Validation of a simple and economic HPLC-UV method for the simultaneous determination of vancomycin, meropenem, piperacillin and tazobactam in plasma samples

Paola Milla; Fiorenza Ferrari; Elisabetta Muntoni; Marco Sartori; Claudio Ronco; Silvia Arpicco

doi:10.1016/j.jchromb.2020.122151

Validation of a simple and economic HPLC-UV method for the simultaneous determination of vancomycin, meropenem, piperacillin and tazobactam in plasma samples

J Chromatogr B Analyt Technol Biomed Life Sci. 2020 May 11:1148:122151. doi: 10.1016/j.jchromb.2020.122151. Online ahead of print.

Authors

Paola Milla¹, Fiorenza Ferrari², Elisabetta Muntoni³, Marco Sartori⁴, Claudio Ronco⁵, Silvia Arpicco⁶

Affiliations

¹ Department of Drug Science and Technology, University of Turin, Via P. Giuria 9, I-10125 Turin, Italy. Electronic address: paola.milla@unito.it.
² Intensive Care Unit, I.R.C.C.S. Fondazione Policlinico San Matteo di Pavia, Viale C. Golgi 19, I-27100 Pavia, Italy. Electronic address: fioreferrari28@gmail.com.
³ Department of Drug Science and Technology, University of Turin, Via P. Giuria 9, I-10125 Turin, Italy. Electronic address: elisabetta.muntoni@unito.it.
⁴ International Renal Research Institute of Vicenza and Department of Nephrology, Dialysis and Transplant of San Bortolo Hospital, Viale F. Rodolfi 37, I-36100 Vicenza, Italy. Electronic address: sartorimarco.sm@gmail.com.
⁵ International Renal Research Institute of Vicenza and Department of Nephrology, Dialysis and Transplant of San Bortolo Hospital, Viale F. Rodolfi 37, I-36100 Vicenza, Italy; Department of Medicine, University of Padova, Via N. Giustiniani 2, I-35128 Padova, Italy. Electronic address: cronco@goldnet.it.
⁶ Department of Drug Science and Technology, University of Turin, Via P. Giuria 9, I-10125 Turin, Italy. Electronic address: silvia.arpicco@unito.it.

PMID: 32417718
DOI: 10.1016/j.jchromb.2020.122151

Abstract

Critically ill patients are often affected by several pathophysiological conditions requiring antibiotic administration and, frequently, extracorporeal therapy that significantly alter the normal pharmacokinetics of drugs. Therapeutic drug monitoring (TDM) may assist to establish the correct antibiotic dosage, but a TDM service is usually available only for some aminoglycosides and glycopeptides. The aim of this study is the validation of an HPLC-UV method for the simultaneous quantification of meropenem, vancomycin, piperacillin and tazobactam in human plasma samples. The analytes were extracted from 250 μL of human plasma by the addition of acetonitrile for protein precipitation. After evaporation to dryness of the solvent, samples were reconstituted with 250 μL of mobile phase, and 100 μL were injected in HPLC. Chromatographic analysis was performed using a Kinetex C18 column and an UV/Vis detector set at 220 and 298 nm. The mobile phase was a mixture of phosphate buffer 0.1 M pH 3.15 and methanol in gradient, delivered at 1 mL/min. The method was validated over clinical concentration ranges. For all the analytes, the lower limit of quantification was 1 μg/mL, and the calibration curves were linear between 1 and 100 μg/mL, with coefficients of determination ≥ 0.999. Intra-day precision was < 4%, while inter-day precision was < 7% for each analyte. The applicability of the method has been evaluated by analysing plasma samples collected from 4 critically ill patients undergoing continuous renal replacement therapy. Moreover, the analysis of vancomycin with VANC Flex® confirmed a good correlation between the results of HPLC-UV and commercially available kits usually used by TDM service. The method we developed only requires a small volume of plasma and uses the same sample preparation protocol, stationary phase and elution conditions for all analytes. This method offers the additional advantages of simple and rather inexpensive sample preparation and instrumentation, features that make this method an easy implementation for a general TDM laboratory.

Keywords: HPLC-UV; Intensive care unit; Meropenem; Piperacillin /tazobactam; Therapeutic drug monitoring; Vancomycin.