title: A rapid UPLC-MS/MS assay for the simultaneous measurement of fluconazole, voriconazole, posaconazole, itraconazole, and hydroxyitraconazole concentrations in serum.
abstract: BACKGROUND
  Triazole antifungals are essential to the treatment and prophylaxis of fungal infections. Significant pharmacokinetic variability combined with a clinical need for faster turnaround times has increased demand for in-house therapeutic drug monitoring of these drugs, which is best performed using mass spectrometry-based platforms. However, technical and logistical obstacles to implementing these platforms in hospital laboratories have limited their widespread utilization. Here, we present the development and validation of a fast and simple ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method to measure fluconazole, voriconazole, posaconazole, itraconazole, and hydroxyitraconazole in human serum suitable for incorporation into a hospital clinical laboratory.
  
  
  METHODS
  Serum samples (20 µL) were prepared using protein precipitation in the presence of deuterated internal standards. Chromatographic separation was accomplished using reversed phase UPLC and analysis was performed using positive-mode electrospray ionization and collision-induced dissociation MS.
  
  
  RESULTS
  Total analytical run time was 3 min. All analytes demonstrated linearity (r2>0.998) from 0.1 to 10 µg/mL (1-100 µg/mL for fluconazole), acceptable accuracy and precision (%DEV<15% and %CV<15% at all levels tested), suitable stability under relevant storage conditions, and correlated well with reference laboratory results.
  
  
  CONCLUSIONS
  A simple and rapid UPLC-MS/MS method for monitoring multiple triazole antifungals was developed with a focus on the needs of hospital laboratories. The assay is suitable for clinical utilization and management of patients on these medications.
authors.affiliation: Brigham and Women's Hospital, Boston, MA.
authors.forename: Sankha S
authors.initials: SS
authors.lastname: Basu
authors.affiliation: Brigham and Women's Hospital, Boston, MA.
authors.forename: Athena
authors.initials: A
authors.lastname: Petrides
authors.affiliation: Waters Corporation, Milford, MA.
authors.forename: Donald S
authors.initials: DS
authors.lastname: Mason
authors.affiliation: Harvard Medical School, Boston, MA.
authors.forename: Petr
authors.initials: P
authors.lastname: Jarolim
mesh_terms: D000935:Antifungal Agents / Q000097:blood*; D001774:Blood Chemical Analysis / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D015725:Fluconazole / Q000097:blood; D006801:Humans; D017964:Itraconazole / Q000031:analogs & derivatives / Q000097:blood; D057230:Limit of Detection; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors; D014230:Triazoles / Q000097:blood; D065819:Voriconazole / Q000097:blood
chemical_list: D000935:Antifungal Agents; D014230:Triazoles; C084794:hydroxyitraconazole; D017964:Itraconazole; C101425:posaconazole; D015725:Fluconazole; D065819:Voriconazole
keywords: liquid chromatography; mass spectrometry; method development; therapeutic drug monitoring; triazole antifungals