Study On The Active Ingredients From Nerium Indicum Mill Leaf Of Medicinal Plant

In this paper, nutritional components of Nerium indicum Mill leaf (NL) were analyzed. The optimize extraction technique of Nerium indicum Mill leaf Polysaccharides (NLPS) and Nerium indicum Mill leaf glycoprotein (NLGP) were obtained. Pure NLPS and NLGP were obtained by purify techniques from crude NLPS and NLGP. The structure and monosaccharide composition of NLPS were studied. The isoelectric point and molecular weight of NLGP were also studied in this paper.The nutritional components of NL were analyzed. The result showed that water could up to about 65.11% in fresh leaf.The percentage of diet fiber, carbohydrates, proteins, ash, polysaccharides, lipids, reducing sugar and others are 36.05%,9.65%,7.26%,6.32%,6.69% 3.25%, 2.69% and 27.82%,respectively.Response surface methodology (RSM) was used to optimize the extraction technique of NLPS. On the basis of single factor experiments, using statistical techniques such as a three factor, three level Box–Behnken factorial design (BBD) and RSM, with a view to maximize the extraction yield under optimal extraction parameters combination. The optimal extraction conditions were as follows: extraction temperature of 88.2℃,extraction time of 3.0h,ratio of water to raw material of 45,under such conditions the yield of polysaccharides could be up to 6.73% at twice extraction.NLPS was separated and purified by deposition,protein and pigment moving, column chromatography and analyzed by ultraviolet spectrum, thin layer chromatography(TLC) and infrared spectrum. It showed that 4 times ethanol,8h precipitation,protein and pigment moved by TCA and macroporous resin AB-8 were the optimize conditions.One single peak was obtained after DEAE-cellulose 52 and Sephadex G-200 column chromatography, which showed that NLPS was homogeneous.Ultraviolet spectrum showed that NLPS contained little DNA and protein,infrared spectrum showed that NLPS was main pyranopolysaccharides and hadβ-glycosidic bouds in its molecule structure. Studies by TLC showed that NLPS was mainly made up of Glucose, probably made up of Arabinose, Fucose and Fructose.Based on the Plackett-Burman experiment and the path of steepest ascent experiment, BBD and RSM were used to optimize the extraction technique of NLGP. The optimal extraction conditions were as follows: ratio of water to raw material of 18.5;salt concentration 0.03;pH 8.4, under such conditions the yield of NLGP could be up to 3.424mg/g.NLGP was deposited by ammonium sulfate and dialyzed against water and PBS buffer before column chromatography.One single peak was obtained after DEAE-cellulose 52 and Sephadex G-200 column chromatography,which showed that NLGP was homogeneous. Isoelectric experiment showed that isoelectric point of NLGP was pH 5.9. Three bonds were obtained after SDS-PAGE and the molecular weight of NLGP was about 53kDa, 29kDa and 26kDa.