Study On The Content Determination Method Of Green Tea Water-soluble Polysaccharide

It was well known that the tea polysaccharide had the strong biological activity, but its content determination method still had the deficiency. The present paper had studied the content determination method of water-soluble green tea polysaccharide. First puried-TPS was get from corse green tea and puried by DEAE-52 and Sephacryl S-400 to get 0.25-Ⅱ-DEAE and 0.25-Ⅱ-DEAE-400. The high-purity component 0.25-Ⅱ-DEAE was choosed and purified by the Sephacryl S -400 column chromatographic. Through examination its purity, it was obvious that the high-purity component 0.25-Ⅱ-DEAE was homogeneous component. The designation purified TPS, 0.25-Ⅱ-DEAE and 0.25-Ⅱ-DEAE-400 were choosed for standard to analysis tea polysaccharide monosaccharide composition and the biggest absorption wavelength of anthrone color method. PMP derived and liquid chromatogram analysis indicated that the mole content of Galactose and Arabinose was highest in monosaccharide composition of the purifies TPS, 0.25-Ⅱ- DEAE and 0.25-Ⅱ-DEAE-400. The 460nm～700nm scanning of anthrone color method indicated that the absorption wavelength of tea polysaccharide was 670nm. Therefore a 1/1 mole ratio of the Galactose/ Arabinose as the comparison, and 670nm of determination wavelength were designated.On the base of single factor experiment, the best tea polysaccharide content determination optimum condition of anthrone color method by orthogonal experiment was obtained, which including 2.0g/L of developer concentration, 5min of boils water bath, 40min of ice water cools. By the determination condition of Galactose:Arabinose =1: 1 (a mole of ratio) for the comparison, the developer amount 2.0g/L, boils water bathing 5min and examination wavelength 670nm, the tea polysaccharide specification curve regression equation was Y=0.2043X+0.0028, R2=0.9924 and its linear-range was 20ug/mL~200ug/mL (Y was the concentration, unit mg/mL; X was the extinction value).Including tea Polyphenol, pigment, protein, amino acid, more disturbance factor in determination was analyzed. And it was discovered that tea Polyphenol and pigment obviously disturbed the mensuration of tea polysaccharides, otherwise the interferences of protein, caffeine and amino acids were less comparatively. In view of eliminations of Polyphenol, protein and amino acid, the eliminated mixed effects of acetone immersion, the ethyl alcohol immersion and the ethyl acetate soaked were compared. It was discovered that acetone has more eliminates effect.In view of the Polyphenol residual question, the Polyphenol eliminated mixed effects by ethyl alcohol precipitation method or PVPP processing method were compared. It was discovered that not only the polysaccharide and the Polyphenol were separated by ethyl alcohol precipitation method, but also it might realize the dissociation the separation of monosaccharide and polysaccharide.And it also simplified the processing step. Although PVPP processing method only could eliminated Polyphenol, but the Polyphenol residual quantity was still high after PVPP processing. Thus the superior sample preparation method of"dried tea→smashing→acetone immersion→boil water extraction→filtration→ethyl alcohol process→centrifugal→constant volume"was brought forward.Through synthesized the determination condition, the standard working curve and the sample superior preparation method, the content determination method of water-soluble tea polysaccharide was established. This method was stable in 4h, and its average returns-ratio, RSD and picked out limits was 96.20%, 1.76% (n=6), 0.0033mg/mL. The tea polysaccharide content determinations of 18 tea samples were analyzed by this method.The results obvioued that the famous tea polysaccharide content of the Xihu Longjing tea and the Yandang Maofeng had reached as high as 4.5%, and the change rule of the Longjing tea series polysaccharide content was not obviously. For the top grade series of Huangshan Maofeng, while the series elevated the polysaccharide content reduced. However for the one to third-level series, while the series elevated, the polysaccharide content also elevated. The polysaccharide content of Longjing 43 tea fresh leaves reached 1.35%. This method had the good accuracy, reproduction quality and stability. And it could be fast and accurate to application for water-soluble tea polysaccharide content determination of tea.