Study On Digestion And Metabolism Characteristics And Stability Of Wheat Embryo Functional Proteins In Vitro And In Vivo

Functional foods cover with food and functional regulation attributes.However,some functional food ingredients show poor gastrointestinal stability,which limits utilization of functional factors in the human body.The wheat embryo is rich in a variety of biologically active ingredients.Among them,wheat embryo globulin(WEG)and wheat embryo albumin(WEA)are the main functional proteins in wheat embryos.The previous research showed that wheat embryo protein has a variety of physiological functions such as immune regulation,anti-inflammatory,and anti-oxidation in our research group.In this study,WEG and WEA were used as raw materials.Firstly,the study clarified the digestion characteristics of protein from three levels of proteolysis,peptide release and free amino acids in vitro by simulating human gastrointestinal digestion,and revealed the influence of gastrointestinal digestion on the structure and function of the protein.Then an immunosuppressive rat model was established by intraperitoneal injection of cyclophosphamide(CTX),and the metabolic dynamics of the protein in healthy rats and immunosuppressed rats were analyzed through big data of digestion,absorption and metabolism in vitro.Finally,the modification of protein was processed by electrostatic interaction between chitosan(CS)and protein to prepare the proteinpolysaccharide complex with excellent dispersion stability through optimization of conditions,and the dispersion characteristics of the complex were characterized.The main contents and results in this study are as follows:1.The simulated digestion characteristics of wheat embryo functional protein in vitro:WEG and WEA showed similar changes during the simulated gastrointestinal digestion.First,most of the proteins were degraded during gastric digestion.About 30% protein(molecular weight was about 50 k Da)was resistant to digestion.However,as the protein entered the intestine digestion stage,all the proteins were degraded.Resistant digestion peptides(amino acid sequence ISQFXX,GTVX and MYPX)were identified by HPLC-MS/MS technology at the endpoint of digestion,and the total free amino acid content was significantly increased.Secondly,the secondary structure of the protein changed.The content of ?-helix and random coil were significantly reduced,and the ?-sheet was significantly increased,while the ?-turn no significantly changed.Finally,although the intact protein was degraded,the antioxidant activity was not negatively affected.2.The nutritional metabolism kinetics of wheat embryo functional protein in vitro and its effect on the gut microbes: the supplementation of WEG and WEA can affect protein levels in tissues and organs,and can restore protein metabolism in CTX-induced immunocompromised rats to normal levels.After supplementing with WEG in healthy rats,there was no significant change in organ quality,the peptide concentration in plasma was significantly reduced,and the free amino concentration in the heart,liver,and brain was significantly increased.After WEA supplementation,the heart quality and plasma peptide concentration were significantly reduced.The free amino concentration in the heart,liver,lungs,brain and muscles increased significantly.After immunosuppressive rats were supplemented with WEG in advance,WEG restored heart quality,liver peptide concentration,and slightly restored the free amino concentration of the brain to normal levels.It also increased the similarity of cecal flora in the number of optical transform unit(OTU)between immunosuppressed rats and normal healthy rats and did not affect the diversity of the cecal flora.There are 2 different bacterial phyla and 56 different bacterial genera,which significantly increase the abundance of probiotics such as Acetatifactor and significantly reduce the abundance of Burkholderia and other pathogenic bacteria.After WEA supplementation,the heart quality and liver peptide concentration returned to normal levels,the kidney peptide concentration showed a trend of recovery,the metabolism of free amino and the diversity of cecal bacteria was not affected,and the similarity in the number of OTU between immunosuppressed rats and normal healthy rats reduced.There are 3 different phyla and 60 different bacterial genera.The abundance of Corynebacterium?1 and other bacteria genus significantly increased,and the abundance of Desulfovibrio and Bilophila significantly reduced.3.Dispersion stability of wheat embryo functional protein: The addition of CS improves the dispersion stability of WEG and WEA,and the protein-polysaccharide compound nutritional drink with relatively long storage can be prepared through optimization of conditions.Firstly,the solubility and zeta-potential of WEG and WEA with p H were measured to gain the basic properties of the protein,showed that the solubility was the lowest between p H 4 and 5,and the isoelectric points of WEG and WEA were 4.70 and 4.06,respectively.The turbidity titration curve determines that the soluble complex formation intervals between WEG or WEA and CS are p H 3?4 and p H 2.6?3.6,respectively.Secondly,the p H value,complex ratio and ionic strength were regarded as single factors to optimize the preparation conditions of the complex,showing CS-WEG complex: p H 5.903,a ratio of 1.05,and ionic strength of 77.070 m M,and CS-WEA complex: p H of 5.005,a ratio of 1.012 and ionic strength of 400 m M.Under this condition,the compound spanned excellent dispersion stability.Finally,the zeta-potential,particle size,polydispersity index(PDI),rheological properties and fluorescence microscopy of the compound were characterized to clarify the mechanism on the stability between CS and WEG or WEA,showing that the addition of CS significantly increased the zeta-potential and reduced the PDI and reduced the particle size to a certain extent.The rheological properties showed shear-thinning behavior and the apparent viscosity increase,but the dispersion stability decreased with a stabilization time extension.The experimental studies in vivo and in vitro have shown that: WEG and WEA were digested by gastrointestinal and degraded into characteristic peptides and free amino acids,which distributed in tissues and organs,restored the protein level of immunosuppressed rats,regulated their immune levels,and improved their flora disordered.It is speculated that WEG and WEA mainly exert their activity through characteristic peptides.Besides,to maintain the stability of WEG and WEA,CS-WEG and CS-WEA complexes were prepared,which significantly improved the stability of WEG and WEA,revealing the mechanism between them.