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Ultra-performance liquid chromatography combined with time-of-flight mass spectrometry (UPLC-ToF-MS) has been used for screening and quantification of more than 100 veterinary drugs in milk. The veterinary drugs represent different classes including benzimidazoles, macrolides, penicillins, quinolones, sulphonamides, pyrimidines, tetracylines, nitroimidazoles, tranquillizers, ionophores, amphenicols and non-steroidal anti-inflammatory agents (NSAIDs). After protein precipitation, centrifugation and solid-phase extraction (SPE), the extracts were analysed by UPLC-ToF-MS. From the acquired full scan data the drug-specific ions were extracted for construction of the chromatograms and evaluation of the results. The analytical method was validated according to the EU guidelines (2002/657/EC) for a quantitative screening method. At the concentration level of interest (MRL level) the results for repeatability (%RSD<20% for 86% of the compounds), reproducibility (%RSD<40% for 96% of the compounds) and the accuracy (80-120% for 88% of the compounds) were satisfactory. Evaluation of the CCβ values and the linearity results demonstrates that the developed method shows adequate sensitivity and linearity to provide quantitative results. Furthermore, the method is accurate enough to differentiate between suspected and negative samples or drug concentrations below or above the MRL. A set of 100 samples of raw milk were screened for residues. No suspected (positive) results were obtained except for the included blind reference sample containing sulphamethazine (88 μg/l) that tested positive for this compound. UPLC-ToF-MS combines high resolution for both LC and MS with high mass accuracy which is very powerful for the multi-compound analysis of veterinary drugs. The technique seems to be powerful enough for the analysis of not only veterinary drugs but also organic contaminants like pesticides, mycotoxins and plant toxins in one single method.

The use of beta-agonists as growth promoters in cattle breeding is forbidden in many countries for reasons of fair trade and consumer protection. In recent years the use of liquid chromatography (LC) tandem mass spectrometry (MS/MS) has been shown to be the method of choice for the control of beta-agonists. In this study an LC-MS/MS multiresidue analysis method is presented for trace analysis of 22 beta-agonists. A truly generic concept has been designed based on mixed-mode solid-phase extraction and positive electrospray ionisation LC-MS/MS operated in the multiple reaction monitoring mode. This method allows application to a wide variety of sample matrices such as urine, feed and hair, following minor modifications to the analysis procedure only. The method features fit-for-purpose sensitivity in urine as shown by CCalpha and CCbeta values of less than 0.2 and less than 0.5 microg/l respectively, for all beta-agonists studied (terbutaline and reproterol, less than 0.3 and less than 1.0 respectively). Similar but semiquantitative application to feed and hair showed CCbeta values of less than 10.0 and less than 5.0 microg/kg, respectively. A further simplification and improvement is demonstrated using Ultra Performance LC (UPLC) and fast-switching MS/MS. The successful validation of this method following the latest EU requirements and its application to real samples demonstrate that a new versatile tool has been achieved for veterinary control of beta-agonists.

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Synthetic polymers having a polydispersity greater than 1.1 cannot be reliably characterized by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry alone. This complication has been overcome by off-line coupling of size-exclusion chromatography (SEC) with MALDI. Data are presented for a wide variety of synthetic polymers having polydispersities from 1.7 up to 3.0. Polystyrene (15 kDa and 48 kDa), polybutylacrylate (62 kDa), polycarbonate (28 kDa), aromatic polyester resin (8 kDa) and a methyl methacrylate-methacrylic acid copolymer (34 kDa) were injected on a conventional SEC system with refractive index (RI) or UV absorbance detection and 40 fractions collected over the polymer distribution. In a recent comparison between most probable peak values (M p ) as measured for polymer distributions by mass spectrometry and size-exclusion chromatography (SEC) it has been concluded, using theoretical considerations, that, in the case of polydisperse polymers, i.e. in most routine cases, the molecular weight distribution (MWD) data provided by mass spectrometry does not resemble the distribution as obtained by SEC because of the fundamental difference between recording the number fraction versus m/z (as in mass spectrometry) and the weight fraction versus the logarithm of the molecular mass (as in SEC with refractive index (RI) detection).1 As a practical consequence, the high-mass tail of the polydisperse distribution disappears much earlier into the baseline noise of the mass spectrometer system than into the noise of the SEC-RI system, thus yielding incorrect calculation of the so-called molecular averages: This polydispersity complication has limited the application of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry 3 to the characterization of synthetic polymers; it has been concluded that MALDI-MS is only applicable to narrowlydistributed synthetic polymers (polydispersity less than 1.1). 4 Recently, Montaudo et al. suggested a procedure to obtain improved MALDI-MS data from polydisperse polymers.5 In our experience, however, data thus obtained are extremely sensitive to the baseline subtraction procedure; even minor changes in baseline level cause huge deviations in the final MWD calculation which becomes inaccurate and irreproducible. Moreover, it is hard to believe that detector sensitivity does not alter over a range of say m/z 500 up to 300 000.Several groups have investigated experimental parameters in MALDI time-of-flight (TOF) analysis in order to get a better understanding and/or to improve the results for polydisperse synthetic polymers. Different accelerating voltages and detection efficiencies were found to cause severe distortion of the molecular weight distribution in case of polydisperse synthetic polymers. 6 The MWD of polymethylmethacrylate

New anabolic steroids show up occasionally in sports doping and in veterinary control. The discovery of these designer steroids is facilitated by findings of illicit preparations, thus allowing bioactivity testing, structure elucidation using NMR and mass spectrometry, and final incorporation in urine testing. However, as long as these preparations remain undiscovered, new designer steroids are not screened for in routine sports doping or veterinary control urine tests since the established GC/MS and LC/MS/MS methods are set up for the monitoring of a few selected ions or MS/MS transitions of known substances only. In this study, the feasibility of androgen bioactivity testing and mass spectrometric identification is being investigated for trace analysis of designer steroids in urine. Following enzymatic deconjugation and a generic solid-phase extraction, the samples are analyzed by gradient LC with effluent splitting toward two identical 96-well fraction collectors. One well plate is used for androgen bioactivity detection using a novel robust yeast reporter gene bioassay yielding a biogram featuring a 20-s time resolution. The bioactive wells direct the identification efforts to the corresponding well numbers in the duplicate plate. These are subjected to high-resolution LC using a short column packed with 1.7-microm C18 material and coupled with electrospray quadrupole time-of-flight mass spectrometry (LC/QTOFMS) with accurate mass measurement. Element compositions are calculated and used to interrogate electronic substance databases. The feasibility of this approach for doping control is demonstrated via the screening of human urine samples spiked with the designer anabolic steroid tetrahydrogestrinone. Application of the proposed methodology, complementary to the established targeted urine screening for known anabolics, will increase the chance of finding unknown emerging designer steroids, rather then being solely dependent on findings of the illicit preparations themselves.