Spectroscopic Study On Interaction Of Some Drugs With Bovine Serum Albumin

The research actuality of interaction between small molecules with serum albumin in recent years has been reviewed and the interaction between some drugs with bovine serum albumin (BSA) was studied. Spectroscopy method has become firmly entrenched, particularly for the interaction of small molecules and macromolecules. However, the spectroscopy investigation combined with chemometrics in this field is limited. The application of chemometrics method- multivariate curve resolution-alternating least squares (MCR-ALS) in processing of spectral data also had been developed. And the application of chemometrics in complicated biochemical system can solve some problems and obtain the equilibrium concentration and pure spectra of each component.This thesis is divided into five parts:Part IIn part one, firstly, the structures, functions and natures of proteins were introduced briefly. Then the methods and contents of interaction of different kind of small ligands with serum albumin were reviewed in turns. Finally, the application of chemometrics in the field of protein research was discussed.Part IIIn part two, the binding of trazodone hydrochloride (TZH), generally used as an antidepressive agent, to BSA has been investigated by fluorescence method. The quenching mechanism of fluorescence of BSA by TZH was investigated, and the binding sites number n and apparent binding constant K was measured by fluorescence quenching method. Thermodynamic parameters obtained from measured data at different temperatures showed the binding force of TZH to BSA. Phenylbutazone and ibuprofen were used as the site I and site II binding markers on the BSA in this work, respectively. According to the displacement procedure of the site markers on albumins, the binding site of TZH on the BSA has been found to be site I.Part IIIIn part three, the binding of pyrazinamide, generally used as an antitubercular drug, to BSA has been investigated by spectrometry. The quenching mechanism of fluorescence of BSA by pyrazinamide was found to be static quenching, and the binding sites number n and apparent binding constant K was measured by fluorescence quenching method. Thermodynamic parameters obtained from measured data at different temperatures showed that the binding of pyrazinamide to BSA involves hydrogen bond and Van der Waals' interation. The binding distance was obtained according to the theory of Forster's non-radiation energy transter. The effect of pyrazinamide on the conformation of BSA was also analyzed using synchronous fluorescence spectrometry and absorption spectra.Part IVIn part four, the interaction of quercetin, which is a bioflavonoid, with BSA was investigated under physiological conditions by the application of UV-vis spectrometry, spectrofluorimery and cyclic voltammetry (CV). The spectrofluorimetry was divided into two parts: fluorescence quenching and fluorescence enhancement method. UV-vis spectrometry and spectrofluorimetry were applied to investigate the nature of the binding site of quercetin on BSA with the use of warfarin as site I marker. These studies indicated a cooperative interaction between the quercetin-BSA complex and warfarin, which produced a ternary complex, i.e. quercetin-BSA-warfarin. It was found that quercetin and warfarin were located in site I. However, the spectra of these three components overlapped and chemometrics method MCR-ALS was applied to resolve the spectra. Complex ratio and equilibrium constant were obtained by this technique.Part VIn part five, the interaction of tetracycline (TC), which is a antibiotic, with BSA was investigated under physiological conditions by the application of fluorescence spectroscopy with the use of chemometric method MCR-ALS. In order to break through the limit of traditional site marker, we try to exploit new fluorescence dye probe eosin (EY). Fluorescence competitive binding was divided into two parts: first, the classical site I marker warfarin (WF) was used to determine the binding site of EY which we tried to used as fluoresce probe on BSA. The chemometrics technique MCR-ALS was applied to the two-way fluorescence spectra to extract more information. Then the binding site of EY on BSA was confirmed to be site I. Second, the exploited probe EY was used as site marker of the interaction between TC and BSA. MCR-ALS was also used to resolve the two-way fluorescence spectra. The competitive binding studies showed that the binding site of TC on BSA was site I. In addition, the results of fluorescence polarization was also demonstrated the conclusion of the site I binding site of TC on BSA. Synchronous fluorescence technique was used to study the conformation transformation, which showed that the conformation of BSA was not changed by the addition of TC. In comparison with traditional protein site markers, EY exploited as site markers shows more superiority.