Journal article
Aggressive dereplication using UHPLC–DAD–QTOF: screening extracts for up to 3000 fungal secondary metabolites
Department of Systems Biology, Technical University of Denmark1
Metabolic Signaling and Regulation, Department of Systems Biology, Technical University of Denmark2
Center for Microbial Biotechnology, Department of Systems Biology, Technical University of Denmark3
Network Engineering of Eukaryotic Cell Factories, Department of Systems Biology, Technical University of Denmark4
Natural Product Chemistry, Department of Systems Biology, Technical University of Denmark5
Metabolomics Platform, Department of Systems Biology, Technical University of Denmark6
In natural-product drug discovery, finding new compounds is the main task, and thus fast dereplication of known compounds is essential. This is usually performed by manual liquid chromatography-ultraviolet (LC-UV) or visible light-mass spectroscopy (Vis-MS) interpretation of detected peaks, often assisted by automated identification of previously identified compounds.
We used a 15 min high-performance liquid chromatography–diode array detection (UHPLC–DAD)–high-resolution MS method (electrospray ionization (ESI)+ or ESI−), followed by 10–60 s of automated data analysis for up to 3000 relevant elemental compositions. By overlaying automatically generated extracted-ion chromatograms from detected compounds on the base peak chromatogram, all major potentially novel peaks could be visualized.
Peaks corresponding to compounds available as reference standards, previously identified compounds, and major contaminants from solvents, media, filters etc. were labeled to differentiate these from compounds only identified by elemental composition. This enabled fast manual evaluation of both known peaks and potential novel-compound peaks, by manual verification of: the adduct pattern, UV–Vis, retention time compared with log D, co-identified biosynthetic related compounds, and elution order.
System performance, including adduct patterns, in-source fragmentation, and ion-cooler bias, was investigated on reference standards, and the overall method was used on extracts of Aspergillus carbonarius and Penicillium melanoconidium, revealing new nitrogen-containing biomarkers for both species.
Language: | English |
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Publisher: | Springer Berlin Heidelberg |
Year: | 2014 |
Pages: | 1933-1943 |
ISSN: | 16182650 and 16182642 |
Types: | Journal article |
DOI: | 10.1007/s00216-013-7582-x |
ORCIDs: | Klitgaard, Andreas , Andersen, Mikael Rørdam , Larsen, Thomas Ostenfeld , Frisvad, Jens Christian and Nielsen, Kristian Fog |
Analytical Chemistry Biochemistry, general Biological Products Characterization and Evaluation of Materials Chemistry Chemistry and Materials Science Chromatography, High Pressure Liquid Databases, Factual Drug Discovery Environmental Monitoring/Analysis Food Science Fungi LC–MS Laboratory Medicine Metabolomics Mycotoxin NRPS Nonribosomal peptide Polyketide Secondary Metabolism Spectrometry, Mass, Electrospray Ionization UPLC