Quantification of Cefepime, Meropenem, Piperacillin, and Tazobactam in Human Plasma Using a Sensitive and Robust Liquid Chromatography-Tandem Mass Spectrometry Method, Part 2: Stability Evaluation.

Although the stability of ß-lactam antibiotics is a known issue, none of the previously reported bioanalytical methods had an adequate evaluation of the stability of these drugs. In the current study, the stability of cefepime, meropenem, piperacillin, and tazobactam under various conditions was comprehensively evaluated. The evaluated parameters included stock solution stability, short-term stability, long-term stability, freeze-thaw stability, processed sample stability, and whole-blood stability. When stored at -20°C, the stock solution of meropenem in methanol was stable for up to 3 weeks, and the stock solutions of cefepime, piperacillin, and tazobactam were stable for up to 6 weeks. All four antibiotics were stable in human plasma for up to 3 months when stored at -80°C and were stable in whole blood for up to 4 h at room temperature. Short-term stability results indicated that all four ß-lactams were stable at room temperature for 2 h, but substantial degradation was observed when the plasma samples were stored at room temperature for 24 h, with the degradation rates for cefepime, meropenem, piperacillin, and tazobactam being 30.1%, 75.6%, 49.0%, and 37.7%, respectively. Because the stability information is method independent, our stability results can be used as a reference by other research groups that work with these antibiotics.

Quantification of Cefepime, Meropenem, Piperacillin, and Tazobactam in Human Plasma Using a Sensitive and Robust Liquid Chromatography-Tandem Mass Spectrometry Method, Part 1: Assay Development and Validation.

The highly variable pharmacokinetics of ß-lactam antibiotics and ß-lactamase inhibitors poses a significant challenge to clinicians in ensuring appropriate antibiotic doses in critically ill patients. Therefore, routine monitoring of plasma concentrations is important for individualization of antimicrobial therapy. Accordingly, a simple and robust analytical method for the simultaneous measurement of multiple ß-lactam antibiotics and ß-lactamase inhibitors is highly desirable to ensure quick decisions on dose adjustments. In this study, a sensitive, simple, and robust method for the simultaneous quantification of cefepime, meropenem, piperacillin, and tazobactam in human plasma was developed and rigorously validated according to FDA guidance. Sample extraction was accomplished by simple protein precipitation. Chromatographic separation of analytes was achieved using stepwise gradient elution. Analytes were monitored using tandem mass spectrometry (MS/MS) with a turbo ion spray source in positive multiple-reaction-monitoring mode. The calibration curve ranged from 0.5 to 150 µg/ml for cefepime, 0.1 to 150 µg/ml for meropenem and piperacillin, and 0.25 to 150 µg/ml for tazobactam. Inter- and intraday precision and accuracy, sensitivity, selectivity, dilution integrity, matrix effect, extraction recovery, and hemolysis effect were investigated for all four analytes, and the results met the acceptance criteria. Compared to other reported methods, our method is more robust because of the combination of the following features: (i) a simple sample extraction procedure, (ii) a short sample run time, (iii) a wide dynamic range, and (iv) the small plasma sample volume needed. Since our method already covers ß-lactams and a ß-lactamase inhibitor with highly heterogeneous physicochemical properties, further antibiotic candidates may easily be incorporated into this multianalyte method.