A wide variety of dry-fermented sausages are produced in European countries, where are considered valued traditional products. An intense proteolysis takes place during the processing of dry-fermented sausages due to the combined action of muscle and microbial peptidases, generating large amounts of peptides and free amino acids. These compounds participate in the development of the characteristic flavor of dry-fermented products, but some peptides can also exert certain bioactivities such as antioxidant and ACE inhibitory activities. This study has evaluated the changes in peptide profile and amino acid contents of three European dry-fermented sausages produced in Spain, Italy and Belgium, proving the intense degradation of proteins, mainly myofibrillar, and the generation of high amounts of different size peptides and free amino acids. The changes observed between the profiles of European sausages could be due to differences in product formulation, processing conditions and starter cultures used, which influence the activity of enzymes, both from muscle and bacterial origin. On the other hand, the bioactivity profile of each type of dry-fermented sausage was evaluated through the measurement of the ACE inhibitory and antioxidant activities in water-soluble peptide extracts fractionated by size-exclusion chromatography. Spanish and Belgian dry-fermented sausages showed values of ACE inhibition around 85%, whereas Belgian samples presented the highest DPPH radical-scavenging activity and ferric reducing power capacity. These results evidence the potential of Spanish, Italian and Belgian dry-fermented sausages as natural sources of bioactive peptides, giving an added-value to these traditional products.

30Angiotensin converting enzyme (ACE) inhibitory peptides are been extensively studied 31as an alternative to synthetic drugs for the treatment of hypertension. Recent studies 32 have shown that dry-cured ham is an important source of naturally generated bioactive 33 peptides, especially showing ACE inhibitory activity. However, due to their excessive 34 degradation by digestive and brush-border enzymes, it is not clear whether these 35 peptides resist intestinal absorption and reach the blood stream where they may exert 36 their antihypertensive effect. So, dDry-cured ham extracts and specific pure peptides 37 naturally generated during the dry-curing process, showing ACE inhibitory activity, 38 have been studied for their stability during transepithelial transport in a Caco-2 cell 39 monolayer. The ACE inhibitory activity of transport samples was assayed, reaching the 40 highest values in apical samples after 15 min of incubation. In basal solutions, the 41 highest ACE inhibition was observed for peptides AAPLAP and KPVAAP after 60 min 42 of cellular transport. However, when basal samples were four times concentrated, a 43 considerable increased ACE inhibitory activity was observed in these peptides from 15 44 min of incubation. Fragments generated by cellular activity were detected by using 45 tandem mass spectrometry MStechniques, showing that AAATP, AAPLAP, and 46 KPVAAP were hydrolysed during the transport, although KPVAAP was also absorbed 47 intactly. This study highlights the potential of intact dry-cured ham peptides as well as 48 their fragments to be absorbed across the intestinal epithelium and reach the blood 49 stream to exert an antihypertensive action. 50 52Keywords: Dry-cured ham, ACE inhibitory peptides, Caco-2 cell monolayer, intestinal 53 transport, mass spectrometry.

There is a wide variety of peptides released from food proteins which are able to exert a relevant benefit for human health like angiotensin-converting enzyme (ACE) inhibition, antioxidant, anti-inflammatory, hypoglucemic or antithrombotic activity, among others. This manuscript is reviewing the recent advances on enzymatic mechanisms for the hydrolysis of food proteins, including the types of enzymes and mechanisms of action involved, the strategies followed for the isolation and identification of bioactive peptides through advanced proteomic tools, the assessment of bioactivity and its beneficial effects. Specific applications in fermented and/or ripened foods where a significant number of bioactive peptides have been reported with relevant in vivo physiological effects on laboratory rats and humans, as well as the hydrolysis of food proteins for the production of bioactive peptides are also reviewed.