INVESTIGADORES
CABRERA Gabriela Myriam
congresos y reuniones científicas
Título:
Secondary metabolite profile of Fusarium oxysporum by LCMS using QTOF, different sources, ion modes and metal solution post-column addition
Autor/es:
ADRIANA M CIRIGLIANO; B. V. BERTINETTI; M. ALEJANDRA RODRÍGUEZ; A. GODEAS; GABRIELA M. CABRERA
Reunión:
Conferencia; 19th International Mass Spectrometry Conference; 2012
Resumen:
Metabolite profiling is crucial for many aspects of fundamental research and industrial applications of filamentous fungi and other microorganisms. The use of a quadrupole - time-of-flight mass spectrometer has proved to be very suitable for this purpose due to its mass high resolution capability, and ESI is the most widely used ionization method in both positive and negative ion modes. In this work, we report the metabolite profiling using HPLC-MS in a QTOF, of a strain of Fusarium oxysporum that has significant antifungal activity against pythopathogens. After optimization of LC conditions such as different solvent systems and phase columns( C18, C8, PFP,HILIC) and in order to explore the whole universe of chemical structures that may be present in the extract, different sources were employed like ESI, APCI and APPI in both positive and negative ion modes. C18 chromatographic phase was selected for the analysis of the mycelium extract, PFP for medium analysis and HCOOH 0.1 % -MeOH as solvent system in both cases. A complex metabolite pattern was observed in all LCMS runs. When ESI was employed, a greater number of metabolites were observed compared to APCI and APPI runs, while APCI and APPI gave similar results. Using the obtained molecular formulas and MS/MS data, a search in different libraries and literature 1 was performed allowing the identification of seven cyclosporines, sansalvamide, N-methyl sansalvamide, a related cyclodepsipeptide not previously described, fusarubin and related compounds. APCI and APPI resulted more sensitive for cyclic peptides, and MS/MS analysis of [M+H] + was especially useful for these compounds, which give predominantly sodiated forms in the ESI LCMS runs. The use of negative ion mode allowed the identification of some compounds such as the antibiotic F244, where MS/MS fragments were very informative for its identification. Many of the identified structures were confirmed by NMR spectroscopy after the isolation of the compounds. Based on our previous results in the application of ESI-LCMS with post-column addition of metal solutions for the differentiation of isomers, as in the case of hydroxypyridine N-oxides 2 and dihydroxyarenes 3 , we used this methodology to obtain further information about the metabolites produced by this fungus. The use of metal complexation by post-column addition of a solution of Ca( II), Cu( II) or Zn( II) allowed the differentiation of isomeric Cyclosporine A from Isocyclosporine A, which are known to be indistinguishable by MS/MS 4 . In the MS spectra, complexes of cyclosporine A with the corresponding divalent metal were observed, whereas complexes were not observed for isocyclosporine A. In conclusion, the use of different sources, ion modes and post-column addition of metal solutions allowed the complete analysis of the extracts of this fungal strain, providing complementary information.