The time required for trypsin restoration in the pyloric ceca-pancreatic tissue of yellowtail Seriola quinqueradiata (average body weight 942 g) was estimated. Fish were collected after starvation for 24, 48 or 72 h, and 6 h after being fed after each of the three tested starvation periods. Pyloric ceca isolated together with the pancreas were used for measurements of trypsin activity, quantity and expression rate. In the starved group, trypsin activities after 72 and 48 h starvation were significantly higher than after 24 h. Trypsin quantity, which was estimated by a Western blot test, showed similar results. The expression rate, estimated by reverse transcription polymerase chain reaction, was high after 24 h, but decreased after 48 h starvation. Thus, it was concluded that trypsin reaches the maximum storage level after 48 h starvation. In the fed group, the reduction in trypsin activity was greater when fish were fed after 48 and 72 h starvation periods than after 24 h. Trypsin expression rates 6 h after feeding were higher than before feeding. This indicates that trypsin synthesis is initiated after feeding or trypsin secretion and continues until 48 h of food deprivation. These results indicate that yellowtail of this size require feeding intervals of about 48 h to maximize trypsin storage.