This study examined the biochemical characteristics of key digestive enzymes in artificially produced Pacific bluefin tuna juveniles and revealed details of the digestive physiology of juveniles fed a commercial pellet diet and raw fish over long-term (225 days) and short-term (response to feeding) periods. pH and thermal profiles of key digestive enzymes of pepsin, trypsin, chymotrypsin, lipase and amylase were examined in juvenile bluefin tuna and the characteristics differed from those previously reported for larval and adult tuna. Long-term feeding (225 days) of the commercial diet markedly increased stomach and pyloric caeca size as well as storage/synthesis levels of the digestive enzymes. Highly increased gene expression levels of cholecystokinin (cck) and its receptor (cck1r) were also found in juveniles fed the commercial diet, suggesting cck might be involved in the gastrointestinal hypertrophy and digestion enhancement seen in the present study. In the short-term experiment (response to feeding), juveniles fed the commercial diet displayed faster gastric emptying, higher pH in the intestinal content and increased digestive enzymes storage/synthesis whereas the enzymes secretion was similar with those of juveniles fed raw fish. These indicate that juvenile bluefin tuna morphologically and physiologically adapted their digestive system to the artificial diet, however the adaptation was not sufficient to obtain a comparable performance as tuna fed raw fish feed. Biochemical characteristics of the digestive enzymes and the observed effects of artificial diet on digestive physiology could be good indicators to improve feeding strategies such as diet composition, feeding ration and frequency in juvenile tuna.