Development of a simplified, sensitive high-performance liquid chromatographic method using fluorescence detection to determine the concentration of UCN-01 in human plasma

UCN-01 is a naturally derived anticancer agent isolated in the culture brot h of actinomyces streptomyces. We have developed a sensitive high-performan ce liquid chromatographic method for the determination of UCN-01 in human p lasma. UCN-01 was isolated from human plasma after intravenous administrati on, by using 100% ice-cold acetonitrile liquid-liquid phase extraction. Liq uid chromatographic separation was achieved by isocratic elution on a pheny l analytical column. The mobile phase consisted of acetonitrile-0.5 M ammon ium acetate (45:55) with 0.2% triethylamine added as a modifier. The UCN-01 peak was identified from other peaks using fluorescence excitation energy and emission energy wavelengths of 310 and 410 nm, respectively. Retention time for UCN-01 was 4.2 +/-0.5 min. The UCN-01 peak was baseline resolved, with nearest peak at 2.6 min distance. No interfering peaks were observed a t the retention time of UCN-01. Peak area amounts from extracted samples we re proportional over the dynamic concentration range used: 0.2 to 30 mug/ml . Mean recoveries of UCN-01 at concentrations of 0.5 and 25 mug/ml were 89 and 90.2%, respectively. Relative standard deviations for UCN-01 calibratio n standards ranged from 1.89 to 2.31%, with relative errors ranging from 0. 3 to 11.6%. Assay precision for UCN-01 based on quality control samples of 0.50 mug/ml was +/-4.86% with an accuracy of +/-5.7%. For drug extracted fr om plasma the lowest limit of detection was 0.1 mug/ml, with the lowest lim it of quantitation being 0.2 mug/ml. This method is suitable for routine an alysis of UCN-01 in human plasma at concentration from 0.2 to 30 mug/ml. (C ) 2001 Elsevier Science BV All rights reserved.