Vinblastine (VLB) is an antineoplastic agent contained in leaves of Catharanthus roseus. One-pot synthesis of VLB from vindoline and catharanthine, biosynthetic precursors in the plant, was achieved under biomimetic conditions. FMN-mediated coupling of vindoline and catharanthine under irradiation with near-ultraviolet light was followed by NAD(P)H-mediated direct conversion of the coupling product to VLB in the dark. The yield of VLB was governed by the level of NAD(P)H added, and the highest yield was obtained by incubation with 10 mM NAD(P)H for 5 h. It is postulated, as one concept of VLB biosynthesis, that similar non-enzymic VLB synthesis may occur in the plant leaves.