Desarrollo y validación de un método para la determinación de darunavir en plasma mediante LC-MS/MS

Abstract

Introduction Darunavir is a latest generation of protease inhibitors (PI) drugs used as a treatment of HIV infection owing to its improved efficacy and its better tolerance among other characteristics. Several studies have demonstrated a correlation between the dose and the effect of darunavir, although a therapeutic range for the drug concentrations has not yet been established. These factors, besides the high inter-individual variability and the adverse effects resulting from drug interactions, reinforce the need for having a sensitive method for monitoring its plasma concentrations in treated patients. Objective Technical validation of a new method for darunavir plasma concentration monitoring by LC/MS/MS. Materials and methods The method validation procedure was based on the recommendations published in the guidelines ICH Topic Q 2B Validation of Analytical Procedures: Methodology ( CPMP/ICH/281/95). Darunavir was extracted from plasma samples by a protein precipitation procedure. The chromatographic separation was achieved with a gradient program (acetonitrile/buffer) on an X-Bridge TM C18 3.5 μm 2.1×100 mm (Waters ®). Analytes quantification is performed by electrospray ionization in positive mode, a Quattro Micro triple quadrupole mass spectrometer. Results Linearity (0.1-10 μg/mL): y = 18.85×-150.4 r 2: 0.997. Precision: Intra-assay CV: 1.07-4.62%. Inter-assay CV: 2.72-4.70%. Accuracy: Intra-assay: 1-9%. Inter-assay: 2-7%. LOD: 0.05 μg/mL, LLOQ: 0.15 μg/mL. Conclusion The developed method based on LC/MS/MS has an adequate sensibility and reproducibility for the determination of plasma concentrations of darunavir in treated patients.