D. H. Jadhav and C. S. Ramaa
An aim of the study is development and validation of a method for the simultaneous estimation of raloxifene (RAL) and its two active metabolites, raloxifene-4-glucuronide (R4G) and raloxifene-6-glucuronide (R6G) in human plasma samples using raloxifene-D4 as an Internal Standard. Sample preparation was performed by solid phase extraction (SPE) and was followed by separation of the analytes on a UPLC system with a linear gradient and a mobile phase consisting of acetonitrile and ammonium formate. Detection was achieved by tandem mass spectrometry operated in the electrospray positive ion mode. The method had a short sample preparation time, as well as a chromatographic run time of just 4.2 min, the shortest so far reported for RAL, R4G and R6G determination. It was validated and fulfilled all preset criteria for sensitivity, specificity, linearity, within inter- and intra-accuracy and precision, stability studies for all molecules. The method was linear in the concentration range of 0.040 to 1.5 ng/mL, 0.6 to 50.0 ng/mL and 0.6 to 50.0 ng/mL for RAL, R4G and R6G, respectively. The proposed method could be applied to the rapid and reliable simultaneous determination of RAL, R4G and R6G in a bioequivalence study.
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