HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC ASSAY OF A NEW HIV-1 PROTEASEINHIBITOR, LB71350, IN THE PLASMA OF DOGS

A reliable reversed-phase high-performance liquid chromatographic meth od has been developed for the determination of LB71350 in the plasma o f dogs. The analyte was deproteinized with 1.5 volumes of methanol and 0.5 volumes of 10% zinc sulfate, and the supernatant was injected int o a 5-mu m Capcell Pak C-18 column (150x4.6 mm I.D.). The mobile phase was a stepwise gradient mixture of acetonitrile and 0.2% triethylamin e-HCl with a flow-rate of 1 ml/min and detection at UV 245 nm. The pro portion of acetonitrile was kept at 52% for the first 6 min, increased to 100% for the next 0.5 min, kept at 100% for the next 2 min, decrea sed to 52% for the next 0.5 min, and finally kept at 52% for the next 7 min. The retention time of LB71350 was 6.9 min. The calibration was linear over the concentration range of 0.1-100 mg/l for dog plasma (r> 0.997) and the limit of quantitation was 0.1 mg/l using 0.1 ml plasma. The quality control samples were reproducible with acceptable accurac y and precision at 0.1, 1, 10 and 100 mg/l concentrations. The within- day recover); (n=5) was 90.2-93.9%, the between-day recovery (n=5) was 89.5-93.5%, and the absolute between-day recovery (n=5) was 77-81%. T he within-day precision (n=5) and between-day precision (n=5) were 2.5 9-5.82% and 3.17-4.55%, respectively. No interferences from endogenous substances were observed. Taken together, the above HPLC assay method by deproteinization and UV detection was suitable for the determinati on of LB71350 in the preclinical pharmacokinetics. (C) 1997 Elsevier S cience B.V.