The Study On Extraction, Purification And Antioxidative Activity Of Polysaccharides From Allium Macrosttemon Bge.

Allium macrosttemon Bge.is a traditional Chinese native medicine,it is dry bulb of lily branch plant Allium macrosttemon Bge.or Allium chinensis G. Don. It has the characteristic of spicy, bitter, warm and the effect of warm center passes pasitive,regulates the flow of vital energy the wide chest,passes positive relieves congestion. Its original plant not only is the medicine,but also can be edible. It is one of 88 kinds of Chinese herbal medicines having dual purpose of medicine and food, which was announced by Ministry of Health in 2002. Modern research proves, Allium macrosttemon Bge. has the effect of anti-oxidation, preventing and controlling atherosclerosis,and so on. Allium macrosttemon Bge.is distributed extensively in our country, and its resource is abundant. It has great potentiality to be exploited. Chemical compositions such as the volatile oil, soap glucoside, compound containing nitrogen, the prostaglandin and so on have been separated from Allium macrosttemon Bge, but there are few reports on polysaccharide from Allium macrosttemon Bge. In this paper, in order to probe undiscovered polysaccharides and their active functions, and lay the foundation for further development and utilization polysaccharide of Allium macrosttemon Bge. is studied as our object.In this thesis, we study mainly the extraction, separation, purification, components, properties of polysaccharides from Allium macrosttemon Bge., moreover, we also study their antioxidative activities by H2O2/Fe2+ system and pyrogallol system.Through single factor test and orthogonal experiment design method[ L9(3)4 ] we analyze the influence of factors such as temperature(A), solid-liquid ratio(B) and time(C) on the extraction percent of the polysaccharides. Results indicated that the influence of the three factors on extraction percent of the polysaccharides is A>C>B and temperature is the main factor. The optimum extraction technology was A3B2C3 (temperature is 90℃,solid-liquid ratio is 1:12 and the extraction time is 90 min). Under these conditions, the extraction percent of soluble polysaccharides from Alluim macrosttemon Bge. is 13.93%. In addition, the method of removing protein was studied, and results indicated that Sevag method combined with the enzyme was best. Polysaccharide after removing protein is dialysed, we obtain part-pure product of polysaccharide(PAM).PAM is separated into three fractions(PAM-I,PAM-II and PAM-III) by DEAE-cellulose chromatography. Then they are purified by sephadex G-150 gel chromatography into PAM-Ib, PAM-IIa and PAM-III',which are tested by cellulose acetate pellicle electrophoresis and gel filtration on sephadex G-150. The result of electrophoresis assumes a single spot. There is a single symmetric peak respectively by gel filtration. The identification of purity implies that they are homogeneous components.Molecular weight of the polysaccharide is determined by gel filtration. The result indicates their average molecular weights are respectively estimated to be 12.18kD, 51.58kD and 291.610kD,molecular weight of PAM-III'is the maximum and the molecular weight of PAM-Ib is the minimum.PAM-Ib, PAM-IIa and PAM-III'are hydrolyzed by 12 mol·L-1 hydrochloric acid, then their component are measured by means of thin-layer chromatography(TLC). The result shows PAM-Ib is composed of Galactose and Glucose. PAM-IIa is consisted of Galactose ,Glucose , Fructose, Xylose and Rhamnose. The main compositions of PAM-III'are Galactose, Glucose and Xylose.The experiment mainly studies antioxidative activities of four kinds of polysaccharides (PAM,PAM-Ib, PAM-IIa and PAM-III')from Allium macrosttemon Bge., and analyses their effects of clearing oxygen radical. Hydroxid radical(·OH)and superoxide anion radical (O2—·) are produced respectively by H202/Fe2+ system and pyrogallol system. The result shows that·OH and O2—·can be inhibited by PAM and PAM-Ib, the scavenging effect of PAM is stronger than PAM-Ib's, their activities and amount are relative. It is proved by the test that PAM-III'is more effective in scavenging·OH, but it can't inhibit O2—·.PAM-IIa shows no evident effect in scavenging O2—·or·OH.