A simple and sensitive assay for determining plasma tipranavir concentration in the clinical setting by new HPLC method

A simple method for the quantification of tipranavir, the first non-peptidic HIV protease inhibitor, was developed and validated. Quinoxaline, as internal standard, was added to 50 μl of plasma before a liquid–liquid extraction by 600 μl of protein precipitation solution. The extracts were diluted before being injected in the chromatographic system. Chromatographic separation was made on a C18 column using potassium phosphate buffer (pH 3.2) and acetonitrile with gradient. Detection was performed by an UV detector at 260 nm. Relative error at three control quality concentrations ranged from −1.81 to 1.72%. Intra-day (CV%) and inter-day (CV%) precision ranged from 0.94 to 2.55% and from 3.07 to 4.24%, respectively. LOQ and LOD were 0.090 μg/ml and 0.035 μg/ml, respectively. Mean recovery was 87.1% ± 2.4%. Calibration curve was linear up to 180 μg/ml. Concentration range when optimized (0.703–180 μg/ml) proved to be adequate to measure tipranavir concentration in HIV-1-positive patients, therefore this method could be suitable for therapeutic drug monitoring of this drug.