Study On Roasting Effect On Allergenicity Of Peanut And Undergoing Mechanism

Peanut allergy is accountable for 59% of death cases caused by food allergy and is one of the major threats to human health.Recent studies have shown a higher prevalence of peanut allergy in western populations,possibly due to their preference for peanut butter made from roasted peanuts.Different processing conditions,such as roasting,can alter the structure of peanut allergens to varying degrees,including unfolding,aggregation,cross-linking between ingredients and chemical modification such as oxidation and glycosylation,and may have a profound effect on peanut allergenicity.In order to explore the effect and mechanism of roasting on peanut allergenicity,in this paper,roasted peanuts was used as material,the changes of roasting temperature and roasting time in peanut protein content and glycosylation degree were analyzed.BALB/c mice model was employed to evaluate the effects of different roasting treatments on peanut protein allergenicity.Furthermore,Ara h 3,one of the main allergens in peanut,was selected for further study.The Caco-2 cell model and dendritic cell model were chosen to investigate the effect of roasting treatment on Ara h 3 in the initial phase of allergy,with attempt of revealing the mechanism for roastingenhanced allergenicity of Ara h 3.The main research contents and results are as follows: 1 Effects of peanut roasting conditions on protein content,glycosylation and allergenicityPeanut kernels were roasted for 20 min at 110 ℃,130 ℃,150 ℃ or 170 ℃;or the kernels were placed in 150 ℃ oven for 10 min,20 min,30 min or 40 min,respectively,and roasted peanut protein was extracted.With the increase of roasting temperature or the prolongation of roasting time,the protein concentration of peanut decreased gradually.Compared with protein from raw peanut,the content of free amino groups in protein from roasted peanut(150 ℃ for different duration or 110 ℃,130 ℃,150 ℃,170 ℃ for 20 min),decreased significantly,accompanied with escalation of bound sugar content,fructosamine and formation of small molecule fluorescence products,showing peanut proteins were glycosylated to different degrees during roasting.Ground peanuts emulsion was used to sensitize BALB/c mice by gavage.The results showed that degranulation degree of mast cell and basophil in roasted peanutsensitized mice exhibited more severe,and inflammatory infiltration and hyperemia of jejunum and lung tissue were more serious too.Levels of Th2 cytokines(IL-4,IL-5,IL-13)in the serum of roasted peanut-sensitized mice was elevated and were accompanied by increased levels of specific antibodies(Ig E,Ig G)and histamine.Thymic Stromal Lymphopoietin(TSLP)gene expression was up-regulated in small intestine and lung tissues,Ig E binding capacity to roasted peanut proteins was augmented as well.Overall,peanut allergenicity was enhanced with the increase in temperature and duration of roasting.2 Ara h 3 purification and roasting effect on Ara h 3 structure,glycosylation and potential allergenicityAra h 3-Raw and Ara h 3-Roasted were purified from raw peanut and roasted peanut(150 ℃,20 min),respectively,through anion exchange chromatography.The purified Ara h 3 was used to detect roasting effect on protein molecule microstructure.The results showed that,compared with Ara h 3-Raw,the secondary structure of Ara h 3-Roasted exhibited more disordered,less hydrophobic,and aggregated to some degree.The glycosylation level showed that,compared with Ara h 3-Raw,Maillard reaction of Ara h 3-Roasted reached an advanced stage.Competitive inhibition ELISA results showed that roasting enhanced the Ig E and Ig G binding capacity to Ara h 3,suggesting that roasting enhanced the potential allergenicity of Ara h 3.3 The mechanism of roasting-enhanced allergenicity of Ara h 3Caco-2 monolayers model and dendritic cell model were set in order to simulate effect of Ara h 3-Roasted on the gastrointestinal immune system in this paper.The results showed that,Ara h 3-Roasted reduced Caco-2 cell viability,inhibited cell proliferation,and promoted ROS formation.The imbalance of oxidative stress activated the NF-κB signaling pathway,leading proinflammatory factors IL-6,IL-8 and MCP-1 to an elevated level.Ara h 3-Roasted subsided transepithelial electrical resistance(TEER)of Caco-2 monolayers,and down-regulated the expression of tight junction proteins such as zonula occluden-1(ZO-1),Occludin and Junctional Adhesion Molecule-1(JAM-1).These changes together would destroy ZO-1 distribution in Caco-2 monolayers and destroy compactness or integrity of Caco-2 monolayers.In this way,transport of Ara h 3-Roasted into or through Caco-2 cells would be promoted and Ara h 3-Roasted absorption by the gastrointestinal become facilitated.Besides,Ara h 3-Roasted up-regulated the expression of LRP-1 gene,whose protein product is the scavenger receptor component of dendritic cells,and in turn,helped dendritic cells with Ara h 3-Roasted uptake.Ara h 3-Roasted also up-regulated the expression of IL-10 gene of Th2 type inflammatory factor,and down-regulated the expression of IL-12 gene of Th1 type cytokine,and then trigged more severe Th2 type allergic reaction.