Isolation, Purification And Antioxidative Activity Of Polysaccharide From Misgurnus Anguillicavdatu

Loach is a kind of fish which belongs to the family of Misgurnus anguillicaudatus. It is recorded in A Dictionary of Chinese Traditional Medicine that its meat or whole body can be used as-Chinese traditional medicine. The main pharmaceutical functions of the loach seen in folk recipes are to nourish spleen and enrich vital energy, to protect liver and bring down jaundice (or icterus index), to relieve internal heat and detoxify, to eliminate carbuncles and loosen scabs. Recently, live loach or its dry powder has been used in folk thrapy to treat infectious hepatitis, and found very effective. But no investigations on the bioactivities of the loach and its effective substances by modern pharmacological methods have been seen reported in literature.This thesis is designed to study the isolation, purification and pharmacological activities of polysaccharides from the skin mucus, scarfskin and all body of the loach. In this thesis, we study mainly the extraction, purification of polysaccharides from the skin mucus, scarfskin and all body of the loach. Moreover, we also study their antioxidative activities. The main results in this work are as follows:In this paper, we adopt traditional polysaccharide extraction method, that is being extracted by alkaline solution, and precipitated by ethanol, then washed several times with 100% ethanol and acetone, at last, dried in vacuum. The crude polysaccharide of Loach obtained is light yellow. In the course of drawing, the effects of several main factors on extraction rate, e. g: volume of water, temperature, time and times of extraction have been investigated. Through orthogonal design, several main affecting factors and levels are discussed, and then the super condition of extraction is determined. The removal of protein in the product is compared by three different methods, Sevag method, TCA method and enzyme combined with Sevag method. The third method has its own obvious advantage. It may increase the removal rate of protein. At the same time, the loss rate of polysaccharides is reduced greatly. Enzyme can cut off the protein chain, then use classical chemical method (Sevag method) to deal with three times. The method is an ideal method for eliminating protein. Polysaccharide after removing protein is dialysed, we obtain pure product of polysaccharide-MAP.Misgurnus anguillicaudatus polysaccharides, a novel type of free neutr polysaccharide never reported before, was extracted from the skin mucus, scarfskin and all body of the loach, Purified MAP were obtained by gel filtration gel permeation chromatography (GPC). Through the procedure of total acidic hydrolysis, PC and GC analysis, MAP was shown to be a neutral heteropolysaccharide. According to IR data, periodate oxidation and smith degradation, the maintype of bond existing in the molecules.The experiment studies antioxidative activities of polysaccharides from the skin mucus, scarfskin and all body of the Loach, and analyses theireffects of clearing oxygen radical. Hydroxid radical(DPPH·) and superoxide anion radical(O2-·)are produced respectively by H2O2/Fe2 system and AP-TEMED system. The result shows that DPPH·and O2-·can be inhibited by the polysaccharides from the skin mucus, scarfskin and all body of the Loach, the scavenging effect of DPPH·by polysaccharides from the skin mucus, scarfskin and all body of the Loach is 65.1%,81.2%,91.2% at the concentration of 0.6ug/mL, the scavenging effect of O2-·by polysaccharides from the skin mucus, scarfskin and all body of the Loach is 72.1%,85.7%,86.3% at the concentration of 1.2ug/mL.The antioxidative activities of polysaccharides from the skin mucus, scarfskin and all body of the Loach should offer scientific basis for their application.