Development of a UHPLC-UV/Vis Method for Simultaneously Determining Six Beta-Lactam Antibiotics in Plasma: A Tool for the Clinical Implementation of Therapeutic Monitoring of Beta-Lactams.

Beta-lactam antibiotics are among the most frequently prescribed drugs in clinical practice, yet their therapeutic drug monitoring remains underutilized despite high interindividual pharmacokinetic variability, especially in critically ill patients. To address this, we developed and validated an ultra-high-performance liquid chromatography (UHPLC-UV/Vis) method for the simultaneous quantification of six beta-lactams (cefepime, ceftolozane, ceftazidime, meropenem, ampicillin, and ertapenem) in plasma. This method uses a single gradient mobile phase and a photodiode array detector, ensuring accurate separation, minimal interference, and robust analyte identification. Validation followed EMA bioanalytical guidelines, demonstrating selectivity, precision, accuracy, and linearity within clinically relevant ranges (1.0-50.0 mg/L). Stability tests showed that the analytes were stable in plasma for up to seven days at 4 °C and one month at -20 °C. Pilot clinical implementation in 35 patients revealed significant interindividual variability, supporting the need for routine beta-lactam monitoring. Approximately 26% of trough concentrations were below the minimal inhibitory concentration, while others exceeded thresholds associated with potential toxicity. : This study represents the first UHPLC-UV/Vis method for the simultaneous determination of these six beta-lactams, overcoming limitations of prior methods that required different mobile phases or excluded clinically relevant antibiotics. The method is universally applicable and easily transferable to routine clinical practice. These findings underline the importance of beta-lactam monitoring in optimizing treatment outcomes and combating antibiotic resistance in vulnerable populations. Further studies to assess free drug concentrations are warranted to enhance clinical applicability.