Abstract: OBJECTIVE To study the metabolism of gastrodin (Gas) in rat brain, liver, kidney and different brain regions. METHODS Gas was incubated in different tissue homogenate at 37℃ in vitro. An HPLC method was established to determine the metabolite concentration after incubation. RESULTS The results showed the velocity constants of formation metabolite, HBA, were 0.030 5, 0.012 1 and 0.140 9 g·mL-1 min-1 in brain, liver and kidney, respectively. The clearance rates of Gas were (29.7 2.1), (10.4 0.8) and (299.5 22.7) L·min-1·g-1 in the brain, liver and kidney, respectively. The metabolism rate of Gas was slow in brain and liver, while it was fast in kidney. In brain homogenate, the formation velocity of HBA was fast in the cerebellum, thalamus, pons and medulla oblongata, and slow in the frontal cortex, hippocampus and striatum. The velocity constants of the formers were about 1.5 times higher than those of the latters. Clearance rates of Gas were (45.9 7.4), (39.6 5.6) and (24.4 3.0) L·min-1·g-1in cerebellum, thalamus and cortex, respectively. CONCLUSION Gas is stable in non-enzyme system and can be metabolized to HBA in brain, liver and kidney homogenate in vitro. And Gas is metabolized most rapidly in the kidney. The metabolism rates of Gas in the cerebellum, thalamus, pons and medulla oblongata are faster than other three brain regions.