The Interaction Of Quinolone Drugs And Biological Macromolecules

The essential biological materials—protein, nucleic acid, have been playing vital roles in all kinds of biological phenomena. Exploring the interaction mechanisms on these bio-macromolecules with small molecules as well as effect of ions on these reactions, especially for those drug molecules, at the molecular level is of current interest in many research areas such as biology, clinical medicine chemistry, chemistry and so on. the present work consists of the following four parts:The pefloxacin mesylate (PFLX), Gatifloxacin(GTFX), Fleroxacin (FLRX) are broad-spectrum antibacterial fluoroquinolone, which are active against some microorganisms including Gram-positive and Gram-negative bacteria. In the first part, The interaction between PFLX, GTFX, FLRX and ions, such as copper(Ⅱ) or iron(Ⅲ) was investigated and their reaction equilibrium constant and the number of binding sites were determined with the fluorescence spectroscopy in aqueous solution. The binding characteristics of PFLX, GTFX, FRLX and Human Serum Albumin have been studied by fluorescence spectroscopy in aqueous solution, and the interaction influenced by ions was also explored in the paper. The quenching mechanism of fluorescence of HSA by PFLX, GTFX or FRLX is a static quenching procedure. The binding distance between PFLX, GTFX or FRLX and HSA and the energy transfer efficiency are obtained based on the theory of Forster spectroscopy energy transfer. The effect of PFLX, GTFX or FRLX on the conformation of HSA was also been analyzed by using synchronous fluorescence spectroscopy. The interaction of fluoroquinolone and HSA have been studied by flow-mixed microcalorimetry in the absence and presence of ions, such as copper(Ⅱ) or iron(Ⅲ), and their thermodynamic parameters were obtained. The enthalpy changes and the entropy changes were calculated to be ΔH ≈ 0, ΔS > 0 in the absence of copper( Ⅱ) or iron(Ⅲ),which indicated that hydrophobic forces played major role in the interaction of GTFX and HSA, and to be ΔH < 0, ΔS > 0 in the presence of ions, which indicated that the static forces also played major role on the reaction. The molar free energy changes of the two reactions are identical with each other because the entropy-enthalpy compensation happened between the two reactions. The orientation sites of the PFLX for HSA have been studied. Further more, the experiment results were explained by molecular modeling.In the second part, The interaction between PFLXn GTFXn FRLX and calf thymus DNA(CT-DNA) has been studied by spectral and electrochemical methods. Though there exist hypochromic effect on the reaction, no strong red shifts in the absorption spectra were observed when fluoroquinolone binds to CT-DNA. The quenching mechanism of fluoroquinolone fluorescence by DNA is a static quenching procedure and their equilibrium constant was determined. In addition, the binding characteristics of fluoroquinolone and DNA have been further studied by fluorescence method and spectral experiments showed that the quenching of fluorescence from fluoroquinolone by single strand (ssDNA) was larger than that by double strand (dsDNA), which indicated that the interaction between fluoroquinolone and DNA not belong to intercalation. The binding constant decreased with the [Na~+] concentration increasing, which indicated that there exists electrostatic force on the reaction. Theinfluence of copper (11^ iron(III) and magnesium (II) on the fluoroquinolone binding to DNA also investigated and it is found that ions played a media role between fluoroquinolone and DNA. The electrochemical characteristics of fluoroquinolone at solid electrode were also studied by cyclic voltammetry in the presence and absence of DNA based on the diffusion control theory.In the third part, the electrochemical interaction of sparfloxacin(SPFX) and DNA was investigated by adsorptive theory, and the electrode reaction mechanism of the interaction was also discussed in the paper. If the preconcentration time is longer enough, the electrode reaction will change from diffusion control to be adsorptive control. Cyclodextrins, containing peculiar hydmphobic cavum, could form inclusion complexes with SPFX. By means of the inclusion complexes, we concluded the structure of SPFX which could interact with DNA.The interaction of rivanol with calf thymus DNA(CT DNA) has been investigated by absorption, DNA melting experiment and fluorescence. When bound to CT DNA, rivanol shows hypochromism and red shift in absorption spectra, DNA melting temperature increasing and fluorescence quenching. These results strongly support intercalation of rivanol into CT DNA. The modified Scatchard plots constructed from fluorescence titration data give binding constant and binding size of base pairs per bound drug molecules at different ionic strength, further more, by means of the polyelectrolyte theory, the polyelectrolyte contribution, AGpe, to free energy was calculated. Isothermal titration calorimetry(ITC) has been used to determine the binding enthalpy and heat capacity(ACp) for rivanol. The ACP terms were used to estimate the hydrophobic contribution (AGhyd) to intercalative binding free energies. Comparison of the rivanol structure and doxorubicin structure allowed us to understand the value of AGhyd and entropy difference between the two molecules.