A rapid and reliable method has been optimized and established for the analysis of the metabolites from a marine actinomycete by high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (HPLC/QTOF MS/MS). From MS/MS spectra, the product ions of [M + H](+) were recorded to provide abundant structural information of the mother nucleus and peptide moieties. Using the QTOF MS/MS and in-source collision-induced dissociation (in-source CID) techniques, three main metabolites including actinomycin D, actinomycin V and actinomycin I were determined and characterized by elemental compositions of precursor and product ions (<7 ppm).
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