title: Identification of the major ACE-inhibitory peptides produced by enzymatic hydrolysis of a protein concentrate from cuttlefish wastewater.
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.
authors.affiliation: Group of Recycling and Valorisation of Waste Materials (REVAL), Marine Research Institute (IIM-CSIC), r/Eduardo Cabello, 6. Vigo, Galicia 36208, Spain. sabelara@iim.csic.es.
authors.forename: Isabel Rodríguez
authors.initials: IR
authors.lastname: Amado
authors.affiliation: Group of Recycling and Valorisation of Waste Materials (REVAL), Marine Research Institute (IIM-CSIC), r/Eduardo Cabello, 6. Vigo, Galicia 36208, Spain. jvazquez@iim.csic.es.
authors.forename: José Antonio
authors.initials: JA
authors.lastname: Vázquez
authors.affiliation: Group of Recycling and Valorisation of Waste Materials (REVAL), Marine Research Institute (IIM-CSIC), r/Eduardo Cabello, 6. Vigo, Galicia 36208, Spain. pgonzalez@iim.csic.es.
authors.forename: Pilar
authors.initials: P
authors.lastname: González
authors.affiliation: Department of Chemistry, Humboldt-Universitaet zu Berlin, Brook-Taylor Strasse 2, Berlin 12489, Germany. esteband@chemie.hu-berlin.de.
authors.forename: Diego
authors.initials: D
authors.lastname: Esteban-Fernández
authors.affiliation: Institute of Molecular Systems Biology (IMSB), ETH Zürich, Zürich 8093, Switzerland. mcarrera@iim.csic.es.
authors.forename: Mónica
authors.initials: M
authors.lastname: Carrera
authors.affiliation: Scientific Instrumentation Service (SICIM), Marine Research Institute (IIM-CSIC), r/ Eduardo Cabello, 6. Vigo, Galicia 36208, Spain. cpineiro@iim.csic.es.
authors.forename: Carmen
authors.initials: C
authors.lastname: Piñeiro
mesh_terms: D000465:Algorithms; D000806:Angiotensin-Converting Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D049832:Decapodiformes / Q000737:chemistry / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D019649:Food Industry; D006868:Hydrolysis; D007220:Industrial Waste / Q000032:analysis*; D008970:Molecular Weight; D017711:Nonlinear Dynamics; D010455:Peptides / Q000302:isolation & purification / Q000494:pharmacology*; D021241:Spectrometry, Mass, Electrospray Ionization; D014462:Ultrafiltration; D062065:Wastewater / Q000032:analysis*
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D007220:Industrial Waste; D010455:Peptides; D062065:Wastewater