Study On The Preparation Of Salvianolic Acid B And Alkaloids From Nelumbo Nucifera And The Interaction Between Them And Bovine Serum Albumin

Several natural active substances were separated and purified by high-speedcountercurrent chromatography. Then the interaction between small molecules andbiomacromolecules was investigated using fluorescence spectroscopy, synchronousfluorescence spectroscopy, circular dichroism and ultraviolet spectroscopy at differentconditions. This article is divided into four chapters, each abstract is as follows:1、 too much references at home and abroad were cited. Functions and natures ofproteins and methods of how to extract, analyse, separate and purify the natural activesubstances were introduced. The methods, contents and significances of theinteraction between small molecules and biomacromolecules were summarized briefly.Then the topic of this article was presented.2、 the methods and steps of how to extract, analyse, separate and purify thenatural active substances were elaborated in detail. A two-phase solvent system whichwas applied to preparative pH-Zone-refining CCC, composed of n-hexane–ethylacetate–methanol–water (5:5:2:8, v/v), added to the organic stationary phase10mMtriethylamine (TEA) and5mM HCl in aqueous mobile phase, was the optimum forthree alkaloids from Nelumbo nucifera Gaertn. A two-phase solvent system which wasapplied to preparative pH-Zone-refining CCC, composed of MtBE–water (1:1, v/v),added to the organic stationary phase10mM trifluoroacetic acid and10mM ammoniawater in aqueous mobile phase,was the optimum for salvianolic acid B. Four highpurity monomer substance were isolated, which were liensinine, isoliensinine,neferine and salvianolic acid B, respectively. And identified their structures. Thisexperiment provide the material basis for future experiments.3、 the interaction between the three alkaloids from Nelumbo nucifera Gaertnand bovine serum albumin (BSA) was investigated using fluorescence spectroscopy, synchronous fluorescence spectroscopy and ultraviolet spectroscopy at differentconditions. The binding constant (K), the numbers of binding sites (n), the bindingdistance (r), and the thermodynamic parameters enthalpy change H0, the entropychange S0, Gibbs free energy change G0, were calculated. The conformation ofBSA changed in the presence of alkaloids on synchronous fluorescence spectroscopy.The results show that alkaloids-BSA system is a spontaneous reaction, and the mainforce respectively is Van’t Hoff force, hydrogen bond and electrostatic force.According to the comprehensive characterization parameter Y and the chemicalstructures of the three alkaloids, the power of the combination of alkaloids isliensinine neferine isoliensinine.4、Under simulated physiological conditions, the interaction between salvianolicacid B and BSA was investigated using fluorescence spectroscopy, synchronousfluorescence spectroscopy, circular dichroism and ultraviolet spectroscopy at differentconditions. As a result, the binding constant (K), the numbers of binding sites (n), thebinding distance (r), and the thermodynamic parameters enthalpy change H0, theentropy change S0, Gibbs free energy change G0, are calculated in a simulatedhuman body conditions. The conformation of BSA changed in the presence ofsalvianolic acid B on circular dichroism and synchronous fluorescence spectroscopy.The results show that the static quenching and non-radiative energy transfer lead tothe fluorescence quenching of bovine serum albumin, and salvianolic acid B-BSAsystem is a spontaneous reaction, the main electrostatic force is hydrophobic effect. Inaddition, the addition of external common metal ions Cu2+and K+decreased thebinding constant between salvianolic acid B and BSA.5、The effect of different drying methods (microwave-drying, hot-air drying andshade-drying) on the antioxidant activity and chemical ingredient of Honeysuckle wasresearched. Compared with ascorbic acid, antioxidant activity of Honeysuckle driedby three different methods was evaluated and calculated by Luminolchemiluminescence based on superoxide anion scavenging, hydrogen peroxidescavenging and hydroxyl radical scavenging in vitro models, the content of total phenolic and total flavonoid were also determined. The results show that Honeysucklehad remarkable antioxidant activity. There were significant effects on both theantioxidant activity and the content of the active components among the threedifferent drying methods. Microwave-drying at70C,10min, showed the best results,the content of total phenolic and total flavonoid were30.402.75mg/g,61.247.62mg/g respectively, the IC50of the three index were9.44%,0.029%,0.12%respectively. Shade-drying at natural conditions,4days, showed the worst results, thecontent of total phenolic and total flavonoid were10.220.81mg/g,17.502.28mg/grespectively, the IC50of the three index were11.97%,0.53%,0.37%respectively.