Extraction, Purification, Structure Analysis And Biological Activity Of Polysaccharides From The Roots Of Saposhnikovia Devaricata

Polysaccharide,a kind of natural polymers in tissue of plant,animal and microbe,is one of basic materials to maintain normal operation of the lives,and has many physiological functions such as anti-oxidation,anti-decrepitude,immune regulation and so on while poses no poisonous effect to the body,so it has been applied to clinical service and sold in health product market.Saposhnikovia devaricata(Turcz.) Schischk,one of most critical wild Chinese medicinal materials,belongs to Umbelliferous,perennial and herbaceous plant.Its dry root with no florescence could be treated cold, acute conjunctivitis,high blood pressure,adiposity,apoplexy and hemorrhoid for the physiological functions of anti-febrile,balsamic,antalgic,anti-agnail,anti-tumour,immune regulation and so on.As for most of application in clinic,there is a lack of systemic and in-depth research on specific active ingredient and functionary mechanism,only according to recordation of ancient document.As one of main active ingredients of Saposhnikovia devaricata,polysaccharide is involved in its many physiological functions,so these research on Saposhnikovia devaricata polysaccharides will provide necessary theory for farther exploiture of Saposhnikovia devaricata.In the present paper,we studied the polysaccharides from the roots of Saposhnikovia devaricata,including process of extraction, isolation,purification,molecular modification,structure analysis and biological activity.The primary results were as follows:1.Polysaccharides from Saposhnikovia divaricata(Turez.) Schischk were obtained through traditional hot water extraction,ultrasonic-intensified extraction and microwave-assisted extraction to optimize the best extraction process in view of yield of polysaccharide by single factor test and orthogonal experiment.The results showed that the optimal conditions of traditional hot water extraction were temperature 75℃,extraction time three hours once,solid-liquid ratio 1:25 and extracting three times,the optimal conditions of ultrasonic-intensified extraction were ultrasonic power of 1000 W,extraction time of 30 min and solid-liquid ratio of 1:25,the optimal conditions of microwave-assisted extraction were microwave power of 560 W,extraction time of 10 min and solid-liquid ratio of 1:30.Compared to traditional hot water extraction,both of ultrasonic-intensified extraction and microwave-assisted extraction could increase the yield of polysaccharide,effect of microwave-assisted extraction was more significant,under the optimal conditions,its yield of polysaccharide increased from 4.913%to 7.639%,while extraction time shortened from 3h to 10min. Therefore,microwave-assisted extraction was one more quick and efficient method to extract Saposhnikovia devaricata polysaccharides with fewer time and lower cost.2.Two kinds of homogeneous Saposhnikovia devaricata polysaccharides SPS-â… and SPS-â…¡were attained from the crude polysaccharide through de-coloring by active carbon adsorption method, de-proteinizating by Sevag method,purifying by DEAE-52 cellulose coltmm chromatography and Sephadex G-200 gel filtration chromatography.In SPS-â… and SPS-â…¡,the contents of potysaccharides and uronic acids,determined by phenol-sulfuric acid method and sulfuric acid carbazole method, were 93.1790%,71.0493%and 6.4525%,29.1153%,respectively.The molecular weight of SPS-â… and SPS-â…¡were 32700 and 68100Da by the determination of high performance gel permeation chromatography(HPGPC).Gas chromatography(GC) analysis showed that SPS-â… was composed of rhamnose,arabinose,mannose,glucose,galactose in molar ratios of 0.80:9.34:1.42:3.98:7.40 and SPS-â…¡was also composed of rhamnose,arabinose,mannose,glucose,galactose in molar ratios of 2.34:5.49:1.22:2.40:9.04.After the methods of partial hydrolysis with acid,periodate oxidation and Smith degradation analysis,the results indicated that the main chain of SPS-â… was made up of galactose,arabinose and rhamnose,and the side chain of SPS-â… was composed of glucose,galactose, arabinose and mannose,while in SPS-â…¡,galactose,arabinose,glucose and rhamnose formed not only the main chain but also the side chain.In SPS-â… and SPS-â…¡,the main linking of were 1→2,1→6 and 1→3 linkage,the minority was 1→4 linkage,the glycosidic conformation was mostlyα-linked glycosidic bond and characteristic of pyranose.3.Sulfated Saphoshnikovia devaricata polysaccharide(S-SPS) was gamed from Saphoshnikovia devaricata polysaccharide(SPS) by the method of sulfur trioxide pyridine complex with the yield of 143.4513%,the conten of SO₄^2- was 19.3853%and the degree of substituent(DS) was 0.412.Gas chromatography(GC) analysis results showed that both of SPS and S-SPS were composed of rhamnose,arabinose,mannose,glucose and galactose,only proportion of each monosaccharide has a little difference.4.The results of antioxidative experiments revealed that Saposhnikovia devaricata polysaccharides had some functions of scavenging free radicals and inhibiting lipid peroxidation and scavenging effects on DPPH.and·OH were especially significant.S-SPS and SPS-â…¡have better effect among the four polysaccharides of SPS,SPS-â… ,SPS-â…¡,S-SPS and the scavenging rate of·OH could exceed 90%when the experimental concentration was 8mg/mL.The results of antibacterial experiments showed that Saposhnikovia devaricata polysaccharides inhibited selectively growth of some microbes and had better inhibitory effect on Bacillus subtilis,while had no visible inhibitory effects on Escherichia coli,Penicillum chrysogenum and Mucor mucedo.Among different Saposhnikovia devaricata polysaccharides,S-SPS had best antibacterial effect and its minimal inhibitory concentration(MIC) to Bacillus subtilis was 1mg/mL,greatly lower than that of SPS(4mg/mL).In conclusion,the sulfation of SPS was a considerable molecular modification method for improving soluble property in water and enhancing antioxidative activities and antibacterial ability, providing some reference for farther exploiture and using of the resource of Saposhnikovia devaricata.