Systematical Investigation on Anti-Fatigue Function and Underlying Mechanism of High Fischer Ratio Oligopeptides from Antarctic Krill on Exercise-Induced Fatigue in Mice.

High Fischer ratio oligopeptides (HFOs) have a variety of biological activities, but their mechanisms of action for anti-fatigue are less systematically studied at present. This study aimed to systematically evaluate the anti-fatigue efficacy of HFOs from Antarctic krill (HFOs-AK) and explore its mechanism of action through establishing the fatigue model of endurance swimming in mice. Therefore, according to the comparison with the endurance swimming model group, HFOs-AK were able to dose-dependently prolong the endurance swimming time, reduce the levels of the metabolites (lactic acid, blood urea nitrogen, and blood ammonia), increase the content of blood glucose, muscle glycogen, and liver glycogen, reduce lactate dehydrogenase and creatine kinase extravasation, and protect muscle tissue from damage in the endurance swimming mice. HFOs-AK were shown to enhance Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities and increase ATP content in muscle tissue. Meanwhile, HFOs-AK also showed significantly antioxidant ability by increasing the activities of superoxide dismutase and glutathione peroxidase in the liver and decreasing the level of malondialdehyde. Further studies showed that HFOs-AK could regulate the body's energy metabolism and thus exert its anti-fatigue effects by activating the AMPK signaling pathway and up-regulating the expression of p-AMPK and PGC-α proteins. Therefore, HFOs-AK can be used as an auxiliary functional dietary molecules to exert its good anti-fatigue activity and be applied to anti-fatigue functional foods.

Preparation, Identification, Molecular Docking Study and Protective Function on HUVECs of Novel ACE Inhibitory Peptides from Protein Hydrolysate of Skipjack Tuna Muscle.

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.

Purification, Identification, Activity Evaluation, and Stability of Antioxidant Peptides from Alcalase Hydrolysate of Antarctic Krill (Euphausia superba) Proteins.

For utilizing the largest source of marine proteins, Antarctic krill (Euphausia superba) proteins were defatted and hydrolyzed separately using pepsin, alcalase, papain, trypsin, and netrase, and alcalase hydrolysate (EPAH) showed the highest DPPH radical (DPPH·) and hydroxyl radical (HO·) scavenging activity among five hydrolysates. Using ultrafiltration and chromatography methods, fifteen antioxidant peptides were purified from EPAH and identified as Asn-Gln-Met (NQM), Trp-Phe-Pro-Met (WFPM), Gln-Asn-Pro-Thr (QNPT), Tyr-Met-Asn-Phe (YMNF), Ser-Gly-Pro-Ala (SGPA), Ser-Leu-Pro-Tyr (SLPY), Gln-Tyr-Pro-Pro-Met-Gln-Tyr (QYPPMQY), Glu-Tyr-Glu-Ala (EYEA), Asn-Trp-Asp-Asp-Met-Arg-Ile-Val-Ala-Val (NWDDMRIVAV), Trp-Asp-Asp-Met-Glu-Arg-Leu-Val-Met-Ile (WDDMERLVMI), Asn-Trp-Asp-Asp-Met-Glu-Pro-Ser-Phe (NWD-DMEPSF), Asn-Gly-Pro-Asp-Pro-Arg-Pro-Ser-Gln-Gln (NGPDPRPSQQ), Ala-Phe-Leu-Trp-Asn (AFLWA), Asn-Val-Pro-Asp-Met (NVPDM), and Thr-Phe-Pro-Ile-Tyr-Asp-Tyr-Pro-Gln (TFPIYDPQ), respectively, using a protein sequencer and ESI/MS. Among fifteen antioxidant peptides, SLPY, QYPPMQY and EYEA showed the highest scavenging activities on DPPH· (EC50 values of 1.18 ± 0.036, 1.547 ± 0.150, and 1.372 ± 0.274 mg/mL, respectively), HO· (EC50 values of 0.826 ± 0.027, 1.022 ± 0.058, and 0.946 ± 0.011 mg/mL, respectively), and superoxide anion radical (EC50 values of 0.789 ± 0.079, 0.913 ± 0.007, and 0.793 ± 0.056 mg/mL, respectively). Moreover, SLPY, QYPPMQY and EYEA showed strong reducing power, protective capability against H2O2-damaged plasmid DNA, and lipid peroxidation inhibition ability. Furthermore, SLPY, QYPPMQY, and EYEA had high stability under temperatures lower than 80 °C, pH values ranged from 6-8, and simulated GI digestion for 180 min. The results showed that fifteen antioxidant peptides from alcalase hydrolysate of Antarctic krill proteins, especially SLPY, QYPPMQY and EYEA, might serve as effective antioxidant agents applied in food and health products.

