Abstract
In drug discovery, reliable and fast dereplication of known compounds is essential for identification of novel bioactive compounds. Here, we show an integrated approach using ultra-high performance liquid chromatography-diode array detection-quadrupole time of flight mass spectrometry (UHPLC-DAD-QTOFMS) providing both accurate mass full-scan mass spectrometry (MS) and tandem high resolution MS (MS/HRMS) data. The methodology was demonstrated on compounds from bioactive marine-derived strains of Aspergillus, Penicillium, and Emericellopsis, including small polyketides, non-ribosomal peptides, terpenes, and meroterpenoids. The MS/HRMS data were then searched against an in-house MS/HRMS library of ~1300 compounds for unambiguous identification. The full scan MS data was used for dereplication of compounds not in the MS/HRMS library, combined with ultraviolet/visual (UV/Vis) and MS/HRMS data for faster exclusion of database search results. This led to the identification of four novel isomers of the known anticancer compound, asperphenamate. Except for very low intensity peaks, no false negatives were found using the MS/HRMS approach, which proved to be robust against poor data quality caused by system overload or loss of lock-mass. Only for small polyketides, like patulin, were both retention time and UV/Vis spectra necessary for unambiguous identification. For the ophiobolin family with many structurally similar analogues partly co-eluting, the peaks could be assigned correctly by combining MS/HRMS data and m/z of the [M + Na]+ ions.
Highlights
Due to the cosmopolitan occurrence of many bioactive compounds, most natural product extracts contain compounds that have previously been characterized, despite intelligent selection of new organisms
Fifteen marine-derived strains from different species belonging to Penicillium, Aspergillus, and Emericellopsis were fractionated and screened for their anti-microbial [48], anti-inflammatory [49], central nervous system (CNS) [50], and anticancer activity, that resulted in 35 active fractions to be evaluated for their chemistry
In this work we demonstrate that mass spectrometry (MS)/HRMS search in a library is a robust and reliable way of tentatively identifying known bioactive compounds on a single instrument
Summary
Due to the cosmopolitan occurrence of many bioactive compounds, most natural product extracts contain compounds that have previously been characterized, despite intelligent selection of new organisms. Back integration/deconvolution of raw data to find corresponding full scan data and linking MS/MS spectra of adducts belonging to the same molecular feature as well as retention time still needs to be done manually and is very time consuming In this current study, we demonstrate the use of our MS/HRMS library search to dereplicate known compounds in bioactive extracts from marine-derived Aspergillus, Penicillium, and Emericellopsis strains. Ultra-high performance liquid chromatography-diode array detection-quadrupole time of flight mass spectrometry (UHPLC-DAD-HRMS) with auto- tandem high resolution mass spectrometry (MS/HRMS) analysis was used to screen the extracts and subsequently, MS/HRMS data was matched against a newly constructed library of 1300 compounds (10, 20, and 40 eV spectra) using the Agilent search algorithm. Comparison with UV/Vis detection was done for a number of poorly ionizing compounds showing the value of this additional cheap detector
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