The passage from shore to marine life is a critical step in the development of juvenile penguins and is characterized by a fuel selection towards lipid oxida- tion concomitant to an enhancement of lipid-induced ther- mogenesis. However, mechanisms of such thermogenic improvement at fledging remain undefined. We used two different groups of pre-fledging king penguins (Apteno- dytes patagonicus) to investigate the specific contribution of cold exposure during water immersion to lipid metabo- lism. Terrestrial penguins that had never been immersed in cold water were compared with experimentally cold-water immersed juveniles. Experimentally immersed penguins underwent ten successive immersions at approximately 9–10 °C for 5 h over 3 weeks. We evaluated adaptive ther- mogenesis by measuring body temperature, metabolic rate and shivering activity in fully immersed penguins exposed to water temperatures ranging from 12 to 29 °C. Both never-immersed and experimentally immersed penguins were able to maintain their homeothermy in cold water, exhibiting similar thermogenic activity. In vivo, perfusion of lipid emulsion at thermoneutrality induced a twofold larger calorigenic response in experimentally immersed than in never-immersed birds. In vitro, the respiratory rates and the oxidative phosphorylation efficiency of isolated muscle mitochondria were not improved with cold-water immersions. The present study shows that acclimation to cold water only partially reproduced the fuel selection towards lipid oxidation that characterizes penguin acclima- tization to marine life.