title: Assessment of the Dissociation Energetics of Some Selected Ligand Drugs Bound on Human Serum Albumin by Differential Scanning Calorimetry.
abstract: Drug-protein binding may play a role in the thermal energetics of protein denaturation and could lead to the determination of its equilibrium dissociation parameter. The aim of this study was to assess the energetics of a drug that was bound to human serum albumin (HSA) during thermal denaturation. Drugs that were bound at a single high-affinity primary binding site on HSA, including diazepam and ibuprofen, were employed. Commercial HSA was treated with charcoal to remove stabilizers and adjusted to 20% w/v in a pH 7.4 buffered solution. Serial concentrations of individual drugs up to 0.16 mmole/g-protein were added to the cleaned HSA solutions whereas diazepam was added to a commercial HSA solution. Samples were subjected to differential scanning calorimetry (DSC) set to run from 37 to 90°C at 3.0°C/min. Binding of the drug slightly increased the denaturing temperature of the cleaned HSA due to a shift in the equilibrium toward the native protein bound with the drug. Diazepam depressed the denaturing temperature of the commercial HSA by competing with the stabilizers already bound to the primary site of the HSA. This yielded not only the HSA-stabilizer but also the HSA-diazepam type complexes that exhibited a different denaturation process. A rational approximation of the Lumry-Eyring protein denaturation model was used to treat the DSC endotherms. The approximated scheme: [Formula: see text] was successfully fitted to the data. It was used to determine the dissociation parameters for diazepam and ibuprofen bound to the HSA. These results were comparable to those obtained from other methods.
authors.affiliation: Drug Delivery System Excellent Center, Department of Pharmaceutical Technology, Faculty of Pharmaceutical Sciences, Prince of Songkla University, Hat Yai, 90112, Thailand. damrongsak.f@psu.ac.th.
authors.forename: Damrongsak
authors.initials: D
authors.lastname: Faroongsarng
mesh_terms: D001665:Binding Sites; D002152:Calorimetry, Differential Scanning / Q000379:methods; D003975:Diazepam / Q000737:chemistry*; D006801:Humans; D007052:Ibuprofen / Q000737:chemistry*; D008024:Ligands; D011485:Protein Binding; D011489:Protein Denaturation; D012709:Serum Albumin / Q000737:chemistry*; D013696:Temperature
chemical_list: D008024:Ligands; D012709:Serum Albumin; D003975:Diazepam; D007052:Ibuprofen
keywords: Lumry-Eyring model; differential scanning calorimetry; dissociation energetics; human serum albumin