Study On The Extraction, Purification And Antioxidant Activity In Vitro Of Polysaccharide From Lycoris Aurea

Lycoris aurea is a kind of perennial herb with high medicinal and ornamental value. Currently alkaloids of Lycoris aurea is a major aspect by the researcher, but there are few domestic and international research results about the polysaccharide structure and biological activities. In this thesis, The extraction, purification and antioxidant activity of polysaccharide from Lycoris aurea were studied.（1） The method of anthrone-sulfuric acid was used to measure the content of polysaccharide from Lycoris aurea. The maximum absorption wavelength（λ max）= 625 nm was determined by ultraviolet spectrophotometer（UV） at 800⁴00 nm. The linear regression equation was y = 8.878 x + 0.015, R2 = 0.9990, and the detection range was 0.01⁰.1 mg/m L. The methodological evaluation showed that precision, stability, repeatability and recovery were accurate and reliable.（2） Response surface methodology was used to optimize water and ultrasonicassisted extraction of polysaccharide. The optimal water extractions were that the liquid/solid ratio was 23:1（m L:g）, extracted for 7.7 h at 70℃. Under the optimized conditions, the yield of polysaccharide was 42.00%. The optimal ultrasonic-assisted extractions were that the liquid/solid ratio was 22:1（m L:g）, ultrasonic pretreatment for 15 min, then extracted for 6.9 h at 84℃. Under the optimized conditions, the yield of polysaccharide was 40.95%.（3） The purification of polysaccharide from Lycoris aurea was studied. The polysaccharide was decolorized by 732-positive ion exchange resin. The results showed that resin dosage added at 10%, p H of 4, then decolored 2 h at 50℃. The decolorization rate was 65.76% and the retention rate of polysaccharide reached 98.61%. When deproteination for seven times, the protein elimination rate could be up to 94.89% and the retention rate of polysaccharide was 78.57%. The preliminary purification of polysaccharide was applied by DEAE-52 cellulose column chromatography. Then the column was stepwise eluted with 0, 0.1, 0.3, 0.5 mol/L Na Cl solutions, and four fractions of polysaccharide（PBL1-1, PBL1-2, PBL1-3, PBL1-4） were obtained.（4） The structures of PBL1-1、PBL1-2 and PBL1-3 were analyzed and determined by GPC-MALS-RI, HPLC, IR and UV. The results showed that weight average molecular weight（Mw） of PBL1-1, PBL1-2 and PBL1-3 were 3.642×10³, 1.206×10⁴ and 3.992×10⁵ g/mol. PBL1-1 was composed of D-Mannose, D-Glucose and D-Galactose with the mole ratio of 1:0.34:0.036. PBL1-2 was composed of D-Mannose, Rhamnose, D-Galacturonic acid, D-Glucose, D-Galactose and Arabinose with the mole ratio of 1:0.29:0.78:21.89:2.69:0.63. PBL1-3 was composed of D-Mannose, Rhamnose, D-Glucuronic acid, D-Galacturonic acid, D-Glucose, D-Galactose and Arabinose with the mole ratio of 1:2.63:0.47:1.63:0.83:6.92:1.34. IR spectra showed that pyranoside of sugar residues configuration exists within PBL1-1, PBL1-2 and PBL1-3. UV scanning spectrum showed that PBL1-2 and PBL1-3 contains a small amount of proteins.（5） Through estimated the scavenging ability of DPPH·, ·OH and O₂^-· in vitro about PBL, PBL1, PBL1-1 and PBL1-2. The results showed that these four components had antioxidant activities. The strongest antioxidant capacity was PBL and the second was PBL1. PBL1-1 was the weakest to eliminate free-radicals.