人造沸石对甘薯不同链长直支链淀粉的分离效果

Separation effect of sweet potato amylose and amylopectin with different chain length by artificial zeolite

柱层析法是根据混合物中各组分的理化性质差异,利用混合物中各组分在固定相和流动相中分配系数不同,经过多次分配将组分分离纯化。该文采用回生法分离甘薯直、支链淀粉后再进行二次回生,以窄化其分子链长,并采用柱层析法分离出不同分子量分布范围的淀粉组分,直、支链淀粉组分总制备率均大于2.4%。可见光谱及分子量分布研究表明,粒径为1~3 mm的人造沸石适合分离支链淀粉,粒径为4~6 mm的人造沸石适合分离直链淀粉。经大沸石分离,聚合度大的直链淀粉分子依赖沸石表面吸附,洗脱速度快,先被洗脱下来,聚合度较小的直链淀粉分子进入沸石孔隙内部,吸附力大,后被洗脱下来;经小沸石分离,甘薯支链淀粉组分F1b中分子量小、均一度高的组分先被洗脱下来,F2b中分子量大、均一度低的组分先被洗脱下来,说明甘薯直、支链淀粉的分支度对淀粉组分的分离也有一定影响。X射线衍射表明,甘薯直链淀粉组分在18.9°、23.4°、27.2°、29.3°、32.3°、33.7°附近出现强衍射峰,甘薯支链淀粉组分在21.6°、22.9°、23.9°、26.5°、27.1°、29.3°、34.1°、35.8°、39.5°附近也出现明显的衍射峰,此时甘薯直、支链淀粉的分子量分布指数(PDI,polydispersity index)均接近1.0。人造沸石柱层析可制备出分子量分布范围极窄的甘薯直、支链淀粉,该类淀粉表现出类似金属盐的X射线衍射峰,可作为深入研究淀粉大分子空间结构变化的材料。显微图片显示直链淀粉是由多个线型分子聚集在一起,呈典型的"柳条"状,而支链淀粉呈典型的"树枝"状。该研究提供了一种大量制备分子量分布范围极窄的直、支链淀粉的简单、高效方法,为深入探索淀粉大分子聚集过程中的形态变化创造了有利条件。

Column chromatography is based on the difference of physicochemical properties of components in the mixture.The mixture is separated after multiple distributions by using the different distribution coefficients of each component in the stationary and mobile phase.The starch was hydrolyzed by amylase after retrogradation under high pressure and humidity.Then,the retrograde starch was dissolved by 4 mol/L potassium hydroxide and the solution was adjusted to neutral next.The precipitate of amylose was obtained by adding n-butanol to the solution.While,amylopectin was prepared by adding ethanol to the supernatant.Both of amylose and amylopectin in sweet potato were retrograded for the second time to narrow their molecular weight distribution.And column chromatography was used to separate the components furtherly.The total yields of amylose and amylopectin were more than 2.4%respectively.The results of visible spectrum and molecular weight distribution showed that Mwof amylose and amylopectin narrowed to 1641-2069 g/mol and 1671-2167 g/mol,respectively,while Mnnarrowed to 1516-1710 g/mol and 1526-1678 g/mol.The corresponding PDI of amylose ranged from1.082-1.209,and PDI of amylopectin ranged from 1.095-1.291.Artificial zeolite with 1-3 mm diameter was suitable for the separation of amylopectin,while artificial zeolite with 4-6 mm diameter was suitable for amylose.In the course of separation,amylose with higher DP adsorbed on the macrozeolite surface.It was eluted out from the mixture first for the weaker adsorption force.Amylose with lower DP entered into the small holes of macrozeolite,and was eluted out subsequently for the stronger adsorption force.When separated by small zeolite column chromatography,the amylopectin components of F1 b with small molecular weight and high homogeneity were eluted out first.While,amylopectin components of F2 b with large molecular weight and low homogeneity were eluted out first,indicating that the branching degree of amylopectin also played a certain role in the separation of starch components by zeolite column chromatography.X-ray diffraction showed distinctive spectra.Amylose components revealed strong sharp peaks at the diffraction angles(2θ)of 18.9°,23.4°,27.2°,29.3°,32.3°,33.7°.Amylopectin components showed obvious sharp peaks at around 21.6°,22.9°,23.9°,26.5°,27.1°,29.3°,34.1°,35.8°,39.5°.The molecular weight distribution index(PDI)of sweet potato amylose was close to 1.0 at the same time.Amylose and amylopectin with extreme narrow molecular weight exhibited sharp peaks like metallic salts.That could be used for the study of the spatial structure changes of starch macromolecules in depth.Micrographs showed that amylose was composed of many linear molecules and presented typical linear"wicker-like"morphology,while amylopectin showed"branch-like"shape.The results provide a simple and efficient method for the preparing of amylose and amylopectin with extreme narrow molecular weight distribution.It brings about favorable conditions for further exploring morphological changes of starch macromolecules during aggregation progress.