Aconitum carmichaelii Debeaux is a widely used traditional Chinese medicine and an important source of clinical drugs, of which the parent and lateral roots are known as ‘Chuanwu’ and ‘Fuzi’, respectively. Four new C19‐diterpenoid alkaloids, carmichasines A – D (1 – 4), were isolated from the roots of Aconitum carmichaelii, together with twelve known compounds (5 – 16). Their structures were elucidated via spectroscopic analyses, including HR‐ESI‐MS, IR, and NMR. Carmichasine A (1) is the first natural C19‐diterpenoid alkaloid possessing a cyano group. Most of the diterpenoid alkaloids isolated were C19‐category, which might provide further clues for understanding the chemotaxonomic significance of this plant. The cytotoxicity of the new compounds was also investigated against several human cancer cell lines, including MCF‐7, HCT116, A549, and 786‐0, and none of them showed considerable cytotoxic activity.