Angiotensin I-converting enzyme inhibitory peptides from hydrolyzed cowpea flour

Lightly roasted cowpea flour was hydrolyzed with Alcalase for 6 h and the resulting hydrolysate with IC50 value being 123.6 mu g/ml was used to isolate angiotensin I-converting enzyme (ACE) inhibitory peptides. The hydrolysate was first filtered through a polyethersulphone membrane with a molecular weight cutoff (MWCO) of 10 kDa and the resulting permeate had an IC50 value of 95.7 mu g/ml. After separation by primary reverse-phase high performance liquid chromatography (RP-HPLC) into six fractions, the most potent fraction with eluting time 50-60 min had an IC50 value of 58.5 mu g/ml and was used for further purification. After isolation by secondary RP-HPLC, four peaks were found to have the strong ACE inhibition, and their IC50 values were determined to be 22.0, 25.0, 31.1 and 28.4 mu g/ml, respectively. Peptide mass for the most potent peak I was obtained by electrospray ionization (ESI) mass spectrometer and sequence was determined by matrix-assisted laser desorption and ionization (MALDI) tandem TOF-TOF (time-of-flight) mass spectrometer (MS/MS) to be Phe-Phe. ACE inhibition pattern of the dipeptide was found to be competitive. We suggest that products from different purification steps of cowpea protein hydrolysate may be used as health promoting ingredients in functional foods.