Analysis of binding interaction between pegylated puerarin and bovine serum albumin by spectroscopic methods and dynamic light scattering

The interaction between bovine serum albumin (BSA) and pegylated puerarin (Pur) in aqueous solution was investigated by UV–Vis spectroscopy, fluorescence spectroscopy and circular dichroism spectra (CD), as well as dynamic light scattering (DLS). The fluorescence of BSA was strongly quenched by the binding of pegylated Pur to BSA. The binding constants and the number of binding sites of mPEG5000-Pur with BSA were 2.67 ± 0.12 and 1.37 ± 0.05 folds larger after pegylating, which were calculated from the data obtained from fluorescence quenching experiments. The enthalpy change (ΔH) and entropy change (ΔS) were calculated to be 4.09 kJ mol−1 and 20.01 J mol−1 K−1, respectively, according to Van’t Hoff equation, indicating that the hydrophobic force plays a main role in the binding interaction between pegylated Pur and BSA. In addition, the negative sign for Gibbs free energy change (ΔG) implies that the interaction process is spontaneous. Moreover, the results of synchronous fluorescence and CD spectra demonstrated that the microenvironment and the secondary conformation of BSA were changed. Comparing with Pur, all our data collected indicated that pegylated Pur interacted with BSA in the same way as that of Pur, but docked into the hydrophobic pocket of BSA with more accessibility and stronger binding force. DLS measurements showed monomethoxy polyethylene glycol (mPEG) have an effect on BSA conformation, and revealed that changes in BSA size might be due to increases in binding constant and the absolute values of ΔG after Pur pegylation.

The radius change models of BSA/Puerarin-mPEG prodrug complex and its probable mechanism in different solvents. (a) BSA model; (b) BSA solvent; (c) mPEG5000-containing solvent with CmPEG5000/CBSA=1CmPEG5000/CBSA=1; (d), (e) and (f) are mPEG5000-Pur with CmPEG5000-Pur/CBSA=1CmPEG5000-Pur/CBSA=1, 2, 10, respectively.Figure optionsDownload as PowerPoint slideHighlights
► mPEG5000-Pur-BSA interaction model was established by spectroscopic methods and dynamic light scattering methods.
► The pegylated puerarin interacted with BSA in the same way as that of free puerarin, but pegylation enhanced its accessibility and binding force to BSA.
► The PEG chain extending from BSA surface may conceal antigen sites formed by puerarin and BSA, thereby prolonging the systemic half-life and ameliorating or extinguishing the hemolytic side effect of puerarin.