Methanol meoh

All laboratory reagents used in the experiments were of analytical grade. The deionised water (d.i.H₂O) utilised for the experiments was procured from a MilliQ^TM system (Millipore Australia Pty Ltd., Sydney, NSW, Australia). Phen, 5-Mephen, 5,6-Me₂phen, NPX, ACE, N,N′-dicyclohexylcarbodiimide (DCC), dimethyl sulfoxide (DMSO), acetonitrile (CH₃CN) and trifluoroacetic acid (TFA) were purchased from Sigma-Aldrich, Sydney, NSW, Australia. Sep-Pak^® C₁₈-reverse phase columns were purchased from Waters Australia Pty Ltd., Sydney, NSW, Australia. Deuterated oxide 99.9% (D₂O) and deuterated DMSO (DMSO-d₆) were purchased from Cambridge Isotope Laboratories, Andover, MA, USA. Methanol (MeOH) was obtained from Honeywell Research Chemicals, NJ, USA. Diethyl ether (Et₂O) and acetone (C₃H₆O) were purchased from ChemSupply Australia, Gillman, SA, Australia. Additional laboratory reagents were acquired from commercial sources.

DON was obtained from Romer Labs (Tulln, Austria) and CER was purified as previously described [28 (link)]. Cell culture media and supplements were obtained from Gibco® Life Technologies (Karlsruhe, Germany). KH₂PO₄, KCl, NaCl, Na₂HPO₄, D-glucose, HEPES, CaCl₂, MgCl₂, glycin, formaldehyde, sodium dodecyl sulfate (SDS), NaOH and Triton X-100 were purchased from Carl Roth (Karlsruhe, Germany) and Lucifer Yellow CH di-lithium salt from Santa Cruz Technologies (Dallas, TX, USA). For LC-MS/MS measurements, MS grade water was purchased from VWR (Fontenay-sous-Bois, France), acetonitrile (ACN) and methanol (MeOH) from Honeywell (Seelze, Germany). Acetic acid was obtained from Merck (Darmstadt, Germany) and ammonium acetate (both LC-MS grade) from Sigma Aldrich (Vienna, Austria). DON-3-sulfate, DON-15-sulfate and DON-3-glucuronide were kindly provided by Dr. Philipp Fruhmann (Technical University of Vienna) and synthesized as previously described [44 (link)].

All chemicals and reagents were of spectroscopic grade and used without further purification. The deionized water (d.i.H₂O) used for the experiments was obtained from a MilliQ^TM system (Millipore Australia Pty Ltd, Sydney, NSW, Australia). Potassium tetrachloroplatinate(II) (K₂PtCl₄), S,S-DACH, 1,10-phenanthroline, 5-methyl-1,10-phenanthroline, 5,6-dimethyl-1,10-phenanthroline, CLB, N,N′-dicyclohexylcarbodiimide (DCC), dimethyl sulfoxide (DMSO), acetonitrile (CH₃CN), trifluoroacetic acid (TFA), silver nitrate (AgNO₃⁻), ascorbic acid (AsA) and 30% hydrogen peroxide (H₂O₂) were purchased from Sigma-Aldrich, Sydney, NSW, Australia. Sep-Pak^® C₁₈-reverse phase columns were purchased from Waters Australia Pty Ltd, Sydney, NSW, Australia. Deuterated solvents such as deuterium oxide 99.9% (D₂O) and deuterated acetonitrile (CD₃CN) were purchased from Cambridge Isotope Laboratories, Andover, MA, USA. Methanol (MeOH) was obtained from Honeywell Research Chemicals, NJ, USA. Ethanol (EtOH), diethyl ether (Et₂O), ethyl acetate (EtOAc) and acetone were purchased from ChemSupply, Gillman, SA, Australia. All other chemicals, reagents, and lab consumables were purchased from commercial sources.

Certified mycotoxin standards were purchased as follows: AOH from Alternaria sp. (purity >96%), AME from Alternaria alternata (>98%), TeA copper salt from Alternaria alternata (>98%), and TEN from Alternaria tenuis (>95%) were obtained from Sigma-Aldrich (St. Louis, MO, USA); ALT (>98%) was purchased from ChemFaces (Wuhan, China), and ATX- I (>97%) was obtained from Cayman (Ann Arbor, MI, USA). All mycotoxin standards were prepared at 1 mg/mL in acetonitrile (ACN) and kept at −20 °C. HPLC-grade water, ACN, and methanol (MeOH) from Honeywell Burdick and Jackson (Muskegon, MI, USA) were used for sample preparation. LC/MS-grade water and methanol from Fisher Scientific (Cleveland, OH, USA) were used as the mobile phase. LC/MS-grade ammonium acetate was purchased from Merck (Darmstadt, Germany) and glacial acetic acid (≥99.7%, HPLC grade) for pH adjustment was obtained from Fisher Scientific (Cleveland, OH, USA).

Water was purchased from Th. Geyer (Renningen, Germany), methanol (MeOH) from Honeywell Riedel-de Haën (Seelze, Germany), and acetonitrile (ACN) and the ammonia solution (25%) were purchased from VWR (Ismaning, Germany), all at least in analytical grade. Reference standards for TEN, ALT, and TeA were bought from Merck (Darmstadt, Germany), while AOH, AME, ATX I, ATX II, STTX III, ALTP, AOH-3-G, and AOH-9-G, AME-3-G as well as AOH-3-S and AME-3-S were either isolated out of fungal extracts or synthesized at our chair as described previously (Liu and Rychlik 2015 (link); Scheibenzuber et al. 2020 (link), 2021 (link)). The stable isotope–labelled standards [²H₄]-AOH, [²H₄]-AME, and [¹³C₆, ¹⁵ N]-TeA were synthesized in our laboratory as reported in literature (Asam et al. 2009 (link), 2011a (link)).
Fig. 1

Chemical structures of the six most frequent Alternaria mycotoxins AOH, AME, TeA, TEN, ALTP, and ATX I