Conformational space of cinchonidine has been explored by means of ab initio potential and free energy surfaces, and the temperature-induced changes of conformational populations were studied by a combined NOESY-DFT analysis. The DFT-derived potential energy surface investigation identified four new conformers. Among them, Closed(7) is substantially relevant to fully understand the conformational behavior. The energy surfaces gave access to the favored transformation pathways at different temperatures (280-320 K). They also revealed the reasons for the negligible presence of energetically stable conformers and explained the experimentally observed temperature dependence of the populations.