Enzymatic hydrolysis of silkworm pupa and its allergenicity evaluation by animal model with different immunization routes

Silkworm pupa is a nourishing food with high nutritional value,but its consumption has been greatly limited given its allergenicity.Enzyme hydrolytic technique is recognized as an effective method to reduce the allergenicity of protein.In this study,we aimed to investigate the effect of enzymolysis on the allergenicity of silkworm pupa.Crude silkworm pupa protein was extracted through alkali extraction and acid precipitation,which included 5 proteins with the molecular weights ranging from 34 kDa to 76 kDa,and silkworm pupa were then hydrolyzed by alkaline protease.The allergenicity of silkworm pupa protein and its enzymatic hydrolysates was evaluated by establishing BALB/c mice model,and the mice were immunized via intragastric gavage and intraperitoneal injection,respectively.The results indicated that the intraperitoneal inj ection immunization route induced more by detecting with antibodies,histamine and Th2-related cytokines.Moreover,mice treated with silkworm pupa protein peptide displayed no obvious allergic symptoms,indicating that enzyme hydrolytic technique could significantly reduce the allergenicity of silkworm pupa.

Effect of thermal processing and fermentation with Chinese traditional starters on characteristics and allergenicity of wheat matrix

Wheat allergy has become a serious health threat worldwide and its prevalence has increased alarmingly in the past few years.Factors such as food matrix and food processing may alter the structure of wheat proteins,and hence affect its allergenic properties.However,few reports have focused on the influence of Chinese traditional starter fermentation on wheat allergy.In this study,5 starters from different regions of China were used for fermentation,and protein characteristics were monitored by sodium dodecyl sulfate polyacrylamide gel electropheresis,and immunoreactivity analyzed by immunoassay with allergenic serum was obtained from New Zealand white rabbits.The allergenicity of steamed and baked matrices was also evaluated.The results showed that the allergenicity of wheat dough was basically increased at the beginning and then decreased during fermentation,but specific trends depend on different starters.With the progress of fermentation,especially as pH value decreased to 3.0-4.0,the allergenicity decreased significantly.Baking and steaming can reduce the allergenicity of wheat matrix,but fermentation is not a key factor affecting the allergenic activity of proteins.Our results can provide a theoretical basis for controlling wheat allergenicity in food proces sing or producing hypoallergenic food.

Structure and allergenicity of a-lactalbumin:effects of ultrasonic prior to glycation and subsequent phosphorylation

Bovineα-lactalbumin(BLA)induced severe cow's milk allergy.In this study,a novel strategy combining ultrasonication,performed before glycation,and phosphorylation was proposed to reduce BLA allergenicity.Result showed that IgE-and IgG-binding capacities and the release rates of histamine and interleukin-6 from RBL-2 H3 were reduced.Moreover,intrinsic fluorescence intensity and surface hydrophobicity were decreased,whereas glycated sites(R10,N44,K79,K108,N102 and K114)and phosphorylated sites(Y36 and S112)of BLA were increased.Minimum allergenicity was detected during BLA treatment after ultrasonic prior to glycation and subsequent phosphorylation because of considerable increase in glycated and phosphorylated sites.Therefore,the decrease in allergenicity of BLA,the effect correlated well with the shielding effect of glycated sites combined with phosphorylated sites and the conformational changes.This study provides important theoretical foundations for improving and using the ultrasonic technology combined with protein modification in allergenic protein processing.

Study on mechanism of increased allergenicity induced by Ara h 3 from roasted peanut using bone marrow-derived dendritic cells

Little information was so far available about allergenic mechanism of the roasted peanut allergens during initial stages of allergy.The purpose of this study was to determine the influence of roasting(150℃,20 min)on biochemical and biological properties of Ara h 3,a major peanut allergen.Allergenicity of roasted peanut emulsion to mice,differences in uptakes between Ara h 3 purified from raw peanuts(named as Ara h 3-Raw)and that purified from roasted peanuts(named as Ara h 3-Roasted)by bone marrow-derived dendritic cells(BMDCs)and the implication of cell surface receptors involving in uptake,and changes in glycosylation and structure of Ara h 3 after roasting were analyzed in this study.This study suggested that roasting increased allergenicity of peanut to BALB/c mice.Maillard reaction and structural changes of Ara h 3 induced by roasting significantly altered the uptake of Ara h 3-Roasted by BMDCs,and modified Ara h 3 fate in processes involved in immunogenicity and hyper allergenicity,indicating that food processing pattern can change food allergenicity.

