Study On Preparation Of Wheat Embryo Albumin And Its Peptide And Analysis Of Its Anti-exercise Fatigue Effect

Exercise fatigue seriously affects people’s quality of life and work efficiency,long-term accumulation can lead to chronic fatigue syndrome,even karoshi.It is an effective way to relieve fatigue by supplementing exogenous nutrition or drugs.However,many clinical drugs contain side effects and are not suitable for long-term use.Therefore,it is of great practical significance to extract anti-fatigue active ingredients from natural food.Wheat embryo albumin(WEA)is one of the components of WEA,which is regarded as the "natural treasure house of human nutrition".WEA is rich in amino acids,and the proportion of amino acids is closest to that in human body.Studies have shown that WEA has antioxidant,anti-tumor and immune activities.In this paper,WEA was prepared by water extraction from wheat germ.Antioxidant peptides were prepared by enzymolysis and the enzymolysis process was optimized.The structure and function of wheat germ albumin and its peptides were characterized,and gastrointestinal digestion characteristics of wheat germ albumin were investigated.The effects of wheat germ albumin and its peptide on acute exercise fatigue were investigated by establishing a mouse model of acute exercise fatigue.The effects of wheat germ albumin on chronic exercise fatigue and intestinal microbe were studied by establishing a mouse model of chronic exercise fatigue.The aim of this study was to provide a new method and idea for the development and utilization of wheat germ albumin and the alleviation of exercise fatigue.The main research contents and results of this paper are as follows:1.Preparation,characterization and digestion characteristics of Wheat embryo albumin was prepared by water extraction method.The structure and function of WEA were characterized by transmission electron microscopy,scanning electron microscopy,circular chromatograph and amino acid analyzer,and its polypeptide sequence was analyzed by LCMS/MS.The changes of hydrolytic activity and biological activity of WEA during digestion were studied by in vitro digestion model.In addition,the digestibility,antioxidant activity,free amino acid and microstructure of the digested products were also studied.And it turns out,WEA has certain free radical scavenging ability and was rich in amino acids,including essential amino acids and antioxidant activities of Aromatic amino acid(AAA)and Hydrophobic amino acid(HAA).In addition,most of the peptide sequences in WEA contain amino acid residues with antioxidant activity.The shape of WEA was irregular and flaky,with smooth surface and uneven thickness and some holes.The secondary structure of WEA mainly includes β-folding,random crimp,β-turning and α-helix,among which β-folding was the most important.During digestion,the digestibility of WEA showed a decreasing trend.With the progress of digestion,WEA was gradually degraded and its antioxidant activity increased.Meanwhile,the morphology of WEA changed significantly during digestion.2.Preparation,characterization and antioxidant activity analysis of wheat embryo albumin antioxidant peptide: The enzymatic hydrolysis conditions of WEA were optimized by single factor experiment,and the products with the most antioxidant activity were isolated by ultrafiltration.Its structure and function were characterized by transmission electron microscopy,scanning electron microscopy,circular dichromatic spectrometer and amino acid analyzer,and its polypeptide sequence was analyzed by LC-MS/MS.And it turns out,the optimal enzymatic hydrolysis conditions of WEA were as follows: substrate concentration 2%,p H 6,enzyme dosage 8000 U/g,temperature 55 ℃.In the five ultrafiltration components,F3(3k Da < MW < 5 k Da)had the best antioxidant effect.The microstructure of F3 was spherical and granular with some pores and uneven thickness.The secondary structure of F3 mainly includes β-folding,irregular crimp,β-turning and α-helix,among which β-folding was the most important.F3 was rich in amino acids,including essential amino acids and antioxidant activity of AAA and HAA.In addition,most of the peptide sequences in F3 contained amino