Rapid quantification of polymyxin B in human pulmonary epithelial lining fluid by LC-MS/MS and its clinical application

Abstract

Pulmonary epithelial lining fluid (ELF) was commonly used for the pharmacokinetic study in lower respiratory tract infections. To characterize the intrapulmonary pharmacokinetic properties of polymyxin B following aerosol delivery, we developed and fully validated a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for quantifying polymyxin B in human bronchoalveolar lavage fluid (BALF). The ELF concentrations were calculated by the BALF values of polymyxin B using urea as a volume normalizer. Chromatographic separation was achieved on a Phonomenex Kinetex XB-C18 column(100 mm × 2.1 mm I.D., 2.6 μm)in acetonitrile and water both containing 0.2 % formic acid. The flow rate was set as 0.4 mL/min for a 3.5 min running time. Protein precipitation was used in preparing BALF samples with polymyxin E1 as an internal standard. Polymyxin B was detected under multiple reaction monitoring conditions using the electrospray ionization interface running in the positive ionization mode. The assay showed a good linear relationship over the tested concentration ranges of 0.0300/0.00306–––10.0/1.02 mg/L for polymyxin B1/B2 in bronchoalveolar lavage fluid (R² > 0.99). The inter- and inter-day precisions (RSD, %) were < 12.2 %(15.2 % for LLOQ samples)and the accuracies (%) were within the range of 94.3 ∼ 110.4 %. This reliable LC–MS/MS method for detection of polymyxin B was successfully applied to conduct a pulmonary penetration study in patients following aerosol administration.