Fast-atom-bombardment Tandem Mass-spectrometric Analysis of Hydroxamate Siderophores

This paper reports a study of the fast atom bombardment (FAB) tandem mass spectrometric analysis of six trihydroxamate siderophores as the free ligand or as iron complexes in several matrixes. It demonstrates that the [M + H - 53]+ and [M + H - 16]+ ions always detected in the FAB spectra are not fragments, but for the first the free ligand resulting from a decomplexation in the matrix, and for the second a reduction product. The tandem mass spectrometric study of the fragmentation shows that the main fragments result from the break of either peptide or hydroxamate C-N bonds, or (when present) of an ester linkage. When the iron is present in the complex, the heterolytic break of the hydroxamate-iron bond creates a positive charge on the iron, and a negative charge on the hydroxamate group. This causes positive fragments of the free ligand to become corresponding neutrals in the complex, and inversely, when a hydroxamate bond is broken. As the free ligand pseudo-molecular ion is always present in the FAB spectra of the complex, this feature is useful for structural elucidation. Iron has an Fe-54 isotope representing 6.35% of the Fe-56 isotope. This allows identification of iron complexes by a comparison of the daughter spectra of both isotopic pseudo-molecualr ions. As most of the positive fragments in the daughter spectra contain iron, they are shifted two mass units lower by the presence of Fe-54. No fragments or neutral losses typical of all the siderophores were identified, and thus there is no selective scan mode for this compound class. Unambiguous identification of the pseudo-molecular ion of siderophore iron complexes can be achieved by the presence in the FAB spectrum of the free ligand, the reduction product and the iron isotopic peak. Clear confirmation can be obtained by a comparison of the daughter spectra of these ions.