LC–MS/MS method for simultaneous determination of ramelteon and its metabolite M‐II in human plasma: Application to a clinical pharmacokinetic study in healthy Chinese volunteers

Abstract

A sensitive liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of ramelteon and its active metabolite M‐II in human plasma. After extraction from 200 μL of plasma by protein precipitation, the analytes and internal standard (IS) diazepam were separated on a Hedera ODS‐2 (5 μm, 150 × 2.1 mm) column with a mobile phase consisted of methanol–0.1% formic acid in 10 mm ammonium acetate solution (85:15, v/v) delivered at a flow rate of 0.5 mL/min. Mass spectrometric detection was operated in positive multiple reaction monitoring mode. The calibration curves were linear over the concentration range of 0.0500–30.0 ng/mL for ramelteon and 1.00–250 ng/mL for M‐II, respectively. This method was successfully applied to a clinical pharmacokinetic study in healthy Chinese volunteers after a single oral administration of ramelteon. The maximum plasma concentration (Cmax), the time to the Cmax and the elimination half‐life for ramelteon were 4.50 ± 4.64ng/mL, 0.8 ± 0.4h and 1.0 ± 0.9 h, respectively, and for M‐II were 136 ± 36 ng/mL, 1.1 ± 0.5 h, 2.1 ± 0.4 h, respectively.

Method development and validation for simultaneous determination of ebastine and its active metabolite carebastine in human plasma by liquid chromatography–tandem mass spectrometry and its application to a clinical pharmacokinetic study in healthy Chinese volunteers

“…Protein precipitation with acetonitrile was selected to achieve a short pre‐treatment time. Acetonitrile was utilized as a precipitant in addition to methanol to achieve higher precipitation efficiency (Zhu, 2019). The gradient elution method was applied for all analytes and ISs due to its ability to analyze two different compounds of significantly different polarity while improving run time significantly (Llupià et al, 2003).…”