Changes in wheat protein digestibility and allergenicity: role of Pediococcus acidilactici XZ31 and yeast during dough fermentation

Wheat flour,as the most important source of food globally,is one of the most common causative agents of food allergy.This study aimed to investigate the effects of fermentation on wheat protein digestibility and allergenicity.Protein digestibility were evaluated using the sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and enzyme-linked immunosorbent assay.The effect of protein on intestinal permeability was investigated by Caco-2 cell monolayers.Co-culture fermentation with Pediococcus acidilactici XZ31 and yeast leads to improvement in digestibility of wheat protein compared to single strain fermentation.Fermentation leads to a decrease in albumin/globulin antigenicity and an increase in gluten R5 reactivity,with the most significant changes in the co-culture group.Digestion strengthen the decrease of protein antigenicity and counteracts the difference in antigenicity induced by fermentation between groups.However,pretreatment with P.acidilactici XZ31 reduces the amount of allergens across Caco-2 monolayer and attenuates the gluten-induced increase in permeability of Caco-2 cell monolayer by reducing actin polymerization and villous atrophy.Co-culture fermentation reduces gluten-induced cell monolayer damage to a greater extent than P.acidilactici XZ31 monoculture.These results gives valuable insight into the effects of P.acidilactici XZ31 fermentation on the allergenicity and toxicity of wheat proteins,which contribute to promoting the application of multi-strain leavening agent in hypoallergenic and gluten-free wheat products.

Effect of lipid peroxidation on the allergenicity and functional properties of soybeanβ-conglycinin(7S)and glycinin(11S)

This study aimed to analyze the effect of lipid peroxidation on the allergenicity and functional properties of soybeanβ-conglycinin(7 S)and glycinin(11 S).Oxidation complexes were determined using the lipid peroxidation method.Functional properties were analyzed based on emulsifying and foaming properties.The potential allergenicity was evaluated by in vitro and in vivo methods.The results found that oxidation altered structures of the proteins and resulted in the formation of cross-linked protein polymers.The emulsion and foaming properties of the proteins were improved after oxidation.The IgE-binding capacity of 7 S and11 S reduced after oxidation.KU812 cell assays showed that both histamine and IL-4 release decreased after oxidation treatment.A mouse model showed that oxidation reduced the IgE,IgG,and IgG1 levels,as well as reduced histamine and mMCP-1 release in serum,which might suppress the allergic reaction.In conclusion,the lipid peroxidation treatment likely causes changes to the functional properties of soybean,decreasing the potential allergenicity of 7 S and 11 S.

Interactive effect of hot air roasting processes on the sensory property,allergenicity,and oil extraction of sesame(Sesamum indicum L.)seeds

Sesame seeds are a healthy food ingredient and an oil crop for sesame oil production;however,it has recently been recognized as an essential allergenic food by FAO/WHO.This research investigated the relationship between the hot air roasting process(at 120,150,and 180℃ for 10,20,and 30 min)and several quality attributes of sesame seeds since roasting is the key process for preparing sesame seeds for both consumption and oil production.The hot air process followed the central composite design.The changes of sesame in terms of color,sensory properties(odor,texture,color,and taste),allergenicity caused by oleosins(ses i 4 and ses i 5),as well as oil extraction and quality were monitored using a colorimeter,sensory evaluation panelists,ELISA,as well as oil yield and acid value,respectively.Roasting temperature influenced the product quality more than roasting time,although the two processing parameters significantly interacted with each other(P<0.001).Sensory evaluation indicated medium roasting generated attractive flavor,order,appearance,and crispy texture.Allergenicity was high in sesame seeds after high-temperature roasting,according to IgE binding capacity test.Sesame oil extraction was favored by high-temperature roasting,which,however,adversely affected the oil quality.The optimal roasting conditions were 150.5℃ for 15 min for optimized sesame seeds quality in terms of sensory properties and allergenicity,while roasting at 158℃ for 10 min was optimal for sesame oil production.The finding will benefit the sesame seed industry.

Potential of high hydrostatic pressure in reducing the allergenicity of seafood

Seafood,as a primary high-quality protein source,plays an increasingly vital role in diets around the world,while seafood allergy is a worldwide health problem that affects the quality of life and may even threaten lives.High hydrostatic pressure(HHP),a novel non-thermal processing technology,shows the unique potential in alleviating seafood allergenicity.This comment provides a brief introduction of potential of high hydrostatic pressure in reducing the allergenicity of seafood.

