Structural Identification Of Immunoactive Peptides Derived From Hazelnut(corylus Heterophylla Fisch.) Protein And Its Immunoregulatory Effects On Inflammation

Hazelnut contains oil,protein,carbohydrates,unsaturated fatty acids,fiber and other nutrients.The content of protein in hazelnut is about 20%,and the proportion of amino acids in protein is balanced,while with plenty of glutamic acid and arginine to meet the needs of adults.It is safe to use plant-derived peptides to develop a variety of functional products.The objective of this research is to isolate,purify and identify the hazelnut peptide ased on preliminary study in laboratory.Using cell inflammation model to explore the mechanism of action of NF-?B and MAPK signaling pathways the study can provide basis for the development and utilization of hazelnut immunoreactive peptides,thus conducive to the research on the active component of hazelnuts.The results are as follows:In this study,the hazelnuts peptides were separated and purified by using reversed-phase high-performance liquid chromatography(RP-HPLC),and the components of F1,F2,F3,F4,F5 and F6 were obtained.The above 6 components were then identified by using electrospray tandem mass spe-ctrometer(ESI MS/MS),and 10 hazelnut peptides were obtained after screening the identification results.The amino acid sequences were Val-Asp-Glu-Asp-Thr-Ala-Arg(VDEDT AR),Val-Leu-Leu-Ala-Leu-Val-Leu-Leu-Arg(VLLALVLLR),Pro-Glu-Asp-Glu-Phe-Arg(PED EFR),Arg-Asn-Val-Glu-Asp-Glu-Leu-Ala-Arg(RNVEDELDAPGHR),Val-Glu-Asp-Glu-LeuArg-Val-Leu(VEDELRVL),Asp-Glu-Glu-Thr-Arg-Pro-Trp-Arg(DEETRPWR),His-Leu-ArgAsp-Pro-Gln(HLRDPQ),Ala-Asp-Leu-Tyr-Asn-Pro-Arg(ADLYNPR),His-Leu-Arg-Asp-ProThr-His(HLRDPTH)and Leu-Asp-Ala-Pro-Gly-His-Arg(LDAPGHR),respectively.Finally,the 10 peptides were synthesized.LDAPGHR had higher immunomodulatory capacity aft-er evaluation of immunocompetence.The results of the immunology experiments showed that LDAPGHR could enhance the phagocytosis of mouse macrophages and the prolifera-tion rate of spleen lymphocytes.In this study,the in vitro model was established by LPS-stimulated mouse RAW264.7 cells,and the effects of LDAPGHR on Mitogen-activated protein kinase(MAPK)and Nuclear transcription factor-?B(NF-?B)signaling pathway were investigated.There was no toxic effect on the cells when LDAPGHR and Lipopolysaccharide(LPS)co-stimulated in the concentration of 12.5-50 ?mol/L.Compared with LPS group,the expression of NO,PGE2,IL-1?,IL-6 and their mRNA in RAW264.7 cells were inhibited by LDAPGHR(P <0.05),but no significant effect on TNF-?.The protein expression of p-JNK,p-ERK and p-p38 in MAPKs pathway and the expression of p-I?B-? and p-65 in nucleus in NF-?B pathway were inhibited.The experimental results showed that LDAPGHR could inhibit the inflammatory response induced by LPS in mouse RAW264.7 cells by inhibiting the NF-?B and MAPK pathways.