Identification of Binding Epitopes of a Fluorinated Compound Bound to Proteins Using ¹H and ¹⁹F NMR Spectroscopy

¹H/¹⁹F NMR-based screening methods were applied to a human serum albumin-fleroxacin complex. Fleroxacin contains three fluorine atoms in a molecule, which is suitable as a model fluorinated compound for NMR analysis with ¹H and ¹⁹F detection. The ¹⁹F{¹H} and ¹H{¹H} saturation transfer difference spectra were acquired and the ¹H/¹⁹F spin-lattice relaxation rates were measured with and without any selective irradiation of protein resonance to identify the binding epitopes of fleroxacin. Because several ¹H signals of fleroxacin resonated close to water, its precise signal intensities were unavailable. The ¹⁹F NMR-based screening methods successfully provide complementary information, indicating its importance in the analysis of fluorinated compounds.