Novel angiotensin-converting enzyme inhibitory peptides from tuna byproducts-milts: Preparation, characterization, molecular docking study, and antioxidant function on H2O2-damaged human umbilical vein endothelial cells.

To prepare peptides with high angiotensin-converting enzyme (ACE) inhibitory (ACEi) activity, Alcalase was screened from five proteases and employed to prepare protein hydrolysate (TMH) of skipjack tuna (Katsuwonus pelamis) milts. Subsequently, 10 novel ACEi peptides were isolated from the high-ACEi activity TMH and identified as Tyr-Asp-Asp (YDD), Thr-Arg-Glu (TRE), Arg-Asp-Tyr (RDY), Thr-Glu-Arg-Met (TERM), Asp-Arg-Arg-Tyr-Gly (DRRYG), Ile-Cys-Tyr (ICY), Leu-Ser-Phe-Arg (LSFR), Gly-Val-Arg-Phe (GVRF), Lys-Leu-Tyr-Ala-Leu-Phe (KLYALF), and Ile-Tyr-Ser-Pro (IYSP) with molecular weights of 411.35, 404.41, 452.45, 535.60, 665.69, 397.48, 521.61, 477.55, 753.91, and 478.53 Da, respectively. Among them, the IC50 values of ICY, LSFR, and IYSP on ACE were 0.48, 0.59, and 0.76 mg/mL, respectively. The significant ACEi activity of ICY, LSFR, and IYSP with affinities of -7.0, -8.5, and -8.3 kcal/mol mainly attributed to effectively combining with the ACEi active sites through hydrogen bonding, electrostatic force, and hydrophobic interaction. Moreover, ICY, LSFR, and IYSP could positively influence the production of nitric oxide (NO) and endothelin-1 (ET-1) secretion in human umbilical vein endothelial cells (HUVECs) and weaken the adverse impact of norepinephrine (NE) on the production of NO and ET-1. In addition, ICY, LSFR, and IYSP could provide significant protection to HUVECs against H2O2 damage by increasing antioxidase levels to decrease the contents of reactive oxide species and malondialdehyde. Therefore, the ACEi peptides of ICY, LSFR, and IYSP are beneficial functional molecules for healthy foods against hypertension and cardiovascular diseases.

Preparation, Characterization, and Cytoprotective Effects on HUVECs of Fourteen Novel Angiotensin-I-Converting Enzyme Inhibitory Peptides From Protein Hydrolysate of Tuna Processing By-Products.

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.

Seventeen novel angiotensin converting enzyme (ACE) inhibitory peptides from the protein hydrolysate of Mytilus edulis: isolation, identification, molecular docking study, and protective function on HUVECs.

In the study, seventeen angiotensin converting enzyme (ACE) inhibitory peptides were isolated from the protein hydrolysate of blue mussel (Mytilus edulis) and identified as MFR, MFV, FV, KP, QP, QVK, IK, YKV, IRK, MLKV, NFRPQ, YEGDP, WF, GPE, SWISS, SVEWK, and FKWH, respectively. Among them, IK, YEGDP, WF, and SWISS showed the strongest ACE inhibitory activity with IC50 values of 0.77 ± 0.020, 0.19 ± 0.010, 0.40 ± 0.015, and 0.32 ± 0.017 mg mL-1, respectively. Molecular docking study indicated that IK, YEGDP, WF, and SWISS exhibited better inhibitory activity attributed to its effective interaction with the active site of ACE by hydrogen bonding, electrostatic force and hydrophobic interaction. Furthermore, IK, YEGDP and WF perform an important protective function on human umbilical vein endothelial cells (HUVECs) by increasing nitric oxide (NO) content, decreasing endothelin-1 (ET-1) secretion, and antagonizing the adverse impact of norepinephrine on the secretion of NO and ET-1. In addition, YEGDP and WF could provide protection to HUVECs against H2O2 damage by increasing superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and NO levels to decrease the contents of reactive oxygen species (ROS) and malondialdehyde. Therefore, seventeen ACE inhibitory peptides, especially YEGDP and WF, might be used as natural ingredients for the development of products with antihypertensive functions.