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Maraviroc is a CCR5 inhibitor approved in 2007 for treatment of therapy experienced adult patients infected with CCR5-tropic HIV-1 virus. International guidelines for HIV therapy indicate a plasma concentration cutoff of maraviroc for response. We developed and validated a new HPLC-MS method to quantify maraviroc concentrations in human plasma. 6,7-Dimethyl-2,3-di(2-pyridyl)quinoxaline was used as internal standard and added to 100μL of plasma. Samples were then treated with 500μL of acetonitrile for the protein precipitation procedure. An analytical T3 Atlantis column (150mm×4.6mm i.d.) with a particle size of 5μm was used to separate the compounds and ions were detected at m/z 257.5 and 313.3 for maraviroc and quinoxaline respectively. The calibration curve was linear up to 2500ng/mL. The mean recovery of maraviroc was 89.1%. All validation data results were in accordance to Food and Drug Administration and European Medicines Agency requirements. The HPLC-MS method reported here could be used routinely to monitor plasma concentrations of maraviroc in HIV-infected patients. more...

Ribavirin-induced hemolytic anemia is the main cause of discontinuation of the combination therapy with alpha-interferon-2b and ribavirin for the treatment of hepatitis C virus (HCV) infection. The determination of intracellular ribavirin levels in blood, including the levels of its phosphorylated metabolites, might be useful for predicting ribavirin-induced anemia, which could be caused by the accumulation of these molecules within the erythrocytes. In this work, we simplified and validated a previously developed assay method, to make it suitable for routine monitoring of cellular ribavirin. Whole blood diluted with a five-fold volume of ice-cold distilled underwent a process of acid phosphatase digestion to convert phosphorylated ribavirin metabolites to free ribavirin. The resulting mixture, spiked with an internal standard, was treated with a protein precipitation protocol in acetonitrile, followed by reverse-phase high-performance liquid chromatography analysis. The calibration curve for ribavirin levels in whole blood was linear at concentrations from 625 to 320,000 ng/mL (r(2)=0.998). Accuracy, intra-day and inter-day precision for ribavirin and phosphorylated-ribavirin quality controls were all below 9.0%. We tested this method by monitoring blood ribavirin concentrations in 13 HCV+ patients, receiving alpha interferon-plus ribavirin combination therapy. more...