Site-specific N-glycosylation characterization and allergenicity analysis of globulin-1 S allele from wheat

N-glycans in many proteins are of great concern because of their strong association with food allergies. Triticum aestivum(bread wheat), a major food crop, is known as one of the “Big Eight” allergenic groups. However, little research has been done about N-glycans in wheat glycoproteins. In this study, a soluble wheat glycoprotein was purified from wheat and further identified as globulin-1 S allele(GSA). The wheat GSA displayed significant IgE-binding activity. Moreover, one N-glycosylation site and 6 kinds of N-glycans were identified by mass spectrometry, including 3 high mannose types and 3 complex types. Furthermore, the IgE-binding activity of wheat GSA is proved to be reduced by the removal of N-glycan, thermal treatment(temperatures > 80 ℃), and strong acidic treatment(pH 3.0). These findings would provide a better understanding of the effects of N-glycosylation, thermal treatment, and acidic treatment on the molecular characteristics of GSA, and further provide new insights into the development of hypoallergenic wheat products.

Allergenicity studies of Naoluotai injection and H3 injection before and after ultrafiltration

OBJECTIVE To explore the influence of ultrafiltration on the allergenicity of Naoluotai injection and H3 injectable powder in guinea pig.METHODS The ultra filtrated and non-ultra filtrated Naoluotai injection was utilized to do the sensitization test of guinea pig.The guinea pig was sensitized three times by subcutaneous injection of test solution every other day.After 16 dof the first sensitization.All guinea pigs were challenged.The serotonin level in guinea pig plasma before and after the challenge was recognized as the evaluation criteria of allergenicity.RESULTS The increasing rate of serotonin in guinea pigs sensitized by ultra filtrated Naoluotai injection was 5.80% while by non-ultra filtrated Naoluotai injection was 37.03%.The increasing rate in ultra filtrated and non-ultra filtrated H3 injection was 23.59% and 61.94%,respectively.CONCLUSION The animal protein in tradition Chinese medicine injection was thought to be allergized.The ultrafiltration technology could reduce the allergenicity of tradition Chinese medicine injection and improve its safety.

Insights into sensitizing and eliciting capacity of gastric and gastrointestinal digestion products of shrimp(Penaeus vannamei)proteins in BALB/c mice

Shrimp(Penaeus vannamei)proteins have been shown an allergenic potential;however,little information is available on the sensitizing and eliciting capacity of shrimp protein digestion products.In this study,a BALB/c mice model was used to explore the allergenicity of shrimp protein sample(SPS)and their gastric and gastrointestinal digestion products(GDS/GIDS).As compared with the SPS groups,the GDS/GIDS groups caused lower specific immunoglobulins(Ig E/Ig G1)levels(P<0.05),but higher than the control groups,indicating that the digestion products sensitized the mice.Meanwhile,spleen index,mouse mast cell protease-1(m MCP-1)concentration and proportion of degranulated mast cells were significantly reduced in the GDS/GIDS groups(P<0.05);simultaneously,allergic symptoms,vascular permeability and histopathological changes of tissues were alleviated.Nevertheless,the allergenicity of digestion products cannot be eliminated and still cause systemic allergic reactions in mice.The study showed that the digestion products of shrimp still had high sensitizing and eliciting capacity.

The detection and regulation advances of food allergens

Food allergy has become an important food quality and safety issue,posing a challenge to the food industry and affecting consumer health.On the one hand,from the perspective of food processing industry,the diversity of food raw material ingredients,exogenous additives,and processing forms make the presence of allergens in modern food processing more complex.In addition,due to the lack of allergen identification,effective detection and allergenicity evaluation systems,there are serious deficiencies in the current theories and techniques for food allergen screening and detection,tracking and prediction,intervention and control;On the other hand,from the perspective of public health,meeting consumers'right to know whether there are raw materials containing food allergens in processed foods,and improving the credibility of government and people's satisfaction have become urgent matters;In addition,as people come into contact with more and more new borne novel foods,the probability of food allergy is also increasing.The food safety and health problems induced by increasingly complex,widespread and severe food allergy are difficult to avoid.In view of this,in response to the increasingly serious food allergy issues,this paper introduced the detection methods of food allergens,summarized the reduction and control techniques of food allergens,and elaborated hypoallergenic foods,which aims to provide the basis for preventing and controlling food allergy and ensuring the physical health of food allergy patients.

