Development and validation of an ultra-high performance liquid chromatography tandem mass spectrometry method for quantifying thyreostats in urine without derivatisation

• Published in: Journal of Chromatography A Vol. 1217; no. 26; pp. 4285 - 4293
• Main Authors: Vanden Bussche, J., Vanhaecke, L., Deceuninck, Y., Verheyden, K., Wille, K., Bekaert, K., Le Bizec, B., De Brabander, H.F.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 25.06.2010; Amsterdam; New York: Elsevier; Elsevier
• Subjects: 2002/657/EC; Animal productions; Animals; Antithyroid Agents - urine; Biological and medical sciences; Cattle; Chromatography, High Pressure Liquid - methods; Chromatography, High Pressure Liquid - veterinary; Fundamental and applied biological sciences. Psychology; Sheep; Swine; Tandem Mass Spectrometry - methods; Tandem Mass Spectrometry - veterinary; Terrestrial animal productions; Thiouracil; Thiouracil - urine; Thyreostats; Triple quadrupole mass analyzer; UHPLC; Validation; Vertebrates; Validation; Thyreostats; Triple quadrupole mass analyzer; 2002/657/EC; UHPLC; Thiouracil; Urine; Ultra performance liquid chromatography; Biological fluid; Tandem mass spectrometry; Antithyroid agent; Chemical analysis; Farming animal; Liquid liquid extraction; Coupled method; Solvent extraction; Veterinary medicine; Electrospray; Quadrupole spectrometer; Thiouracil derivatives; European directive; Pyrimidine derivatives; Quantitative analysis
• ISSN: 0021-9673; 1873-3778
• DOI: 10.1016/j.chroma.2010.04.030

Thyreostatic drugs, illegally administrated to livestock for fattening purposes, are banned in the European Union since 1981 (Council Directive 81/602/EC). For monitoring their illegal use, sensitive and specific analytical methods are required. In this study an UHPLC-MS/MS method was described for quantitative analysis of eight thyreostatic drugs in urine, this without a derivatisation step. The sample pretreatment involved a reduction step with dithiothreitol under denaturating conditions at 65 °C, followed by liquid-liquid extraction with ethyl acetate. This analytical procedure was subsequently validated according to the EU criteria (2002/657/EC Decision), resulting in decision limits and detection capabilities ranging between 1.1 and 5.5 μg L −1 and 1.7 and 7.5 μg L −1, respectively. The method obtained for all, xenobiotic thyreostats, a precision (relative standard deviation) lower than 15.5%, and the linearity ranged between 0.982 and 0.999. The performance characteristics fulfill not only the requirements of the EU regarding the provisional minimum required performance limit (100 μg L −1), but also the recommended concentration fixed at 10 μg L −1 in urine set by the Community of Reference Laboratories. Future experiments applying this method should provide the answer to the alleged endogenous status of thiouracil.