Abstract: Organic matter (straw (S) and manure (M)) has been widely applied to the black soil. This study aims to explore the effect of mean annual temperature on soil enzyme measurement in the cold region. According to the latitude in western Heilongjiang Province, China, 12 positioning test sites were selected with the application period of organic matter greater than five years. A mixed-effects model was established to analyze the relationship between mean annual temperature and soil enzyme activity that related to carbon (C), nitrogen (N), and phosphorus (P) acquisition. In addition, a systematic analysis was made to determine the relationship among the vector angle and vector length, soil nutrient, microbial biomass C:N ratio, and extracellular enzyme stoichiometry. The results show that the C acquisition enzyme activity of M and S treatment decreased, as the mean annual temperature increased, whereas, there was an increase in the N acquisition enzyme activity. There was a significant difference in the P acquisition enzyme activity of M and S treatment. The P acquisition enzyme activity decreased in the M treatment, as the mean annual temperature increased, whereas increased in the S treatment. Moreover, there was a negative correlation between the vector length and vector angle of soil extracellular enzymes. All soil samples were limited by microbial N, while some ones were limited by microbial C. In addition, the natural logarithmic ratio of NAG+LAP and BG+CBH was significantly negatively correlated in the M and S treatments. The combination of the Mantel test and the heat map showed that the soil C, N, and P acquisition enzyme activities were significantly correlated with the mean annual temperature. Redundancy analysis and hierarchical segmentation showed that the individual factors greater than 10% in the M treatment were the mean annual temperature, pH, and soil C:N ratio, accounting for 30.13%, 22.32%, and 12.27%, respectively. The extracellular enzyme N:P was positively correlated with the mean annual temperature and pH. The extracellular enzymes C: N and C:P were positively correlated with the soil C: N. The individual factors greater than 10% in the S treatment were pH, the application period of organic matter, C input, mean annual temperature, and soil C:P, accounting for 21.3%, 20.01%, 18.48%, 11.46%, and 11.31%, respectively. The extracellular enzyme C: N was positively correlated with pH, soil C:P, and mean annual temperature. The extracellular enzyme C:P was positively correlated with pH and soil C:P. The extracellular enzyme N:P was positively correlated with the C input, the application period of organic matter, and the mean annual temperature. In conclusion, the microbial N mainly limited the organic matter (straw and manure) application into black soil in the northeast cold region, according to the soil extracellular enzyme measurement and vector analysis. Both manure and straw application were driven by mean annual temperature, whereas the straw application was further influenced by carbon input and the application period. This finding can provide the data support and theoretical basis for the C, N, and P fertilization model for the black soil nutrient restoration in the cold region.