Sensitive SPE-HPLC Method to Determine a Novel Angiotensin-AT1 Antagonist in Biological Samples

A high-performance liquid chromatography (HPLC)-method after solid-phase extraction (SPE) has been developed in order to determine a new angiotensin-AT1 antagonist, i.e. CR 3210 (C27H 24N8; MW=460.54), 4-[4-[(2-ethyl-5,7-dimethylimidazo[4,5- b]pyridin-3-yl)methyl]phenyl]-3-(2H-tetrazol-5-yl)quinoline in rat plasma and urine after oral administration to Sprague-Dawley rats. CR 3210 and the internal standard (IS) CR 1505 (loxiglumide), i.e. 4-[(3,4-dichlorobenzoyl) amino]-5-[(3-methoxypropyl)pentylamino]-5-oxopentanoic acid, were isolated from rat urine and plasma by solid-phase extraction. The procedure was optimized regarding the sorbent extraction material, the pH in the conditioning solution, the washing step, the dry time and the type of elution solvent. The separation was performed by reversed-phase high-performance liquid chromatography with ultraviolet detection. The samples were injected onto the analytical column (Tracer Extrasil ODS1) and detected at 238nm, giving a capacity factor of 1.87 for CR 3210 and 1.10 for the internal standard. The selectivity of the method was satisfactory. The mean recovery of CR 3210 from spiked rat plasma was 68.5 at 75ng/ml and 80.9 at 3000ng/ml; the mean recovery of CR 3210 from spiked rat urine was 69.9 at 75ng/ml and 78.6 at 3000ng/ml. The lower limit of detection (LOD) was 14ng/ml in plasma and 22ng/ml in urine samples. The lower limit of quantification (LOQ) was taken as 30ng/ml, the lowest calibration standard using 500μl rat plasma and urine. The procedures were validated according to international standards with a good reproducibility and linear response from 30 to 3000ng/ml, for either plasma or urine. The sensitivity of the method allowed for its application to pharmacokinetic studies. © 2003 Elsevier B.V. All rights reserved.