Effects of different cooking methods on peanut allergenicity

Peanut allergy is a type of serious food allergy worldwide.In this study,peanuts were treated with two different cooking methods.The effect of cooking on digestion stability was evaluated by simulated gastrointestinal digestions.The effects of cooking on the allergenicity were assessed by immunoblotting,passive cutaneous anaphylaxis (PCA) mice model and rat basophilic leukemia (RBL)-2H3 cell model.Results showed that raw peanuts were more stable to digestion than boiled and fried peanuts.SDS-PAGE showed that the quantities of allergens Ara h 1 (64 kDa),Ara h 2 (20 kDa),and Ara h 3 (38 kDa) in the boiled and fried peanuts were all reduced.Meanwhile,immunoblotting revealed the IgE binding capacities of boiled and fried peanuts were decreased significantly,especially the fried peanuts.In the PCA mice model,the vascular permeability of allergic mice,as well as the release of histamine,were both alleviated in mice gavaged with boiled and fried peanuts compared with raw peanuts.Lastly,the release of β-hexosaminidase of RBL-2H3 cells treated with boiled and fried peanuts was significantly decreased compared with the raw peanuts.In addition,the analyses of circular dichroism (CD) spectra showed that the α-helical content of peanut proteins was decreased after boiling and frying,indicating the structure stability of peanut allergens decreased.The analyses of ultraviolet (UV) spectra showed that aromatic amino acid residues were exposed,causing the change of the tertiary structure of peanut allergens after boiling and frying.In conclusion,the allergenicity of peanuts was significantly reduced after boiling and frying,especially frying.

Recent advances in the study of epitopes,allergens and immunologic cross-reactivity of edible mango

Mango(Mangifera indica L.)is a tropical fruit that is widely consumed as both fresh fruits and processed products around the world.The high incidence of mango allergy,on the other hand,has sparked widespread concern.Therefore,a summary and analysis of the current status and issues in mango allergen research can guide in-depth study on the mechanism of mango allergy and reveal effective desensitization methods.We described the incidence of fruit allergy,as well as the mechanism and clinical symptoms of mango allergy,in this review.We also looked into the structural properties of mango allergens,the effect of processing methods on mango allergens,prediction methods for mango allergen epitopes,and the current state of research on mango cross-reactive allergens and preventive measures.Finally,the research directions and ideas for the future are proposed and discussed.

Characterization of physicochemical and immunogenic properties of allergenic proteins altered by food processing:a review

Food allergens are mainly naturally-occurring proteins with immunoglobulin E(IgE)-binding epitopes.Understanding the structural and immunogenic characteristics of allergenic proteins is essential in assessing whether and how food processing techniques reduce allergenicity.We here discuss the impacts of food processing technologies on the modification of physicochemical,structural,and immunogenic properties of allergenic proteins.Detection techniques for characterizing changes in these properties of food allergens are summarized.Food processing helps to reduce allergenicity by aggregating or denaturing proteins,which masks,modifies,or destroys antigenic epitopes,whereas,it cannot eliminate allergenicity completely,and sometimes even improves allergenicity by exposing new epitopes.Moreover,most food processing techniques have been tested on purified food allergens rather than food products due to potential interference of other food components.We provide guidance for further development of processing operations that can decrease the allergenicity of allergenic food proteins without negatively impacting the nutritional profile.

Allergen degradation of bee pollen by lactic acid bacteria fermentation and its alleviatory effects on allergic reactions in BALB/c mice

Food allergy as a global health problem threatens food industry.Bee pollen(BP)is a typical food with allergenic potentials,although it performs various nutritional/pharmacological functions to humans.In this study,lactic acid bacteria(LAB)were used to ferment Brassica napus BP for alleviating its allergenicity.Four novel allergens(glutaredoxin,oleosin-B2,catalase and lipase)were identified with significant decreases in LAB-fermented BP(FBP)than natural BP by proteomics.Meanwhile,metabolomics analysis showed significant increases of 28 characteristic oligopeptides and amino acids in FBP versus BP,indicating the degradation of LAB on allergens.Moreover,FBP showed alleviatory effects in BALB/c mice,which relieved pathological symptoms and lowered production of allergic mediators.Microbial high-throughput sequencing analysis showed that FBP could regulate gut microbiota and metabolism to strengthen immunity,which were closely correlated with the alleviation of allergic reactivity.These findings could contribute to the development and utilization of hypoallergenic BP products.

