Identification of copper(II) binding sites in the aminoglycosidic antibiotic Neomycin B

Protonation and copper(II) coordination properties of neomycin B were studied in solution by potentiometry, NMR, UV/Vis, CD, and EPR spectroscopy, XAS and mass spectrometry. Mono- and dinuclear complexes were found depending on the metal-to-ligand molar ratio. Neomycin B anchors CuII ions above pH 5.0 with an NH2 group from ring B. Simultaneously, the second amino group of the same ring and the hydroxyl group of ring A complete the binding set of donors. With an increase in pH the remaining –NH3+ functional groups in the neomycin B molecule are deprotonated without affecting the complexation pattern. However, these groups, particularly the ones located in the D-ring of the antibiotic, may coordinate the second copper(II) ion when the metal is present in excess. We have proved this process with the use of potentiometry, CD and especially mass spectrometry.