| Radix Astragali membranaceus(Fisch.)Bge.var.mongholicus(Bge.)Hsiao polysaccharides and Radix Astragali membranaceus(Fisch.)Bge polysaccharides Panax ginseng are two important plants of the Radix Astragali.The polysaccharides of Radix Astragali have the same chemical component among which polysaccharides are the most active constituents,which play an important role in the pharmacological activity of Radix Astragali.Therefore,it is an effective method to evaluate the quality of medicinal materials based on polysaccharides.The polysaccharides were depolymerized by complete acid hydrolysis,partial acid hydrolysis,smith degradation and complete methylation.GC-MS and HILIC-ELSD were used to separate and detect the products.In order to further improve the quality evaluation of polysaccharides and simplify the difficult process of traditional analysis methods,the HILIC-UPLC-ESI+-HCD-MSn technology was used to establish the type of glycosidic bonds based on mass spectrometry to determine the structure of Radix Astragali.The resulting bottom-up fingerprints and Sugar spectrometry reflect the structural changes and identification of Radix Astragali polysaccharides.The main research contents and results are as follows:1.Identification of Radix Astragali polysaccharides based on monosaccharide fingerprinting based on completed acid hydrolysisThe optimal trimethylsilyl-alditol derivative method was used to analyze the monosaccharide composition of Radix Astragali polysaccharides According to the similarity analysis of TCM polysaccharides fingerprint,the 7 batches of AEP(S1-S7)and the 13 batches of AOP(S'1-S'13)show Cr values ranging from 0.914 to 0.996 and 0.966 to 0.998.Meanwhile,the GC-MS fingerprint profiles of AEP samples compared with the R' derived from AOP,AOP samples compared with the R derived from AEP,show Cr values ranging from 0.675 to 0.796 and 0.611 to 0.796.In terms of the composition and content of the monosaccharides,it was found that the similarity between Radix Astragali samples of different origins was higher than that across species2.Identification of Radix Astragali polysaccharides based on Polyol fingerprinting based on Smith degradationCombined with GC-MS,the polysaccharides of Radix Astragali were pretreated with periodate oxidation and Smith degradation.As the result,the similarity based on the selected eight variables showed that the Cr values of AEP and AOP ranged between 0.957-0.996 and 0.942-0.999,respectively However,the Cr value between AEP and AOP was less than 0.743.These values also indicated that AEP and AOP had different polyol fingerprints There were 3 degraded products,which were unambiguously identified as glycol,glycerol and erythritol.This fact indicated that there is possible occurrence of,1?6 and/or 1?4 glycosidic linkages for hexose in Radix Astragali polysaccharides.The most dominant glycerol was observed as degraded products along with some small glycol and erythritol.This fact indicated that 1?6 glycosidic linkages may be occurrence in structural skeleton of Radix Astragali polysaccharides rather than 1?4 glycosidic linkages.It also noted that characteristic peak area ratio could be used a feasible parameter for direct discrimination of AEP and AOP.AEP produced the ratios of Man/Gal which are from 0.18 to 0.48,but the AOP produced the value of Man/Gal that ranged from 1.13 to 1.84.This is the most convincing evidence for the successful discrimination of polysaccharides from A membranaceus and A.mongholicus3.Identification of Radix Astragali polysaccharides based on Aldi alcohol acetate fingerprintingThe GC-MS combined with aldi alcohol acetate showed the methylation assay afforded additional evidences that the similarity Cr values of AEP and AOP ranged between 0.969-0.998 and 0.935-0.999,respectively.However,the Cr value between AMP and AHP was ranged from 0.635 to 0.779.These hexose glycopolymers possessed absolutely dominant 1?6 glyosidic linkages(peak 2)along with minor 1?4 glyosidic linkages.Nevertheless,1?4 glycosidic linkages were reported to be the dominant skeletal structure in purified Radix Astragali polysaccharides.For example,some glucans were isolated from the roots of A.membranaceus.The glucans were identified to be the ?-(1?4)-backbone and the ?-(1?6)-linked branch attached to the O-6 of branch points every 10 residues.This discrepancy may arise from crude polysaccharides used in this study4.Identification of Radix Astragali polysaccharides based on mild the acid hydrolysisIn this study,mild acid hydrolysate of Radix Astragali polysaccharides was further investigated using hydrophilic interaction liquid chromatography-evaporative light scattering detectors(HILIC-ELS D).The result showed 11 peaks as mild acid hydrolyzates were observed.The similarity evaluation showed that the Cr values of different origins of AEP and AOP were 0.966 to 0.977 and 0.935 to 0.999,respectively.Meanwhile,the GC-MS fingerprint profiles of AEP samples compared with the R' derived from AOP,AOP samples compared with the R derived from AEP,the Cr values ranged from 0.683 to 0.731 across species.It indicated that these differences made it easier to distinguish different species.However,Mass spectrometric fingerprinting may be a more effective and sensitive tool than ELSD technique,so Electrospray-quadrupole time-of-flight(ESI'-QTOF)was used to record m/z fingerprinting in the negative mode through a simple separation of oligosaccharides in mild acid hydrolyzates.The results showed that principal component analysis(PCA)from m/z fingerprinting data resulted in clear clustering of AEP and AOP.For qualitative analysis of oligosaccharide peaks structure,compare different detection methods based on CID and HCD,and summarized the MS/MS mass spectrometry rule of different disaccharides.Then,with the increase of polysaccharides,the structure analysis ability of the polysaccharides was observed.The HILIC-UPLC-ESI-MSn was attempted for structural identifications in mild acid hydrolyzates of polysaccharides from AEP and AOP.The dominating peaks 2,4,6,8,10,12,14,16,18,20,22,24,26,28,30,32,34,36,and 37-41 should be identified as a series of linear glucans with DPs from 2 to 24,characterized by 1?6 glycosidic linkages.In the same way,the minor peaks 1,3,5,7,9,11,13,15,17,19,21,23,25,27,29,31,33 and 35 were also identified as a series of hexose glycopolymers from DP 2 to 19,with 1?6 glycosidic linkages,which should be a mixture of glucosyl,galactosyl,and/or mannosyl oligosaccharide,with a diverse ratio. |
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