Fig 1.
Molecular structures of salvianolic acid B (A) and rosmarinic acid (B).
Fig 2.
(A) UV-visible absorbance spectra of Au NPs. Inset is the photo taken from gold hydrosols, (B-D) Representative TEM images and the corresponding size distributions (inset) of Au NPs synthesized at different molar ratio of sodium citrate/chloroauric acid. (B) 4/1, (C) 3/1, (D) 2/1.
Fig 3.
Fluorescence quenching spectra of BSA–drug systems with and without Au NPs at 298 K, λex = 280 nm, CBSA = 5.0 × 10-6 mol L-1.
(A) BSA-SAB system, the concentrations of SAB (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (B) BSA-RA system, the concentrations of RA (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (C) BSA-SAB-Au4 system, CAu4 = 10.0 × 10-11 mol L-1, the concentrations of SAB (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (D) BSA-RA-Au4 system, CAu4 = 10.0 × 10-11 mol L-1, the concentrations of RA (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (E) BSA-SAB-Au3 system, CAu3 = 10.0 × 10-11 mol L-1, the concentrations of SAB (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (F) BSA-RA-Au3 system, CAu3 = 10.0 × 10-11 mol L-1, the concentrations of RA (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (G) BSA-SAB-Au2 system, CAu2 = 10.0 × 10-11 mol L-1, the concentrations of SAB (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively. (H) BSA-RA-Au2 system, CAu2 = 10.0 × 10-11 mol L-1, the concentrations of RA (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0, and 25.0 × 10-6 mol L-1, respectively.
Fig 4.
The Stern-Volmer plots of BSA–drug systems without and with Au NPs at 298 K.
The Stern-Volmer plots for BSA fluorescence quenching by SAB (A) and RA (B) in the absence of Au NPs (square) and in the presence of Au4 (circle), Au3 (up triangle) and Au2 (down triangle). CBSA = 5.0 × 10-6 mol L-1, the concentration of Au NPs was 10.0 × 10-11 mol L-1, λex = 280 nm, pH = 7.40.
Table 1.
Stern-Volmer quenching constants (KSV) and bimolecular quenching rate constants (kq) of drug-protein systems with and without Au NPs at 298 K and pH = 7.4.
Fig 5.
Plots for calculating the binding constants and the stoichiometry of binding of BSA–drug systems without and with Au NPs.
Double logarithmic curves of SAB (A) and RA (B) quenching BSA fluorescence in the absence of Au NPs (square) and in the presence of Au4 (circle), Au3 (up triangle) and Au2 (down triangle). CBSA = 5.0 × 10-6 mol L-1, the concentration of Au NPs was 10.0 × 10-11 mol L-1, λex = 280 nm, pH = 7.40.
Table 2.
Binding constants (Kb), the stoichiometry of binding (n) and free energy change (ΔG) of drug-protein systems with and without Au NPs at 298 K and pH = 7.4.
Table 3.
The distance parameters of different drug-protein systems at 298 K.
Fig 6.
Schematic representation of mechanism of drug binding to BSA in the binary complex systems and the ternary complex systems.
(A) The BSA-drug interaction in the binary complex systems. (B) Competitive binding: competitive displacement between the third substance and drug from the same binding site on the BSA in the ternary complex systems. (C) Synergistic reaction: a new complex is formed by the third substance and drug, and then it interacts with BSA in the ternary complex systems. (D) Noncompetitive binding: the interaction of the third substance with BSA at a different binding region induces the conformational change of drug binding site on the BSA in the ternary complex systems.
Fig 7.
UV-visible absorbance spectra of free drug (A: SAB and B: RA) and their corresponding Au NPs complexes.
CSAB = CRA = 2.5 × 10-5 mol L-1, the concentration of Au NPs was 10.0 × 10-11 mol L-1.
Fig 8.
Effect of site marker probes on the fluorescence intensities of BSA-drug systems (A: BSA+SAB, B: BSA+RA).
Site marker competitive experiments were carried out using three site markers (warfarin, ibuprofen, and digitoxin). CBSA = 5.0 × 10-6 mol L-1, CSAB = CRA = 2.0 × 10-5 mol L-1, λex = 280 nm, T = 298 K.
Fig 9.
The synchronous fluorescence spectra of BSA–drug systems without and with Au NPs at different Δλ values (15 and 60 nm).
(A) BSA-SAB system, (B) BSA-RA system, (C) BSA-SAB-Au4 system, (D) BSA-RA-Au4 system, (E) BSA-SAB-Au3 system, (F) BSA-RA-Au3 system, (G) BSA-SAB-Au2 system and (H) BSA-RA-Au2 system. The concentration of BSA was fixed at 5.0 × 10-6 mol L-1 and the concentrations of SAB/RA (1–9) were 0, 2.5, 5.0, 7.5, 10.0, 12.5, 15.0, 20.0 and 25.0 × 10-6 mol L-1, respectively. The concentration of Au NPs was 10.0 × 10-11 mol L-1.
Table 4.
Three-dimensional fluorescence spectral characteristics of BSA-drug systems.