Effects of gastrointestinal digestion and heating on the allergenicity of the kiwi allergens Act d 1, actinidin, and Act d 2, a thaumatin-like protein

Kiwifruit is a significant elicitor of allergy both in children and adults. Digestibility of two kiwifruit allergens, actinidin (Act d 1) and thaumatin-like protein (Act d 2), was assessed using an in vitro digestion system that approximates physiological conditions with respect to the passage of food through the stomach into the duodenum. Act d 1 precipitated in simulated gastric fluid at pH 2 and digestion of the aggregated protein proceeded slowly. The residual precipitate redissolved completely in simulated duodenal fluid at pH 6.5 and was partially digested. Forty percent of Act d 2 remained intact during gastric digestion and were cleaved by duodenal proteases into large fragments covalently linked by disulfide bonds. Both digested allergen samples displayed nearly unchanged IgE binding abilities. Circular dichroism spectra were used to analyze heat and acid-induced unfolding. Thermal stability of both allergens was strongly pH dependent. While Act d 1 was irreversibly destabilized in acidic solutions, heat-induced denaturation of Act d 2 at pH 2 was fully reversible. IgE binding to Act d 2 but not Act d 1 was detected in processed food products. The stability of Act d 1 and Act d 2 provides one explanation for the allergenic potency of kiwifruit.     

Cysteine proteinase inhibitor Act d 4 is a functional allergen contributing to the clinical symptoms of kiwifruit allergy

Kiwifruit has become a frequent cause of fruit allergy in the recent years. The molecular basis of type I hypersensitivity to kiwifruit is attributed to 11 IUIS allergens, with Act d 1, Act d 2 and Act d 5 characterized in extenso. Evaluation of the allergenic properties of Act d 4, a cysteine proteinase inhibitor from green kiwifruit (Actinidia deliciosa) was performed in this study. Identity of the purified glycoprotein was determined by Edman degradation and by mass fingerprint whereby more than 90% of the primary structure of the mature kiwifruit cystatin was confirmed. Using MALDI TOF analysis, molecular masses of 10902.5 and 11055.2 Da were detected for Act d 4, respectively. Positive skin prick reactivity with Act d 4 was induced in three kiwifruit allergic patients, as well as the upregulation of CD63 and CD203c molecules in the basophile activation assay. The IgE reactivity was detected in dot blot analysis while Western blot analysis was negative using sera from six kiwifruit patients, suggesting the presence of conformational IgE epitopes on the Act d 4 molecule. As activator of effector cells in type I hypersensitivity Act d 4 is a functional allergen contributing to the clinical symptoms of kiwifruit allergy.     

Characterization of Bet v 1-related allergens from kiwifruit relevant for patients with combined kiwifruit and birch pollen allergy

Allergy to kiwifruit appears to have become more common in Europe and elsewhere during the past several years. Seven allergens have been identified from kiwifruit so far, with actinidin, kiwellin and the thaumatin-like protein as the most relevant ones. In contrast to other fruits, no Bet v 1 homologues were characterized from kiwifruit so far. We cloned, purified, and characterized recombinant Bet v 1-homologous allergens from green (Actinidia deliciosa, Act d 8) and gold (Actinidia chinensis, Act c 8) kiwifruit, and confirmed the presence of its natural counterpart by inhibition assays. Well-characterized recombinant Act d 8 and Act c 8 were recognized by birch pollen/kiwifruit (confirmed by double-blind placebo-controlled food challenge) allergic patients in IgE immunoblots and ELISA experiments. The present data point out that Bet v 1 homologues are allergens in kiwifruit and of relevance for patients sensitized to tree pollen and kiwifruit, and might have been neglected so far due to low abundance in the conventional extracts used for diagnosis.     

