Abstract: The aim of this work was the purification and identification of the major angiotensin converting enzyme (ACE) inhibitory peptides produced by enzymatic hydrolysis of a protein concentrate recovered from a cuttlefish industrial manufacturing effluent. This process consisted on the ultrafiltration of cuttlefish softening wastewater, with a 10 kDa cut-off membrane, followed by the hydrolysis with alcalase of the retained fraction. Alcalase produced ACE inhibitors reaching the highest activity (IC50 = 76.8 ± 15.2 μg mL−1) after 8 h of proteolysis. Sequential ultrafiltration of the 8 h hydrolysate with molecular weight cut-off (MWCO) membranes of 10 and 1 kDa resulted in the increased activity of each permeate, with a final IC50 value of 58.4 ± 4.6 μg mL−1. Permeate containing peptides lower than 1 kDa was separated by reversed-phase high performance liquid chromatography (RP-HPLC). Four fractions (A–D) with potent ACE inhibitory activity were isolated and their main peptides identified using high performance liquid chromatography coupled to an electrospray ion trap Fourier transform ion cyclotron resonance-mass spectrometer (HPLC-ESI-IT-FTICR) followed by comparison with databases and de novo sequencing. The amino acid sequences of the identified peptides contained at least one hydrophobic and/or a proline together with positively charged residues in at least one of the three C-terminal positions. The IC50 values of the fractions ranged from 1.92 to 8.83 μg mL−1, however this study fails to identify which of these peptides are ultimately responsible for the potent antihypertensive activity of these fractions.
Keywords: ultrafiltration; proteolysis; ACE inhibitory peptides; cuttlefish byproducts; peptide identification; HPLC-ESI-MS
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Amado, I.R.; Vázquez, J.A.; González, P.; Esteban-Fernández, D.; Carrera, M.; Piñeiro, C. Identification of the Major ACE-Inhibitory Peptides Produced by Enzymatic Hydrolysis of a Protein Concentrate from Cuttlefish Wastewater. Mar. Drugs 2014, 12, 1390-1405.
Amado IR, Vázquez JA, González P, Esteban-Fernández D, Carrera M, Piñeiro C. Identification of the Major ACE-Inhibitory Peptides Produced by Enzymatic Hydrolysis of a Protein Concentrate from Cuttlefish Wastewater. Marine Drugs. 2014; 12(3):1390-1405.
Amado, Isabel R.; Vázquez, José A.; González, Pilar; Esteban-Fernández, Diego; Carrera, Mónica; Piñeiro, Carmen. 2014. "Identification of the Major ACE-Inhibitory Peptides Produced by Enzymatic Hydrolysis of a Protein Concentrate from Cuttlefish Wastewater." Mar. Drugs 12, no. 3: 1390-1405.