| Research Background:Hericium erinaceus is a kind of classic fungus used as both medicine and food,which has come into people's lives for centuries because not only it was precious and had excellent taste but also it was associated with in favor of organs,promoting digestion and nourishing your bodies by traditional Chinese medicine.With the domestication of wild H.erinaceus and the development of artificial cultivation,the amount of studies on effective components in H.erinaceus has been increasing.Among them,polysaccharides of H.erinaceus attract numerous researchers for high content and tiny side effects.It's reported that the polysaccharides have multiple pharmacological properties,such as antitumor and immuno-regulation,protecting gastrointestinal mucosa,protecting and regenerating neurons,antihyperlipidemic and hypoglycemic effects.Especially,glucan with ?-(1?3)and ?-(1?6)linkages has remarkable antitumor and immunoregulatory activities.Whereas,different sources of H.erinaceus and various methods to prepare and purify polysaccharides result in huge diversities on purity and structure.What' more,the general lack of sequence analysis sets barriers to the understanding of structure-activity relationship and clinical application.Purposes:To optimize a series of procedures to prepare ?-glucan with high purity and to illuminate the sequence of ?-glucan fraction in depthMethods:(1)We optimized conditions of papin digestion,molecular cut-off,methods to depigment and chromatographic separation through single-factor experiments;(2)The types of glycosidic bonds,configuration and molecular weight were judged by NMR,methylation and HPSEC-MALLS-RI;(3)Coupled HPAEC-PAD with Q-TOF-MS/MS to analyze oligosaccharides digested from ?-glucanResults:(1)We acquired four fractions,named HEP1?HEP4;(2)The ?-configuration of fucose,galactose and ?-configuration of glucose were confirmed in HEP1 and HEP2,which contained terminal fucose,?-1,6-Ga1 linkage with low branched ?-1,2,6-Ga1 linkage,?-1,6-Glc linkage as main chain with numbers of branches on position O-3.(3)The results of sequence analysis showed that there were oligosaccharides with various dps and several isomerides with the same dp in aliquots.After digestion of most 1-6 linked fractions,remnant fractions of oligosaccharide were composed of two or three short 1?6 linked sugar chains hooked up by 1?3 linkages,which seemed like products digested from the net-like glucan.It can be imagined that the net-like glucan is composed of numerous ?-1,6-linked glucose linear chains hooked each other on the position O-3 by 1?2 ?-1,3-linked glucosyl residue(s).What' more,PAD semi-quantitatively calculated the ratio of ?-1,3-G1c and ?-1,6-Glc linkages as 1:6.Conclusions:In our project,we established a set of procedures to prepare polysaccharides from H.erinaceus with high purity and elucidated detailed structure information of four fractions.It is worth noted that we established the hyphenated method,which combined HPAEC-PAD with Q-TOF-MS/MS,to analyze the sequence of ?-glucan from H.erinaceus and characteristics of digested oligosaccharides.The results laid a solid foundation on structure to explore structure-bioactivities of ?-glucan in H.erinaceus. |
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