Sub-critical water extraction of thyreostats from bovine muscle followed by liquid chromatography-tandem mass spectrometry

Thyreostats can be used fraudulently to promote a rapid weight increase of breeding animals at low cost. Their severe toxicological effects impose the development of reliable analytical methods to be used in monitoring plans. This work describes an alternative approach to isolate residues of thiouracil, methyl-thiouracil, propyl-thiouracil, phenyl-thiouracil, tapazole and mercaptobenzimidazole from bovine muscle tissue. The developed procedure is based on three steps: <i>i</i>) matrix solid phase dispersion with C₁₈ for the preliminary sample preparation; <i>ii</i>) subcritical water extraction (SWE) at 160°C and 100 bar; <i>iii</i>) clean-up on an Oasis HLB cartridge. The quantitative determination was performed by liquid chromatography-electrospray-tandem mass spectrometry (LC-MS/MS) in dual polarity ionization by using internal standardization. The SWE-LC-MS/MS method was validated according to the identification criteria of the Commission decision 2002/657/EC. The relative recoveries ranged from 72% to 97%; within-lab reproducibility was less than 18%. The decision limit and the detection capability of all analytes were below the recommended concentration, set at 10 µg kg⁻¹, but the validation results demonstrated that this method could only be applied for screening of thiouracil and methyl-thiouracil. Besides the analytical advantages related to the use of water as solvent extraction, the procedure allowed significant removal of lipids, whose detrimental effects on instrumentation and MS sensitivity are well-known.