Abstract: In this paper, the metabolism of homoharringtonine has been studied in vitro with liver microsomes from rats and rabbits. A method of HPLC-DAD (photodiode array detector)with gradient elution was developed for screening the metabolites from the microsomal incubation system. Only one main metabolite, whose formation was indepen-dent of NADPH and NADH, was found. The metabolite was extracted and purified with preparative HP LC technique. Its chemical structure was identified as 2'-hydroxy-2'(α-acetic acid )-6 '-hydroxy- 6 '-methyl- heptanoyl cephalotaxine by UV, IR, NMR, M S analysis, and confirmed further by comparison with the authentic compound,the spectra and chromatographic behavior were all the same.The rates of the metabolite formation was markedly increased in the phenobarbital pretreated liver microsome system.