Heterocyclic nitrogen compounds containing two adjacent nitrogen atoms generally have a significant effect on soil nitrification inhibition, and 3,5-dimethylpyrazole (DMP) is a typical representative of this structure. However, the inhibitory effect and the regulatory mechanism of DMP on soil N transformation are unclear. In this study, a microcosm with different concentrations of DMP was carried out in brown soil to detect the dynamic changes of soil NH₄⁺–N, NO₃–N and related soil enzyme activities. Results showed that DMP inhibited soil nitrification effectively and decreased soil nitrate reductase activity, while increasing nitrite reductase and dehydrogenase activities. The inhibition effects were dose dependent, and DMP at the rate of 0.025 g kg⁻¹ dry soil showed the strongest inhibitory effect on N transformation in brown soil. The soil dehydrogenase activity was increased with an increasing DMP application rate. The changes in the soil’s chemical and biological properties caused by DMP application provided a new idea for systematically explaining how DMP participated in the soil N transformation process. This study further implied that DMP would play positive roles in alleviating environmental pressure by delaying nitrate-N formation and decreasing the activity of nitrate reductase.