The effects of pH, temperature and protein concentration on thein vitrobinding of flutamide to human serum albumin

Abstract

Human serum albumin (HSA) is the most abundant protein in plasma with a high ligand-binding capacity. This capacity impacts the pharmacokinetic properties of therapeutic drugs. In the present study, the binding properties of flutamide to HSA at different temperatures, pHs and percentages of HSA were investigated. Thermodynamic parameters were also determined to describe the nature of binding interaction. A modified ultrafiltration method was used for accurate determination of flutamide-HSA parameters. Ultra filtrate samples containing free flutamide were extracted and analyzed by developed HPLC-UV method. Analysis of binding data was performed in terms of Scatchard, Klotz and Hill plots. Kinetic parameters (n, K) were found to be affected by temperature, pH and HSA concentration. However, flutamide-HSA-binding level did not show significant differences under different experimental conditions. The negative value of Gibbs free energy (ΔG) indicated that the binding was spontaneous. Moreover, the negative value for enthalpy and entropy changes suggested that hydrogen bonding and van der Waals forces played major role in the binding of flutamide to HSA. The results suggested a positive cooperation behavior of flutamide-HSA-binding that was affected by pH, temperature and percentage of HSA.

Enantioresolution and Binding Affinity Studies on Human Serum Albumin: Recent Applications and Trends

“…Additionally, by using a chiral stationary phase (CSP), it is possible to analyze the behavior of both enantiomers of racemic or enantiomeric mixtures in the same run [80][81][82]. Another advantage is the possibility to perform other type of analyses in addition to bioaffinity binding studies, including: displacement studies with the presence of a competitive agent [83], thermodynamic and variability studies of chromatographic conditions [84], quantitative structure-retention relationship studies [85], and enantioseparation of racemic or enantiomeric mixtures [2].…”

Evaluation of pH change effects on the HSA folding and its drug binding characteristics, a computational biology investigation

“…Many factors such as type of drug binding (competitive/noncompetitive), 5 pH, 3,6,7 temperature, salt concentration, 8 and polymorphisms of HSA 9 affect the binding of pharmaceutical agents to this blood protein. Regarding pH change effects, it was revealed that this physiological factor can affect conformation, net charge, sedimentation rate, denaturation temperature, and drug binding potency of HSA 6,10–12 …”

“…Regarding pH change effects, it was revealed that this physiological factor can affect conformation, net charge, sedimentation rate, denaturation temperature, and drug binding potency of HSA. 6,[10][11][12] The interaction of drugs with HSA is mainly mediated by hydrogen, hydrophobic and electrostatic bonds. 2,13 Currently, different types of laboratory approaches include high-performance affinity chromatography, 14 spectrophotometry, 15 topological substructural molecular descriptors/design (TOPS-MODE), 16 nuclear magnetic resonance (NMR), 17 X-ray, 18 equilibrium dialysis and ultrafiltration 19 and tandem affinity systems 20 developed to study of drugs interaction with plasma proteins.…”