This study investigated the effect of aerial exposure duration (0, 24, 48, and 72 h) on the juvenile Chinese mitten crab, Eriocheir sinensis. The effects on histological structure, energy metabolism, non-specific immunity, and expression of cytochrome P450 (CYP2) and protein kinase B (AKT) genes in the anterior gill and hepatopancreas were analyzed. Longer durations of air exposure led to loss of anterior gill filament shape, and enlargement and deformation of the hepatopancreatic tubules with vacuolization. Pyruvate kinase activity in the hemolymph peaked, and was the lowest in the anterior gill, at 72 h. Lactate dehydrogenase activity and lactic acid content in the hemolymph and anterior gills peaked at 72 h of air exposure. Triglyceride content in hemolymph and anterior gill first increased, then decreased. Superoxide dismutase activity and total antioxidant capacity increased in the hemolymph but initially increased and then decreased in the hepatopancreas, peaking at 24 h. Hemolymph and hepatopancreas acid phosphatase activity peaked at 48 h and in these same tissues lysozyme activity declined with prolonged air exposure. The relative expression of CYP2 and AKT increased during exposure to air in both gill and hepatopancreas. Air exposure exceeding 48 h may affect tissue morphology, energy metabolism, oxidative stress, and immunity of juvenile E. sinensis. Juvenile crabs alter their energy utilization, antioxidant, and metabolic capacities to acclimate to air exposure stress. Therefore, the air exposure time for juvenile E. sinensis should be limited to 48 h during transportation.