生化分離程序中膜過濾機制之界面現象解析

Membrane Filtration Mechanisms for Bioseparation Using Interfacial Phenomena Analysis

胡哲嘉(Che-Chia Hu)

指導教授 ： 童國倫 ； 莊清榮

中原大學/工學院/化學工程研究所/博士(2007年)

https://doi.org/10.6840/cycu200700085

摘要

本研究探討薄膜過濾生物粒子時粒子阻塞和其界面現象關係，以瞭解利用薄膜進行生物粒子微過濾和蛋白質混合溶液超過濾之過濾機制，期能得知粒子的結垢、阻塞行為並藉由粒子-粒子和粒子-薄膜間的界面現象探討對分離的效能，進而開發出更有效率的操作程序以降低分離的成本。
研究的第一部份即使用可變形酵母菌粒子並改變其不同粒子變形度在不同微過濾薄膜孔徑和操作壓力下，探討不同變形度粒子在恆壓過濾和掃流過濾時的薄膜阻塞機制。結果發現可變形粒子和改質後粒子的過濾機制並不完全相同，因為可變形粒子在過濾時受壓產生變形，使得濾餅的孔隙度降低而形成較緻密的濾餅層，造成過濾阻力的快速上升；而隨著改質程度的增加，改質後的粒子因較不易變形，所以過濾阻力主要來自於濾餅層的厚度，在相同過濾時間下通常都有較高的濾液通量且過濾阻力也較低。
第二部份為蛋白質 (bovine serum albumin、ovalbumin和lysozyme) 混合溶液的超過濾分離和吸附性掃流過濾的分離探討。結果指出單成分蛋白質過濾行為明顯受到靜電作用力的影響，若蛋白質和薄膜帶相反電性時，操作在接近蛋白質的等電點，有較高的回收率；若蛋白質和薄膜帶相同電性時，適當的靜電作用力有助於蛋白質穿透率提升。在雙成分蛋白質分離，親水化改質之PES膜，可以有效分離分子量相近之BSA-OV，濾液端可獲得100% 純度之OV；而對於分子量相差大之BSA-LY系統，選擇適合的薄膜孔徑和溶液環境亦可將兩蛋白質完全分離。由吸附性掃流過濾結果得知，利用該方法可以將BSA-LY混合蛋白質完全分離而獲得與層析相同純度的LY。比較利用吸附性掃流微過濾、掃流超過濾和層析三種方式對雙成分蛋白質的分離，顯示利用三種方式皆可獲得高純度的產物，而在相同的操作時間下以分離效率來比較，吸附性掃流微過濾的回收量是最高的而層析法則為最低。
綜觀本研究，由結果可得知不同變形度粒子在不同膜孔徑下的過濾阻塞機制也不盡相同；而利用吸附性掃流微過濾和掃流超過濾能有效將蛋白質混合溶液進行分離和純化且可提升分離的效率，若純化程度不及層析時，亦可做為層析法的前處理步驟可大幅減輕後段的純化分離程序。

關鍵字

超過濾 ； 蛋白質分離 ； 蛋白質吸附 ； 離子交換吸附 ； 電漿改質 ； 酵母菌 ； 薄膜過濾 ； 微過濾

並列摘要

In this study, membrane filtration mechanisms of bio-particle blocking in microfiltration and protein mixture separation with ultrafiltration were analyzed in order to develop efficient and economically viable processes. The investigation was done by exploring the particle blocking behaviors and separation performance of the interfacial phenomena between particle-particle and particle-membrane system.
In the first part of this study, membrane blocking mechanisms of yeast particles with different degree of deformability were studied by analyzing the different filtration behavior due to the pore sizes of microfiltration membranes, applied pressures on dead-end filtration, and crossflow filtration. The results show that there are different filtration mechanisms between soft yeast particles and which were treated with glutaraldehyde more rigid yeast particle. Due to the compression of yeast particles, the lower porosity particle cake layers on the membrane surface formed a dense skin cake layer which significantly increased the filtration resistance during filtration. On the other hand, the filtration resistance of treated yeast particle was mainly caused by the growing in cake thickness because the treated particle could not be compressed as easily by pressure, and as a result, more filtrate was collected during filtration. At the same filtration time, the treated yeast particle had higher filtrate flux and lower filtration resistance compared to the soft yeast particles.
The protein mixture (bovine serum albumin, BSA; ovalbumin, OV; lysozyme, LY) separation mechanisms by crossflow ultrafiltration and adsorption crossflow filtration were conducted in the second part of this study. The results show that the filtration behaviors are affected by electrostatic interactive force during single protein filtration. If the protein and the membrane carry opposite charges, higher protein recovery is observed when the solution pH approaches protein’s isoelectric point. If the protein and the membrane carry the same charge, electrostatic interaction increases the protein transmission during filtration. For protein mixture separation, the hydrophilic treated polyethersulfone membrane effectively separates BSA-OV mixture which have similar molecular weight in each component. For a system (BSA-LY mixture) with large molecular weight difference, selecting the suitable membrane molecular weight cut-off and solution condition were also shown to be effective to achieve 100% purity target protein in the filtrate. The results of adsorption crossflow filtration show that the method can produce high purity target protein as obtained by chromatography in BSA-LY mixture system. Comparing the performance and efficiency of crossflow ultrafiltration, adsorption crossflow filtration, and chromatography, all methods can achieve high purity target protein after separation. However, the adsorption crossflow filtration show higher target recovery rate than crossflow ultrafiltration and chromatography.
In conclusion, the results indicate that in microfiltration different filtration mechanisms exist for yeast particles with different deformability. The use of adsorption crossflow filtration and crossflow ultrafiltration can enhance the separation efficiency and product purity of protein mixture. If the protein purity by the crossflow ultrafiltration is not as good as what a chromatography method can provide, filtration process can serve as a pre-treatment step in separation and purification procedure to reduce the loading of the final chromatography step.

並列關鍵字

protein separation ； ion exchange adsorption ； protein adsorption ； membrane filtration ； Saccharomyces cerevisia ； ultrafiltration ； plasma modification ； microfiltration

參考文獻

李佳玲，蛋白質混合溶液膜過濾之界面現象研究，碩士學位論文，私立中原大學化學工程研究所，中壢 (2004)。

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