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

Different Spanish cheeses (Cabrales, Idiazábal, Roncal, Manchego, Mahón 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, Mahón 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 β-and α_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 β-CN f(47–51)], a sequence detected in all samples except Mahón cheese, showed the highest inhibitory activity with an IC₅₀ value of 113.1 μM.