Determination of 9-nitrocamptothecin and its metabolite 9-aminocamptothecin in human plasma using high-performance liquid chromatography with ultraviolet and fluorescence detection

KW-2170, 5-(3-aminopropyl) amino-7,10-dihydroxy-2-(2-hydroxethyl)-aminoethyl-6H-pyrazolo [4,5,1-de] acridin-6-one dihydroxychloride, is a novel anticancer agent under clinical development. We have established a highly sensitive method which can simultaneously quantitate KW-2170 and its two metabolites, a carboxylic (M1) and hydroxylated (M2) derivative involving the 5-position, in human and dog plasma. KW-2170 and its metabolites were extracted from plasma using a weak cation-exchange cartridge and then determined by HPLC using an electrochemical detector (ED). Over the concentration range 0.1-50 ng/ml, precision and accuracy of intra- and inter-day assay were within 11% in human plasma. In dog plasma, they were within 17% at the lower quantitation limit and within 11% at other concentrations. These three compounds were stable during the assay procedure, freeze-thawing cycles and during long-term storage. Using this methodology, the pharmacokinetics of KW-2170 in a dog could be monitored over 24 h. This method is suitable for evaluation of the detailed pharmacokinetics of KW-2170 and its metabolites in humans and dogs.