Characterization of Solute Binding at Human Serum Albumin Site II and its Geometry Using a Biochromatographic Approach

Abstract

Chiral recognition mechanism relationships for binding at site II on human serum albumin (HSA) were investigated using D, L dansyl amino acids. Sodium phosphate salt was used as a solute-HSA interaction modifier. A new model was developed using a biochromatographic approach to describe the variation in the transfer equilibrium constant with the salt concentration, i.e., the nature of the interactions. The solute binding was divided into two salt concentration ranges c. For the low c values, below 0.03 M, the nonstereoselective interactions constituted the preponderant contribution to the variation in the solute binding with the salt concentration. For the high c values, above 0.03 M, the solute binding was governed by the hydrophobic effect and the stereoselective interactions. The different contributions implied in the binding process provided an estimation of both the surface charge density (sigma/F) and the surface area of the site II binding cavity accessible to solvent, which were found to be equal to around 10.10(-7) mol/m(2) and 2 nm(2). As well, the excess of sodium ions excluded by the solute transfer from the surface area of the pocket were about(-0.7) for dansyl norvaline and (-0.8) for dansyl tryptophan.

Interactions between D,L dansyl amino acids and immobilized teicoplanin: Study of the dual effect of sodium citrate on chiral recognition

“…When the chromatographic data were fitted to Eq. (4) (the model taking into account anion binding at the solute interaction site), however, using a non-linear procedure [19], very good correlation (r 2 > 0.99) was obtained between predicted and experimental retention. This result shows that the sodium citrate buffer affects dansyl amino acid binding as a result of two antagonist effects: (i) an increase in affinity as a result of enhancement of the hydrophobic effect; and (ii) competitive displacement of the solute from its binding site by the citrate anion.…”

“…All k values increased when c was increased. This can be attributed to enhancement of the hydrophobic effect [19]. It is well known that adding salting-out agents to an aqueous medium increases its surface tension and the energy required for cavity formation [28].…”

“…Plots of retention factor against amount of sample had a plateau at a sample concentrations below 5 ixg mL 1 followed by a small decrease at higher solute concentrations. Each solute was, therefore, injected at a concentration of 2.5 ixg mL 1, at which retention was independent of sample concentration, i. e. under linear elution conditions [19].…”

Effect of sodium chloride on herbicide-fluorinated stationary phase association-Evidence of a dual mechanism

“…Similar behaviour was observed for the other derivatives. When the NaCI concentration was increased beyond xo selectivity remained constant, confirming well that hydrophobic interactions were not specific [37][38][39]. Figure 10 shows the chromatogram obtained from separation of 2-CPPA and 3-CPPA when the NaC1 concentration was 0.4 M. At this NaCI concentration, the column efficiency, i.e.…”

Chiral separation of enantiomers of amino acid derivatives by high-performance liquid chromatography on a norvancomycin-bonded chiral stationary phase

“…Plots of retention factor against amount of sample had a plateau at a sample concentration lower than 4 g ml −1 followed by a small decrease at higher solute concentration. Each solute was, therefore, injected at a concentration of 2.5 g ml −1 , at which retention was independent of sample concentration, i.e., under linear elution conditions [13,14]. No apparent changes in the stability of the NVC column were noted over the course of this study, as determined by periodically measuring the retention factors for the compounds tested under the same operating conditions.…”