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

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 to 200 µg kg , according to the analyte. Depending on the analyte sensitivity, decision limits achieved ranged from 0.12 µg kg for arbidol to 34.7 µg kg 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.