Fully automated determination of a new anthracycline N-l-leucyldoxorubicin and six metabolites in plasma by high-performance liquid chromatography with on-line sample handling.
N-l-Leucyldoxorubicin (Leu-Dox) was developed as a prodrug of doxorubicin (Dox) in order to diminish the cardiotoxic side-effect associated with repeated anthracycline treatment. To study the pharmacokinetics of Leu-Dox, Dox and other metabolites a sensitive and selective assay was needed. Leu-Dox and six of its known metabolites were extracted from plasma using an in-line reversed-phase precolumn (40-50 microns C8 particles). The trapped analytes were subsequently flushed to the analytical column (3 microns C18) using 0.5 ml of phosphate buffer (pH 3.5)-acetonitrile (2:1, v/v), which also served as the isocratic mobile phase. Within 12 min, a clean baseline-resolved chromatogram is obtained by fluorescence detection. Recoveries were almost quantitative and highly reproducible, with standard deviations less than or equal to 5.4% and less than or equal to 2.7% at spiked concentrations of 10 and 100 nM. Using 300 microliters of plasma, detection limits ranged from 0.3 to 0.8 nM at a signal-to-noise ratio of 3. The calibration curves were linear from 1 to 300 nM (r2 greater than or equal to 0.999) for each of the seven compounds. The between-day accuracy was in the range 91-99% and 99-105% at 10 and 100 nM, respectively, with standard deviations of 1-4%. Application of the assay to the analysis of plasma from patients after administration of Leu-Dox proved successful.
de Jong
J
J
Department of Oncology, Free University Hospital, Amsterdam, Netherlands.
Munniksma
C N
CN
Guérand
W S
WS
Bast
A
A
van der Vijgh
W J
WJ
D000903:Antibiotics, Antineoplastic / Q000097:blood / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004317:Doxorubicin / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism; D006801:Humans; D011355:Prodrugs; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence
D000903:Antibiotics, Antineoplastic; D011355:Prodrugs; C073409:N-leucyldoxorubicin; D004317:Doxorubicin