Purification And Characterization Of The Glycoprotein Isolated By Alkaline Extraction From Common Yam Rhizome

Yam glycoprotein is traditionally considered as a fighter to diabetes mellitus, high blood-fat,chronic gastritis, chronic enteritis, and has a high antioxidant activity and efficient immunizing action. At present, the development of supplement food from yam has made some encouraging achievements.However, the yam glycoprotein has a large development space. In order to adapt to the needs of markets, the paper seeks a new extraction and separation method to obtain higher rate of yam glycoprotein. The alkaline processing method is used for glycoprotein isolation. The physical and chemical characteristics of the glycoprotein extracted are identified. The main investigative conclusions of the paper are as follows:The pH value of the solution is adjusted with Na2CO3to adopt the alkaline processing method for glycoprotein isolationn.In the single factor experiments, the six factors of extraction time, extraction temperature, pH value, the ratio of material to water, powder size, and extraction times are evaluated the influence on extraction rate of glycoprotein. During the experiment of single factors, the reseach chooses the suitable four factors (extraction temperature, extracting time, extraction pH value, material-water ratio)and three levels for orthogonal test, considering to low extractionbn cycle, low energy consumption. After the analysis of range and variance analysis, the test results are as follows:The best extraction condition for the yam glycoprotein is extracting temperature60℃, extracting time2.0h, material-water ratio1:15, pH=10. Extraction factors on the influence of the extraction rate is as follows:extracting temperature> pH value> extracting time material-water ratio. The verification test shows that, the average extraction rate of yam glycoprotein is0.74%and is highest one under the best extraction condition.DEAE-52cellulose and SephadexG-75column chromatography are applied to purify the crude glycoprotein extracted. After the gradient elution, GLP-P1is obtained from the fraction of water, and GLP-P2from the fraction of0.05M NaCl eluant. The test mainly determines the properties of GLP-P2, owning to the limit of the yield of the glycoprotein.By means of infrared spectrum of the two kinds of glycoprotein, the absorption peaks react the structures of glycoprotein; Through the Sephadex-75column chromatography and SDS-PAGE, the result shows that GLP-P2is the single component, and the molecular weight is approximately30kDa; Determine the carbohydrate moiety through phenol-sulfuric analysis and protein moiety by the method of coomassie brilliant blue, respectively. The rate of protein moiety:to carbohydrate moiety is approximately20. GLP-P2is a kind of compound protein with short oligosaccharide chain; The monosaccharide compositions of the GLP-P2is distinguished by thin-layer chromatography, the result shows that the oligosaccharide chain contains D-glucose, L-galactose and mannose; By the automatic amino acid analyzer, the result shows that the GLP-P2consists of17kinds of amino acids and contains a high percentage of glutamic and aspartic acid. The glycoprotein extracted contains affluent amino acids including both the essential amino acid and non-essential amino acid. It indicates that the glycoprotein extract has a high nutritive value to human health; The test indicates short oligosaccharide chain of the GLP-P2connects to threonine of polypeptide chain by the O-cardohydrata-peptide linkage.