AbstractThe structural isomerization of scopine into scopoline (oscine) has been observed in a supersonic jet expansion using microwave spectroscopy. The rotational spectrum evidences a single structure in the gas phase, providing a first description of the (three‐ring) structurally distorted tropane in scopoline. The absence of rotational signatures of any scopine conformation suggests a practically quantitative isomerization at the vaporization temperatures of the experiment (ca. 90 °C). The determined rotational parameters of scopoline reveal the structural consequences of the intramolecular cyclation of scopine, which breaks the original epoxy group and creates a new ether bridge and a 7β‐hydroxytropane configuration. The hydroxy group further stabilizes the molecule by an OH⋅⋅⋅N intramolecular hydrogen bond, which, in turn, forces the N‐methyl group to the less stable axial form. Supporting ab initio (MP2) and DFT (B3LYP, M06‐2X) calculations are included.