High‐throughput salting‐out‐assisted liquid–liquid extraction for the simultaneous determination of atorvastatin, ortho‐hydroxyatorvastatin, and para‐hydroxyatorvastatin in human plasma using ultrafast liquid chromatography with tandem mass spectrometry
Yong Yang, Qiufang Xu, Lei Zhou, Dafang Zhong, Xiaoyan Chen · Journal of Separation Science · 2015
A high-throughput, specific, and rapid liquid chromatography with tandem mass spectrometry method was established and validated for the simultaneous determination of atorvastatin and its two major metabolites, ortho-hydroxyatorvastatin and para-hydroxyatorvastatin, in human plasma. A simple salting-out-assisted liquid-liquid extraction using acetonitrile and a mass-spectrometry-friendly salt, ammonium acetate, was employed to extract the analytes from human plasma. A recovery of more than 81% for all analytes was achieved in 1 min extraction time. Chromatographic separation was performed on a Kinetex XB C18 column utilizing a gradient elution starting with a 60% of water solution (1% formic acid), followed by increasing percentages of acetonitrile. Analytes were detected on a tandem mass spectrometer equipped with an electrospray ionization source that was operated in the positive mode, using the transitions of m/z 559.3 → m/z 440.2 for atorvastatin, and m/z 575.3 → m/z 440.2 for both ortho- and para-hydroxyatorvastatin. Deuterium-labeled compounds were used as the internal standards. The method was validated over the concentration ranges of 0.0200-15.0 ng/mL for atorvastatin and ortho-hydroxyatorvastatin, and 0.0100-2.00 ng/mL for para-hydroxyatorvastatin with acceptable accuracy and precision. It was then successfully applied in a bioequivalence study of atorvastatin.