Automated HPLC-MS/MS assay for the simultaneous determination of ten plasma antibiotic concentrations

Therapeutic drug monitoring (TDM) of antibiotics (ATB) in patients with serious bacterial infections allows optimization of the efficacy of the treatment while reducing the risk of toxicity. Notably, early measurement of plasma beta-lactam concentration has been shown to be associated with reduced mortality in intensive care patients. In this context, a rapid, robust, and accurate assay method is essential for daily TDM. A fully automated procedure for quantification of the plasma concentrations of ten ATB was developed. The ATB were divided into two calibration pools, with Pool 1: aztreonam, ceftobiprole, cefoxitin, avibactam, tazobactam and Pool 2: metronidazole, ceftriaxone, daptomycin, ceftolozane, moxifloxacin. Sample preparation consisting of acetonitrile plasma protein precipitation and H20 dilution was applied to all analytes. This procedure was carried out by an automated sample preparation system directly coupled to a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. Since the instrument extracts sample n while sample n-1 is in the LC-MS/MS system, the delay between obtaining the results for two samples corresponds to the analytical run time, which is less than 7 min. The method was validated according to the Food and Drug Administration guidelines. The method was sensitive (lower limit of quantification 0.1-1 mg/L, depending on the ATB), accurate (intra/inter-assay bias -14.8 to 14.2 %) and precise (intra/inter-assay CVs 1.27 to 16.3 %). Application of the TDM assay was illustrated by the report of an intensive care patient treated with the ceftazidime/aztreonam/avibactam combination. Four assays were performed in 8 days with results returned within 24 h to quickly manage the dose regimen in this patient. An automated, simple, rapid, robust LC-MS/MS analysis was developed and validated for the simultaneous quantification of plasma concentrations of 10 ATB and was applied with success to perform TDM. This method provides a shorter turnaround time than classic sample batch-based analytical methods.