Spectrometric and voltammetric studies of the interaction between quercetin and bovine serum albumin using warfarin as site marker with the aid of chemometrics

The interaction of quercetin, which is a bioflavonoid. with bovine serum albumin (BSA) was investigated under pseudo- physiological conditions by the application of UV-vis spectrometry,spectrofluorometry and cyclic voltammetry (CV). These studies indicated a cooperative interaction between the quercetin-BSA complex and warfarin,which produced a ternary complex, quercetin-BSA-warfarin. It was found that both quercetin and warfarin were located in site I. However, the spectra of these three components overlapped and the chemometrics method-multivariate curve resolution-alternating least squares (MCR-ALS) was applied to resolve the spectra. The resolved spectra of quercetin-BSA and warfarin agreed well with their measured spectra, and importantly, the spectrum of the quercetin-BSA-warfarin complex was extracted. These results allowed the rationalization of the behaviour of the overlapping spectra. At lower concentrations ([warfarin] < 1 x 10(-5) mol L-1), most of the site marker reacted with the quercetin-BSA, but free warfarin was present at higher concentrations. Interestingly, the ratio between quercetin-BSA and warfarin Was found to be 1:2, suggesting a quercetin-BSA-(warfarin)(2) complex, and the estimated equilibrium constant was 1.4 x 10(11) m(-2). The results suggest that at low concentrations, warfarin binds at the high-affinity sites (HAS). while low-affinity binding sites (LAS) are occupied at higher concentrations. (C) 2008 Elsevier B.V. All rights reserved.