Study On The Antioxidant Activity And Antifatigue Effect Of Mackerel Peptides

Marine low-value fishes had high productivity and high protein content, it wouldbe of great importance in processing the low-value fishes into special bioactivepeptides. In our study, the antioxidant active peptides were prepared from mackerelprotein which with high protein content through enzymatic technology andultrafiltration. Their antioxidant activity stabilities were also studied, andion-exchange chromatography and gel filtration chromatography were used toseparate and purify the target peptides. Finally, chemical models in vitro and animalmodels in vivo were used to evaluate the antioxidant activity and antifatigue effect oftarget peptides, and the relationship between antioxidant activity and antifatigue effectof target peptides were discussed. The research results of this study were as follows:1. Determine the optimum enzymatic hydrolysis technology of mackerel peptides:Mackerel was a material with high protein content (16.40%), low fat content and avariety of amino acids, especially Glu, Lys and Leu which had been reported tocontribute greatly to the potency of antioxidant peptides, had high content. Themackerel was appropriate to prepare antioxidant peptides. Five kinds of proteases, i.e.trypsin, acid protease, papain, flavourzymeand neutrase were used for hydrolysis toproduce the mackerel protein hydrolysate, considering the degree of hydrolysis,nitrogen recovery, DPPH radical scavenging activity and the relative molecularweight distributions, neutrase was the best enzyme for the hydrolysis. On the basis ofsingle factor experiment, the suggested hydrolysis conditions were performed by theresponse surface methodology (RSM): enzyme concentration1726.85U/g,pH7.00,temperature39.55℃, time5.50h and water to substrate at25:1. Themaximum DPPH scavenging activity was78.43%0.32, which was similar to theexperimental value of79.19%.2. The research on the separation and purification of mackerel peptides: TheMPH was fractionated into five portions using ultrafiltration, the results of antioxidantactivities showed that the MPHs with molecular weight below2.5kDa (MPH-III, MPH-IV and MPH-V) exhibited higher antioxidant activities. Ion-exchangechromatography separated the peptides on the basis of different number of charge andgel filtration chromatography separated the peptides on the different size of molecularweight. Finally, the MPHs (<2.5kDa) was isolated and purified through SP SephadexC-25ion-exchange chromatography and Sephadex G-25gel filtration chromatography,respectively, and the peptide which had the highest DPPH radicals and hydroxylradicals scavenging activities was identified as the molecular weight of1664Da usingHPLC.3. The stabilities of mackerel peptides: The antioxidant activity stabilities ofMPHs were investigated, and the results showed that the mackerel protein peptideshad good stabilities of temperature even though the high temperature of100℃could not reduce their antioxidant activities. In addition, the freeze-melt frequencyand UV had no significant effect on DPPH radicals and hydroxyl radicals scavengingactivities of MPHs (P<0.05). The MPHs had bad stabilities of pH, the DPPH radicalsand hydroxyl radicals scavenging activities of MPHs were reduced with the pH valuewas increase, when the pH value was9.2, the DPPH scavenging activity wasdecreased by89%. The MPHs was sensitive to metal ions, especially Fe2+、Fe3+、Zn2+and Cu2+could significantly decreased the antioxidant activities of MPHs, especiallywhen the concentrations of Fe2+and Fe3+were5mM, the DPPH scavenging activitieswere only1.1%and0.6%，respectively. while when the Cu2+with concentration of5mM could completely inhibited the DPPH scavenging activity of MPHs, on the otherhand, the K+、Mg2+had no significant effect on the antioxidant activities of MPHs(P<0.05).4. The evaluation on antifatigue effect of mackerel peptides: the antifatigue effectsof MPHs were evaluated and the results showed that the MPHs had no effect on bodyweight and growth of mice, and could increase the viscera index of mice. The MPHscould significantly prolong the exhaustion swimming time, the MPHs-H, MPHs-Mand MPHs-L groups were4.8-fold,2.8-fold and3.0-fold of the control group, and allMPHs groups were showed a dose-dependent behavior; the liver glycogen level was markedly increased by3335%and the BUN (1617%), LA (1419%) and MDA(1631%) levels were decreased in MPHs groups compared to that of the controlgroup. The activities of SOD and GSH-Px were increased by1115%and913%,respectively, the result showed that the MPHs had in vitro antioxidant activities. Inaddition, the correlation coefficient between the activity of GSH-Px and exhaustionswimming time was0.912, which indicated that the in vivo antioxidant activities ofMPHs had a good relationship with their antifatigue effect.