Study of the gastrointestinal biotransformation of zearalenone in a Caco-2 cell culture system with liquid chromatographic methods

A high‐performance liquid chromatography method with fluorescence detection (HPLC‐FLD) was developed and validated for the detection of zearalenone (ZON), α‐zearalenol (α‐ZOL) and β‐zearalenol (β‐ZOL) in in vitro biological samples. Furthermore, a liquid chromatography–tandem mass spectrometry (LC‐M...

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Veröffentlicht in:Journal of applied toxicology 2008-11, Vol.28 (8), p.966-973
Hauptverfasser: Schaut, A., De Saeger, S., Sergent, T., Schneider, Y-J., Larondelle, Y., Pussemier, L., Van Peteghem, C.
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container_issue 8
container_start_page 966
container_title Journal of applied toxicology
container_volume 28
creator Schaut, A.
De Saeger, S.
Sergent, T.
Schneider, Y-J.
Larondelle, Y.
Pussemier, L.
Van Peteghem, C.
description A high‐performance liquid chromatography method with fluorescence detection (HPLC‐FLD) was developed and validated for the detection of zearalenone (ZON), α‐zearalenol (α‐ZOL) and β‐zearalenol (β‐ZOL) in in vitro biological samples. Furthermore, a liquid chromatography–tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the detection of ZON, α‐ZOL, β‐ZOL, α‐zearalanol (α‐ZAL) and β‐zearalanol (β‐ZAL) in in vitro biological samples. Zearalanone (ZAN) was used as internal standard in both methods. The limit of detection/limit of quantitation (LOD/LOQ) values for ZON, α‐ZOL and β‐ZOL were 2/7, 2/7 and 4/13 µg l−1, respectively, for the HPLC‐FLD method. For the LC‐MS/MS method LOD/LOQ values for ZON, α‐ZOL, β‐ZOL, α‐ZAL and β‐ZAL were 6/20, 5/17, 4/14, 9/30 and 6/19 µg l−1, respectively. Within‐day and between‐day precision were less then 11 and 14%, respectively for the HPLC‐FLD method, and both less then 20% for the LC‐MS/MS method. The recovery of ZON and its metabolites ranged between 73 and 89% for the HPLC‐FLD method and between 69 and 112% for the LC‐MS/MS method. The methods were used for the detection of the compounds in in vitro biological samples, obtained with human intestinal Caco‐2 cells culture experiments. The 8‐days post‐confluent Caco‐2 cells were treated with ZON or a mixture of ZON and imazalil (IMA). After an incubation time of 24 h the samples were analysed with the HPLC‐FLD method. Neither ZON nor its derivatives were detected in the samples. The disappearance of ZON could possibly point out the formation of phase II metabolites like glucuronide conjugates. Therefore, samples were pretreated with β‐glucuronidase before LC‐MS/MS analysis. The LC‐MS/MS results showed that ZON, α‐ZOL and β‐ZOL could only be detected in the β‐glucuronidase pretreated samples. This confirmed the formation of glucuronide conjugates and the hydroxylation of ZON during the incubation with Caco‐2 cells. Copyright © 2008 John Wiley & Sons, Ltd.
doi_str_mv 10.1002/jat.1362
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Furthermore, a liquid chromatography–tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the detection of ZON, α‐ZOL, β‐ZOL, α‐zearalanol (α‐ZAL) and β‐zearalanol (β‐ZAL) in in vitro biological samples. Zearalanone (ZAN) was used as internal standard in both methods. The limit of detection/limit of quantitation (LOD/LOQ) values for ZON, α‐ZOL and β‐ZOL were 2/7, 2/7 and 4/13 µg l−1, respectively, for the HPLC‐FLD method. For the LC‐MS/MS method LOD/LOQ values for ZON, α‐ZOL, β‐ZOL, α‐ZAL and β‐ZAL were 6/20, 5/17, 4/14, 9/30 and 6/19 µg l−1, respectively. Within‐day and between‐day precision were less then 11 and 14%, respectively for the HPLC‐FLD method, and both less then 20% for the LC‐MS/MS method. The recovery of ZON and its metabolites ranged between 73 and 89% for the HPLC‐FLD method and between 69 and 112% for the LC‐MS/MS method. The methods were used for the detection of the compounds in in vitro biological samples, obtained with human intestinal Caco‐2 cells culture experiments. The 8‐days post‐confluent Caco‐2 cells were treated with ZON or a mixture of ZON and imazalil (IMA). After an incubation time of 24 h the samples were analysed with the HPLC‐FLD method. Neither ZON nor its derivatives were detected in the samples. The disappearance of ZON could possibly point out the formation of phase II metabolites like glucuronide conjugates. Therefore, samples were pretreated with β‐glucuronidase before LC‐MS/MS analysis. The LC‐MS/MS results showed that ZON, α‐ZOL and β‐ZOL could only be detected in the β‐glucuronidase pretreated samples. This confirmed the formation of glucuronide conjugates and the hydroxylation of ZON during the incubation with Caco‐2 cells. 