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Development and in-house validation of an LC-MS/MS method for the determination of stilbenes and resorcylic acid lactones in bovine urine

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Abstract

Stilbenes and zeranol are nonsteroidal estrogenic growth promoters which are banned in the European Union (EU) for use in food-producing animals by Council Directive 96/22/EC. A liquid chromatography–tandem mass spectrometry (LC-MS/MS) method was developed for the screening and confirmation of stilbenes (diethylstilbestrol, dienestrol, hexestrol) and resorcylic acid lactones (zeranol and its metabolites taleranol and zearalanone as well as the mycotoxins α-zearalenol, β-zearalenol and zearalenone) in bovine urine. The method permits the confirmation and quantification of stilbenes and resorcylic acid lactones at levels below 1 µg L−1 and 1.5 µg L−1, respectively. The validation was carried out according to Commission Decision 2002/657/EC, Chap. 3.1.3 “alternative validation” by a matrix-comprehensive in-house validation concept. Decision limit CCα, detection capability CCβ, recovery, repeatabiliy, within-laboratory reproducibility and the uncertainty of measurement were calculated. Furthermore, a factorial effect analysis was carried out to identify factors that have a significant influence on the method. Factors considered to be relevant for the method in routine analysis (e.g. operator, matrix condition, storage duration of the extracts before measurement, different cartridge lots, hydrolysis conditions) were systematically varied on two levels. The factorial analysis showed that different cartridge lots, storage durations and matrix conditions can exert a relevant influence on the method.

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Acknowledgements

The authors wish to thank Mrs. M. Grunow and Mr. S. Zimmermann for their technical assistance in the in-house validation study. Dr. Frank Hamann is acknowledged for making available the samples of bovine urine. Dr. Steffen Uhlig is acknowledged for helpful discussions on statistical issues.

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Authors and Affiliations

  1. National Reference Laboratory for Residues Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832, Berlin, Germany

    Kathrin Schmidt, Carolin Stachel & Petra Gowik

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  1. Kathrin Schmidt
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  2. Carolin Stachel
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  3. Petra Gowik
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Correspondence to Petra Gowik.

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Schmidt, K., Stachel, C. & Gowik, P. Development and in-house validation of an LC-MS/MS method for the determination of stilbenes and resorcylic acid lactones in bovine urine. Anal Bioanal Chem 391, 1199–1210 (2008). https://doi.org/10.1007/s00216-008-1943-x

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  • DOI: https://doi.org/10.1007/s00216-008-1943-x

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