Detection and quantification of ezetimibe and its major glucuronide in patients with hepatic impairment via liquid chromatography-tandem mass spectrometry.

Following administration of ezetimibe (EZE), levels of the glucuronidated metabolite ezetimibe-glucuronide (EZEG) are altered in patients with hepatic impairment. A rapid and sensitive liquid chromatography-tandem mass spectrometry method (LC-MS/MS) method was developed and validated to quantify EZE and EZEG in human plasma and urine. Samples were prepared using a protein precipitation procedure with methanol containing stable isotope-labeled internal standards (EZE-d4 and EZEG-d4). Separation of the analytes was achieved using acetonitrile-water (0.1% formic acid) as the mobile phase at a flow rate of 0.5 mL/min on a C18 column. The analytes were detected using negative ionization in multiple reaction monitoring (MRM) mode. The mass transition pairs of m/z 408.4→271.0 and m/z 584.5→271.0 were used to quantify EZE and EZEG, respectively. The method was linear over a concentration range of 1.5 ng/mL - 1 µg/mL for EZE and EZEG in the plasma. In the urine, EZE was detectable a linear range of 5 ng/mL - 1 µg/mL and EZEG ranged from 3 ng/mL - 1 µg/mL. The method was validated in accordance with U.S. Food and Drug Administration and European Medicines Agency regulatory standards, including specificity, sensitivity, stability, repeatability and reproducibility. This ensures accuracy and reliability of test results which thereby enhances patient safety and helps support clinical decisions.