Simultaneous Quantification of Levofloxacin and Linezolid in human Plasma and Saliva Using Liquid Chromatography Quadrupole‐Orbitrap Mass Spectrometry

ABSTRACT Levofloxacin and linezolid are key antibiotics used for the treatment of multidrug‐resistant Mycobacterium tuberculosis , and therapeutic drug monitoring is recommended to guide prescribing to ensure optimal drug exposure. The aim of this study was to validate a rapid, selective, and reliable method for simultaneous quantification of levofloxacin and linezolid with simple sample preparation. After a simple protein precipitation with 1‐to‐3 methanol of plasma and saliva samples, the chromatographic separation was performed using a liquid chromatography system coupled with quadrupole‐Orbitrap mass spectrometry in positive ionization mode. The mobile phase consisted of a gradient elution of water‐formic acid (99.9:0.1, v/v) (A) and methanol‐formic acid (99.9:0.1, v/v) (B) at 0.2 mL/min. The stationary phase was the Agilent Poroshell 120 CS‐C18 (100 mm × 2.1 mm, 2.7 µm). Analyte transitions were m/z 362.151 → 318.161 for levofloxacin and m/z 338.151 → 296.141 for linezolid. [ 13 C 3 , 2 H 3 ]‐levofloxacin and [ 2 H 8 ]‐linezolid were used as internal standards. The quality control (QC) concentrations were 1, 10, and 20 mg/L. The analysis time was 7 min, and all analytes eluted within 4 min. The calibration curves were quadratic (0.5–20 mg/L) in plasma and saliva for both analytes, with regression coefficients >0.99 for both analytes. The intraday and inter‐day coefficients of variation were <15% for all QC concentrations. The method was successfully applied to drug monitoring in patients with multidrug‐resistant tuberculosis.