Quantification of the thaumatin-like kiwi allergen by a monoclonal antibody-based ELISA

Thaumatin-like proteins (TLPs) have been established as a new family of fruit and pollen allergens. The aim of this study was to develop a two-site ELISA for the quantification of the thaumatin-like kiwi allergen (Act d 2) in kiwifruit extracts and kiwifruit-containing food products. Genomic DNA (gDNA) of Act d 2 was amplified and the deduced amino acid sequence was determined to obtain a primary structure. Act d 2 purified from kiwifruit extract by HPLC was identified by Edman degradation and MS. Balb/c mice were immunized with Act d 2 for the production of mAbs by hybridoma technology. The optimized ELISA measured Act d 2 concentrations ranging from 0.2 to 9.0 ng/mL, with intra- and interassay coefficients of variation of 3.65 and 10.44%, respectively. The developed ELISA is a useful method for the quantification of the thaumatin-like kiwi allergen in kiwifruit extracts as well as the allergen level in kiwifruit-containing food products. It may be a helpful analytical tool for the evaluation of the stability (integrity) of fruit allergen extracts for in vitro diagnosis.     

Impact of dietary factors and food processing on food allergy

Allergic reactions to food can significantly reduce the quality of life and even result in life‐threatening complications. In addition, the prevalence of food allergy has increased in the last decades in industrialized countries and the mechanisms underlying (increased) sensitization are still not fully understood. It is believed that the development and maintenance of oral tolerance to food antigens is a process actively mediated by the immune system and that this reaction is essential to inhibit sensitization. Ongoing research indicates that different dietary factors also may contribute to immune homeostasis and oral tolerance to food and that food processing modulates allergenicity. One of the major questions in food allergy research is therefore which impact nutrition and food processing may have on allergenicity of food and perhaps on sensitization. We summarize in this review the different dietary factors that are believed to contribute to induction of oral tolerance and discuss the underlying mechanisms. In addition, the functional consequences of allergen modification will be emphasized in the second part as severity of allergic reactions and perhaps sensitization to food is influenced by structural modifications of food allergens.     

A food matrix reduces digestion and absorption of food allergens in vivo

Scope: Food allergy is caused by primary (class 1) food allergens, e.g. Bos d 5 (cow's milk) and Cor a 8 (hazelnut) or secondary (class 2) food allergens, e.g. Mal d 1 (apple). The latter cannot sensitize susceptible individuals but can cause allergy due to immunological cross‐reactivity with homologous respiratory allergens. Here, we studied the effects of food matrix on gastrointestinal proteolysis, epithelial transport and in vivo absorption of class 1 and class 2 food allergens. Methods and results: Mal d 1 lost its IgE‐reactivity immediately after simulated gastric digestion whereas Bos d 5 and Cor a 8 did not. Only Cor a 8 maintained IgE‐binding capacity after simulated intestinal proteolysis. The presence of hazelnut and peanut extracts, which served as protein‐rich model food matrices, delayed gastrointestinal degradation and reduced epithelial transport rates of all allergens through CaCo‐2 monolayers. Finally, IgE‐reactive allergens were assessed at different time points in sera from rats fed with all three allergens with or without hazelnut extract. The levels of all allergens peaked 2 h after animals were fed without matrix and increased over 8 h after feeding. Conclusions: A protein‐rich food matrix delays gastrointestinal digestion and epithelial transport of food allergens and thereby may affect their sensitizing capacity and clinical symptoms.     

热加工花生的蛋白分离及其过敏原纯化方法研究进展

花生不仅本身是一种营养丰富的食品,而且作为原料或配料广泛应用于食品加工中。然而花生及其制品是FAO/WHO认定的八大类食物过敏原之一,可导致严重的过敏反应,通常伴随终身,甚至危及生命。不同地区的人们食用花生的加工方式不同,其花生过敏的患病率也有所不同,热加工是花生的主要加工方式,因此各类热加工导致的花生致敏性变化成为研究热点。过敏原蛋白的分离作为花生热加工研究中的重要步骤,也变得十分重要。本文主要对常见的3种热加工花生(水煮、油炸和烘烤)中的花生蛋白分离及其过敏原纯化的方法研究进行综述。现有的花生热加工研究中蛋白分离技术主要是通过溶剂浸提;而过敏原纯化技术主要是借助层析法,根据各组分在物理化学性质上的差异进行纯化;此外还可以根据最终研究目的的不同采用其他的辅助方法达到分离纯化效果。通过对现有分离纯化方法进行了解和比较,可为热加工花生过敏原蛋白的分离纯化甚至进一步的分析检测提供理论参考和指导。     

