Simultaneous determination of seven β-lactam antibiotics in human plasma for therapeutic drug monitoring and pharmacokinetic studies

Abstract

This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. AbstractThere is strong evidence in literature supporting the benefit of monitoring plasma concentrations of -lactam antibiotics in the critically ill to ensure appropriateness of dosing.The objective of this work was to develop a method for the simultaneous determination of total concentrations piperacillin, benzylpenicillin, flucloxacillin, meropenem, ertapenem, cephazolin and ceftazidime in human plasma. Sample preparation involved protein precipitation with acetonitrile containing 0.1% formic acid and subsequent dilution of supernatant with 0.1% formic acid in water. Chromatographic separation was achieved on a reversed phase column (C18, 2.6µm, 2.1* 50 mm) via gradient elution using water and acetonitrile, each containing 0.1% formic acid, as mobile phase. Tandem mass spectrometry (MSMS) analysis was performed, after electrospray ionization in the positive mode, with multiple reaction monitoring (MRM). The method is accurate with the inter-day and intra-day accuracies of quality control samples (QCs) ranging from 95%-107% and 95%-108%, respectively. It is also precise with intra-day and inter-day coefficient of variations ranging from 4 to 12 % and 5-14% respectively. The lower limit of quantification was 0.1g/mL for each antibiotic except flucloxacillin (0.25g/mL). Recovery was greater than 96% for all analytes except for ertapenem (78%). Coefficients of variation for the matrix effect were less than 10% over the six batches of plasma. Analytes were stable over three freeze-thaw cycles, and for reasonable hours on the bench top as well as post-preparation. This novel liquid chromatography tandem mass spectrometry method proved accurate, precise and applicable for therapeutic drug monitoring and pharmacokinetic studies of the selected -lactam antibiotics.Page 5 Highlights  We present a method for simultaneous determination of seven beta-lactams in plasma  The selected antibiotics are those commonly used in critically ill patients  The method is accurate, precise and meets validation requirements by guidelines  It proved applicable for therapeutic drug monitoring and pharmacokinetic studies

The use of liquid chromatography-tandem mass spectrometry for therapeutic drug monitoring of antibiotics in cancer patients

“…Although as compared to mass spectrometry less sophisticated analysis is performed with the use of the other conventional detectors, the speed and sensitivity of the former one surpass those offered by the latter ones (UV, CL, EC). The current interest focuses on LC-MS/MS [60,[64][65][66][67][68][69][70][71][72][73][74]. This is not surprising as it is well acknowledged that LC-MS/ MS has the power of combining the sensitivity of tandem MS and the selectivity of chromatography thus improving the accuracy of quantitating low drug concentrations from different matrices.…”

“…In many cases the use of LC-MS/ MS offers an increase in the sensitivity by several orders of magnitude. For instance, reported lower limit of detection of some β-lactams achieved with LC-UV or LC-EC detectors is 5 mg/L [28,61], while it reached 10-50 times lower levels (0.1-0.5mg/L) when LC-MS/MS is used [64,74]. As in the case of critically ill patients very low levels can be anticipated methods that offer higher sensitivity are preferred.…”

“…Although different analytical methods have been developed for the quantification of individual antibiotics, in the last 5 years several methods have been published for the simultaneous analysis of subsets of antibiotics usually given to critically ill patients [64][65][66][67][68][69][70][71][72]74], and continuous attempts for improved method development are ongoing. This stems from the need for rapid and accurate methods for the detection in one run of as many as possible of the antibiotics used as first and second lines of treatment in critically ill patients.…”

“…: methanol or acetonitrile [64,72,74]. Even though the use of acidic solutions such as trichloroacetic acid (TCA) has been used successfully, care must be taken due to the instability of the four member β-lactam ring and its consequent degradation in high acid concentration [73,75].…”

“…The stability of β-lactams is also significantly affected in the presence of zinc sulfate and, therefore, its use as a precipitant should be avoided [76]. Though protein precipitation with methanol and acetonitrile is frequently used, it seems that acetonitrile is preferred due to the great reproducibility it offers along with its lowest ionization suppression effect [64,68,74,77]. Furthermore, due to the inherent instability of this class of antibiotics especially ring hydrolysis the use of stabilizing buffers has been recommended and in some cases was found as a necessity [23].…”