Liquid chromatography–tandem mass spectrometry methods for clinical quantitation of antibiotics

The accurate quantification of antibiotics in biological matrices is pivotal for supporting individualized medication, optimizing antibiotics therapy and addressing the global crisis of antimicrobial resistance (AMR). LC-MS/MS has emerged as the gold standard for clinical antibiotic quantification, particularly valued for its accuracy, multiplexing efficiency, and compatibility with complex sample types. This review provides a comprehensive overview of the principles, methodologies, and clinical applications of LC-MS/MS in antibiotic quantification, with a focus on its role in therapeutic drug monitoring (TDM), pharmacokinetics (PK), and epidemiological studies. Key challenges, including sample preparation and matrix effects, are critically discussed. Tailored strategies for analyzing diverse antibiotic classes (e.g. β-lactams, aminoglycosides, colistins, macrolides, and glycopeptides) are highlighted by considering their physicochemical properties, including polarity, acid-base characteristics, and structural complexity. Furthermore, emerging trends in automation, high-resolution mass spectrometry, and quality control frameworks are outlined to guide future advancements. By bridging technological innovation with clinical needs, LC-MS/MS continues to be a cornerstone of precision medicine and a vital tool in global AMR mitigation efforts.