Study Of The Interaction Of Flavonoids With Serum Albumin

In this thesis, fluorescence quenching spectrums, ultra-visible absorptionspectrum as well as other multi-spectral spectrum analysis means were used toinvestigate the interaction between several flavonoids with similar structure andbovine serum albumin (BSA) at molecule level in simulated physiological conditions.By the study on interaction parameters, we contrasted different spectral characteristicsof different flavonoids binding to BSA, which are important for the transport andmetabolism of flavonoids in body, new drug discovery and design as well as thefunctional food research and development.This thesis consists of four parts as follows:1. In chapter I, I reviewed the molecule structure of serum albumin, classificationand physiological functions of flavonoids, related background knowledge as well ascommon research methods of interaction between biological macromolecules andfunctional small molecules. We also introduced the research progress in the field andsome of the problems that need to be addressed.2. In chapter II, I discussed the effect of measure method, mixing and standingtime on fluorescence quenching system. We also studied the multi-spectralcharacteristics of the interaction between quercetin and BSA at simulatedphysiological environment. We obtained the Stern-volmer quenching constant (Ksv)and binding constants and proved that the interaction between quercetin and BSA wassingle static quenching, the binding distance between the binding site of quercetin onBSA and 212 tryptophan was 1.783 nm. The dominant force of quercetin and BSAwas proved as electrostatic attraction. Synchronous fluorescence and Fouriertransform infrared spectroscopy (FT-IR) spectra showed that quercetin could affectthe secondary structure of BSA. Metal ions experiment results showed that differentmetal ions may affect differently on different binding parameters. 3. In Chapter III, I studied on both the interaction of luteolin-BSA andhesperetin-BSA with UV absorption spectra, fluorescence quenching spectra,synchronous fluorescence spectra and FT-IR spectra. The fluorescence quenching ofluteolin-BSA was mixed quenching of dynamic quenching and static quenching whilethe quenching of hesperetin-BSA was single dynamic quenching. In addition, thequenching degree of luteolin to BSA was significantly higher than that of hesperetinto BSA. Calculated results of thermodynamic parameters showed that electrostaticattraction exist in both luteolin-BSA and hesperetin-BSA quenching system.Synchronous fluorescence spectra indicated that the binding sites of luteolin andhesperetin on BSA may be in the vicinity of the tryptophan residues. UV absorptionspectra showed that BSA could interact with luteolin and hesperetin, but in differentdegrees of changes on the structure of BSA. FT-IR spectra further illustrated theluteolin and hesperetin have different effects on the secondary structure of BSA.4. In chapterⅣ, I studied on the effects of Ca2+，K+，Cu2+，Mg2+，Fe3+on Ksv,Kaand n of apigenin-BSA system, the result of which was that Mg2+showed thegreatest impact. The pH value, concentration of NaCl and Mg2+concentration wereselected as the single factors to explore the relationship of Ksv, Ka, n and the threesingle factors. The effect of the interaction of the independent variables on Ksv, Ka,nwas investigated by response surface methods (RSMs). Prediction models were fit toa quadratic polynomial regression equation and the results showed that both Ksvand ndisplayed a second-order model. Under these experimental conditions, [NaCl] was themost significant impact factor on Ksvand Ka, whereas n was affected most by pH. Thisconclusion may provide some help in the choice of experimental conditions whenother researchers study on the interaction between functional small molecules andbiological macromolecules by fluorescence quenching spectroscopy methods.