Measurement of posaconazole, itraconazole, and hydroxyitraconazole in plasma/serum by high-performance liquid chromatography with fluorescence detection.

Abstract

BACKGROUND:

Itraconazole and posaconazole are used in the prevention and treatment of invasive fungal infections. However, the oral bioavailability of both compounds varies widely, and dose-serum concentration relationships are poorly defined for these analytes. The aim of this work was to develop and validate a simple assay that could be implemented in most laboratories for the purpose of therapeutic drug monitoring.

METHODS:

Calibrators (n = 7) and internal quality control solutions (n = 3) were prepared in pooled human serum. Sample (100 μL), internal standard solution (25 μL), Tris solution (2 mol/L; pH 10.6), and extraction solvent (methyl tert-butyl ether, 600 μL) were vortex mixed and centrifuged. The solvent layer was removed and evaporated to dryness and the residue reconstituted in water:methanol (1 + 3, 50 μL). A portion (5 μL) of the reconstituted extract was analyzed using a 3-μm Gemini C6 phenyl column with fluorescence detection (excitation 260 nm, emission 350 nm). The method was used to measure itraconazole and hydroxyitraconazole, or posaconazole, in serum samples taken 1-2 hours before the next dose, from patients forming part of a study into management and diagnostic strategies for invasive aspergillosis.

RESULTS:

Response was linear over the calibration ranges. Accuracy and imprecision were 92-111.4% and 3.2-13.4% (relative standard deviation), respectively. No interferences were noted. There was a good agreement with nominal values of each analyte in an external quality assessment scheme. In patients prescribed either 400 mg/d of itraconazole (n = 46) or 600-800 mg/d of posaconazole (n = 28) only 24% and 7% of samples, respectively, had serum itraconazole or posaconazole concentrations above the target threshold suggested in published guidelines.

CONCLUSIONS:

A simple, sensitive high-performance liquid chromatographic method has been developed for the analysis of itraconazole, hydroxyitraconazole, and posaconazole in serum/plasma. Few of the samples measured from patients participating in the clinical study attained concentrations of the drug/metabolite in serum that have been recommended for effective antifungal therapy.

• Authors: Sara L Buckner, M Mansour Ceesay, Antonio Pagliuca, Phillip E Morgan, Robert J Flanagan
• Journal: Therapeutic drug monitoring (Ther Drug Monit) Vol. 33 Issue 6 Pg. 735-41 (Dec 2011) ISSN: 1536-3694 [Electronic] United States
• PMID: 22105591 (Publication Type: Clinical Trial, Journal Article, Validation Study)

Chemical References

• Antifungal Agents
• Triazoles
• hydroxyitraconazole
• Itraconazole
• posaconazole

Topics

• Adult
• Aged
• Antifungal Agents (administration & dosage, blood)
• Calibration
• Chromatography, High Pressure Liquid
• Cohort Studies
• Dose-Response Relationship, Drug
• Drug Monitoring (methods)
• Female
• Humans
• Invasive Pulmonary Aspergillosis (blood, complications, drug therapy)
• Itraconazole (administration & dosage, analogs & derivatives, blood)
• Limit of Detection
• Male
• Middle Aged
• Neutropenia (complications)
• Reproducibility of Results
• Spectrometry, Fluorescence
• Triazoles (administration & dosage, blood)
• Young Adult