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Simultaneous determination of five novel luteinizing hormone-releasing hormone antagonists by LC–MS and pharmacokinetics in rats following cassette dosing

Yao, Jin-Feng, Zhou, Ning, Bai, Lu, Xu, Ping-Xiang, Liu, Ke-Liang, Xue, Ming
Journal of chromatography 2014 v.962 pp. 94-101
acetonitrile, antagonists, drugs, formic acid, gonadotropin-releasing hormone, hormone antagonists, intravenous injection, liquid chromatography, liquid-liquid extraction, luteinization, mass spectrometry, models, monitoring, pharmacokinetics, rapid methods, rats, screening
Long acting luteinizing hormone-releasing hormone (LHRH) antagonists designed to be protease-resistant were a series of novel decapeptides structurally similar to LHRH. In the present work, a high-throughput method based on a LC–MS/MS has been developed for the simultaneous determination of pharmacokinetics of five LHRH antagonists in rat via cassette dosing. The method was performed under selected reaction monitoring (SRM) in positive ion mode. The analytes were extracted from rat plasma by liquid–liquid extraction with acetonitrile. Chromatographic separation of the analytes was successfully achieved on a Hypersil Gold (100mm×2.1mm, 3μm) using a mobile phase composed of acetonitrile–water (30:70) containing 0.05% (v/v) formic acid. The result showed good linearity and selectivity were obtained for all antagonists. The limits of quantification of the five LHRH antagonists were from 5 to 10ng/mL. The average extract recoveries in the rat plasma were all over 72%. The intra-day and inter-day precisions (R.S.D. %) were all within 10% and the accuracy was ranged from 92.54 to 109.05%. This method has been successfully applied to the pharmacokinetic studies of the five LHRH antagonists. The results indicated that the plasma drug concentrations versus time curves after intravenous injection of five antagonists via cassette dosing were all fitted to a two-compartment model. The pharmacokinetic parameters of five LHRH antagonists suggested that LY616 could be the more stable candidate drugs and optimized as the candidate drug for further study. Our studies enabled high-throughput rapid screening for pharmacokinetic assessment of new peptide candidates, and provided abundant information on the metabolic properties of these LHRH antagonists.