Validation of a multi-residue UHPLC-HRMS method for antibiotics screening in milk, fresh cheese, and whey

The aim of this study was to develop a UHPLC-HRMS methodology as a suitable tool for the screening of antibiotics in milk, fresh cheese, and whey. Compounds were analysed using an Orbitrap Exactive (TM) analyser working at a resolving power of 50,000 FWHM in full scan, both in positive and negative electrospray ionization mode, and a high mass accuracy (<5 ppm) and a retention time deviation of +/- 2.5 % were set as identification criteria. The multiclass method was developed for the simultaneous determination of 36 substances from different antibiotic groups (beta-lactams, lincosamides, macrolides, quinolones, sulfonamides, and tetracyclines) and the quantification was performed by using matrix-matched calibration curves corrected by the corresponding internal standards. Matrix effect and recovery percentages were also calculated prior to the selection of internal standard. After comparing different sample preparation procedures, a quick and easy extraction with acetate buffer at pH 5.2 and acetonitrile (20/80, v/v), followed by C18 dispersive Solid Phase Extraction (dSPE) bulk sorbent was selected. The detection capabilities (CC beta s) were established at 25% or 50% of the Maximum Residue Limit (MRL) set in milk (no regulation for cheese and whey), and precision (repeatability and reproducibility) and trueness (recovery) were satisfactory according to Commission Decision 2002/657/EC.

Automated summary

This study aimed to develop a UHPLC-HRMS methodology for the screening of antibiotics in milk, fresh cheese, and whey. A multiclass method was developed for the simultaneous determination of 36 substances from different antibiotic groups. A quick and easy extraction method was selected after comparing different sample preparation procedures. The developed method showed satisfactory precision and trueness.