Study On Extraction, Isolation, Purification And Physicochemical Property Of Opuntia Dillenii Haw. Polysaccharides

To explore an extraction method which is suitable to the industrialization and to provide a reference for the further study of the structure-activity relationships of Opuntia dillenii Haw. poly sac charides (ODPs), different extraction methods and physicochemical property of the ODPs were studied.Methods:Method for determination of polysaccharide content, phenol-sulfuric acid method, was optimized. The pectinase and cellulase were used to increase the productivity of ODPs; The extraction conditions of ODPs by sulfuric acid or hot water were optimized by the single-factor test and the orthogonal test; The antioxidant activities of ODPs extracted by sulfuric acid, hot water or enzyme were studied by the modified orthophenanthroline-Fe2+-H2O2 system; The components of ODPs were isolated by Gel chromatography and the contents of ODPs were determined by the optimized phenol-sulfuric acid method; The monosaccharide compositions of ODPs were analysed by HPLC-ELSD and sulfuric acid-carbazole method; The contents of protein and nitrogen in ODPs were determined by Coomassie brilliant blue method and micro Kjeldahl method; The optical activities of ODPs were determined by spectropolarimeter.The main results are as follows:(1) The optimal conditions for the determination of ODPs with phenol-sulfuric acid were as follows:2 mL of ODPs samples,0.5 mL of 4% phenol and 5 mL of sulphuric acid, shaken up and placed stationarily for 5 min. The samples were measured by spectrophotometer at the wavelength of 482 nm. The calibration curve was linear in the concentration range of 7.5 37.5 g/mL. The optimal conditions of the extraction method by two kinds of enzymes were as follows:pectinase 0.7%, cellulase 0.3%, enzymolysis temperature 40℃, enzymolysis time 1.5 h and pH 3.5. Under these conditions, the yield of polysaccharides was 20.08%, with purity of 83.0% and protein content of 2.2%.(2) The optimal conditions of the extraction method by sulfuric acid were as follows:the concentration of sulfuric acid 0.10 mol/L, ratio of liquid to material 20:1, extraction temperature 70℃and time 2 h for 2 times. Under these conditions, the yield of polysaccha-rides was 39.92%, with purity of 87.11% and protein content of 1.06%. The optimal conditions of the extraction method by hot water were as follows:ratio of liquid to material 60:1, extraction temperature 95℃and time 3 h for 2 times. Under these conditions, the yield of polysaccharides was 14.67%, with purity of 78.11% and protein content of 0.36%. The antioxidant activities of ODPs (7.5 g/L) extracted by hot water and enzyme were significantly higher (P<0.01) than those of the ODPs extracted by sulfuric acid, but no significant difference was found between the antioxidant activities of the ODPs extracted by hot water and enzyme, and the 2 g/L Vc (P> 0.05).(3) The average polysaccharide contents of ODPs, ODP-I, ODP-II, ODP-Ia, ODP-Ib and ODP-II'were 78.69%,92.44%,88.78%,94.60%,87.43%, and 90.36%, respectively. ODP-Ia was composed mainly of rhamnose, arabinose, galactose and glucose, with 15.13%(w/w) of uronic acid; ODP-Ib was composed mainly of rhamnose, arabinose, and galactose, with 52.66%(w/w) of uronic acid; ODP-IF was composed mainly of rhamnose, glucose, with 26.38%(w/w) of uronic acid. The main components of ODPs did not contain protein nearly and their optical activities were dextrorotation. The ODP-I and ODP-Ia were nitrogenous compound.The phenol-sulfuric acid method optimized was simple, delicacy, accurate and repeatable. The extraction method by two kinds of enzymes was the best craft, its yield, purity and the antioxidant activities of polysaccharides were high, and the conditions were moderate. It should be studied further. The purity of the main components of ODPs isolated by Gel chromatography was high, they were composed of 5 kinds of monosaccharides at least and their contents of uronic acid were high. Their structure should be studied further on the basis of this thesis. This study can provide reference for the structure-activity relationships and the industrialization of ODPs.