Abstract: Glyciphosphoramide (GPA) is one of the anticancer agents belonging to the group of phosphoramide mustard. It has apparent antitumor effects in some animal models and in clinical trials against breast cancer, lymphosareoma, uterocervical cancer and cancerous ulcer with good resuits. In this paper, the determination procedure of GPA and its metabolite using nitrobenzylpyridine(NBP) method is reported, The absorbanee of the eoloured products from the reaction of hydrolyzedor metabolized GPA with NBP was measured at 570 nm and 564 nm, respectively. The linearity of the reaction for GPA and its metabolite was established over the range of 6.25～100 μtg/ml water or plasma. The plasma of rats and miee was found to be able to metabolize GPA to form alkylating agent (s) which react with NBP, but that of rabbits cannot. The plasma concentration- time curve of metabolite obtained after oral administration of GPA (100 mg/kg) in rats was shown to fit a two compartment open model with the following parameters: T_1/2β = 44.5 min, T_1/2β= 3. 16 min, T_1/2Ka=2.14 min, T_1/2Km=0.0644 rain, T_max=7.57 min, C_max= 55.8 μg/ml, AUC= 2827. 39 μg/ml·min, K₂₁=0.09663/min, K₁₀= 0.03535/rain, K₁₂=0. 1030/min, Vc=1.00 L/kg, Vd=2.07 L/kg, CLt=2.12 L/h. Kidney was found to be the main organ for GPA metabolite elimination. About one fourth of the given dose was excreted in urine within 24 h with the main portion exereted in the first 2 h.