Isolation And Purification,Structure Characterization And Bioactivities Of Polysaccharides From Hylocereus Undatus Flowers

Hylocereus undatus?Haw.?Britton et Rose,belonging to the family of Cactaceae,is a medicinal and edible plant flower widely distributed in tropical and subtropical regions.The flowerof H.undatus was first recorded as a traditional Chinese medicine in the book of"Lingnan Herb Collection"with the effects of relieving cough,detoxification and swelling.The flowers of H.undatus are often used as a adjuvant theraphy for cerebral arteriosclerosis,cardiovascular disease,tuberculosis,bronchitis,cervical lymphatic tuberculosis,mumps and other diseases.Phytochemical studies have shown that the flowers are rich in polysaccharides with many benefical bioactivities.However,there are few studies on the purification,structural characteristics,antioxidant and immunomodulatory activities of the polysaccharides.Therefore,the present work preliminarily studied the purification,structure identification,and in vitro antioxidant and immunomodulatory activity of the polysaccharides from H.undatus flowers.In addition,the effects of H₂O₂-Fe²⁺degradation on the polysaccharides from H.undatus flowers were investigated.The results will provide theoretical basis for further research and development of the polysaccharides from H.undatus.The main findings are listed as follows:?1?The crude polysaccharide,designated as FHRP,was prepared by hot water extraction and ethanol precipitation.Chemical analysis indicated that the contents of total sugar and galacturonic acid were 71.08±1.71%and 5.12±0.12%,respectively.In addition,FHRP contained a little amount of protein,sulfate and glucuronic acid.Monosaccharide analysis showed that FHRP was composed of fucose,arabinose,galactose and glucose with a molar ratio of 32.86:60.46:6.09:0.58.High performance gel permeation chromatography?HPGPC?analysis showed that FHRP had four different molecular-weight fractions.Infrared spectroscopy?FT-IR?showed that FHRP was a pyran acid polysaccharide with?-and?-configurations.Scanning electron microscopy?SEM?analysis revealed that FHRP presented irregular fragmentation.In vitro antioxidant and immunomodulatory assays showed that FHRP had good ABTS radical scavenging activity and oxygen radical absorption activity?ORAC?,and also could significantly promote the secretion of NO and IL-6 of macrophages RAW264.7 cells.?2?FHRP was fractionated by DEAE-Sepharose fast flow column.Three main polysaccharide fractions,designated as FHRP-1,FHRP-2 and FHRP-3,were obtained.The total sugarcontents were 84.06±1.01%,80.63±1.97%,and 80.12±1.25%,respectively.HPGPC analysis indicated that FHRP-1 had two components with the average molecular weights of 5535.99 and 116.89 kDa,respectively.FHRP-2 and FHRP-3 showed only one peak.Their average molecular weights were 55.03 and 134.26 kDa,respectively.Monosaccharide composition analysis indicated that FHRP-1,FHRP-2 and FHRP-3 were composed of arabinose,galactose,glucose,and fucose with the molar ratios of37.55:54.78:1.31:6.36,18.11:70.64:3.72:7.53,and 36.90:33.16:0.24:29.70,respectively.FT-IR analysis showed that the three polysaccharides exhibited characteristic peaks of uronic acid and pyranose ring.All of them contained?-configurational glycosidic bonds,but FHRP-1 also had?-configurational glycosidic bonds.The Congo red experiment showed that FHRP-1 and FHRP-2 had no triple-helix structurein aqueous solution.All three polysaccharides had triple helix structure in weakly alkaline solution.The results of periodate oxidation-Smith degradation and methylation analyses showed that the main glycosidic bonds of FHRP-1 were?6)-Galp-?1?,?6?-Glcp-(1?,Galp-?1?,?3,4?-fucp-?1?,?4?-Arap-?1?,and?4?-Glcp-?1?.FHRP-2 contained?6?-Galp-?1?,?4?-Arap-?1?and?4?-Glcp-?1?.FHRP-3 was mainly composed of?4?-Arap-?1?,?3?-Glcp-?1?,?6?-Glcp-?1?,?3,4?-fucp-(1?,Galp-?1?,?6?-Galp-?1?and?4?-Glcp-(1?.In vitro antioxidant experiments showed that FHRP-1,FHRP-2 and FHRP-3 could reduce the content of MDA and improve the activities of SOD and CAT in the cell supernatant.In vitro immunoregulation experiments showed that FHRP-1,FHRP-2 and FHRP-3 could significantly improve the proliferation and pinocytosis of mouse macrophages RAW264.7cells,and promote the secretion of NO,IL-6 and TNF-?.FHRP-2 and FHRP-3 exhibited better immunomodulatory than FHRP-1.?3?Two degraded polysaccharides,designated as DFHRP-1 and DFHRR-2,were obtained from FHRP by using the Fe²⁺-H₂O₂ degradation method.Chemical composition analysis indicated that H₂O₂-Fe²⁺degradation had no significant effect on the chemical composition and monosaccharide composition of the polysaccharides.HPGPC analysis showed that the molecular weights of DFHRP-1 and DFHRP-2 were significantly decreased compared with FHRP.FTIR analysis showed that they have similar functional groups and structures.SEM results showed that DFHRP-1 and DFHRP-2 had irregular pores on their surfaces,and the surface was smoother than that of FHRP.The original surface morphology of polysaccharides was destroyed by degradation.The functional assaysindicated that DFHRP-1 and DFHRR-2 showed better oil-holding,but weaker water-holding and emulsifying capacities than FHRP.In vitro antioxidant experiments showed that DFHRP-1showed the strongest ABTS radical scavenging activity at the same concentration point,followed by DFHRP-2 and FHRP.The ORAC values of DFHRP-1 and DFHRP-2 were higher than that of FHRP.In addition,in vitro immunoregulation experiment showed that DFHRP-1and DFHRP-2 could significantly improve the secretion contents of NO and IL-6 in RAW264.7 cells.These results indicated that the functional and biological properties of the degraded polysaccharides from H.undatus flowers were greatly determined by its molecular weight.DFHRP-1 and DFHRP-2 might have potential applications as multi-functionaladditives in foods,pharmaceuticals and cosmetics.