title: Egg-yolk protein by-product as a source of ACE-inhibitory peptides obtained with using unconventional proteinase from Asian pumpkin (Cucurbita ficifolia).
abstract: In the present study angiotensin I-converting enzyme (ACE) inhibitory peptides were isolated from egg-yolk protein preparation (YP). Enzymatic hydrolysis conducted using unconventional enzyme from Cucurbita ficifolia (dose: 1000 U/mg of hydrolyzed YP (E/S (w/w)=1:7.52)) was employed to obtain protein hydrolysates. The 4-h hydrolysate exhibited a significant (IC₅₀=482.5 μg/mL) ACE inhibitory activity. Moreover, hydrolysate showed no cytotoxic activity on human and animal cell lines which makes it a very useful multifunctional method for peptide preparation. The compiled isolation procedure (ultrafiltration, size-exclusion chromatography and RP-HPLC) of bioactive peptides from YP hydrolysate resulted in obtaining peptides with the strong ACE inhibitory activity. One homogeneous and three heterogeneous peptide fractions were identified. The peptides were composed of 9-18 amino-acid residues, including mainly arginine and leucine at the N-terminal positions. To confirm the selected bioactive peptide sequences their analogs were chemically synthesized and tested. Peptide LAPSLPGKPKPD showed the strongest ACE inhibitory activity, with IC₅₀ value of 1.97 μmol/L.
  
  
  CONCLUSIONS
  Peptides with specific biological activity can be used in pharmaceutical, cosmetic or food industries. Because of their potential role as physiological modulators, as well as theirhigh safety profile, they can be used as natural pharmacological compounds or functional food ingredients. The development of biotechnological solutions to obtain peptides with desired biological activity is already in progress. Studies in this area are focused on using unconventional highly specific enzymes and more efficient methods developed to conduct food process technologies. Natural peptides have many advantages. They are mainly toxicologically safe, have wide spectra of therapeutic actions, exhibit less side effects compared to synthetic drugs and are more efficiently absorbed in the intestinal tract. The complexity of operation of large scale technologies and high cost of purification techniques are limiting factors to the commercialization of food-derived bioactive peptides. Research on the isolation of bioactive peptides in order to reduce the processing time and costs is continuously developing. Bioactive peptides can also be released from protein by-products of the food industry, which reduce the substrate expense and production cost as well as provide the added advantage of an efficient waste disposal. Moreover, proteins as precursors of food-derived peptides are well-tolerated by the human body and therefore their application in drug development may reduce costs and duration of toxicological studies during research, development and clinical trials.
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Ewelina
authors.initials: E
authors.lastname: Eckert
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland. Electronic address: aleksandra.zambrowicz@up.wroc.pl.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Zambrowicz
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Marta
authors.initials: M
authors.lastname: Pokora
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Bartosz
authors.initials: B
authors.lastname: Setner
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Dąbrowska
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Marek
authors.initials: M
authors.lastname: Szołtysik
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Zbigniew
authors.initials: Z
authors.lastname: Szewczuk
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Antoni
authors.initials: A
authors.lastname: Polanowski
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Tadeusz
authors.initials: T
authors.lastname: Trziszka
authors.affiliation: Department of Animal Products Technology and Quality Management, Wroclaw University of Environmental and Life Sciences, Chełmońskiego 37/41, 51-630 Wrocław, Poland.
authors.forename: Józefa
authors.initials: J
authors.lastname: Chrzanowska
mesh_terms: D000595:Amino Acid Sequence; D000806:Angiotensin-Converting Enzyme Inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D028464:Cucurbita / Q000145:classification / Q000201:enzymology*; D004527:Egg Proteins / Q000737:chemistry* / Q000302:isolation & purification; D004789:Enzyme Activation; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification; D013045:Species Specificity
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D004527:Egg Proteins; D010455:Peptides; D010940:Plant Proteins; D010447:Peptide Hydrolases
keywords: ACE inhibitory activity; Cucurbita ficifolia; Egg yolk protein; Enzymatic hydrolysis