| Some functional groups of biopolymers can bind with the molecular of drugs, which can keep some density for harmfulless, efficient and controlled-release action. Drugs bind with biopolymers can prolong drug existing in blood.Albumin, a typical biopolymer, is an important transport carrier for drugs. The binding of drugs with protein has a great influence on not only the distribution of the drugs in the body but also their patterns of metabolism. Thus, studies on supramolecular interaction of drugs with albumin have significantly contributed to the understanding of the structures and functions of biomacromolecules and some biophysical processes.Objective: In this article, the interactions of five flavones active components with bovine serum albumin (BSA) and influence of metal ions or alcohol were investigated under physiological condition in vitro. The work according to several studies such as quenching, conformation of protein, forces, binding distance and the effect of metal ions or alcohol on the interactions between small medicinal molecules and protein is worthy to be done, which provides important information on the design, and synthesis of new drugs.Methods:1. The interactions between flavones (Luteolin, Luteolin-7-o-β-D-glucoside, Acacetin, Acacetin-7-o-β-D-glucoside and Apigenin) with BSA in physiological condition were studied by fluorescence and UV-visible spctrophotometry. The quenching constant Ksv, the binding constant K, the number of binding sites n and thermodynamic parameters△H,△G and△S were calculated. The distance r between donor (BSA) and acceptor (flavones) was obtained according to the theory of F?ster's nonradioactive energy transfer. The effects of flavones on the conformation of BSA were analyzed by UV-vis absorbance and synchronous fluorescence spectroscopy.2. Add certain density of metal ions liquor to the Flavone-BSA reaction system. The influences of metal ions Mn2+, Sr2+, Co2+, Ni2+ on the interactions of BSA-flavones, such as the changes of quenching mechanism of fluorescence, the interaction forces, the conformation of BSA, the binding constants and the number of binding sites, were investigated by fluorescence at different temperature (298 K, 310 K).3. Add a certain content of alcohol to the Flavone-BSA reaction system, make the concentration of alcohol reaching different levels of drunk driving, which are according to the highway code in China. The influence of alcohol on the interactions between BSA and flavones, such as the binding constants and the number of binding sites were investigated by fluorescence at 310 K.Results:1. The binding mechanism was proved to be the static quenching preocess according to the binding constant decreasing along with the temperatures increasing (298 K, 303 K and 310 K). Five flavone compounds formed 1: 1 complex with BSA. The binding constants, were 1.17×106, 7.62×105, 1.34×106, 4.99×104, 2.54×105 L·mol-1 at 298 K, 7.20×104, 8.47×104, 2.61×105, 1.77×104, 1.66×105 L·mol-1 at 303 K, and 2.62×103, 8.53×104, 2.26×104, 1.32×104, 1.5×105 L·mol-1 at 310 K respectively. And the dominant binding forces were hydrophobic forces according to△H>0,△G<0 and△S>0. There was nonradioactive energy transferring between them. The binding distances were 3.6, 4.16, 4.29, 4.07 and 3.8 nm respectively. The conformation of BSA was changed due to the presence of five compounds.2. The results showed that the binding constants of Flavone-BSA appeared significantly changed because of the existence of metal ions (Mn2+, Sr2+, Co2+ and Ni2+). At 298 K, the influence of Co2+ was the most remarkable in the four metal ions for Luteolin, Luteoloside, Acacetin and Acacipetalin, KA increasing from 1.17×106, 7.62×105, 1.34×106, 0.50×105 L·mol-1 to 26.49×106, 26.77×105, 6.64×106, 5.48×105 L·mol-1 respectively. Sr2+ was the most remarkable for Apigenin, KA increasing from 2.54×105 to 41.57×105 L·mol-1. At 310 K, the metal ions increased KA of Luteolin, Luteoloside, Acacipetalin with BSA, while decreased the others. Ni2+ was the most remar Kable for Luteolin, KA increased from 0.03 to 55.59×105 L·mol-1. KA of Apigenin-BSA decreased from 1.60 to 0.06×105 L·mol-1 due to the existence of Mn2+. The quenching mechanism of the Flavone-BSA reaction system appears different changes. The dominant binding forces still were hydrophobic forces. The variation of synchronous fluorescence peak of tryptophan residues in the ternary system of Me2+-Flavone-BSA was opposite to the system without Me2+, fluorescence peak red shift occuring, it showed that metal ions influenced the spectrum characteristic of synchronous fluorescence observably and intensified the changes of the comformation of BSA.3. The results showed that the binding constants and the number of binding sites were decreased due to adding alcohol when the final contents of alcohol were 0.4 mg/ml and 0.8 mg/ml. The binding constants KA were 1.21×103, 1.13×104, 8.32×103, 1.05×104, and 1.47×104 L·mol-1 respectively, when the concentration of alcohol was 0.4 mg/ml. While it was 0.8 mg/ml, KA was much lower and they were 1.10×103, 6.62×103, 6.20×103, 8.89×103, and 1.34×104 L·mol-1 respectively. Binding sites were decreased, but they maintained around 1.Conclusion: The results showed that the interactions of five flavones with bovine serum albumin in physiological condition was studied by fluorescence and UV-visible spctrophotometry, the binding mechanism was proved to be the single static quenching process, the quenching constant Ksv, the binding constant K and the number of binding sites n were calculated. The distance r between donor (BSA) and acceptor (flavones) was obtained according to the theory of F?ster's nonradioactive energy transfer. The dominant binding force was hydrophobic forces. The conformation of BSA was changed due to the presence of five compounds. Metal ions influenced the intereaction of Flavone-BSA and intensified the change of the forming of BSA. The binding constants and the number of binding sites were decreased due to adding alcohol.
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