A fast and sensitive LC–MS/MS method for the quantification of fosfomycin in human urine and plasma using one sample preparation method and HILIC chromatography

Abstract

Fosfomycin is an old antibiotic that is increasingly prescribed because of emergence of the antibiotic resistance and the growing incidence of multi-drug resistant infections. Surprisingly, little is known about its pharmacokinetics (PK) and the pharmacodynamics (PD). Quantification of fosfomycin in both urine and plasma provides insight into the PK/PD characteristics of fosfomycin, which is crucial for the optimization of the therapy and the prevention of the emergence of resistance. An analytical method is therefore needed for the quantification of fosfomycin in both urine and plasma. A fast and sensitive tandem mass spectrometry method in combination with HILIC chromatography for the quantification of fosfomycin with a universal sample preparation method for urine and plasma was developed and validated according to FDA guidelines. The universal sample preparation method only requires 100μL of a sample, the addition of the internal standard fosfomycin-13C benzylamine and an ultrafiltration step. The method is applicable for the concentration range of 0.75-375mg/L (R of 0.9998 in both matrices) encompassing the clinically relevant concentration range based on the susceptibility of possible (uro)pathogens in the clinical setting. The validation results for urine and plasma for all QC levels, were <2.1% and <3.2% for accuracy, <1.5% and <1.7% for within day precision and <5.0% and <3.8% for between day precision, respectively. No matrix effects were encountered and the total recovery in urine and plasma was high (102.5% and 99.4%). Prepared samples were stable at 4°C and 15°C for at least 72h and stored samples at -80°C were stable for at least 6 months. Selectivity and sensitivity were confirmed and no carry-over was observed. The method was successfully applied in two pharmacokinetic studies in healthy volunteers and patients respectively.

Oral Fosfomycin Treatment for Enterococcal Urinary Tract Infections in a Dynamic In Vitro Model

“…Samples for fosfomycin concentration quantification, which first underwent a 1:10 dilution with saline when concentrations were expected to be above the upper limit of the assay, were frozen at -80°C until testing. An ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) method was used (57). The UPLC-MS-MS method was validated for urine and plasma samples of fosfomycin, but additional tests confirmed its applicability for fosfomycin in SHU samples.…”

Development and validation of a LC‐MS/MS method for quantitation of fosfomycin – Application to in vitro antimicrobial resistance study using hollow‐fiber infection model

“…Although LC‐MS/MS‐based quantification methods of FOS are published in the literature in both preclinical and clinical biological matrices such as plasma, urine and tissues (Rezende, Almeida, Brito, Nonaka, & Leite, ; Parker et al, 2015a, b; El‐Najjar, Jantsch, & Gessner, ; Wijma et al, ), here we present a novel method for accurate quantification of FOS in lysogeny broth to support the antimicrobial resistance study using the in vitro hollow‐fiber infection model.…”

“…Thus, to effectively monitor the kinetics of the emergence of resistant bacteria (especially Gram-negative bacteria) to treat underlying bacterial infection, specific assays for measuring antimicrobial drugs are required. To address this issue, we developed and validated a liquid (Rezende, Almeida, Brito, Nonaka, & Leite, 2012;Parker et al, 2015a, b;El-Najjar, Jantsch, & Gessner, 2017;Wijma et al, 2017), here we present a novel method for accurate quantification of FOS in lysogeny broth to support the antimicrobial resistance study using the in vitro hollow-fiber infection model.…”

Physicochemical and Antimicrobial Properties of Thermosensitive Chitosan Hydrogel Loaded with Fosfomycin

“…Due to fosfomycin’s optical inactivity, direct quantification is difficult to achieve. A method for quantification via LC-MS has been reported [ 31 ]. From a preliminary experiment using these methods, it is believed the low molecular weight chitosan (degradation products, i.e., chitosan monomer/oligomer) used here may have interfered with the hydrophilic interaction liquid chromatography column, causing retention time peaks of the eluent to broaden and vary drastically compared to the standard curve samples, preventing accurate quantification of the released FOS.…”