以逆流層析分離幾丁寡醣可行性之探討

Abstract

目前用來分離幾丁聚醣的方法有高效能液相層析、毛細管電泳，但此兩種方法皆無法收集大量樣品。本研究以逆流層析儀(countercurrent chromatography)結合雙水相溶劑系統(aqueous two phase solvent system)或有機溶劑/水相系統分離並收集幾丁寡糖。試驗的條件包括，改變溶劑的組成、比例、濃度與pH值。其結果顯示，在雙水相溶劑系統中，此類分子量較小、極性較高的幾丁寡醣分子的分離效果不佳，即醣類分子大都留在上層聚乙二醇相( Polyethylene Glycol，PEG )中，儘管提高PEG的分子量、濃度，改變溶劑中所含鹽的種類(包括: 磷酸鹽、檸檬酸鹽)及其所佔的比例，或將溶劑系統的pH值降低，均無法將寡醣流析分離出，此可能是因為幾丁寡醣分子與聚乙二醇間的親和力較大所造成的。而在有機溶劑/水相系統中，無論如何提高溶劑系統的極性或pH值，亦無法提高寡糖分子在兩相間的分配係數，即醣類分子均留在水相當中。

Two methods which are used to separate chitooligosaccharides are HPLC and CE, but they cannot handle great sample volume either. In this study, we try to separate and collect chitooligosaccharides in either of aqueous two phase solvent system and organic/aqueous solvent system in countercurrent chromatography. Experimental factors are composition of solvent, sample concentration and pH value. As the result, in aqueous two phase solvent system, chitooligosaccharides with low molecular weight and high polarity are hardly separated while they almost stay in the upper layer, PEG(polyethylene glycol) - rich phase. Even though we increase the molecular weight or concentration of PEG, add different kinds of salts including phosphate and citrate, change the concentration of salt, or lower the pH value in solvent system, chitooligosaccharides cannot be eluted out. It is suggested that a good affinity exists between chitooligosaccharides and PEG. In organic/aqueous solvent system, however, no matter how greatly we try to increase the polarity or the pH value of our solvent systems, the partition coefficient of chitooligosaccharides between the two phase cannot be increased, so they remain in the aqueous phase.