的高容量分离纯化性能。
This thesis is concerned with the studies of ion-exchange electrochromatography with an oscillatory electric field perpendicular to mobile-phase flow driven by pressure (pIEEC). The electrochromatography is composed of a three-compartment column developed in the laboratory. The work has investigated heat and mass transport phenomena systematically by both experiments and mathematical modeling.
First of all, an experimental system for in-column temperature measurements was constructed, and the dynamic processes of the in-column temperature in electrochromatography were examined. With the experimental system, the effect of electric current strength and mobile-phase ionic strength on the in-column temperature was investigated. Then, a heat transfer model for the pIEEC was established for the dynamic process modeling. It was confirmed that the model was in good agreement with the measurements. By the model calculations, we could also obtain the in-column voltage distribution, effective voltage applied to the central compartment and the efficiency of energy consumption. Moreover, the model was used for scale-up analysis and for investigation of temperature distribution under various conditions.
Protein ion-exchangers of different ion-exchange capacities were prepared to study the dynamic protein adsorption processes of the pIEEC. The results indicated that electric field for the pIEEC was more effective for the high ion-capacity resin. Namely, there existed more significant electroosmotic flow in the high ion-capacity resin, which enhanced the intraparticle mass transfer of protein, leading to the increase of dynamic protein adsorption capacity.
A mathematical model of the pIEEC was developed, and the model parameters were determined by independent experiments or calculations. The model was found in good agreement with the above experimental results. The model was then used to analyze the pIEEC process. It was found that protein adsorption affected the intraparticle electroosmotic flow, which decreased exponentially with increasing protein adsorption. Moreover, protein distribution in adsorbents was found to present excursion along the electric field direction. At the beginning of the pIEEC, intraparticle convection caused by the electric field contributed more to the enhancement of dynamic binding capacity.
The pIEEC (2 mL) was finally used to separate hen egg-white (HEW) proteins. The results were compared with those of normal ion exchange chromatography (IEC). Q Sepharose FF was packed into the central compartment as the chromatographic bed. It was confirmed that the dynamic binding capacity (DBC) of different proteins (ovotransferrin and ovalbumin) in the HEW solution increased 2.3 times when an oscillatory electric current of 30 mA at 1/20 Hz was applied in the transverse column direction. Then, the HEW proteins were separated by the pIEEC at loading amounts 2.3-fold higher than those by the IEC, and similar separation efficiencies of the two chromatographic modes were achieved. Both the recovery yield and purity reached 73% to over 90%. The results indicate that the pIEEC is promising for high-capacity purification of proteins.
首先构建了横向电场电色谱测温系统,测定了电色谱冷却和施加电场的动态升温过程,研究了电流强度和流动相离子强度对柱内温度分布的影响。建立了横向电场电色谱过程的传热模型,模拟从非稳态操作到稳态操作的温度分布。利用该模型可同时获得柱内电压分布、有效电压和功耗等参数,分析电色谱放大的温度效应,以及各种条件下的温度分布行为。
制备了不同配基修饰密度的离子交换介质,研究了电色谱的蛋白质动态吸附行为。结果表明,由于高配基密度介质的表面电荷密度较高,孔内电渗流对促进传质的作用更明显,故蛋白质动态吸附容量随电场强度增加较多。
建立横向交变电场离子交换电色谱过程的理论模型,利用上述蛋白质动态吸附实验结果,验证了模型的可靠性。系统分析了电色谱过程的传质和色谱行为。理论分析发现,电渗引起的孔内对流受蛋白质不可逆吸附的影响,孔内对流速度对吸附饱和度呈负指数下降趋势;在电场作用下,吸附剂内蛋白质浓度分布呈现出沿电场方向的偏移,在操作的初始时刻电场作用效果有利于提高动态吸附容量。
利用填充阴离子交换介质Q Sepharose Fast Flow的横向电场色谱,研究了卵清蛋白质的分离纯化,获得了高纯度卵铁传递蛋白和卵清白蛋白。与普通离子交换色谱相比,横向电场电色谱的分离能力提高一倍以上,证明了该横向电场电色谱的高容量分离纯化性能。
This thesis is concerned with the studies of ion-exchange electrochromatography with an oscillatory electric field perpendicular to mobile-phase flow driven by pressure (pIEEC). The electrochromatography is composed of a three-compartment column developed in the laboratory. The work has investigated heat and mass transport phenomena systematically by both experiments and mathematical modeling.
First of all, an experimental system for in-column temperature measurements was constructed, and the dynamic processes of the in-column temperature in electrochromatography were examined. With the experimental system, the effect of electric current strength and mobile-phase ionic strength on the in-column temperature was investigated. Then, a heat transfer model for the pIEEC was established for the dynamic process modeling. It was confirmed that the model was in good agreement with the measurements. By the model calculations, we could also obtain the in-column voltage distribution, effective voltage applied to the central compartment and the efficiency of energy consumption. Moreover, the model was used for scale-up analysis and for investigation of temperature distribution under various conditions.
Protein ion-exchangers of different ion-exchange capacities were prepared to study the dynamic protein adsorption processes of the pIEEC. The results indicated that electric field for the pIEEC was more effective for the high ion-capacity resin. Namely, there existed more significant electroosmotic flow in the high ion-capacity resin, which enhanced the intraparticle mass transfer of protein, leading to the increase of dynamic protein adsorption capacity.
A mathematical model of the pIEEC was developed, and the model parameters were determined by independent experiments or calculations. The model was found in good agreement with the above experimental results. The model was then used to analyze the pIEEC process. It was found that protein adsorption affected the intraparticle electroosmotic flow, which decreased exponentially with increasing protein adsorption. Moreover, protein distribution in adsorbents was found to present excursion along the electric field direction. At the beginning of the pIEEC, intraparticle convection caused by the electric field contributed more to the enhancement of dynamic binding capacity.
The pIEEC (2 mL) was finally used to separate hen egg-white (HEW) proteins. The results were compared with those of normal ion exchange chromatography (IEC). Q Sepharose FF was packed into the central compartment as the chromatographic bed. It was confirmed that the dynamic binding capacity (DBC) of different proteins (ovotransferrin and ovalbumin) in the HEW solution increased 2.3 times when an oscillatory electric current of 30 mA at 1/20 Hz was applied in the transverse column direction. Then, the HEW proteins were separated by the pIEEC at loading amounts 2.3-fold higher than those by the IEC, and similar separation efficiencies of the two chromatographic modes were achieved. Both the recovery yield and purity reached 73% to over 90%. The results indicate that the pIEEC is promising for high-capacity purification of proteins.
引文
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