Development of a sensitive LC–MS/MS method for quantification of linezolid and its primary metabolites in human serum

Souza, Ernane; Felton, Jeremy; Crass, Ryan L.; Hanaya, Kengo; Pai, Manjunath P.

doi:10.1016/j.jpba.2019.112968

Cited by 11 publications

(12 citation statements)

References 11 publications

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“…A summary of patient demographic and clinical characteristics is included in Table S1 in the supplemental material. The validated LC-MS/MS method applied to quantify serum concentrations of linezolid and its major metabolites showed precision and accuracy within 15%, extraction recoveries between 78% and 103%, and no significant matrix effect (9). Standard curves were linear (r 2 Ն 0.99) for linezolid concentrations within 0.1 to 50 mg/liter and for concentrations of both metabolites within 0.1 to 25 mg/liter.…”

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confidence: 93%

“…In an effort to establish a valid methodology for determining the concentration profiles of the parent drug and metabolites in infected patients who received linezolid treatment, our group recently developed the first liquid chromatography-tandem mass spectrometry (LC-MS/MS) procedure for the simultaneous quantification of linezolid, PNU-142300, and PNU-142586 in human serum (9). This was made possible thanks to development of a synthesis methodology of these metabolites that has not been readily available (10).…”

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confidence: 99%

“…Sample processing and the assays of linezolid and its metabolites by LC-MS/MS are detailed in a publication by our group (9). Briefly, acetonitrile containing a deuterated internal standard was added to the serum samples, which were centrifuged for protein precipitation; the resulting supernatants were removed, and the analytes were separated using a Waters X-bridge column (C18, 150 by 4.6 mm, 3.5 m).…”

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confidence: 99%

“…The current linezolid product label specifically states that "given the absence of information on the clinical significance of accumulation of the primary metabolites, use of linezolid in patients with renal insufficiency should be weighed against the potential risks of accumulation of these metabolites." Despite 20 years of clinical use, knowledge on this risk potential has not been defined because chemical standards have not been readily available (9,10,12). Linezolid and its major metabolites share a chemical feature with other myelotoxic drugs, i.e., tedizolid, oxyphenbutazone, metamizole, 4-aminoantipyrine, and aminopyrine, all of which are aminophenazone analogs (see Fig.…”

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Accumulation of Major Linezolid Metabolites in Patients with Renal Impairment

Souza

Crass

Felton

et al. 2020

Antimicrob Agents Chemother

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In patients with renal impairment (n = 22 of 39), the median serum concentrations of linezolid, PNU-142300, and PNU-142586 were 1.6-, 3.3-, 2.8-fold higher, respectively, than in patients without renal impairment. Metabolite concentrations in paired samples were poorly correlated with linezolid concentrations (r2 = 0.26 for PNU-142300 and 0.06 for PNU-142586). Linezolid and its metabolites share potential toxicophores that deserve characterization to mitigate higher myelosuppression risk in patients with renal impairment.

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Accumulation of Major Linezolid Metabolites in Patients with Renal Impairment

Souza

Crass

Felton

et al. 2020

Antimicrob Agents Chemother

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“…Through poorly described oxidation pathways, linezolid is primarily metabolized into PNU-142300 ( Figure 1B ) and PNU-142586, and both of them exceeded 10% of the dose of the parent drug excreted in the urine, with specific value of 10 and 40% respectively ( Stalker and Jungbluth, 2003 ). Although it has been reported that renal function is not an important influencing factor of the overall exposure of linezolid in plasma, the two main metabolic products in patients with renal insufficiency have been identified and determined to be higher than normal patients ( Stalker and Jungbluth, 2003 ; Souza et al, 2020a ; Souza et al, 2020b ). Therefore, it is urgent to validate and establish a bioanalytical assay for the simultaneous measurement of linezolid, PNU-142300, and PNU-142586 in serum to investigate the contribution of each analyte to toxicity in patients with different renal function.…”

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A Novel UPLC-MS/MS Assay for the Measurement of Linezolid and its Metabolite PNU-142300 in Human Serum and its Application to Patients With Renal Insufficiency

Wang

et al. 2021

Front. Pharmacol.

