Determination of a novel alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor antagonist (LY300164) and its N-acetyl metabolite in mouse, rat, dog, and monkey plasma using high-performance liquid chromatography with ultraviolet detection.

Abstract

A method for the analysis of the AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate) receptor antagonist LY300164 (compound I) and its N-acetyl metabolite (compound II) in plasma was developed. The assay utilized solid-phase extraction on a C18 Bond Elut cartridge followed by reversed-phase HPLC with UV detection at 310 nm. The method exhibited a large linear range from 0.05 microgram/ml to 50 micrograms/ml with an intra-assay accuracy for compound I and compound II ranging from 89.0% to 114.5% and intra-assay precision ranging from 0.5 to 15.3% in mouse, rat, dog, and monkey plasma. The inter-assay accuracy of compound I and compound II was 93.3% to 101.8% and the inter-assay precision was 1.6% to 11.2% in dog plasma. The lower limit of quantitation was 0.05 microgram/ml for compound I in plasma from all species tested. The lower limit of quantitation for compound II was 0.05 microgram/ml in dog and monkey plasma and 0.1 microgram/ml in mouse and rat plasma. Extracts of compound I and II from dog plasma were shown to be stable for 24 h at room temperature, and both compounds were stable when spiked into rat and monkey plasma frozen at -70 degrees C for 27 days. The method has shown to be useful in the investigation of the pharmacokinetics of the parent compound (I) and metabolite (II) in preclinical studies.