The kinetics of angiotensin-I converting enzyme inhibitory peptide production from bonito muscle in an artificial gastric juice was studied. Produced peptides were purified with an affinity column coupled with an antibody against an inhibitory peptide found in tuna (PTHIKWGD), and further with an HPLC column, and then characterized. Although the inhibitory effect was observed immediately after the start of digestion in samples before affinity purification, most activities were due to generated peptide fragments as competitive substrates of ACE. Potent inhibitory activities as real inhibitors were mostly found in the affinity purified peptide fragments. After 24-48 h digestion, two peptides HERDPTHIKWGD and PTHIKWGD, which contained the same sequence as the tuna inhibitory peptide, were major components in the affinity purified samples and their IC50 values were about 8 mu M. (c) 2005 Elsevier Ltd. All tights reserved.