Development and validation of a liquid chromatography-tandem mass spectrometry method for the simultaneous determination of nine corticosteroid residues in bovine liver samples.

A liquid chromatography tandem mass spectrometry (LC-MS/MS) confirmatory method for the simultaneous determination of nine corticosteroids in liver, including the four MRL compounds listed in Council Regulation 37/2010, was developed. After an enzymatic deconjugation and a solvent extraction of the liver tissue, the resulting solution was cleaned up through an SPE Oasis HLB cartridge. The analytes were then detected by liquid chromatography-negative-ion electrospray tandem mass spectrometry, using deuterium-labelled internal standards. The procedure was validated as a quantitative confirmatory method according to the Commission Decision 2002/657/EC criteria. The results showed that the method was suitable for statutory residue testing regarding the following performance characteristics: instrumental linearity, specificity, precision (repeatability and intra-laboratory reproducibility), recovery, decision limit (CCα), detection capability (CCß) and ruggedness. All the corticosteroids can be detected at a concentration around 1 µg kg(-1); the recoveries were above 62% for all the analytes. Repeatability and reproducibility (within-laboratory reproducibility) for all the analytes were below 7.65% and 15.5%, respectively.

Confirmatory method for the determination of nandrolone and trenbolone in urine samples using immunoaffinity cleanup and liquid chromatography-tandem mass spectrometry.

A confirmatory method for the simultaneous determination of nandrolone (alpha and beta) and trenbolone (alpha and beta) in urine samples by liquid chromatography electrospray mass spectrometry (LC-MS-MS) was developed. After an enzymatic deconjugation, the urine was subjected to a one-step cleanup on a commercially available immunoaffinity chromatography cartridge. The analytes were detected by liquid chromatography-positive ion electrospray tandem mass spectrometry using deuterium labelled internal standards. The analytical procedure was applicable to bovine and swine urine samples. The procedure was validated as a quantitative confirmatory method according to the Commission Decision 2002/657/EC criteria. The results obtained showed that the method was suitable for statutory residues testing regarding the following performance characteristics: instrumental linearity, specificity, precision (repeatability and intra-laboratory reproducibility), recovery, decision limit (CCalpha), detection capability (CCbeta) and ruggedness. The decision limits (CCalpha) obtained, were between 0.54 and 0.60 microg L(-1); the recovery was above 64% for all the analytes. Repeatability was between 1.6% and 5.7% and within-laboratory reproducibility between 1.6% and 6.0% for all the steroids.

Confirmatory method for the determination of resorcylic acid lactones in urine sample using immunoaffinity cleanup and liquid chromatography-tandem mass spectrometry.

The presence of zeranol (alpha-zearalanol) in urine samples due to natural contamination or illegal treatment is under debate within the European Union. The simultaneous determination of zeranol, its epimer taleranol (beta-zearalanol), zearalanone and the structurally related mycotoxin zearalenone with the corresponding alpha- and beta-zearalenol metabolites appears to be critical in deciding whether an illegal use has occurred. The aim of this study is to develop and validate a simple analytical procedure applicable to bovine and swine urine samples for the determination of all six resorcylic acid lactones. After an enzymatic deconjugation, the urine was subjected to a one-step cleanup on a commercially available immunoaffinity chromatography cartridge. The analytes were detected by liquid chromatography-negative-ion electrospray tandem mass spectrometry using deuterium-labelled internal standards. The method was validated as a quantitative confirmatory method according to European Commission Decision 2002/657/EC. The evaluated parameters were: linearity, specificity, precision (repeatability and intra-laboratory reproducibility), recovery, decision limit, detection capability and ruggedness. The decision limits (CCalpha) obtained, were between 0.56 and 0.68 microgL(-1); recovery above 66% for all the analytes. Repeatability was between 1.4 and 5.3% and within-laboratory reproducibility between 1.9 and 16.1% for the six resorcylic acid lactones.

Development and validation of a liquid chromatography-tandem mass spectrometry method for the separation of conjugated and unconjugated 17alpha- and 17beta-boldenone in urine sample.

Natural occurrence or illegal treatment of boldenone (BOLD) presence in cattle urine is under debate within the European Union. Separation of conjugated and unconjugated forms of 17alpha-boldenone (alpha-BOLD) and 17beta-boldenone (beta-BOLD) and presence of related molecules as androsta-1,4-diene-3,17-dione (ADD) appear critical points for the decision of an illegal use. The aim of this study is a new analytical approach of BOLD and ADD confirmation in cattle urine. The separation between conjugated and unconjugated forms of BOLD was obtained by a preliminary urine liquid-liquid extraction step with ethyl acetate. In this step the organic phase extracts only unconjugated BOLD and ADD, while BOLD in conjugated form remain in urine phase. Afterwards the urine phase, contains conjugated BOLD, was subjected to an enzymatic deconjugation. Solid-phase extraction (OASIS-HLB Waters) was used for the purification and concentration of analytes in organic and urine phases and liquid chromatography ion electrospray tandem mass spectrometry (LC-MS-MS) was applied for the confirmation of BOLD and ADD, using deuterium-labelled 17beta-boldenone (BOLD-d3) as internal standard. The method was validated as a quantitative confirmatory method according to the Commission Decision 2002/657/CE. The results obtained demonstrate that the developed method show very high specificity, precision, trueness and ruggedness. Decision limits (CCalpha) smaller than 0.5 ng mL(-1) were obtained for each analyte.