Interaction of Febuxostat with Human Serum Albumin and Its Application as a Circular Dichroism Probe for Monitoring Albumin Conformational Changes.

Febuxostat is a nonpurine selective xanthine oxidase inhibitor used chronically, often in combination with other medications, to treat hyperuricemia and gout. Its high plasma protein binding necessitates a thorough understanding of its interactions with human serum albumin (HSA) in order to assess potential protein-binding-mediated drug interactions. In this study, the interaction between febuxostat and HSA was investigated using equilibrium dialysis and circular dichroism (CD) spectroscopy. Equilibrium dialysis indicated that approximately six febuxostat molecules bind to one HSA molecule. Febuxostat induced Cotton effects in the presence of HSA, and the febuxostat concentration-dependent change in CD spectra supported the involvement of multiple binding regions with distinct microenvironments on HSA. Febuxostat caused only minor displacement of site-specific ligands, suggesting a limited impact on protein-binding-mediated drug interactions. Additionally, while fatty acids and pH did not significantly affect the free fraction of febuxostat, myristate and pH markedly altered the induced CD spectra, indicating sensitivity to conformational and microenvironmental changes in HSA. These findings further demonstrate that the febuxostat-HSA-induced CD signal provides a practical probe for monitoring albumin conformational dynamics.