Homoharringtonine (HHT), first isolated from the Chinese evergreen Cephalotaxus harringtonia, has been demonstrated to have a broad antitumor activity in rodents and antileukemic effects in humans. We found that HHT was metabolized to an acid product [HHT-acid; 2'-hydroxy-2'-(alpha-acetic acid)-6'-hydroxy-6'-methylheptanoyl cephalotaxine] when incubated with either human plasma or mouse plasma in vitro. The conversion was faster, however, in mouse plasma, and was both time- and temperature-dependent. Boiled plasma prevented the conversion of HHT to HHT-acid, suggesting that the conversion was enzymatically mediated. When mice were given an intravenous (i.v.) injection of HHT (4 mg/kg), the HHT-acid metabolite was found in both plasma and urine. In mice, HHT-acid was detected in the plasma within 5 min of the i.v. injection of HHT and declined rapidly thereafter. The initial half-lives (t 1/2 alpha) of HHT and HHT-acid were 9 and 17 min, respectively. Twenty-four hours after HHT dosing in mice, approximately 29% of the dose was excreted in the urine as HHT and 20% as HHT-acid. High-pressure liquid chromatography and mass spectrometry were used to confirm the identity and quantify HHT and its metabolite, HHT-acid. The HHT concentration inhibiting 50% of the growth of human leukemic HL-60 cells was 20 ng/ml, while for HHT-acid it was 14,500 ng/ml, indicating that the acid form was more than 700 times less cytotoxic than HHT. The lethal dose of HHT affecting 50% (LD50) of mice was 6.7 mg/kg, but HHT-acid produced no apparent toxic effects at doses up to 280 mg/kg.