使用逆流層析及離子液體製備分離蛋白質中藥藥材和奈米粒子(II)

Abstract

本計畫預定使用離子液體，以及逆流層析法，來製備分離蛋白質、中藥藥材，以及 奈米粒子。 近年來，離子液體之發展迅速，一般認為離子液體的研究屬於綠色化學的一環，也 被使用於液相/液相的萃取，本計畫擬以離子液體和染料合成為加成物，由於此種染料 對於特定蛋白質之結合，有選擇性的效果，若將這種加成物質溶於離子液體中，和水 溶液混和可形成兩相，水溶液中之特定蛋白質會轉移到離子液體相中，兩相分離之後， 離子液體相和另一高鹽濃度之水溶液混和，蛋白質又會回到水相而達成和其他蛋白質 分離之效果。計畫中，我們擬合成數種離子液體和染料合成為加成物，並進行選擇性 之萃取，以試驗此種加成物質是否可應用於蛋白質之純化。 本實驗長年從事逆流層析製備分離之研究，本計畫中之後兩年預定研究中藥有效成 分和奈米粒子之分離。中藥標準品的製備對於下游的品管以及研究部門重要性極高，由 於逆流層析對於天然物之分離，相較於填充矽膠之分離管柱，可免於永久吸附以致於破 壞管柱之狀況發生，很適合於中草藥之分離，我們預定製備分離龍膽和無患子，發展出 最適化之溶劑系統。至於奈米粒子之合成以及應用，近年來也已成為顯學，一般合成時 都會尋求能夠製出粒子大小分佈很窄的範圍，然而並非都能達成，因此使用層析分離的 手段也是值得發展的研究。使用反微胞萃取，可以粗略的將合成的奈米粒子依大小分 離，但是效果並非十分好，逆流層析分離法就如同進行數千次的萃取，因此分離效果會 大幅改進，我們將先嘗試分離金奈米粒子，然後嘗試其他物質，並研究分離相關最適化 之實驗條件。

We plan to achieve preparative separations for proteins, Chinese herbs, and nano-particles using ionic liquids and counter-current chromatography (CCC) in this proposal. We have seen rapid development of ionic liquids in recent years. The researches in this area have been generally considered as one branch of green chemistry. We plan to synthesize several adducts of ionic liquids with dyes. Due to the selective interaction between certain proteins with these dyes, the adducts can be used in selective extraction of proteins. The solution made by dissolving the adduct in ionic liquid would be immiscible with aqueous solution. Protein molecules in the aqueous phase are ready to be transferred to the ionic liquid phase by mixing the two phases. While mixing with another aqueous solution with high electrolyte concentration, protein molecules would be transferred back to the aqueous solution. Thus, the protein with unique interaction with the dye can be selectively separated from other proteins. We plan to investigate the feasibility of using this technique for protein purifications. We have been engaged in preparative separations using CCC for many years. In the last two years of this multi-years proposal, we plan to carry out separations of Chinese herbs and nano-particles. Preparation of standard samples for the active components is an important subject in research of Chinese herbs. Compared with column chromatography packed with silica gel, no permanent adsorption would occur to shorten the column lifespan in CCC. As a result, it is very suitable in natural product separations. We will study two herbs, i.e. Gentiana scabra and Sapindus mukorossi, in this proposal. Syntheses and applications of nano-particles have drawn great attention in recently years. Researchers usually try to synthesize particles with as narrow size distribution as possible. While this can not be achieved, chromatographic techniques would be helpful to obtain narrow-sized portions. Extraction using reverse micelles to fractionate particles in different size has been found in the literature. Efficiency can be greatly improved while thousands of extractions are carried out in one CCC experiment compared with stepwise separatory-funnel extractions. Gold nano-particles would be investigated first, followed by other nano-particles. Experimental parameters will be investigated to optimize separations.