Validated UHPLC–MS/MS assay for quantitative determination of etoposide, gemcitabine, vinorelbine and their metabolites in patients with lung cancer

A fully valid ultra-high performance liquid chromatography coupled with mass spectrometry (UHPLC- MS/MS) method was developed for the determination of etoposide, gemcitabine, vinorelbine and their metabolites (etoposide catechol, 2’,2’-difluorodeoxyuridine and 4-O-deacetylvinorelbine) in human plasma. Multiple reaction monitoring (MRM) mode was performed with an electrospray ionization (ESI) interface operating in both the positive and negative ion modes per compound. The method required only 100 μL plasma with a one-step simple de-proteinization procedure, and a short run time of 7.5 min per sample. A Waters ACQUITY UPLC HSS T3 column (2.1 × 100 mm, 1.8 µm) provided chromatographic separation of analytes using a binary mobile phase gradient (A, 0.1% formic acid in acetonitrile, v/v; B, 0.1% formic acid in water, v/v). Linear coefficients of correlation were >0.995 for all analytes. The relative deviation of this method was <10% for intra- and inter-day assays and the accuracy ranged between 86.35% and 113.44%. The mean extraction recovery and matrix effect of all the analytes were 62.07%-105.46% and 93.67%-105.87%, respectively. This method was successfully applied to clinical samples from lung cancer patients.