Rapid simultaneous quantification of immunosuppressants in transplant patients by turbulent flow chromatography combined with tandem mass spectrometry

Abstract

Background: Immunosuppressant therapeutic drug monitoring (TDM) is an important requirement in the management of post-transplant patients. Our aim was to develop and evaluate a robust high-throughput method using turbulent flow chromatography (TFC) coupled with liquid chromatography–tandem mass spectrometry (LC–MS/MS) for the simultaneous quantification of cyclosporin A (CsA), tacrolimus (FK 506) and sirolimus. Methods: A total of 1483 EDTA-blood pre-dosage samples from 147 kidney, 67 liver, 15 kidney/pancreas, and 48 bone marrow recipients were collected. After hemolysis and protein precipitation of 50 μl blood, fast and efficient on-line matrix elimination was achieved using turbulent flow chromatography. Tandem mass spectrometric detection and quantification was performed using multiple reaction monitoring (MRM). Results: The total analysis time of the column switching method was 3 min. The method was linear from 4.5 to 1500 ng/ml for cyclosporin A, from 0.2 to 100 ng/ml for tacrolimus, and from 0.4 to 100 ng/ml for sirolimus. The accuracy was >95%. Within and between-run assay variation coefficients ranged from 2.4% to 9.3%. Excellent correlation with other standard methods (immunoassay, HPLC) was observed. Conclusions: The presented turbulent flow chromatography–tandem mass spectrometric platform offers a very fast, simple and economical method with an excellent validation profile and is well suited for daily pre- and post-dosage immunosuppressant monitoring.