Validation of Molecular Detection Methods for Gluten Allergens in Infant Formula

[Objectives]To verify the specificity,sensitivity,precision and negative-positive deviation of the foodproof gluten component de-tection kit for the detection of gluten allergens in milk powder matrix,and to establish a real-time fluorescent PCR legal method for the detec-tion of gluten allergens in milk powder.[Methods]The specificity,sensitivity,precision and negative-positive deviation of the detection method of foodproof gluten component detection kit(PCR-probe method)were verified by artificially adding different concentrations of wheat bran and extracting sample DNA by kit method,and applied to sample detection.[Results] The specific detection results of two kinds of milk powder with wheat bran and buckwheat added showed that the foodproof gluten component detection kit(PCR-probe method)had good speci-ficity for wheat gluten.The results of artificially added wheat bran positive samples showed that the false positive rate and false negative rate of the kit in the milk powder matrix were O,and the sensitivity and precision were high.[Conclusions]The kit is simple to operate and has high accuracy,which is suitable for the detection of gluten allergen components in milk powder.

Assessment of the allergenicity and antigenicity potential of enzymatically hydrolyzed cow milk

Cow milk allergy is a major cause of food hypersensitivity in infants and children.However,to the best of our knowledge,there has been minimal research on the antigenicity of cow milk(CM).As it is currently unknown if enzymatically hydrolyzed cow milk(HM)induces severe immunogenic responses,the potential allergenicity of HM was systematically evaluated in this work.HM effectively attenuated symptoms of weight loss and diarrhea,suppressed levels of mast cell tryptase,and relieved allergic inflammation of the intestine.Moreover,HM reduced the B220+cell count in mouse blood.In addition,in vitro studies demonstrated that gastrointestinal digestion of HM was not resistant to pepsin or trypsin.Thus,HM effectively alleviated allergic reactions and exhibited lower potential allergenicity.These results provide practical information for the future development of hypoallergenic CM products.

Antarctic krill antioxidant peptides show inferior IgE-binding ability and RBL-2H3 cell degranulation

Enzymatic hydrolysis,isolation,and purification might make a great deal of difference in antioxidant activity and antigenicity of peptide components.This study aimed to isolate and purify antioxidant peptide components from Antarctic krill and evaluate their allergenicity of them.Electron paramagnetic resonance(EPR)spectroscopy results indicated 3-10 kDa Antarctic krill hydrolysates(AKHs)had higher DPPH and·OH radical scavenging rates.And the second component(N2-2)purified 3-10 kDa hydrolysate showed better ability to scavenge DPPH and·OH radicals(P<0.05),which were(47.43±2.18)%and(34.33±1.25)%,respectively.Additionally,indirect-ELISA results revealed that N2-1 had a weaker ability to bind specific IgE and that N2-2 had a lower binding capability to specific IgG1(P<0.05).And N2-2 had a higher EC50 value of(5.29±0.95)ng/mL(P<0.05)in cell degranulation assay,which was about 13.80 times that of Antarctic krill.Therefore,N2-2 might be the potential source of the antioxidant peptides with lower allergenicity.

Composition analysis of acid hydrolysates from Cucurbita moschata Duch.polysaccharides and their effect on oxidative stress resistance of Caenorhabditis elegans

Pumpkin polysaccharides(PPe)have a variety of bioactive effects and our previous research showed the acid hydrolysate(PPe-S,a mixture)from PPe had an antioxidative capacity both in vitro and in viro.The aim of this study was to purify PPe-S and investigate the antioxidant stress effects of 2 purified components(PPe-S-1 and PPe-S-2)using Caenorhabditis elegans as model organism.The results showed that PPe-S-2 had a notable antioxidant effect,and could significantly enhance the activities of antioxidant enzymes including superoxide dismutase(SOD)(P<0.01),catalase(CAT)(P<0.01)and glutathione reductase(GR)(P<0.05),and increase the level of glutathione(GSH)(P<0.01),and decreased the content of malondialdehyde(MDA)(P<0.05).PPe-S-2 could significantly extend the survival time of C.elegans(P<0.01),which were stress-induced by hydrogen peroxide and methyl viologen.PPe-S-2 was a heteropolys accharide composed of glucose,arabinose,rhamnose and galactose with the molar ratio of 1.00:0.03:0.02:0.14.The molecular weight of PPe-S-2 was 0.73 kDa detected by high performance liquid chromatography.These studies demonstrated that PPe-S-2 obtained by the acid hydrolysis of PPe had a prominent protective effect to the damage induced by the intracellular free radical generating agents.