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Appl. Toxicol</addtitle><description>A high‐performance liquid chromatography method with fluorescence detection (HPLC‐FLD) was developed and validated for the detection of zearalenone (ZON), α‐zearalenol (α‐ZOL) and β‐zearalenol (β‐ZOL) in in vitro biological samples. Furthermore, a liquid chromatography–tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the detection of ZON, α‐ZOL, β‐ZOL, α‐zearalanol (α‐ZAL) and β‐zearalanol (β‐ZAL) in in vitro biological samples. Zearalanone (ZAN) was used as internal standard in both methods. The limit of detection/limit of quantitation (LOD/LOQ) values for ZON, α‐ZOL and β‐ZOL were 2/7, 2/7 and 4/13 µg l−1, respectively, for the HPLC‐FLD method. For the LC‐MS/MS method LOD/LOQ values for ZON, α‐ZOL, β‐ZOL, α‐ZAL and β‐ZAL were 6/20, 5/17, 4/14, 9/30 and 6/19 µg l−1, respectively. Within‐day and between‐day precision were less then 11 and 14%, respectively for the HPLC‐FLD method, and both less then 20% for the LC‐MS/MS method. The recovery of ZON and its metabolites ranged between 73 and 89% for the HPLC‐FLD method and between 69 and 112% for the LC‐MS/MS method. The methods were used for the detection of the compounds in in vitro biological samples, obtained with human intestinal Caco‐2 cells culture experiments. The 8‐days post‐confluent Caco‐2 cells were treated with ZON or a mixture of ZON and imazalil (IMA). After an incubation time of 24 h the samples were analysed with the HPLC‐FLD method. Neither ZON nor its derivatives were detected in the samples. The disappearance of ZON could possibly point out the formation of phase II metabolites like glucuronide conjugates. Therefore, samples were pretreated with β‐glucuronidase before LC‐MS/MS analysis. The LC‐MS/MS results showed that ZON, α‐ZOL and β‐ZOL could only be detected in the β‐glucuronidase pretreated samples. This confirmed the formation of glucuronide conjugates and the hydroxylation of ZON during the incubation with Caco‐2 cells. 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Appl. Toxicol</addtitle><date>2008-11</date><risdate>2008</risdate><volume>28</volume><issue>8</issue><spage>966</spage><epage>973</epage><pages>966-973</pages><issn>0260-437X</issn><eissn>1099-1263</eissn><coden>JJATDK</coden><abstract>A high‐performance liquid chromatography method with fluorescence detection (HPLC‐FLD) was developed and validated for the detection of zearalenone (ZON), α‐zearalenol (α‐ZOL) and β‐zearalenol (β‐ZOL) in in vitro biological samples. Furthermore, a liquid chromatography–tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the detection of ZON, α‐ZOL, β‐ZOL, α‐zearalanol (α‐ZAL) and β‐zearalanol (β‐ZAL) in in vitro biological samples. Zearalanone (ZAN) was used as internal standard in both methods. The limit of detection/limit of quantitation (LOD/LOQ) values for ZON, α‐ZOL and β‐ZOL were 2/7, 2/7 and 4/13 µg l−1, respectively, for the HPLC‐FLD method. For the LC‐MS/MS method LOD/LOQ values for ZON, α‐ZOL, β‐ZOL, α‐ZAL and β‐ZAL were 6/20, 5/17, 4/14, 9/30 and 6/19 µg l−1, respectively. Within‐day and between‐day precision were less then 11 and 14%, respectively for the HPLC‐FLD method, and both less then 20% for the LC‐MS/MS method. The recovery of ZON and its metabolites ranged between 73 and 89% for the HPLC‐FLD method and between 69 and 112% for the LC‐MS/MS method. The methods were used for the detection of the compounds in in vitro biological samples, obtained with human intestinal Caco‐2 cells culture experiments. The 8‐days post‐confluent Caco‐2 cells were treated with ZON or a mixture of ZON and imazalil (IMA). After an incubation time of 24 h the samples were analysed with the HPLC‐FLD method. Neither ZON nor its derivatives were detected in the samples. The disappearance of ZON could possibly point out the formation of phase II metabolites like glucuronide conjugates. Therefore, samples were pretreated with β‐glucuronidase before LC‐MS/MS analysis. The LC‐MS/MS results showed that ZON, α‐ZOL and β‐ZOL could only be detected in the β‐glucuronidase pretreated samples. This confirmed the formation of glucuronide conjugates and the hydroxylation of ZON during the incubation with Caco‐2 cells. Copyright © 2008 John Wiley &amp; Sons, Ltd.</abstract><cop>Chichester, UK</cop><pub>John Wiley &amp; Sons, Ltd</pub><pmid>18548745</pmid><doi>10.1002/jat.1362</doi><tpages>8</tpages></addata></record>
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subjects analytical methodology
Biological and medical sciences
Biotransformation
Caco-2
Caco-2 Cells
Chromatography, High Pressure Liquid
Estrogens, Non-Steroidal - pharmacokinetics
Fungicides, Industrial - pharmacology
Gastrointestinal Tract - metabolism
glucuronidation
Glucuronides - metabolism
Humans
imazalil
Imidazoles - pharmacology
Indicators and Reagents
Medical sciences
Plant poisons toxicology
Reference Standards
Reproducibility of Results
Spectrometry, Fluorescence
Tandem Mass Spectrometry
Toxicology
zearalenone
Zearalenone - pharmacokinetics
title Study of the gastrointestinal biotransformation of zearalenone in a Caco-2 cell culture system with liquid chromatographic methods
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