Bovine Milk Allergens: A Comprehensive Review

Cow milk allergy is one of the most common food allergies in early childhood and often persists through adult life, forcing an individual to a complete elimination diet. Milk proteins are present in uncounted food products, such as cheese, yogurt, or bakery item, exposing allergic persons to a constant threat. Many efforts have been made to overcome this global problem and to improve the life quality of allergic individuals. First, proper and reliable food labeling is fundamental for consumers, but the verification of its compliance is also needed, which should rely on accurate and sensitive analytical methods to detect milk allergens in processed foods. At the same time, strategies to reduce milk allergenicity, such as immunotherapy or the use of food processing techniques to modify allergen structure, have to be extensively studied. Recent research findings on the applicability of food processing, such as heat treatment, fermentation, or high pressure, have revealed great potential in reducing milk allergenicity. In this review, significant research advances on cow milk allergy are explored, focusing on prevalence, diagnosis, and therapy. Molecular characterization of cow milk allergens and cross‐reactivity with other nonbovine milk species are described, as well as the effects of processing, food matrix, and digestibility on milk allergenicity. Additionally, analytical methods for the detection of milk allergens in food are described, from immunoassays and mass spectrometry methods for protein analysis to real‐time polymerase chain reaction for DNA analysis.     

三种猕猴桃酒发酵过程中挥发性香气成分的变化

目的:分析不同的果实品种和发酵时间对猕猴桃发酵酒挥发性香气成分的影响。方法:采用顶空固相微萃取/气相色谱-质谱联用技术分析"金艳"、"红阳"和"米良一号"三种猕猴桃发酵酒在发酵过程中的挥发性香气成分。结果:三种猕猴桃酒的主要香气成分包括17种酯类、5种醇类和2种酸类物质,对24种香气成分进行主成分分析,提取出4个主成分。第一主成分包括绝大部分酯类物质,体现猕猴桃酒的花香和果香气息。红阳和米良一号的花香和果香形成于发酵12 d内,金艳的花香和果香形成于发酵5 d内,随后逐渐减弱;在整个发酵过程中金艳的酯类物质总含量高于红阳和米良一号。在第二主成分中主要组分苯乙醇赋予猕猴桃酒玫瑰花香,在整个发酵过程中米良一号的苯乙醇含量明显高于其他两种,具有更为突出的玫瑰花香;红阳、金艳和米良一号的苯乙醇含量分别在发酵5、19和12 d时达到最高。第三主成分反映的是异丁醇和异戊醇,前发酵结束时高级醇含量达到最高,后发酵阶段高级醇含量不断降低;米良一号和红阳的高级醇含量明显高于金艳。第四主成分中乙醇是主要贡献组分,其构成猕猴桃酒的酒香,三种猕猴桃酒在发酵5 d时乙醇含量达到较高水平,12 d时含量略有降低,19 d时含量回升至之前的水平。结论:猕猴桃酒的主体香气成分形成于前发酵阶段,各种挥发性香气成分的含量在后发酵阶段逐渐降低并趋于稳定。金艳猕猴桃酒的酯类物质含量明显高于其他两种,具有更明显的花香和果香,米良一号猕猴桃酒的苯乙醇含量明显高于其他两种,具有更为突出的玫瑰花香。     

食品加工对过敏原活性的影响

近年来, 食物过敏性疾病的发病率明显上升, 已成为影响人类健康最常见的全球性疾病之一。本文对食品加工中常用的几种方法对特定食品过敏原活性的影响进行了综述, 这些加工方法包括热处理、高压处理、研磨、辐照、碱水解、梅拉德反应、酶解、微生物发酵和基因工程等, 并对存在的问题进行了分析。     

Milk allergens, their characteristics and their detection in food: A review

Cow's milk allergy (CMA) is one of the most common food allergies in childhood. This allergy is normally outgrown in the first year of life, however 15% of allergic children remain allergic. Many studies have been carried out to define and characterise the allergens involved in CMA and described two major allergens: casein (αs1-CN) and β-lactoglobulin. In addition to this, many other milk proteins are antigenic and capable of inducing immune responses. Milk from sheep or goats differs from cow's milk (CM) in terms of composition and allergenic properties. Food processing such as heating affects the stability, structure and intermolecular interactions of CM proteins, thereby changing the allergenic capacity. Chemical and proteolytic treatments of milk to obtain milk hydrolysates have been developed to reduce allergic reactions. Prevention of CMA largely relies on avoidance of all food products containing cow's milk. To achieve this, interest has focused on the development of various technologies for detecting and measuring the presence of milk allergens in food products by immunoassays or proteomic approaches. This review describes the technologies implemented for the analysis of milk allergens (allergenicity, biochemistry) as well as their potential detection in food matrices.     

