Selective determination of doxorubicin and doxorubicinol in rat plasma by HPLC with photosensitization reaction followed by chemiluminescence detection.
A highly sensitive and selective high-performance liquid chromatography (HPLC) method was developed for the determination of doxorubicin (DXR) and its metabolite doxorubicinol (DXR-ol) in rat plasma. The method was based on photosensitization reaction followed by peroxyoxalate chemiluminescence detection (PO-CL). DXR and DXR-ol that were fluorescent quinones, served as a photosensitizer in the presence of a hydrogen atom donor such as ethanol under aerobic conditions to produce hydrogen peroxide. Then the generated hydrogen peroxide and DXR or DXR-ol were monitored through PO-CL reaction by mixing with aryloxalate as a single post-column reagent that enabled highly selective and sensitive determination of DXR and DXR-ol. The separation of DXR and DXR-ol by HPLC was accomplished isocratically on an ODS column within 15 min. The method involves a simple one step protein precipitation by methanol and a sample size of 50-microL was sufficient. Besides, it can detect accurately the low plasma concentrations. The detection limits (signal-to-noise ratio=3) were 4.5 and 3.8 fmol for DXR and DXR-ol, respectively. The percentage recovery was found to be 90.7-102.4% and the inter- and intra-assay RSD values in rat plasma were 2.5-8.9%. The method has been successfully used to study pharmacokinetic profiles of DXR and DXR-ol in rats after a single-dose of DXR.
Ahmed
Sameh
S
Graduate School of Biomedical Sciences, Course of Pharmaceutical Sciences, Nagasaki University, Nagasaki, Japan.
Kishikawa
Naoya
N
Ohyama
Kaname
K
Wada
Mitsuhiro
M
Nakashima
Kenichiro
K
Kuroda
Naotaka
N
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004317:Doxorubicin / Q000031:analogs & derivatives* / Q000097:blood*; D008163:Luminescent Measurements; D010777:Photochemistry / Q000379:methods; D051381:Rats
D004317:Doxorubicin; C010013:adriamycinol