Abstract: This study explored the relationship between the substitution rate of organic fertilizer (SROF) and soil fertility under long-term fertilizer application in black soils and thereby laid the basis of theoretical guidance for optimal fertilization in black soils. A 32-year dataset on black soil fertilization from Gongzhuling in Jilin Province experimental site was analyzed to determine the changes in SROF under different rates of organic fertilizer application based on crop removal of nitrogen (N) and soil N supply-demand balance. Results showed that crop yield increased gradually with increasing application years of organic fertilizer, which was slightly higher than for NPK fertilization after 32 years. A significant positive correlation existed between SROF and fertilization years as driven by crop removal of N (P < 0.01). Furthermore, SROF reached 100% and was stable after 29 years of high quantity manure (M2) application, higher than that of the application of conventional manure quality (M1). The correlation coefficients (R²) between SROFs of conventional and high quantity manure (M1 and M2) applications between based on crop N absorption (R_N) and soil N supply-demand balance (R_N') were 0.78 and 0.84 (P < 0.01), respectively. The corresponding root mean square errors (RMSE) were 10.4% and 14.6%, validating the method of calculation of SROF based on soil N supply-demand balance. The results suggested that SROF under high crop yield could be used as a fertility index under current intensive fertilization. Longer applications of organic fertilizers led to higher soil fertility and crop productivity and a gradual increase in SROF.