Abstract: In order to reduce annual chemical fertilizer amount in tobacco-rice rotation areas of Hunan Province, two experiment sites with different levels of soil organic matter were selected in Guiyang County and Anren County of Chenzhou City. Effects of organic fertilizer applied in the preceding rice crop in reducing the application of chemical fertilizer, and the effects on the N supplying capacity of tobacco field soil, N absorption and utilization of tobacco leaves were studied by field experiments with 35 d waterlogged incubation test. The results showed that the application of organic fertilizer to Guiyang paddy with low level of organic matter increased the soil water-soluble total N by 31.0% before tobacco transplanting, significantly increased the cumulative mineralized N(N_m) and organic N mineralization potential(N₀) at tobacco topping stage. And the application of organic fertilizer to Anren paddy with high level of organic matter did not significantly increase the soil water-soluble total N before tobacco transplanting period, but increased N₀ by 17.6%, and had low effect on N_m and N₀ of soils during tobacco growth process. At the same time, the application of organic fertilizer in rice season maintained or even increased the yield and partial productivity of N fertilizer. The N contents and nicotine contents of tobacco leaves decreased, and the ratio of nitrogen to nicotine increased by organic fertilizer application in Guiyang; while the N contents decreased, nicotine contents increased, and the ratio of nitrogen to nicotine decreased by organic fertilizer application in Anren accordingly. Above all, applying 1500 kg/hm² organic fertilizer instead of 150 kg/hm² compound fertilizer and 75 kg/hm² urea in late rice season could reduce 23.6%~27.7% of annual N fertilizer application in tobacco rice rotation areas, which could maintain tobacco yield and improve soil nitrogen supply capacity to some extent.