Identification of ACE-inhibitory peptides in different Spanish cheeses by tandem mass spectrometry

Different Spanish cheeses (Cabrales, Idiazabal, Roncal, Manchego, Mahon and a goat's milk cheese), made with diverse technologies and milk from different species, were studied for their angiotensin converting enzyme (ACE)-inhibitory activity. Among the water-soluble extract WSE < 1000-Da of the different samples, Cabrales cheese was the most active with an inhibitory activity of 76.1%. On the contrary, Mahon cheese showed the lowest ACE-inhibitory index (56.6%). In order to identify the peptides involved in this activity, the WSE < 1000-Da were analyzed by HPLC-MS/MS and off-line MS/MS. A total of 41 major peptides were identified in the different cheeses. Most of them derived from beta-and alpha(s1)-casein. Several of these peptides were selected on the basis of the presence of proline as the C-terminal amino acid, and they were chemically synthesized. All peptides showed moderate or low ACE-inhibitory activity. Peptide DKIHP [beta-CN f(47-51)], a sequence detected in all samples except Mahon cheese, showed the highest inhibitory activity with an IC50 value of 113.1 mu M.