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The contribution of the metabolites of linezolid to the associated myelosuppression is unknown in patients who are renal impairment. In this research, the purpose of our experiment was to explore and develop a quick and robust ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) assay for the determination of linezolid and its metabolite PNU-142300 in human serum simultaneously. The analytes were prepared using a simple and convenient approach with acetonitrile for protein crash, and then separated from the matrix on a Waters Acquity Ultra performance liquid chromatography (UPLC) BEH C18 (2.1 mm × 50 mm, 1.7 μm) column in a program of gradient elution, where the mobile phase was consisted of water with 0.1% formic acid and acetonitrile, and was placed at 0.40 ml/min flow rate. Multiple reaction monitoring (MRM) was employed and conducted for UPLC-MS/MS detection with ion transitions at m/z 338.01 → 296.03 for linezolid, m/z 369.96 → 327.98 for PNU-142300 and m/z 370.98 → 342.99 for tedizolid (Internal standard, IS), respectively. This method had good linearity respectively in the calibration range of 0.01–20 μg/ml for linezolid, and 0.05–100 μg/ml for PNU-142300. In the intra- and inter-day, the precision of linezolid and PNU-142300 was below 14.2%, and the accuracy in this method was determined to be from −9.7 to 12.8%. In addition, recovery and matrix effect of the analytes were all found to be acceptable, and the analytes during the assay and storage in serum samples were observed to be stable. The novel optimized UPLC-MS/MS assay was also successfully employed to determine the concentration levels of linezolid and PNU-142300 in human serum. The results showed that linezolid-associated myelosuppression occurs more frequently in patients with renal insufficiency, and the metabolite-to-parent concentration ratio of PNU-142300 is predicted to reduce this toxicity of myelosuppression.

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A sensitive and high‐throughput quantitative liquid chromatography high‐resolution mass spectrometry method for therapeutic drug monitoring of 10 β‐lactam antibiotics, linezolid and two β‐lactamase inhibitors in human plasma

Vooren

Verstraete

2021

Biomedical Chromatography

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An ultra-high pressure liquid chromatography high-resolution mass spectrometric (UHPLC-HRMS) method was developed for the simultaneous and sensitive quantification of 10 β-lactam antibiotics (cefepime, meropenem, amoxicillin, cefazolin, benzylpenicillin, ceftazidime, piperacillin, flucloxacillin, cefuroxime and aztreonam), linezolid and β-lactamase inhibitors tazobactam and clavulanic acid in human plasma. Validation according to the EMA guidelines showed excellent withinand between-run accuracy and precision (i.e. between 1.1 and 8.5%) and high sensitivity (i.e. lower limit of quantification between 0.25 and 1 mg/L). The UHPLC-HRMS method enables a short turnaround time and high sensitivity and needs only a small amount of plasma, allowing appropriate routine therapeutic drug monitoring.The short turnaround time is obtained by speeding up the protocol on multiple levels, i.e. fast and workload-efficient sample preparation (i.e. protein precipitation and dilution), short (4 min) instrument run time, simultaneous measurement of all relevant β-lactam antibiotics used in the intensive care unit and the use of the same instrument, column and mobile phases as for the other routine methods in our laboratory. K E Y W O R D Sβ-lactam antibiotics, HRMS, therapeutic drug monitoring | INTRODUCTIONSevere infection and associated septic shock persist as significant healthcare concerns as they are the most common cause of mortality in critically ill patients worldwide (Vincent et al., 2009). Since adequate and early administration of antibiotics remains the mainstay of therapy for the treatment of severe infections, there is a high level of consumption of antibiotics in intensive care units (ICU) (Dellinger et al., 2013). A single-day point prevalence study showed that 71% of the ICU patients received antimicrobial therapy (Vincent et al., 2009). However, in specific patient populations like critically ill patients, cystic fibrosis patients, elderly patients, neonates, etc., the pharmacokinetics of these antibiotics may be profoundly disturbed owing to pathophysiological changes in distribution and elimination (Roberts et al., 2014;Taccone et al., 2010;Theuretzbacher, 2012). Consequently, an empirical fixed dose approach, in which every patient is treated according to the same dosing scheme, appears to be inadequate and can lead to excessive drug exposure with an increased risk of toxicity or inadequate drug exposure with clinical failure and/or antimicrobial resistance (Roberts & Lipman, 2006). Optimizing antibiotic dosing may be a key intervention, not only to improve clinical outcome and minimize toxicity, but also to prolong the clinical lifespan of the currently available antibiotics by limiting the emergence of

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Development of a sensitive LC–MS/MS method for quantification of linezolid and its primary metabolites in human serum

Cited by 11 publications

References 11 publications

Accumulation of Major Linezolid Metabolites in Patients with Renal Impairment

Accumulation of Major Linezolid Metabolites in Patients with Renal Impairment

A Novel UPLC-MS/MS Assay for the Measurement of Linezolid and its Metabolite PNU-142300 in Human Serum and its Application to Patients With Renal Insufficiency

A sensitive and high‐throughput quantitative liquid chromatography high‐resolution mass spectrometry method for therapeutic drug monitoring of 10 β‐lactam antibiotics, linezolid and two β‐lactamase inhibitors in human plasma

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