Enzymatic Hydrolysis of Porcine Blood as a Strategy to Obtain a Peptide-Rich Functional Ingredient.

The sustainable revalorization of porcine blood is crucial due to the large volumes daily generated in slaughterhouses. The aim of this study was to obtain a novel ingredient rich in free amino acids and bioactive peptides from the sequential hydrolysis of porcine blood. Porcine blood was hydrolyzed with Alcalase 4.0 L and Protana™ Prime enzymes, followed by molecular weight fractionation (<10 kDa) and spray-drying. The antioxidant, hypoglycemic, and anti-inflammatory bioactivities of the resulting hydrolysate (PBSH) were studied in vitro. Further fractionation by reversed-phase high-performance liquid chromatography (RP-HPLC) was performed to isolate the most bioactive fraction based on polarity. Peptides from fraction 1 (F1) were identified using LC-MS/MS and analyzed in silico. Finally, some peptides were synthesized, and their bioactivity was subsequently assessed. PBSH hydrolysate showed antioxidant activity with IC(50) values of 2.09, 135.05, and 26.73 mg/mL for ABTS, FRAP, and DPPH assays, respectively. Additionally, PBSH exhibited hypoglycemic, anti-inflammatory, and immunomodulatory potential through the inhibition of DPP-IV (82.78%), NEP (84.72%), TACE (50.79%), and MGL (69.08%) enzymes at a concentration of 20, 20, 100, and 20 mg/mL, respectively. Peptides PDDFNPS, FPPKPKD, DNPIPK, GHLDDLPG, and GDL were identified in the most polar and bioactive fraction (F1) and proved a synergistic hypoglycemic effect at a concentration of 1 mmol/L. The peptide PDDFNPS exhibited multifunctional properties with 56.43% inhibition of DPP-IV and 83.54% inhibition of NEP. PBSH resulted in a novel functional ingredient for animal feed as it contains a variety of identified bioactive peptides and a high amount of free amino acids.