title: Combining UHPLC-High Resolution MS and Feeding of Stable Isotope Labeled Polyketide Intermediates for Linking Precursors to End Products.
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 (13)C8-6-methylsalicylic acid (6-MSA) and (13)C14-YWA1, both produced in-house, as well as commercial (13)C7-benzoic acid and (2)H7-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.
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Andreas
authors.initials: A
authors.lastname: Klitgaard
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Rasmus J N
authors.initials: RJ
authors.lastname: Frandsen
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Dorte K
authors.initials: DK
authors.lastname: Holm
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Peter B
authors.initials: PB
authors.lastname: Knudsen
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Jens C
authors.initials: JC
authors.lastname: Frisvad
authors.affiliation: Department of Systems Biology, Technical University of Denmark, DK-2800 Kongens Lyngby, Denmark.
authors.forename: Kristian F
authors.initials: KF
authors.lastname: Nielsen
mesh_terms: D000465:Algorithms; D001230:Aspergillus / Q000737:chemistry; D005670:Fusarium / Q000737:chemistry; D007553:Isotope Labeling; D015394:Molecular Structure; D010365:Patulin / Q000737:chemistry; D061065:Polyketides / Q000737:chemistry*; D011753:Pyrones / Q000737:chemistry; D011809:Quinones / Q000737:chemistry; D012459:Salicylates / Q000737:chemistry
chemical_list: D061065:Polyketides; D011753:Pyrones; D011809:Quinones; D012459:Salicylates; C048670:antibiotic Y; C015497:rubrofusarin; D010365:Patulin; C005236:6-methylsalicylic acid; C009400:terreic acid