Combining UHPLC-High Resolution MS and Feeding of Stable Isotope Labeled Polyketide Intermediates for Linking Precursors to End Products

Andreas Klitgaard, Rasmus John Normand Frandsen, Dorte Koefoed Holm, Peter Boldsen Knudsen, Jens Christian Frisvad, Kristian Fog Nielsen

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    Abstract

    We present the results from stable isotope labeled precursor feeding studies combined with ultrahigh performance liquid chromatography-high resolution mass spectrometry for the identification of labeled polyketide (PK) end-products. Feeding experiments were performed with 13C8-6-methylsalicylic acid (6-MSA) and 13C14-YWA1, both produced in-house, as well as commercial 13C7-benzoic acid and 2H7-cinnamic acid, in species of Fusarium, Byssochlamys, Aspergillus, and Penicillium. Incorporation of 6-MSA into terreic acid or patulin was not observed in any of six evaluated species covering three genera, because the 6-MSA was shunted into (2Z,4E)-2-methyl-2,4-hexadienedioic acid. This indicates that patulin and terreic acid may be produced in a closed compartment of the cell and that (2Z,4E)-2-methyl-2,4-hexadienedioic acid is a detoxification product toward terreic acid and patulin. In Fusarium spp., YWA1 was shown to be incorporated into aurofusarin, rubrofusarin, and antibiotic Y. In A. niger, benzoic acid was shown to be incorporated into asperrubrol. Incorporation levels of 0.7–20% into the end-products were detected in wild-type strains. Thus, stable isotope labeling is a promising technique for investigation of polyketide biosynthesis and possible compartmentalization of toxic metabolites.
    Original languageEnglish
    JournalJournal of Natural Products
    Volume78
    Issue number7
    Pages (from-to)1518-1525
    Number of pages8
    ISSN0163-3864
    DOIs
    Publication statusPublished - 2015

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