| Glycyrrhiza uralensis Fisch.,belongs to leguminous,its roots and rhizomes can be used as medicine,and it has many pharmacological activities.Polysaccharide compounds are one of the active ingredients of Glycyrrhiza urlensis Fisch.Glycyrrhiza polysaccharides（GP）is proved to be acidic pyran polysaccharides.GP possesses lots of health benefits,such as anti-cancer,anti-inflammatory,anti-ulcer,anti-oxidant and immunomodulation effects.To date,the researches of GP mainly focus on total polysaccharides（TGP）and water-soluble polysaccharides（WGP）in G.uralensis,however,alcohol-soluble polysaccharides（EGP）has not been fully studied in publications.In the present study,the extraction processes,structural characteristics and anti-tumor activities of different types of GP have been studied.The research contents are as follows:1.In this study,response surface methodology（RSM）was used to optimize the enzymatic extraction and ultrasonic-assisted enzymatic extraction of GP from G.uralensis.The optimum enzymatic extraction conditions of GP were as follows:solid-liquid ratio of 1:10,pH=5,extraction temperature of 49℃,cellulase concentration of 9.4%（g/g）,and extraction time of 61 min.Under the conditions,the actual yield of GP was 23.85±0.09%（g/g）.The optimum conditions of ultrasonic-assisted enzymatic extraction were as follows:solid-liquid ratio of 1:10,pH=5,extraction temperature of 50℃,cellulase concentration of 8.6%（g/g）,ultrasonic power of 300 W,extraction time of 36 min,and the yield of GP was 25.51±0.10%（g/g）.These results suggested that ultrasonic-assisted enzymatic extraction showed higher GP yield than that of enzymatic extraction,and the cellulase dosage and extraction time were reduced by the extraction method.2.Macroporous resins were used to remove the pigments and proteins in TGP,WGP and EGP.Static adsorption tests showed that X-5,HPD-100 and D101 macroporous resins had better decolorization and deproteinization effects in various polysaccharides.In this study,the decolorization and deproteinization conditions of macroporous resins were further optimized using dynamic adsorption experiments.The optimal conditions of decolorization and deproteinization from TGP using X-5 macroporous resins were as follows:the sample concentration was 1.12 mg/mL and the loading flow rate was 1 BV（bed volume）/h.Under the conditions,the decolorization rate,deproteinization rate and TGP yield were 74.26%,47.24%and 78.56%,respectively.The optimal conditions for decolorization and deproteinization of WGP using HPD-100 macroporous resin were as follows:the sample concentration was 1.12 mg/mL,loading flow rate was 1 BV/h,decolorization rate was 68.86%,deproteinization rate was 58.61%and WGP yield was 85.41%.The optimal concentration for decolorization and deproteinization of EGP using D101 macroporous resin were as follows:sample concentration was 2 mg/mL,and the loading flow rate was 2 BV/h,the decolorization rate was 83.44%,the deproteinization rate was 76.12%,and EGP yield was 92.15%.In conclusion,macroporous resin can efficiently remove pigments and protein impurities from various polysaccharides.3.In this section,the structures of TGP,WGP and EGP were analyzed,including molecular weight determination,Fourier transform infrared spectroscopy（FT-IR）analysis,monosaccharide composition analysis and Congo Red test.According to Congo Red test,all three GP have triple helix structures.The molecular weights of TGP,WGP and EGP were determined using high performance gel permeation chromatography.The molecular weight of TGP,WGP and EGP were 1.11×1056.87×105 Da,2.13×105-4.53×105 Da and 2.46×104-9.67×104 Da,respectively.FT-IR results showed that TGP and WGP were polysaccharides with β-glycosidic pyran ring structures,and EGP was polysaccharide with α-glycosidic bond and β-glycosidic pyran ring.The monosaccharide compositions of various polysaccharides were determined using high performance liquid chromatography（HPLC）.The results showed that TGP was composed of mannose,glucose and galactose（molar ratio 1.00:70.19:2.36）,WGP was composed of mannose,rhamnose,galacturonic acid,glucose,galactose and arabinose（molar ratio 2.51:1.00:1.91:15.57:14.77:5.49）,and EGP was composed of mannose and glucose（molar ratio 1.00:134.73）.These results showed that the structures of different GP are quite different.4.The scavenging abilities of the three GP on DPPH radical,ABTS radical,superoxide anion radical,hydroxyl radical and Fe reducing power were investigated.The results showed that different types of GP have strong free radical scavenging ability.EGP showed better scavenging effects on DPPH radical and ABTS radical and higher Fe reducing power than other GPs.WGP had the best scavenging ability on hydroxyl radical,and TGP had the best scavenging ability on superoxide anion radical.Our results showed that the three GPs could inhibit HepG-2 cells in a time-dependent and dose-dependent manners.The results of cell scratch test showed that EGP has better effect on inhibiting the migration of HepG-2 cells than TGP and WGP.Therefore,different types of GPs showed antioxidant and anti-liver cancer activities. |
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