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Abstract

A liquid chromatography-electrospray ionization-mass spectrometry method was developed to quantitatively and qualitatively analyze thirteen hydroxamate siderophores (ferrichrome, ferrirubin, ferrirhodin, ferrichrysin, ferricrocin, ferrioxamine B, D₁, E and G, neocoprogen I and II, coprogen and triacetylfusarinine C). Samples were pre-concentrated on-line via switch-valve setup prior to analyte separation on a Kinetex C18 column. Gradient elution was performed using a mixture of an ammonium formate buffer and acetonitrile. Total analysis time including column conditioning was 20.5 minutes. Analyte were fragmented by applying collision induced dissociation, enabling structural identification via tandem mass spectrometry. Limit of detection values for the selected ion monitoring method ranged from 71 pM to 1.5 nM with corresponding values of two to nine times higher for the multiple reaction monitoring method. The liquid chromatography-mass spectrometry method resulted in a robust and sensitive quantification of hydroxamate siderophores as indicated by retention time stability, linearity, sensitivity, precision and recovery. The analytical error of the methods, assessed trough random-order, duplicate analysis of soil samples extracted with a mixture of 10 mM phosphate buffer and methanol, appears negligible in relation to between-sample variations.

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