acid residues with antioxidant activity.These results indicated that F3 has good antioxidant capacity.3.Effect evaluation of wheat embryo albumin and its peptide on alleviating acute exercise fatigue: By establishing a mouse model of acute exercise fatigue,the exhaustion-to-death swimming time,exhaustion-swimming time,fatigue-related biochemical indexes and antioxidant enzyme activities of mice were analyzed.And it turns out,WEA and F3 can significantly prolong exhaustive swimming time(p<0.05),increased the levels of glycogen in liver and muscle of mice(p<0.05).Meanwhile,WEA and F3 could significantly reduce the activities of Alanine transaminase(ALT),Aspartate transaminase(AST),Lactate dehydrogenase(LDH)and Blood urea nitrogen(BUN)in serum of model mice(p<0.05).In addition,WEA and F3 significantly reduced the content of in Superoxide dismutase(SOD),Glutathione(GSH)and Malondialdehyde(MDA)in liver and skeletal muscle(p<0.05),significantly increased SOD and GSH activities(p<0.05).4.Effect and mechanism analysis of wheat embryo albumin on chronic exercise fatigue:The effects and mechanism of WEA on chronic exercise fatigue were investigated by establishing a model of chronic exercise fatigue and measuring the exhaustion swimming time,fatigue related biochemical indexes,antioxidant enzyme activity and energy metabolism related gene expression in mice.And it turns out,WEA had no significant effect on organ index and body weight of mice,and did not affect the normal growth of mice(p>0.05).WEA significantly prolonged the exhaustive swimming time of mice(p<0.05),increased the glycogen storage in liver and muscle of mice(p<0.05),WEA was significantly decreased in serum ALT,AST,LDH,Creatine kinase(CK)and BUN(p<0.05).Meanwhile,WEA significantly decreased MDA content in liver and skeletal muscle,and significantly increased SOD and GSH activities(p<0.05).In addition,WEA significantly up-regulated the m RNA expression levels of AMP-activated protein kinase(AMPK),Nuclear Respiratory Factor1(NRF-1)and Mitochondrial transcription factor A(TFAM)in skeletal muscle of mice(p<0.05).5.Regulating effect of on intestinal microflora: Through high-throughput sequencing technology,the effects of WEA on intestinal flora of mice with chronic exercise fatigue were analyzed from three aspects of Alpha diversity,bacterial community similarity and bacterial community difference.And it turns out,Chao1 and Observed species index of WEA group were significantly higher than that of MG group(p<0.05).In addition,the Shannon index in WEA group was higher than that in MG group(p>0.05),indicating that the species diversity in WEA group was higher.However,there was no significant difference in Shannon index among all groups(p>0.05),indicating that species diversity was basically the same among all groups.At the same time,WEA supplementation can improve the similarity of OTU quantity in the intestinal flora of fatigued mice and normal mice.In addition,WEA supplementation can significantly increase the abundance of Ruminiclostridium,Bilophila,Oscillibacter,Mucispirillum,Angelakisella,reduced Ruminococcace AE UCG-013,Odoribacter,Anaerotignum,Streptococcus,Candidatus Saccharimonas,Eubacterium xylanophilum group abundance.Both WEA and F3 have good antioxidant capacity and rich nutritional value,and the antioxidant activity and biological activity of WEA are enhanced after digestion.Both WEA and F3 can alleviate acute exercise fatigue by increasing glycogen storage,reducing metabolic product accumulation and alleviating oxidative stress in liver and skeletal muscle.WEA can reduce the metabolism of energy and protein and protect cells from injury,enhance the exercise ability of mice,prevent lipid oxidation,enhance the activity of antioxidant enzymes,and improve the mitochondrial function of skeletal muscle of mice to play the role of antichronic exercise fatigue.In addition,WEA can regulate the bacterial community disorder of chronic exercise fatigue mice to a certain extent,improve the abundance of beneficial bacteria,and reduce the abundance of pathogenic bacteria.This study provided a new method and idea for the development and utilization of wheat embryo albumin and the alleviation of exercise fatigue.