ABSTRACT
To effectively utilize skipjack tuna (Katsuwonus pelamis) processing by-products to prepare peptides with high angiotensin-I-converting enzyme (ACE) inhibitory (ACEi) activity, Neutrase was selected from five kinds of protease for hydrolyzing skipjack tuna dark muscle, and its best hydrolysis conditions were optimized as enzyme dose of 1.6%, pH 6.7, and temperature of 50°C using single factor and response surface experiments. Subsequently, 14 novel ACEi peptides were prepared from the high ACEi protein hydrolysate and identified as TE, AG, MWN, MEKS, VK, MQR, MKKS, VKRT, IPK, YNY, LPRS, FEK, IRR, and WERGE. MWN, MEKS, MKKS, and LPRS displayed significantly ACEi activity with IC50 values of 0.328 ± 0.035, 0.527 ± 0.030, 0.269 ± 0.006, and 0.495 ± 0.024 mg/mL, respectively. Furthermore, LPRS showed the highest increasing ability on nitric oxide (NO) production among four ACEi peptides combining the direct increase and reversing the negative influence of norepinephrine (NE), and MKKS showed the highest ability on directly decreasing and reversing the side effects of NE on the secretion level of endothelin-1 (ET-1) among four ACEi peptides. These findings demonstrate that seafood by-product proteins are potential ACEi peptide sources and prepared ACEi peptides from skipjack tuna dark muscle, which are beneficial components for functional food against hypertension and cardiovascular diseases.
ABSTRACT
To prepare bioactive peptides with high angiotensin-I-converting enzyme (ACE)-inhibitory (ACEi) activity, Alcalase was selected from five kinds of protease for hydrolyzing Skipjack tuna (Katsuwonus pelamis) muscle, and its best hydrolysis conditions were optimized using single factor and response surface experiments. Then, the high ACEi protein hydrolysate (TMPH) of skipjack tuna muscle was prepared using Alcalase under the optimum conditions of enzyme dose 2.3%, enzymolysis temperature 56.2 °C, and pH 9.4, and its ACEi activity reached 72.71% at 1.0 mg/mL. Subsequently, six novel ACEi peptides were prepared from TMPH using ultrafiltration and chromatography methods and were identified as Ser-Pro (SP), Val-Asp-Arg-Tyr-Phe (VDRYF), Val-His-Gly-Val-Val (VHGVV), Tyr-Glu (YE), Phe-Glu-Met (FEM), and Phe-Trp-Arg-Val (FWRV), with molecular weights of 202.3, 698.9, 509.7, 310.4, 425.6, and 606.8 Da, respectively. SP and VDRYF displayed noticeable ACEi activity, with IC50 values of 0.06 ± 0.01 and 0.28 ± 0.03 mg/mL, respectively. Molecular docking analysis illustrated that the high ACEi activity of SP and VDRYF was attributed to effective interaction with the active sites/pockets of ACE by hydrogen bonding, electrostatic force, and hydrophobic interaction. Furthermore, SP and VDRYF could significantly up-regulate nitric oxide (NO) production and down-regulate endothelin-1 (ET-1) secretion in HUVECs after 24 h treatment, but also abolish the negative effect of 0.5 µM norepinephrine (NE) on the generation of NO and ET-1. Therefore, ACEi peptides derived from skipjack tuna (K. pelamis) muscle, especially SP and VDRYF, are beneficial components for functional food against hypertension and cardiovascular diseases.
Subject(s)
Angiotensin-Converting Enzyme Inhibitors , Muscle, Skeletal/chemistry , Peptides , Tuna , Amino Acid Sequence , Angiotensin-Converting Enzyme Inhibitors/chemistry , Angiotensin-Converting Enzyme Inhibitors/isolation & purification , Angiotensin-Converting Enzyme Inhibitors/pharmacology , Animals , Cell Survival/drug effects , Endothelin-1/metabolism , Functional Food , Human Umbilical Vein Endothelial Cells/drug effects , Human Umbilical Vein Endothelial Cells/metabolism , Humans , Hydrolysis , Molecular Docking Simulation , Nitric Oxide/metabolism , Peptides/chemistry , Peptides/isolation & purification , Peptides/pharmacology , Protein Hydrolysates/chemistry , Subtilisins/chemistryABSTRACT
To look for the collagen alternatives of mammalian cartilages from aquatics and their by-products, acid-soluble collagen (ASC-SC) and pepsin-soluble collagen (PSC-SC) were extracted from cartilages of Siberian sturgeon (Acipenser baerii) with yields of 27.13 ± 1.15 and 14.69 ± 0.85% on dry weight basis. ASC-SC and PSC-SC had glycine as the major amino acid with the contents of 326.8 and 327.5 residues 1000 residues-1, and their contents of proline and hydroxyproline were 205.9 and 208.0 residues 1000 residues-1. ASC-SC and PSC-SC comprised type I collagen ([α1(I)]2α2(I)) and type II collagen ([α1(II)]3) on the literatures and results of amino acid composition, SDS-PAGE pattern, UV, and FTIR spectra. Meanwhile, FTIR spectra data indicated that there were more hydrogen bonds in ASC-SC and more intermolecular crosslinks in PSC-SC. The maximum transition temperature (Tmax) of the ASC (28.3 °C) and PSC (30.5 °C) was lower than those of collagens from mammalian cartilages (> 37 °C). ASC-SC and PSC-SC showed high solubility in the acidic pH ranges and the solubility decreased in the presence of NaCl at concentrations above 3%. Zeta potential studies indicated that both ASC-SC and PSC-SC exhibited a net zero charge at pH 6.30 and 6.32. SEM results indicated that ASC-SC and PSC-SC presented irregular dense sheet-like film linked by random-coiled filaments. Therefore, collagens from Siberian sturgeon cartilages might be the suitable alternatives of the collagens of mammal cartilages as functional ingredient to treat some diseases.
