• Development and validation of a quantitative method for 15 antiviral drugs in poultry muscle using liquid chromatography coupled to tandem mass spectrometry

      Douillet, Clément; Moloney, Mary; Di Rocco, Melissa; Elliott, Christopher; Danaher, Martin; European Union; Chinese Ministry of Science and Technology; 727864 (Elsevier BV, 2022-02)
      The objective of this work was to develop a quantitative multi-residue method for analysing antiviral drug residues and their metabolites in poultry meat samples. Antiviral drugs are not licensed for the treatment of influenza in food producing animals. However, there have been some reports indicating their illegal use in poultry. In this study, a method was developed for the analysis of 15 antiviral drug residues in poultry muscle (chicken, duck, quail and turkey) using liquid chromatography coupled to tandem mass spectrometry. This included 13 drugs against influenza and associated metabolites, but also two drugs employed for the treatment of herpes (acyclovir and ganciclovir). The method required the development of a novel chromatographic separation using a hydrophilic interaction chromatographic (HILIC) BEH amide column, which was necessary to retain the highly polar compounds. The analytes were detected using a triple quadrupole mass spectrometer operating in positive electrospray ionization mode. A range of different sample preparation protocols suitable for polar compounds were evaluated. The most effective procedure was based on a simple acetonitrile-based protein precipitation step followed by a further dilution in a methanol/water solution. The confirmatory method was validated according to the EU 2021/808 guidelines on different species including chicken, duck, turkey and quail. The validation was performed using various calibration curves ranging from 0.1 µg kg−1to 200 µg kg−1, according to the analyte. Depending on the analyte sensitivity, decision limits achieved ranged from 0.12 µg kg−1 for arbidol to 34.7 µg kg−1 for ribavirin. Overall, the reproducibility precision values ranged from 2.8% to 22.7% and the recoveries from 84% to 127%. The method was applied to 120 commercial poultry samples from the Irish market, which were all found to be residue-free.
    • Improving the chromatographic selectivity of β-lactam residue analysis in milk using phenyl-column chemistry prior to detection by tandem mass spectrometry

      Di Rocco, Melissa; Moloney, Mary; Haren, Deirdre; Gutierrez, Montserrat; Earley, Seán; Berendsen, Bjorn; Furey, Ambrose; Danaher, Martin; Department of Agriculture, Food and the Marine; 13/F484 (Springer Science and Business Media LLC, 2020-05-23)
      Analyte isobaric interferences can limit the development of a comprehensive analytical method for the quantitative liquid chromatography-tandem mass spectrometry profiling of an important cohort of veterinary drugs. In this work, a selective chromatographic separation was developed for the analysis of 32 β-lactam antibiotic residues (12 penicillins, 14 cephalosporins, five carbapenems and faropenem) in milk samples. A range of analytical columns with different stationary phases and mobile phases were evaluated for retention and separation of the β-lactam compounds. Results showed that, among the columns tested, only phenyl-hexyl could adequately separate ampicillin from cephalexin and amoxicillin from cefadroxil, which had shown isobaric interferences on a number of stationary phases. Chromatography was performed using a water/acetonitrile binary gradient with formic acid and ammonium acetate. The β-lactam residues were extracted from the milk samples using a water:acetonitrile solution and purified by C18 dispersive solid-phase extraction (d-SPE) clean-up, followed by concentration under nitrogen and ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) determination. Analytes were monitored in positive electrospray ionisation mode (ESI(+)). Possible interfering matrix effects were overcome by using 13 internal standards. The method was fully validated according to 2002/657/EC guidelines, showing satisfactory performance characteristics. Under within-laboratory reproducibility conditions, trueness and precision ranged from 91 to 130% and from 1.4 to 38.6%, respectively. Decision limits (CCα) were in the range 2.1–133 μg kg−1. Limits of detection (LODs) and quantitation (LOQs) ranged between 0.0090 and 1.5 μg kg−1 and from 0.030 to 5.0 μg kg−1, respectively.