5TH CONGRESS OF THE INTERNATIONAL SOCIETY FOR APPLIED PHYCOLOGY Bioassay-guided fractionation approach for determination of protein precursors of proteolytic bioactive metabolites from macroalgae Stéphanie Bondu & Claudie Bonnet & Julie Gaubert & Éric Deslandes & Sylvie L. Turgeon & Lucie Beaulieu Received: 28 July 2014 /Revised and accepted: 5 October 2014 # Springer Science+Business Media Dordrecht 2014 Abstract In the last decades, an upsurge in the occurrence of chronic diseases caused by hypertension and oxidative stress has been observed. The objective of this research was to isolate and characterize groups of antioxidant and anti- hypertensive natural bioactive peptides originating from hy- drolyzed proteins of the red seaweeds Solieria chordalis and Palmaria palmata, the green seaweed Ulva lactuca and the brown seaweed Saccharina longicruris. Enzymatic hydroly- sis by trypsin and chymotrypsin was performed in order to release bioactive algal peptides. Three antioxidant assays, 2,2- diphenyl-1-picrylhydrazyl (DPPH), ferric ion reducing anti- oxidant power (FRAP) and oxygen radical absorbance capac- ity (ORAC), were used to determine the activity of seaweed peptides. Angiotensin-converting enzyme (ACE) inhibition assay was used to evaluate the anti-hypertensive potential. Additionally, liquid chromatography-tandem mass spectrom- etry (LC-MS/MS) analyses and database searches allowed for protein identification, indicating where these bioactive pep- tides originated from. Fractions of red seaweeds, especially S. chordalis, produced by chymotrypsin and trypsin hydrolysis, exhibited higher in vitro antioxidant and ACE inhibitory activities than the parent proteins. Size exclusion chromatography highlighted two groups of bioactive peptides: (i) peptides with molecular weight (MW) between 1,400 and 3,200 Da displaying antioxidant activities and (ii) smaller peptides (MW <1,000 Da) displaying both antioxidant and ACE inhibitory activities. LC-MS/MS analyses of the antiox- idant subfractions of P. palmata and S. chordalis were per- formed, and several peptide sequences were elucidated, using tandem mass spectrometry. Of all identified peptides of S. chordalis, 61 and 43 % came from the hydrolysis of ribulose-1, 5-biphosphate carboxylase/oxygenase (RuBiSCo) enzymes. Phycoerythrin, elongation factors, photosystems and cytochrome oxidase clusters represented between 3 and 14 % of identified peptides in S. chordalis fractions. This is the first time, to the author’ s knowledge, that distinction between several groups of bioactive peptides, within seaweed protein hydrolysates, has been reported. In addition, proteins of origin were also identified. Keywords ACE inhibitory activity . Antioxidant capacity . Hydrolysates . Peptide amino acid identification . RuBisCo enzymes . Seaweeds Introduction Free radicals and reactive oxygen species (ROS) are normally eliminated from the organism by antioxidant enzymes and non-enzymatic factors (Je et al. 2009), although under stress- ful conditions, this defence system may be disrupted. An excess of free radicals in the body can cause oxidation of macromolecules such as DNA, amino acids, proteins, lipids and polysaccharides (Je et al. 2009; Kang et al. 2013; Tierney et al. 2013). Oxidative stress consequently may lead to chronic diseases such as hypertension, diabetes, cancer, atherosclero- sis, arthritis, neurological disorders and inflammatory ill- nesses (Heo et al. 2005; Je et al. 2009; Tierney et al. 2013). S. Bondu : C. Bonnet : J. Gaubert : S. L. Turgeon : L. Beaulieu (*) Département des sciences des aliments et de nutrition, Institut sur la nutrition et les aliments fonctionnels (INAF), Université Laval, 2425 rue de l’Agriculture, Québec, Québec G1V 0A6, Canada e-mail: lucie.beaulieu@fsaa.ulaval.ca S. Bondu : C. Bonnet : J. Gaubert : L. Beaulieu Collectif de Recherche Appliquée aux Bioprocédés et à la chimie de l’Environnement (CRABE), Université du Québec à Rimouski, 300 Allée des Ursulines, Rimouski, QC G5L 3A1, Canada É. Deslandes Laboratoire des Sciences de l’Environnement Marin (LEMAR), Institut Universitaire Européen de la Mer, Université de Bretagne Occidentale, Technopôle de Brest-Iroise, 29280 Plouzané, France J Appl Phycol DOI 10.1007/s10811-014-0425-0