| In this thesis,peptides were prepared from hairtail meat through controlled enzymatic hydrolysis.The antioxidant activity of hairtail meat peptides were firstly evaluated in vitro.Then,the in vivo antioxidant and anti-fatigue effects of hairtail meat peptides were further evaluated in animal models.Next,the capillary HPLC coupled with Q Exactive hybrid quadrupole-orbitrap mass spectrometer was used to identify the amino acid sequence of the peptides.The main research results of this study were as follows:（1）The basic composition and amino acid composition of hairtail meat were analyzed.The results showed that the content of water,ash,fat and protein in hairtail meat were 77.91±0.56%,0.64±0.06%,2.01±0.09%,and 13.51±0.22%,respectively.In addition,the results of amino acid composition showed that glutamic acid was the most abundant amino acid in hairtail meat and followed by lysine,aspartic acid,leucine and alanine.Evidence already showed glutamic acid is good for sports endurance.Aspartic acid can reduce blood lactate content.Leucine is a branched amino acid.Aspartic acid and glutamic acid can be used as hydrogen donors.Histidine can quench free radicals and chelate metal ions.This suggested that it is highly possible hairtail meat peptides possessed good antioxidant and anti-fatigue potential.Pepsin,trypsin,dispase,alkaline proteinase,or bromelain were used to hydrolyze hairtail meat.In view of total antioxidant capacity and degree of hydrolysis,dispase was the best enzyme for the hydrolysis of hairtail meat.Then,the effects of incubation time,incubation temperature,and enzyme concentration on the total antioxidant capacity and hydrolysis degree of hairtail meat peptides were analyzed.Hydrolysis conditions were optimized by response surface methodology.The optimal hydrolysis conditions were as follows:incubation time of 12.5 h,incubation temperature of 45℃,and enzyme concentrate of 1872 U/g.Under optimized enzymatic conditions,total antioxidant capacity was30.6532±0.1746 U/m L and degree of hydrolysis was 20.7787±0.1562%,both of which were not significantly different from their predicted values.（2）Bioactive peptides are susceptible to degradation due to the influence of food processing environment,leading to the loss of biological activity.Thus,the effects of pH,temperature,and salt on the antioxidant stability of hairtail meat peptides were determined in our study.In addition,the effects of gastrointestinal digestion on the antioxidant stability of hairtail meat peptides were studied by simulating human gastrointestinal digestive environment externally.The results demonstrated that DPPH radical scavenging activity,hydroxyl radical scavenging activity,and total antioxidant capacity of hairtail meat peptides were affected significantly only in strong acid/alkaline condition（pH<2 or pH>10）compared with that under neutral pH condition（P<0.05）.When the environment temperature was below 60℃,the hairtail meat peptides did not show significant decrease in DPPH radical scavenging activity compared to unheated sample（P<0.05）.Further increasing the ambient temperature to 80℃only caused the decreases of DPPH radical scavenging activity,hydroxyl radical scavenging activity,and total antioxidant capacity by less than 10%.It showed that hairtail meat peptides had a good heat resistance.Meanwhile,when the concentration of Na Cl was below 6%,DPPH free radical scavenging activity,hydroxyl radical scavenging activity and total antioxidant capacity of hairtail meat peptides were not significantly altered（P<0.05）.After digesting by pepsin and trypsin,DPPH radical scavenging activity,hydroxyl radical scavenging activity and total antioxidant capacity of hairtail meat peptides were all significantly reduced（P<0.05）.（3）The anti-fatigue and antioxidant effects of hairtail meat peptide prepared under optimized parameters were evaluated.Hairtail meat peptides have no significant effect on body weight and food intake in mice.Hairtail meat peptides could extend exhaustive swimming time in mice.The exhaustive swimming time of the mice from control,low dose,middle dose,and high dose group was 12.00±6.52、33.19±19.71、64.74±54.08 and 34.05±21.62 min,respectively.In addition,compared with control group,hairtail meat peptides can increase the content of liver glycogen and muscle glycogen,and reduce the content of blood lactic acid,blood urea nitrogen and creatine kinase in mice at all doses tested.Besides,the glutathione peroxidase,superoxide dismutase,and catalase activities in mice receiving low,middle,and high doses of hairtail meat peptides were all elevated,while statistical significance were in middle and high dose groups.Malondialdehyde was a lipid peroxidation product in the body.Hairtail meat peptides could also significantly reduce malondialdehyde in mice.The contents of malondialdehyde in middle dose group and high dose group was 1.87±0.06 and 1.45±0.26nmol/m L,both of which were significantly different from that in control group.（4）Gel permeation chromatography coupled with multi angle laser light scattering was used to determine the molecular weight distribution of hairtail meat peptides.Components with an average molecular weight of 1.166,2.212,and 2.913k Da accounted for 73.0681,12.1709,and9.4850%of total peptides,respectively.It showed that the most of hairtail meat peptide was identified as small peptides with molecular weight less than 3k Da.Next,hairtail meat peptides were separated into five fractions using ultrafiltration.Results confirmed that fraction with molecular weight below 3k Da（HMP-I）exhibited higher antioxidant activities than other fractions.The fraction A obtained by gel column chromatography showed highest antioxidant activity.Its DPPH radical scavenging capacity,hydroxyl radical scavenging capacity and total antioxidant capacity were 45.40±0.24%,44.55±0.61%,2.93±0.04 U/m L,separately.Then,component A was further separated by RP-HPLC.The purified peptide fraction A2was identified as DLYANTVLSGGTTMYPGIADR、SINPDEAVAYGAAVQAAILSGDK、SLGIETAGGVMTVLIK using capillary HPLC coupled with Q Exactive hybrid quadrupole-orbitrapmass spectrometer. |
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