α zol

The analytical standards of ZEN, α-ZOL, and β-ZOL, used for both in vitro and the animal experiment, was obtained from Sigma-Aldrich, CA, USA. Standards were diluted in acetonitrile to obtain working solutions of ZEN, α-ZOL, and β-ZOL for assays with High Performance Liquid Chromatography (HPLC) and Ultra Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC). All analytical standards were stored at ≤−15 °C. The internal standard (IS) --¹³C₁₈-ZEN, enzyme of β-glucuronidase used for hydrolysis of the conjugate, and QuECHERs used for impurities were purchased from Sigma, CA, USA. Chemicals of chloroform and ethyl acetate, used for ZEN extraction, were purchased from Sinopharm Chemical Reagent, Shanghai, China.

DON-biotransforming microorganisms were isolated from soil samples collected from a wheat field at Huazhong Agricultural University in Wuhan, China, where there are frequent FHB epidemics. DON was purified from F. graminearum 52701^{2 (link)} as previously described^{9 (link)}. Standard chemicals such as DON, 3A-DON, NIV, ZEN, α-ZOL, β-ZOL, GO and MG were purchased from SIGMA (St Louis, MO, USA). MM supplemented with DON as a carbon source was used to screen microbial cultures for DON biotransformation capability^{15 (link)}. Nutrient agar (NA) and 10-fold-diluted NA plates were used to isolate single colonies. Wheat (Triticum aestivum) cv. Sumai 3 was used for seedling inoculation.

The standards of AFB₁, AFB₂, AFG₁, AFG₂, OTA, FB₁, FB₂, ENA, ENA₁, ENB, ENB₁, BEA, STG, ZON, α-ZAL, β-ZAL, α-ZOL, β-ZOL, DON, 3-ADON, 15-ADON, DAS, NIV, FUS-X, NEO, T-2, and HT-2 toxins were purchased from Sigma Aldrich. Individual stocks of all analytes were prepared to obtain 20 mg/L in methanol and multianalyte working solutions of 2 mg/L were also used by diluting the individual stock solutions. The multianalyte working standard solution was used for standard calibration curves, matrix-matched calibration curves, and recovery assays. All standards were stored in darkness and kept at −20 °C.

The studied compounds, their molecular formula, exact mass, CAS numbers and structure are summarized in Table 3. Standards of 3+15-ADON, DON, HT-2, OTA, ROQ-C, T-2, ZEA, AFB₁, AFG₁, AFB₂, AFG₂ were obtained from Biopure (Romer Labs, Tulln, Austria); ENN B, ENN B₁, STER, α-ZOL and β-ZOL from Sigma-Aldrich (Saint Louis, MO, USA); fumonisins FB₁ and FB₂ from Acros Organics (New Jersey, USA); MARC A and AND A from Santa Cruz Biotechnology (Dallas, TX, USA); CPA and MPA from Alpha Aesar (Ward Hill, MA, USA); AOH and PEN A from Cayman Chemical (Ann Harbor, MI, USA). Acetonitrile, water and methanol (both > 99.95% purity) were provided by Carlo Erba (Milan, Italy). Formic acid (99%), acetic acid glacial, ammonium formate and ammonium acetate were all LC-MS grade and were provided by Biosolve (Dieuze, France and Valkenswaard, The Netherlands).
A multitoxin maize reference material (QCM7C1, Biopure, Romer Labs, Tulln, Austria) was used for checking the extraction method accuracy.
Individual stock solutions for each mycotoxin were prepared in acetonitrile. Further dilutions and mixtures were prepared in methanol. All solutions were stored at −20 °C.

The mycotoxin standards of ZEN, α-ZOL, β-ZOL, α-ZAL, β-ZAL, and ZAN were purchased from Sigma-Aldrich (St. Louis, MO, USA). The standards of Z14G, α-ZOL14G, and β-ZOL14G were kindly provided by the Laboratory of Food Analysis, Ghent University (Belgium). Methanol and acetonitrile (HPLC-grade) were purchased from Merck (Darmstadt, Germany). Ultrapure water (18.2 MΩ·cm) used in our experiments was obtained from a Milli-Q System (Bedford, MA, USA). Cleanert MC clean-up columns were purchased from Bonna-Agela Technologies (Tianjin, China). Other chemicals were obtained from Aladdin (Shanghai, China).

HPLC grade acetonitrile and methanol were purchased from Merck (Darmstadt, Germany). Phosphate buffered saline(PBS) was purchased from Sigma (St. Louis, Mo, USA). ZEN(α-ZAL, β-ZAL, α-ZOL, β-ZOL, ZAN, ZON) were purchased from Sigma (St. Louis, Mo, USA). As purification IAC, immunoaffinity columns for Zearalenones (IAC-ZER) was purchased from Clover (Irvine, USA). All other organic chemicals and organic solvents were reagent grade or higher.