Stachybotrys mycotoxins: from culture extracts to dust samples.

The filamentous fungus Stachybotrys chartarum is known for its toxic metabolites and has been associated with serious health problems, including mycotoxicosis, among occupants of contaminated buildings. Here, we present results from a case study, where an ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed for known and tentatively identified compounds characterized via UHPLC-quadruple time-of-flight (QTOF) screening of fungal culture extracts, wall scrapings and reference standards. The UHPLC-MS/MS method was able to identify 12 Stachybotrys metabolites, of which four could be quantified based on authentic standards and a further six estimated based on similarity to authentic standards. Samples collected from walls contaminated by S. chartarum in a water-damaged building showed that the two known chemotypes, S and A, coexisted. More importantly, a link between mycotoxin concentrations found on contaminated surfaces and in settled dust was made. One dust sample, collected from a water-damaged room, contained 10 pg/cm(2) macrocyclic trichothecenes (roridin E). For the first time, more than one spirocyclic drimane was detected in dust. Spirocyclic drimanes were detected in all 11 analysed dust samples and in total amounted to 600 pg/cm(2) in the water-damaged room and 340 pg/cm(2) in rooms adjacent to the water-damaged area. Their wide distribution in detectable amounts in dust suggested they could be good candidates for exposure biomarkers. Graphical abstract Stachybotrys growing on a gypsum board, and some of the compounds it produces.

Accurate dereplication of bioactive secondary metabolites from marine-derived fungi by UHPLC-DAD-QTOFMS and a MS/HRMS library.

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.