Extraction And Purification,Chemical Structure Characterization And Antioxidant Activity In Vitro Of Polysaccharides Extracted From Camellia Oleifera Seed Cake

Camellia oleifera seed cake contains a lot of tea saponin, protein, starch and saccharides. However, most of the researches about Camellia oleifera seed cake are focus on tea saponiam. As an important tea extract and active polysaccharides, tea seeds polysaccharides will certainly become research focus. In this paper, the compositions and contents of nutrients in Camellia oleifera seed cake were estimated, besides, the extraction, purification, structure characterization and external antioxidant activity of polysaccharide were studied. Main results are shown as follows:1. The compositions of the raw material:the water content59.09%, the ash content0.74%, the crude fat6.80%, the crude protein7.61%, the crude fibre2.90%, the saccharides9.84%.2. Ultrasonic-assisted technology was applied in the experiment to extract polysaccharides from Camellia oleifera seed cake. According to the single-factor experiments and response surface methodology, the optimum extraction conditions were obtained as follows:ratio of material to solvent1:15, ultrasonic power150W, extraction temperature80℃, extraction time45min. Under these conditions, the yield of polysaccharides was gotten up to14.22±0.51%.3. By comparing the three methods of protein removal, including trichloroacetic acid, Sevage method and papain-Sevage method, papain-Sevage method was choosed as the optimal method. According to the single-factor experiments and response surface methodology, the optimum conditions for protein removal were obtained as follows:78U/mL of papain concentration, temperature65℃, pH7.0, hydrolysis time92min, and then repeat3times with Sevags method. The amount of protein removed was80.52%, and the polysaccharide remained84.29%.4. DEAE-Sephadex A-25gel filitration was applied to purify the crude tea seeds polysaccharides, and three main components TSP1, TSP2and TSP3were obtained. Both TSP1and TSP2are neutral polysaccharide, while TSP3is acid polysaccharide. The results of Sephacryl S-200gel filitration and High Performance Liquid Chromatograph (HPLC) indecated both TSP1and TSP2were heteroglycan polysaccharides, while TSP3was a homogeneous polysaccharide. The PMP-HPLC results showed TSP1consisted of Glu, Man, Rha and their molar ratio was6.29:1.00:0.15; TSP2consisted of Glu, Man, Gal and their molar ratio was3.13:1.00:0.03; TSP3consisted of Glu, Man, Gal, Ara, GalA, Rha and their molar ratio was0.42:1.00:1.09:0.96:0.43:0.19.The Infrad Spcetrum showed that TSP1, TSP2and TSP3had the characteristic absorption peaks of polysaccharide, and the main structure was α-pyranose. The1H NMR of TSP3indicated that the monosaccharide linkages of Araf, Rhap and GalpA were α-forms, while Galp was β-forms.13C NMR showed TSP3contained six kinds of monosaccharides and uronic acid was included, the results were consistent with that of HPLC.5.The antioxidant activity experiments in vitro indicated that both the scavenging activity of·OH and DPPH· showed concentration-dependant. At the concentration of5mg/mL, the·OH and DPPH scavenging rate were84.6%and73.5%, respectively. Nevertheless, the scavening activity of superoxide anion radical was very low and almost no reducing power activity was observed.