Global changes affect the quality and quantity of litter as well as increased nitrogen deposition, which impact the soil nutrient cycling significantly. Soil ecoenzymatic stoichiometry can reveal the limit of nutrients in microbial growth and metabolism. However, it is still unclear how soil ecoenzymatic stoichiometry responds to the synchronous changes in litter input and nitrogen addition in a mixed temperate forest. This study attempts to explore the effects of litter manipulation and nitrogen addition on soil ecoenzymatic stoichiometry and the major influencing factors in a mixed pine and oak. The results showed that (1) there was no significant interactions between litter manipulation and nitrogen addition. Nitrogen addition was no significant difference on soil ecoenzymatic stoichiometry, while the difference was significant under the litter manipulation, which showed that leaf litter double (L) and mixed litter double (LB) were higher than the fine woody litter double (B) and litter removal (N). Soil ecoenzymatic stoichiometry ratios were not significantly deviated 1：1：1 under litter manipulation and nitrogen addition. (2) Soil microbial carbon use efficiency (CUE_C：N and CUE_C：P) under leaf litter double and mixed litter double treatment was significantly lower than that under fine woody litter double and litter removal treatment, but there no significant differences under nitrogen addition treatment. Soil microbial nitrogen utilization efficiency (NUE_N：C) and microbial phosphorus utilization efficiency (PUE_P：C) were no significant differences under litter manipulation and nitrogen addition. TER_C：N was no significant difference under litter manipulation and nitrogen addition. TER_C：P under leaf litter double and mixed litter double treatment was significantly higher than fine woody litter double and litter removal treatment. (3) RDA analysis showed that soil pH was the main factor affecting soil extracellular enzyme activity and ecoenzymatic stoichiometry. The results showed that the quality and quantity of litters had more significant effects on the soil ecoenzymatic stoichiometry than the addition of nitrogen. It is possible that the forest soil microorganisms of mechanism responding to nitrogen addition is not an instantaneous or simple process. The quality and quantity of litters will change the soil nutrient status, and microorganisms will respond to nutrient variation through ecoenzymatic stoichiometry and nutrient utilization efficiency, the input of leaf litter relatively alleviates the limitation of P. It was found that the abiotic factors had more effects on soil extracellular enzyme activity and ecoenzymatic stoichiometry than biological factors under litter manipulation and nitrogen addition. This study can provide a theoretical basis for the response of soil microorganisms to global change.