Tracing the metabolism of HT-2 toxin and T-2 toxin in barley by isotope-assisted untargeted screening and quantitative LC-HRMS analysis.
An extensive study of the metabolism of the type A trichothecene mycotoxins HT-2 toxin and T-2 toxin in barley using liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS) is reported. A recently developed untargeted approach based on stable isotopic labelling, LC-Orbitrap-MS analysis with fast polarity switching and data processing by MetExtract software was combined with targeted LC-Q-TOF-MS(/MS) analysis for metabolite structure elucidation and quantification. In total, 9 HT-2 toxin and 13 T-2 toxin metabolites plus tentative isomers were detected, which were successfully annotated by calculation of elemental formulas and further LC-HRMS/MS measurements as well as partly identified with authentic standards. As a result, glucosylated forms of the toxins, malonylglucosides, and acetyl and feruloyl conjugates were elucidated. Additionally, time courses of metabolite formation and mass balances were established. For absolute quantification of those compounds for which standards were available, the method was validated by determining apparent recovery, signal suppression, or enhancement and extraction recovery. Most importantly, T-2 toxin was rapidly metabolised to HT-2 toxin and for both parent toxins HT-2 toxin-3-O-β-glucoside was identified (confirmed by authentic standard) as the main metabolite, which reached its maximum already 1 day after toxin treatment. Graphical Abstract Isotope-assisted untargeted screening of HT-2 toxin and T-2 toxin metabolites in barley.
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.|Institute for Biotechnology in Plant Production, IFA-Tulln, BOKU, Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Jacqueline
J
Meng-Reiterer
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.|Christian Doppler Laboratory for Mycotoxin Metabolism, IFA-Tulln, BOKU, Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Elisabeth
E
Varga
Chemistry and Toxicology Unit, Research and Laboratory Department, Finnish Food Safety Authority (Evira), Mustialankatu 3, 00790, Helsinki, Finland.
Alexis V
AV
Nathanail
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Christoph
C
Bueschl
University of Applied Sciences Wr. Neustadt, Degree Programme Biotechnical Processes (FHWN-Tulln), Konrad-Lorenz-Str. 10, 3430, Tulln, Austria.
Justyna
J
Rechthaler
Bacterial Foodborne Pathogens and Mycology Research Unit, National Center for Agricultural Utilization Research, U.S. Department of Agriculture, 1815 N. University Street, Peoria, IL, 61604, USA.
Susan P
SP
McCormick
Department of Applied Genetics and Cell Biology, BOKU, Konrad-Lorenz-Str. 24, 3430, Tulln, Austria.
Herbert
H
Michlmayr
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.|Christian Doppler Laboratory for Mycotoxin Metabolism, IFA-Tulln, BOKU, Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Alexandra
A
Malachová
Department of Applied Genetics and Cell Biology, BOKU, Konrad-Lorenz-Str. 24, 3430, Tulln, Austria.|Institute of Applied Synthetic Chemistry, Vienna University of Technology, Getreidemarkt 9/163, 1060, Vienna, Austria.
Philipp
P
Fruhmann
Department of Applied Genetics and Cell Biology, BOKU, Konrad-Lorenz-Str. 24, 3430, Tulln, Austria.
Gerhard
G
Adam
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.|Christian Doppler Laboratory for Mycotoxin Metabolism, IFA-Tulln, BOKU, Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Franz
F
Berthiller
Institute for Biotechnology in Plant Production, IFA-Tulln, BOKU, Konrad-Lorenz-Str. 20, 3430, Tulln, Austria.
Marc
M
Lemmens
Center for Analytical Chemistry, Department of Agrobiotechnology (IFA-Tulln), University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad-Lorenz-Str. 20, 3430, Tulln, Austria. rainer.schuhmacher@boku.ac.at.
Rainer
R
Schuhmacher
D002853:Chromatography, Liquid / Q000379:methods; D005670:Fusarium / Q000378:metabolism*; D001467:Hordeum / Q000378:metabolism* / Q000382:microbiology*; D013605:T-2 Toxin / Q000031:analogs & derivatives* / Q000378:metabolism*; D053719:Tandem Mass Spectrometry / Q000379:methods
D013605:T-2 Toxin; C012351:HT-2 toxin
Fast polarity switching; Masked mycotoxins; Stable isotopic labelling; Type A trichothecenes; Untargeted metabolite profiling