Study On The Extraction Of Polysaccharide From Chinese Yam And Its Structure Identification And Its Effect On Intestinal Flora

Objective Optimize the extraction and purification of yam polysaccharide.Investigatestructural characterization of yam polysaccharide and the influence on the grown ofintestinal bacteria in vitro.Methods To optimization crude yam polysaccharide (CYPN, CYPA) by thecomposite design-response surface method of extraction rate as index. The crude yampolysaccharide was deproteinated according to the TCA and hydrochloric acid method.Using deproteinization rate and polysaccharide retention rate as index, optimize and filterthe best method of deproteinization. And the crude yam polysaccharide afterdeproteinization was discolored by H2O2method and removed small molecules by dialysismethod. CYPN and CYPA were separated by DEAE-52cellulose columnchromatography, and their purity and molecular weight were determined by highperformance gel chromatography. Polysaccharide monosaccharide composition and molarratio of CYPN and CYPA were detected by HPLC. In addition, the structure of CYPN andCYPA were determined by IR and NMR. Finally, the influence beteween yampolysaccharide and bacteria (Bifidobacterium, Lactobacillus, Escherichia coli,Enterococcus faecalis) were tested by the turbidimetry method.Results We obtained the optimum extraction of yam polysaccharide by cold water:ratio of material to liquid10.30:1; extraction time of12.26h; extraction of three time. Asthis extraction condition, the extraction rate of CYPN is2.89%and content ofpolysaccharide is57.43%. The optimum extraction of yam polysaccharide by lye: ratio ofmaterial to liquid10.23:1; extraction time of7.59h;concentration of lye of0.48mol·L-1.As this extraction condition, the extraction rate of CYPA is24.23%and content ofpolysaccharide is45.66%.Concentration of trichloroacetic acid3%is the optimumdeproteinization of TCA method, the deproteinization rate is95.03%, the polysaccharideretention rate is24.19%. The pH2.0is the optimum deproteinization of hydrochloric acidmethod, the deproteinization rate is93.95%, the polysaccharide retention rate is67.74%.So the hydrochloric acid method is the best method to deproteinization. We obtainedhomogeneous polysaccharides (CYPN-I, CYPN-II, CYPA-I, CYPA-II and CYPA-III) by high performance gel chromatography. And the high performance gel chromatographyresults revealed that all the homogeneous polysaccharides display a single peak, and themolecular weight is7907Da,54200Da,60256Da,76913Da and47753Da, respectively.HPLC analysis showed that the monosaccharide composition: CYPN-I contains glucose,galactose, arabinose; CYPN-II contains mannitol, rhamnose, glucose, galactose, xylose,arabinose; CYPA-I contains rhamnose, galacturonic acid, galactose, xylose; CYPA-IIcontaining rhamnose, galacturonic acid, glucose, galactose, arabinose; CYPA-III containsmannose, rhamnose, galacturonic acid; IR and NMR results display that CYPN-I andCYPN-II are neutral polysaccharide, CYPA-I, CYPA-II and CYPA-III are acidicpolysaccharides, CYPN-I and CYPA-II containing α-type pyranose, CYPN-II, CYPA-I,CYPA-III containing β-type pyranose. Turbidimetry results showed that yampolysaccharides have a foster effect on bifidobacteria and lactobacilli, and a inhibitedeffect on colon coli and Enterococcus faecalis.Conclusions This experiment respectively to optimize the best extraction technologyof yam polysaccharide’s cold water extraction and alkali extraction; Determining thehydrochloric acid method is the best removaling of protein; Yam polysaccharide afterpurificating received five yam polysaccharides (CYPN-I, CYPN-II, CYPA-I, CYPA-II,CYPA-III); Detection and analysis of the five yam polysaccharides molecular weight,monosaccharide composition and its structure; Experiments have established that five yampolysaccharides on fostering effects of bifidobacterium, lactobacillus, dung enterococcushave inhibition on escherichia coli, and different concentrations of yam polysaccharidehave different affect of various bacteria in vitro growth, this experiment laids thefoundation experiment for yam polysaccharide on the intestinal flora in micro ecologicalresearch, provides a theoretical basis for new drug development.