Studies On The Interactions Between The Synthesized Oleanolic Acid And5-fluorouracil Derivatives And Bovine Serum Albumin

Aiming at improving the disadvantages of oleanolic acid which is indissolvable inwater, has short organisms half-life and low absolute bioavailability, cannot maintaineffective blood medicine, this research has proposed grafting methoxypolyethyleneglycols onto hydroxyl of oleanolic acid and obtained the polymer coupling medicinesmPEGylated oleanolic acid （mPEG-OA）. Meanwhile, aiming at5-fluorouracil is awidely and effective antitumor medicines but with large poisonous side effect, thispaper has synthesized5-fluorouracil-1-yl acetic acid （5-FuAc） with low side effectwhich was replaced by N-acetic acid.After that, this paper investigates the interaction between BSA and the twomodified medicines including the action mechanism, action zone, acting force andbinding sites through using several spectroscopic techniques. The result of these re-searches is helpful to understand the absorption, distribution, excretion andpharmacological activity in human body as well as has an important and guidingsignificance to know the structure and function of protein further and to research anddevelop new medicine better.The main research results and conclusions are as follows:1. Activated methoxypolyethylene glycols by succinic anhydride is graftedsecondary-OH of oleanolic acid. It used the FT-IR, UV-Vis and GPC spectrumto confirm structure of mPEG-OA. The water-soluble experiment has provedits water-solubility was increased observably. Meanwhile,5-FuAc issynthesized by connecting-CH2COOH with5-Fu.2. The interaction of mPEG-OA with bovine serum albumin （BSA） have beendetailedly studied by fluorescence quenching technique in combination with UV-visabsorption, circular dichroism （CD） spectroscopies under the simulative physiologicalconditions. The results showed that the quenching of BSA fluorescence bymPEG-OA was found to be a dynamic quenching procedure. Based on measuringfluorescence spectra of protein, the binding constants were calculated to be6.870,5.577and2.925×10³L·mol^-1at293,303, and313K, respectively. According to the van′t Hoff equation, the thermodynamic parameters were estimated, which indicatedthat mPEG-OA interacted with BSA mainly through hydrophobic interactions. Theentropy increase and the free energy reducing in this combining process proved it is aspontaneous procedure. The binding distance between BSA and mPEG-OA wascalculated to be3.54nm according to the theory of F rster′s non-radiation energytransfer. The displacement experiments pointed out that mPEG-OA could bind to thesite I of BSA. Alteration of the secondary protein structure in the presence of thepiracetam was confirmed by CD spectroscopy, but, it is detected by synchronousfluorescence spectroscopy.3. To study the interaction mechanism of BSA with5-FuAc under physiologicalconditions by fluorescence quenching in combination with UV-vis absorption, CDspectroscopy. Fluorescence data revealed that5-FuAc had a weak ability to quenchthe intrinsic fluorescence of BSA through a static quenching procedure; Calculatedthe binding constants of the5-FuAc-BSA system was1.209×10⁴、9.530×10³and1.260×10⁴L·mol^-1respectively. Their binding sites n are about equal to1at threetemperatures. Based on thermodynamics relationship, the thermodynamicparameters for the reaction were calculated, which indicated that5-FuAc binded toBSA mainly by hydrophobic forces and electrostatic force interactions. The entropyincrease and the free energy reducing in this combining process proved it is aspontaneous procedure. The binding distance between BSA and5-FuAc wascalculated to be5.25nm according to the theory of F rster′s non-radiation energytransfer. The displacement experiments suggested that5-FuAc could bind to the siteSite II of BSA. CD spectra and synchronous fluorescence spectroscopy demonstratedthat5-FuAc induced secondary structure of BSA molecules in the micro-change.