Maillard reaction in food allergy: Pros and cons

Food allergens have a notable potential to induce various health concerns in susceptible individuals. The majority of allergenic foods are usually subjected to thermal processing prior to their consumption. However, during thermal processing and long storage of foods, Maillard reaction (MR) often takes place. The MR is a non-enzymatic glycation reaction between the carbonyl group of reducing sugars and compounds having free amino groups. MR may sometimes be beneficial by damaging epitope of allergens and reducing allergenic potential, while exacerbation in allergic reactions may also occur due to changes in the motifs of epitopes or neoallergen generation. Apart from these modulations, non-enzymatic glycation can also modify the food protein(s) with various type of advance glycation end products (AGEs) such as N?-(carboxymethyl-)lysine (CML), pentosidine, pyrraline, and methylglyoxal-H1 derived from MR. These Maillard products may act as immunogen by inducing the activation and proliferation of various immune cells. Literature is available to understand pathogenesis of glycation in the context of various diseases but there is hardly any review that can provide a thorough insight on the impact of glycation in food allergy. Therefore, present review explores the pathogenesis with special reference to food allergy caused by non-enzymatic glycation as well as AGEs.     

食品加工对蛋清致敏性的影响研究进展

摘要：鸡蛋是人们日常生活中不可或缺的营养品，然而，鸡蛋也是最常见的引起过敏反应的食物之一。婴儿和儿童是鸡蛋过敏的高发人群，鸡蛋过敏反应主要是由血清免疫球蛋白E（IgE）介导的，引发皮肤、消化道、呼吸道相关的症状，甚至可导致危及生命的严重过敏反应。鸡蛋过敏目前尚无特效治愈方法，患者严格避免摄入鸡蛋成分被认为是最有效的预防手段，但严格避食会影响患者的生活质量和膳食营养，且因意外摄入鸡蛋过敏原导致过敏症状出现的情况屡屡发生。可见在鸡蛋过敏的应对方面，迫切需要新的有效方法。食品加工技术作为从鸡蛋致敏的源头上控制过敏反应的方法，近年来发展迅速。本文综述了各种常见食品加工方法对蛋清致敏性的影响，为后续开发低致敏鸡蛋产品提供思路。     

猕猴桃品种对猕猴桃脆片品质影响的评价

目的:以6个品种的猕猴桃为原料,筛选出最适合加工成脆片的品种。方法:采用真空冷冻干燥的方法制备猕猴桃脆片,测定猕猴桃脆片的得率、脆片中水溶性果胶含量、酸溶性果胶含量、可溶性糖含量、总酸含量、糖酸比、抗坏血酸含量及脆片的硬度和脆度等9个指标,并对猕猴桃脆片进行感官评价。结果:不同品种猕猴桃脆片的得率、营养物质含量和感官指标不同。其中金艳猕猴桃脆片的得率、水溶性果胶含量、可溶性糖含量和硬度较高,总酸含量和脆度较低;金梅猕猴桃脆片的得率、水溶性果胶含量、可溶性糖含量和抗坏血酸含量较高,总酸含量、硬度和脆度较低;金圆猕猴桃脆片的水溶性果胶、酸溶性果胶、结合果胶、可溶性糖、总酸和抗坏血酸含量较高,脆片的硬度和脆度较低;金桃猕猴桃脆片的得率、糖酸比、抗坏血酸等含量及脆片的硬度和脆度较高,脆片的水溶性果胶和总酸含量较低;翠玉猕猴桃脆片的得率、糖酸比和脆度较高,脆片的酸溶性果胶含量、结合态果胶含量、可溶性糖含量、总酸含量和脆片的硬度较低;东红猕猴桃脆片的得率、酸溶性果胶含量、结合态果胶含量、可溶性糖含量、糖酸比和脆片的硬度较高,脆片中水溶性果胶含量、总酸含量和脆片的脆度较低。结论:品种对猕猴桃脆片的得...     

