Bovine Serum Albumin as a Potential Carrier for the Protection of Bioactive Quercetin and Inhibition of Cu(II) Toxicity

Abstract

Considering the protective ability of proteins and the potential toxicity of free Cu­(II), it was proposed herein that the co-presence of protein could play an important role in suppressing the toxicity of free Cu­(II) to the stability of bioactive quercetin if a flavonoid–protein–Cu­(II) complex could be formed. In this study, the interaction between quercetin (a major flavonoid in the human diet) and bovine serum albumin (BSA) was investigated in the absence and presence of free Cu­(II). The results demonstrated that both quercetin and free Cu­(II) had a strong ability to quench the intrinsic fluorescence of BSA through a static procedure (i.e., formation of a BSA–monoligand complex). Site marker competitive experiments illustrated that the binding of both quercetin and Cu­(II) to BSA mainly took place in subdomain IIA. The quenching process of free Cu­(II) with BSA was easily affected by quercetin, and the increased binding capacity possibly resulted from the generation of a ternary quercetin–BSA–Cu­(II) complex. The stability and free radical scavenging activity of bioactive quercetin during incubation was promoted in the BSA–diligand complex relative to a quercetin–Cu­(II) complex. A quercetin–Cu­(II) system could generate reactive oxygen species such as hydrogen peroxide (H2O2) and hydroxyl radicals (•OH), which were significantly inhibited upon BSA binding. Consistently, the cytotoxicity of the quercetin–Cu­(II) system to endothelial cells was decreased in the BSA–diligand complex, where the co-presence of BSA played an important role. These results suggest the possibility and advantage of developing albumin-based carriers for the protection of bioactive components and suppression of Cu­(II) toxicity in their biomedical and nutritional applications.

Binding of Quercetin to Hemoglobin Reduced Hemin Release and Lipid Oxidation

“…Quercetin was then docked to the crystal X-ray structures of the Hb molecule by AutoDock 4.2 according to previous procedures. 23,24 Based on these molecular docking data, the docking model with the lowest binding free energy was adopted as the most preferential binding mode.…”

“…After the interaction of Hb (6 μM) with flavonoid (0–16 μM) for 60 min at different temperatures (280, 298, and 310 K), fluorescence spectra of proteins were determined with a fluorescence spectrophotometer (FluoroMax-4, Horiba Jobin Yvon, Japan) at λ ex 280 nm. To eliminate the inner filter effects of Hb and quercetin, the fluorescence intensities of protein were corrected according to previous methods. ,, The corrected fluorescence value was applied in this study. The mechanism for fluorescence quenching was elucidated by the well-known Stern–Volmer equation F 0 / F = 1 + k q τ 0 [ Q ] = 1 + K SV [ Q ], where F 0 and F represent the fluorescence intensities of Hb without or with the existence of the quencher (quercetin), respectively.…”

“…To eliminate the inner filter effects of Hb and quercetin, the fluorescence intensities of protein were corrected according to previous methods. 23,24,33 The corrected fluorescence value was applied in this study. The mechanism for fluorescence quenching was elucidated by the wellknown Stern−Volmer equation…”

“…In a previous study, it has been demonstrated that gallic acid could act as a reductant to dose-dependently retard Hb oxidation by reducing met-Hb formation . As one of the most major flavonoids in plants and human diet, quercetin has been found to be an efficient inhibitor of lipid oxidation in vitro and vivo. , In addition, the protein–polyphenol interaction is a widespread phenomenon and some flavonoids could bind to Hb with high affinity. − Thus, it was proposed herein that the binding of flavonoid might have the ability to affect Hb redox activity. However, the effects of the interaction of quercetin with Hb on lipid oxidation were rarely elucidated.…”

“…Quercetin is composed of many active hydroxyl groups in the 3, 5, 7, 3′, and 4′ positions of the benzene ring (Figure ), which enables this compound to interact with various proteins. ,, In this way, quercetin would bind to Hb and affect the redox activity of Hb. The aim of this study was to demonstrate the interactions of bioactive quercetin with bovine (or human) Hb, which was related to the potential antioxidant mechanism.…”