The Application And Study Of The Interaction Mechanism Between Biomacromolecule And Pharmaceutical Molecules By Fluorescent Probe

The combining mechanism and functionary mechanism between pharmaceutical molecules and biomacromolecule, such as DNA and protein, are an important part in pharmaceutical science field. The efficiency of treatment of disease is directly affected by the combining location and functionary mechanism of drugs in body. Therefore, it is necessary to investigate the combining location and functionary mechanism between pharmaceutical molecules and biomacromolecule. It also can make sure the security, reasonability and availability when drugs are used by patients. For the past few years, the spectroscopic analysis has been more and more widely applied in organic chemistry analysis and life sciences research. As can be seen from many literatures, organic dye agents have been applied from the field of inorganic chemistry analysis to the field of life active substance (antibody, nucleic acid, protein, amino acid, etc) and the configuration analysis of environmental pollutant. However, sometimes the required experimental data can not be measured directly or calculated indirectly because there are no experiment phenomena between some pharmaceutical molecules and biomacromolecule, or the experiment phenomena are not distinct. The problem can been effectually settled by using the method of fluorescent probe. The method of fluorescent probe is a technique that investigate the functionary mechanism between pharmaceutical molecules and biomacromolecule through observing the fluorescence spectra changes of biomacromolecule or probe molecule in the interactions of drug molecules and probe molecules. Further, to find out the functionary mechanism of drug molecules in body. It is contradictory with the two sides between curative effects and adverse effects of drugs. It is a closely watched issue that how to improve drug treatment index. Launched in recent years targeting drug research is the effect of a certain drug and the appropriate carrier, the chemical combination of targeted drugs, according to the body of certain physiological functions, selective treatment to drug delivery to the sites. Thus increasing the role of drug treatment and reducing the adverse reaction because of the systemic distribution of the drug. Currently used by the carrier for the most intimate of endogenous synthesis of macromolecules or chemical substances, such as albumin, antibodies, glycoprotein, glucan or Poly-amino acid etc. There are many special requirements in applications, such as solubility, stability, purity, inherent toxicity, degradation, antigenicity, as well as the difficulty of combining with the of small molecule drugs.In recent years, the fluorescent probes are commonly used as the model systems for the investigations of surface-induced conformational changes in protein interfaces. From biopharmaceutical point of view, serum albumin is the most abundant protein in blood plasma, one of the most important biological function of albumins is their ability to carry drugs as well as endogenous and exogenous substances, playing the role of storage and transhipment. Numerous experiments with the aim to characterize the binding capacity and sites of albumins have been carried out. Analysis is based on monitoring the change of a physicochemical property of the protein-probe system upon binding either directly (direct technique) or after separation of the bound and free probe (indirect technique). Among the direct techniques fluorometry is extensively used and is considered to be superior to the indirect techniques (equilibrium and dynamic dialysis, ultrafiltration, gel fitration) because, to a first approximation, they do not disturb the binding equilibrium upon separation.In this paper, the combination location and functionary mechanism between biological macromolecules and drug molecules have been studied using fluorescent probes technique. The drug mechanism and the role played pharmacodynamics way in the body have been further understood and studied. Mainly to study the combination constants, binding sites, binding points, the energy transfer parameter and other parametes by observing the fluorescence spectra changes of the interactions among biomacromolecules, probe molecules and drug molecules and probe molecules, and then get the functionary mechanisms of them.The first chapter of the paper reviews the research and application of the interactions between drug molecules and biological macromolecules in recent years. And the application and prospects of fluorescent probe in the field.In the second chapter of the paper, according to the fluorescence quenching phenomenon of Evans blue (EB) to bovine serum albumin (BSA), the interaction between Evans blue and bovine serum albumin has been studied by the fluorescence spectroscopy and the UV-visible absorption spectroscopy. It was important that the fluorescence quenching was occurred when EB was added in BSA. The static fluorescence quenching process was confirmed based on the Stern-Volmer plot and the measurements of the fluorescence lifetimes. The binding constant K=1.122×106Lmol-1 and the number of binding sites n=0.994 were obtained based on the fluorescence quenching results. By F?rster'non-radiative energy transfer theory, the energy transfer efficiency and the distance between BSA and EB were found, they were 0.276 and 3.14nm, respectively. The conformational changes of BSA were studied. The fluorescence spectra of BSA mainly were originated from the tryptophan residues according to their synchronous fluorescence spectra.From the third chapter to sixth chapter in the paper, the competitive reactions between bovine serum albumin and gentamicin, vancomycin, ampicillin or erythromycin have been respectively studied with Evans blue as fluorescent probe by the fluorescence spectroscopy and the UV-visible absorption spectroscopy. Fluorescence quenching was occurred between the bovine serum albumin and Evans blue. The static fluorescence quenching process was confirmed based on the Stern-Volmer plot and the measurements of the fluorescence lifetimes. The binding constant, and the number of binding sites, were obtained. And the thermodynamic constants and types of forces were determined between BSA and EB. It was interesting that the relative fluorescence intensity of BSA was recovered gradually with an increasing concentration of the drugs. This indicated that the competitive reactions occurred between the drugs and BSA for EB. The reaction mechanisms were studied respectively, and the binding constants of the drugs with EB were obtained, respectively.