Processing effects on tree nut allergens: A review

“Tree nut” is a broad term for classification of nuts that include cashews, almonds, hazelnuts, etc. Reports of mild to adverse immune reactions following the consumption of these nuts has been on a rise in recent years. Currently, about 1.2–2% of the world's population suffer from sensitivity to tree nuts. The only solution is complete abstinence from the allergy causing tree nut which is not feasible in most cases due to issues like cross contamination or their presence in the form of hidden ingredients in processed foods. Various studies have shown that food processing can effectively vary the secondary structures of the allergenic protein which in turn influences their functional properties. But, the impact of these processing methods on tree nuts allergens is mixed. This review gives an update on the recent findings on how conventional and novel processing methods influence the tree nut allergens.     

芝麻过敏原分子特征与检测方法研究进展

食物过敏已成为全世界范围存在的公共健康问题,芝麻是一种常见的食物过敏原,对芝麻过敏原的研究日渐深入。目前,芝麻中已确定的过敏原蛋白有7 种（Ses i 1～Ses i 7）。本文综述了近几年来各国对芝麻过敏原的管理规定、芝麻过敏原蛋白的结构特征、加工过程对其结构及致敏性影响、检测方法等方面的研究进展,以期为采用不同工艺、方法消除或降低芝麻过敏原的致敏性提供理论参考,也为过敏原标签标识的实施提供理论依据。最后,本文总结了芝麻过敏原的研究现状并对未来研究趋势进行了展望。     

甲壳类水产食物致敏原及其分子结构的研究进展

甲壳类水产品是亚洲地区最主要的致敏食物。食物过敏主要由摄入的食品致敏原引起,而食物致敏原的结构不仅与抗原表位的形成有关,也决定了其在食品加工和体内消化过程的稳定性。本文针对甲壳类水产品,首先阐述了甲壳类水产食物过敏及致敏原的研究进展;进一步对甲壳类水产食物致敏原的分子结构及其与致敏性的构效关系展开论述,并基于此提出了致敏原脱敏策略。目前已鉴定的甲壳类水产食物致敏原大部分属于肌动蛋白结合蛋白、磷酸原激酶和EF手型钙离子结合蛋白3个家族。针对不同蛋白质家族致敏原的结构特点,通过食品加工技术消减致敏性或利用分子生物学手段获得低致敏性衍生物都是防控食物过敏的重要途径。但甲壳类水产食物致敏原的结构信息有待完善,其与致敏性的构效关系也仍是未来致敏原研究的重要方向。本文综合阐述了甲壳类水产食物致敏原结构特性及其与致敏性的构效关系,以期为甲壳类水产食物致敏原的系统研究和食品脱敏方法的建立提供思路。     

食物致敏原表位定位技术的研究进展

食物致敏原是引起食物过敏的元凶，多为蛋白质。抗原表位是在抗原分子中与抗体反应或被抗原受体识别，并引发机体免疫应答的特殊化学基团。从表位水平认识食物致敏原，能揭示食物过敏的物质基础，为解决食物过敏问题提供精准的靶标。本文基于表位结构和定位方法的不同，介绍了食物致敏原表位定位技术的发展，并进一步展望了致敏原表位信息对于改进食物过敏鉴定技术和低致敏食物加工方法的应用前景。     

水产品过敏原的研究现状和展望

水产品作为人类食物的重要来源之一,其市场和消费群体不断扩大。与此同时,由水产品引发的食物过敏也日益增多,在联合国粮农组织公布的八大类过敏食物中,水产品就占了两大类。水产品过敏原有小清蛋白(Parvalbumin)、鱼卵蛋白(如鲑鱼的硫酸鱼精蛋白)和胶原蛋白(Collagen)、原肌球蛋白(Tropomyosin,TM)、精氨酸激酶(Arginine kinase)、肌球蛋白轻链(Myosin light chain)、肌钙结合蛋白(Sarcoplasmic calcium binding pro-tein)、血蓝蛋白亚基(Hemocyanin subunits)等。基于此,本文重点概述了国内外水产品过敏原及其加工脱敏技术的研究现状,以及今后水产品过敏原研究的发展趋势。