Development of a high-performance liquid chromatographic method to determine the concentration of karenitecin, a novel highly lipophilic camptothecinderivative, in human plasma and urine

Karenitecin is a novel, highly lipophilic camptothecin derivative with pote nt anticancer potential. We have developed a sensitive high-performance liq uid chromatographic method for the determination of karenitecin concentrati on in human plasma and urine. Karenitecin was isolated from human plasma an d urine using solid-phase extraction. Separation was achieved by gradient e lution, using a water and acetonitrile mobile phase, on an ODS analytical c olumn. Karenitecin was detected using fluorescence detection at excitation and emission wavelengths of 370 and 490 nm, respectively. Retention time fo r karenitecin was 16.2 +/-0.5 min and 8.0 +/-0.2 min for camptothecin, the internal standard. The karenitecin peak was baseline resolved, with the nea rest peak at 3.1 min distance. Using normal volunteer plasma and urine from multiple individuals, as well as samples from the 50 patients analyzed to date, no interfering peaks were detected. Inter- and intra-day coefficients of variance were <4.4 and 7.1% for plasma and <4.9 and 11.6% for urine. As say precision, based on an extracted karenitecin standard plasma sample of 2.5 ng/ml, was +4.46% with a mean accuracy of 92.4%. For extracted karenite cin standard urine samples of 2.5 ng/ml assay precision was +2.35% with a m ean accuracy of 99.5%. The mean recovery of karenitecin, at plasma concentr ations of 1.0 and 50 ng/ml, was 81.9 and 87.8% respectively. In urine, at c oncentrations of 1.5 and 50 ng/ml, the mean recoveries were 90.3 and 78.4% respectively. The lower Limit of detection (LLD) for karenitecin was 0.5 ng /ml in plasma and 1.0 ng/ml in urine. The lower limit of quantification (LL Q) for karenitecin was 1 ng/ml and 1.5 ng/ml for plasma and urine, respecti vely. Stability studies indicate that when frozen at -70 degreesC, karenite cin is stable in human plasma for up to 3 months and in human urine for up to 1 month. This method is useful for the quantification of karenitecin in plasma and urine samples for clinical pharmacology studies in patients rece iving this agent in clinical trials. (C) 2001 Elsevier Science B.V. All rig hts reserved.