Concurrent determination of ezetimibe and its phase-I and II metabolites by HPLC with UV detection: quantitative application to various in vitro metabolic stability studies and for qualitative estimation in bile
| Autor: | Ramesh Mullangi, Nadipalli Prabhakar Rao, Nirbhay Kumar Tiwari, Dhanya Shashikumar, Shaik Abdul Naveed, N. R. Srinivas, Nyavanandi Vijay Kumar, Thammera Ranjith Kumar, Shaik Jafar Sadik Basha, Nuggehally R. Srinivas, Syed Muzeeb |
|---|---|
| Rok vydání: | 2006 |
| Předmět: |
Male
Analyte Formic acid Metabolite Clinical Biochemistry Biochemistry High-performance liquid chromatography Analytical Chemistry chemistry.chemical_compound Glucuronides Tandem Mass Spectrometry Animals Bile Rats Wistar Chromatography High Pressure Liquid Detection limit Chromatography Molecular Structure Elution Reproducibility of Results Cell Biology General Medicine Ezetimibe Rats chemistry Azetidines Spectrophotometry Ultraviolet Glucuronide Quantitative analysis (chemistry) |
| Zdroj: | Journal of chromatography. B, Analytical technologies in the biomedical and life sciences. 853(1-2) |
| ISSN: | 1570-0232 |
| Popis: | Simultaneous separation and quantification of ezetimibe (EZM) and its phase-I metabolite i.e., ezetimibe ketone (EZM-K) and phase-II metabolite i.e., ezetimibe glucuronide (EZM-G) in various matrices was accomplished by gradient HPLC with UV detection. The assay procedure involved deproteinization of 500 microL of either incubation or bile sample containing analytes and internal standard (IS, theophylline) with 75 microL acetonitrile containing 25% perchloric acid. An aliquot of 100 microL supernatant was injected onto a C18 column. The chromatographic separation was achieved by gradient elution consisting of 0.05 M formic acid:acetonitrile:methanol:water at a flow rate of 1.0 mL/min. The detection of analyte peaks were achieved by monitoring the eluate using an UV detector set at 250 nm. Nominal retention times of IS, EZM-G, ezetimibe ketone glucuronide (EZM-KG), EZM and EZM-K were 9.39, 24.23, 27.82, 29.04 and 30.56 min, respectively. Average extraction efficiencies of EZM, EZM-G and IS was >75-80% and for EZM-K was >50% from all the matrices tested. Limit of quantitation (LOQ) for EZM, EZM-K and EZM-G was 0.02 microg/mL. Due to the lack of availability of reference standard of EZM-KG, the recovery and LOQ aspects for this metabolite were not assessed. Overall, the method is suitable for simultaneous measurement of EZM, and its phase-I and phase-II metabolite (EZM-G) in in vitro and in vivo studies. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect