Extraction,Purification,Structural Characterization And Biological Activity Of Polysaccharides From Carex Meyeriana Kunth

Carex meyeriana Kunth,which belongs to Cyperaceae,is a ubiquitous perennial herb plant in northeast China.It has the functions of warmth retention and blood circulation improvement,and has high research value.At present,literature search shows that the research of Carex meyeriana Kunth is mainly focused on volatile oils and flavonoids,while the research of polysaccharides from Carex meyeriana Kunth has not been systematically studied.In this paper,the extraction,purification,separation,structure characterization and biological activity of Polysaccharides from Carex meyeriana Kunth were systematically studied,which provided theoretical basis and reference for deep Carex meyeriana Kunth polysaccharides.The research methods and main results are as follows:The hot water extraction process of polysaccharides was optimized by response surface methodology(RSM),microwave-assisted extraction and ultrasound-assisted extraction were compared simultaneously.The structure of Hot-water extraction polysaccharides(HWEP)and Microwave-assisted extraction polysaccharides(MAEP)and Ultrasound-assisted extraction polysaccharides(UAEP)were characterized by high performance liquid chromatography(HPLC),infrared spectroscopy(FTIR)and ultraviolet spectrophotometry(UV-Vis).The antioxidant activities of three polysaccharides were evaluated by measuring DPPH free radical,hydroxyl free radical and ABTS + free radical scavenging activity.The results showed that the extraction rates of polysaccharides by hot water extraction,microwave-assisted extraction and ultrasound-assisted extraction were 0.47±0.01%?0.28±0.02% and 0.25±0.02% respectively,so the extraction rates of polysaccharides by hot water extraction were the highest.HWEP and MAEP both consist of Rha:Xyl:Ara:Fru:Glu:Gal with the molar ratio of 1.05:1.21:3.86:1:3.61:4.5 and 1:1.95:1.72:1.78:4.36:6.18,respectively.UAEP consists of Rha:Xyl:Ara:Fru:Man:Glu:Gal with the molar ratio of 1:2.31:5.23:1.05:3.17:4.17:7.89.The molecular weight distribution of HWEP,MAEP and UAEP ranged from 16 kDa to 1698 kDa,15 kDa to 913 kDa,and 17 kDa to 1118 kDa,respectively.The absorption peaks in FT-IR and the second derivative of FT-IR proved difference of polysaccharides obtained from different extraction methods.The antioxidant activity investigations shown all three polysaccharides extracts possess high scavenging activity of DPPH radicals,hydroxyl radical and ABTS+ radical,in which HWEP has better antioxidant activity than the other two.Therefore,HWEP has higher research value.Five kinds of macroporous resins AB-8,D101,S-8,X-5 and NKA-II were used to purify the polysaccharides from Uracia uralensis by static adsorption test.Finally,AB-8 macroporous resin was screened out to be more suitable for purifying Uracia uralensis polysaccharides.Then,the macroporous resin was used to purify the polysaccharides from Urapidium,and the purification process of Carex meyeriana Kunth polysaccharides was optimized by analytic hierarchy process(AHP)and response surface methodology(RSM).The results of hierarchical analysis showed that the weight coefficients of polysaccharide recovery,deproteinization ratio and decolorization ratio were 0.515,0.372 and 0.113,respectively.The optimum conditions for polysaccharide purification were as follows: elution volume of 2.74 BV,flow rate of 1.88 BV/h,and sample concentration of 2.10 mL/g,and the maximum predicted comprehensive score was 64.02%.Polysaccharide recovery ratio,deproteinization ratio and decolorization ratio were 49.63 ± 1.24 %,82.06 ± 2.17 % and 66.43 ± 1.51,respectively.The purified polysaccharides were separated by anion exchange DEAE-52 cellulose.PMP pre column derivatization,gel chromatography,UV-Vis and FTIR were used to characterize the structure.RAW264.7 cells were used to study the immunomodulatory activity of polysaccharides.Finally,the polysaccharides of CMKP-1 and CMKP-2 were obtained successfully.CMKP-1 and CMKP-2 were both composed of Man,Rib,Rha,GluA,GalA,Glu,Gal,Xyl,Arab and Fuc in a molar ratio of 8.93:1:4.62:1.41:1.78:10.7:24.69:4.51:15.92:1.27 and 7.85:1:5.05:1.99:3.15:12.47:15.41:1.54:6.91:3.15,respectively.The major molecular weight of CMKP-1 and CMKP-2 was 22.30 kDa and 48.10 kDa,respectively.Fourier Transform infrared(FTIR)spectroscopy spectra revealed the general characteristic absorption peaks of CMKP-1 and CMKP-2.Congo-red test indicated that CMKP-1 and CMKP-2 have triple helix conformation.Immunomodulation experiments showed that CMKP-1 and CMKP-2 had good immunomodulatory effects on RAW264.7 cells in a certain concentration range,and could be used as a potential immunomodulator.