Microbore HPLC method with online microdialysis for measurement of topotecan lactone and carboxylate in murine CSF

We developed a chromatography method to measure lactone and carboxylate forms of topotecan (TPT) in mouse cerebrospinal fluid (CSF) using microdialysis sampling. The chromatography method utilized a microbore (0.8 mm) column. Analytes, which eluted in less than 5 min, were detected with a fluorescence detector. The calibration range was 0.25-100.0 ng/mL for both forms. The within-day and between-day precision was less than or equal to 16% for 0.8 ng/mL and less than or equal to 8.0% for 3,12, and 80 ng/mL. Accuracy was +/- 15% (0.8 and 3 ng/mL) and +/- 10% (12 and 80 ng/mL). TPT lactone hydrolyzes to the carboxylate during sampling, so we developed an equation and parameters to describe the TPT lactone hydrolysis in artificial CSF (aCSF). After TPT administration, CSF dialysate samples (2 muL) were analyzed for lactone and carboxylate using online injection. The hydrolysis of each dialysate sample was then estimated and a correction applied. We conclude that this HPLC method coupled with online microdialysis sampling allows for the rapid measurement of both TPT forms in small volumes of murine CSF dialysate. The system allows for the determination of TPT pharmacokinetics in murine CSF and provides a tool to extend pharmacological studies in this brain compartment. (C) 2004 Wiley-Liss, Inc. and the American Pharmacists Association.