QUANTITATIVE-ANALYSIS OF THE LIPOPHILIC DOXORUBICIN ANALOG ANNAMYCIN IN PLASMA AND TISSUE SAMPLES BY REVERSED-PHASE CHROMATOGRAPHY

A rapid and sensitive HPLC method was developed to detect and quantita te the lipophilic doxorubicin analogue annamycin (Ann) and its metabol ites in biological samples. Reversed-phase chromatography coupled with fluorescence detection was used. The emission and excitation waveleng ths of the fluorescent detector were set at 550 and 472 nm, respective ly. The optimal mobile phase was acetonitrile:methanol:water at a 11 5 :95:90 ratio (v/v/v). The lower limit of detection was 0.35 ng of Ann (7 ng/mL). The retention times of 4-demethoxyadriamycinone (Anone) and Ann were 2.6 and 4.3 min, respectively, and their resolution was 1.3. The precision of the determination of Ann and Anone were 0.5 +/- 0.3 and 0.8 +/- 0.7%, respectively. Ann dissolved in methanol was stable u nder the determination conditions. With chloroform, 60-90% of Ann was extracted from biological samples. Apart from Anone, two Ann metabolit es (retention times 3.4 and 6.0 min) were detected in plasma and tissu es from C57BL/6 mice bearing subcutaneous B16 tumors 6 h after intrave nous administration of a 5-mg/kg suspension of Ann. No peaks were dete cted in blank tissues and plasma.