Determination of doxorubicin and metabolites in murine specimens by high-performance liquid chromatography.
A sensitive and selective reversed-phase high-performance liquid chromatographic method for the quantification of doxorubicin and its metabolites doxorubicinol, 7-deoxydoxorubicinone and 7-deoxydoxorubicinolone was developed and validated for a variety of murine specimens. Daunorubicin was used as internal standard. Sample pretreatment involved liquid-liquid extraction of 200 microl sample with 1 ml of chloroform-1-propanol (4:1, v/v). Chromatographic separation was achieved isocratically on a LiChrosorb RP-8 analytical column at ambient temperature. The mobile phase consisted of acidified water (pH 2.05)-acetonitrile-tetrahydrofuran (80:30:1, v/v/v). The column effluent was monitored fluorimetrically at an excitation wavelength of 460 nm and an emission wavelength of 550 nm. The lower limits of quantitation were in the range 1.8-2.4 nM. Spiked murine specimens and samples from treated mice were subjected to stability studies. The results demonstrated the importance of validation in all relevant specimens, since the accuracy and precision were highly matrix-dependent. Accuracies and precisions of measured drug concentrations in liver, spleen, muscle, gastrointestinal tissues, diluted bile, feces and urine were lower than in the other matrices. Doxorubicin was unstable in diluted bile, but not in the other specimens. The method is suitable for studying the pharmacokinetics of doxorubicin and its metabolites in mice.
Department of Clinical Chemistry, The Netherlands Cancer Institute, Amsterdam.
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van Asperen
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van Tellingen
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JH
Beijnen
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000032:analysis* / Q000097:blood / Q000378:metabolism* / Q000493:pharmacokinetics; D001646:Bile / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004317:Doxorubicin / Q000031:analogs & derivatives / Q000032:analysis* / Q000097:blood / Q000378:metabolism* / Q000493:pharmacokinetics; D005243:Feces / Q000737:chemistry; D005260:Female; D006801:Humans; D008297:Male; D051379:Mice; D009279:Naphthacenes / Q000032:analysis / Q000097:blood; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014018:Tissue Distribution
D000903:Antibiotics, Antineoplastic; D009279:Naphthacenes; C074869:7-deoxy-13-dihydroadriamycinone; D004317:Doxorubicin; C010013:adriamycinol; C049326:7-deoxyadriamycin aglycone