克氏原螯虾新型过敏原血蓝蛋白的鉴定及性质研究

以克氏原螯虾为研究对象,鉴定血蓝蛋白的理化性质及其过敏原性。酶联免疫吸附试验结果表明:6份甲壳类过敏患者血清与克氏原螯虾血淋巴发生特异性IgE反应,利用离心和柱层析方法从血淋巴中分离纯化到目的蛋白。SDS-PAGE分析显示纯化的目的蛋白由6个亚基(依次为68,72,76,82,84和88 ku)组成,采用兔抗凡纳滨对虾血蓝蛋白多克隆抗体的免疫印迹试验证实目的蛋白为血蓝蛋白。免疫印迹试验结果显示,纯化的血蓝蛋白与甲壳类过敏患者血清出现了特异的杂交显色条带,表明血蓝蛋白具有IgE结合活性。与克氏原螯虾主要过敏原(原肌球蛋白)相比,血蓝蛋白的糖含量为1.99%,热稳定性好,pH稳定性及对胃蛋白酶的耐受性不如原肌球蛋白,但比原肌球蛋白更耐胰液消化。综合血清学及理化性质分析结果,提示血蓝蛋白是克氏原螯虾的1种新型过敏原。     

蟹类主要过敏原及其消减技术研究进展

蟹是引起食源性过敏的主要食品之一,蟹类过敏原是引起蟹过敏的根源。近年来,过敏原性质的研究及分离纯化技术备受关注,已发现蟹的主要过敏原包括原肌球蛋白、精氨酸激酶、血蓝蛋白等。利用食品加工技术降低蟹致敏性的研究也日益增多,以热加工法、辐射技术、酶处理法和超高压法等为代表的蟹致敏活性消减技术取得了新进展。本文就蟹类主要过敏原的种类、致敏蛋白的制备、蟹与其他甲壳类的交叉过敏性以及蟹类过敏原消减方法等进行综述,以期为蟹过敏的诊断预防和低致敏产品开发提供参考。     

菲律宾蛤仔过敏原的性质研究

文章以菲律宾蛤仔为对象,对其过敏原进行性质研究。采用丙酮沉淀、等电点沉淀、硫酸铵盐析及加热处理等方法纯化过敏原;采用SDS-PAGE和Western-blot鉴定该过敏原;通过p H稳定性试验、消化稳定性试验进一步分析其性质。结果显示菲律宾蛤仔中主要过敏原的分子量为35k Da,耐酸碱性良好,且具有较高的消化稳定性,其降解产物仍具有抗原性。研究验证了菲律宾蛤仔中的主要过敏原是原肌球蛋白(tropomyosin,TM),并对其性质进行了分析,为过敏原的安全控制提供了基础数据。     

马尾藻多酚的提取纯化工艺优化及其对虾原肌球蛋白致敏性的消减作用研究

目的:随着生活水平的不断提高,食物加工程度日益加深、食品成分日益复杂,食物中的病原物质越来越多,消费者面临的食物过敏反应日趋严重。原肌球蛋白（Tropomyosin, TM）是虾类中主要的过敏原,但由于其具有热稳定性,在加工中不易被消减,本文研究了马尾藻（Sargassum carpophyllum）多酚的提取纯化及其对虾原肌球蛋白致敏性的消减作用。方法:首先,比较了马尾藻和其他5种海藻中多酚的含量,通过透明质酸酶抑制活性实验确定出海藻多酚含量且透明质酸酶抑制活性最高的马尾藻为原材料,并通过单因素优化和响应面优化确定了最佳的马尾藻多酚的提取条件,进一步通过大孔树脂和高效液相色谱进行分离纯化。然后,利用间接竞争ELISA和RBL-2H3肥大细胞模型解析马尾藻多酚对虾原肌球蛋白致敏性的消减作用。结果:马尾藻多酚的最佳提取工艺为:提取温度为40℃,提取时间为10 h,马尾藻粉末:75%乙醇（W/V）为1:16,马尾藻多酚提取得率可达18.18 mg/g,经纯化后获得纯度为97.27%的马尾藻多酚,马尾藻多酚可通过结合原肌球蛋白中的芳香族氨基酸和荧光氨基酸,遮蔽和破坏原肌球蛋白的抗原表位,从而消减虾原肌球蛋白的致敏性。结论:本研究优化了马尾藻多酚提取纯化工艺,得到的马尾藻多酚可以显著消减虾原肌球蛋白的致敏性,表现出较强的抗过敏活性。     

虾类过敏原的识别、纯化和检测技术研究

虾类是人类优质的食用蛋白资源之一,也是联合国粮农组织公布的八大类过敏食物之一。虾类过敏反应严重影响着过敏人群的身体健康和生活质量,为此开展虾类过敏原的识别、纯化和检测技术研究非常必要。通过问卷调查初步了解食物过敏现状,获取自诉虾类过敏患者血清和正常人阴性对照血清,采用特异性IgE检测试剂盒筛选虾类过敏血清。提取南美白对虾蛋白,进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳及免疫印迹识别虾类过敏蛋白,并对患者识别率最高的虾类的主要过敏原进行分离纯化。采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳、免疫印迹和酶联免疫吸附测定等方法对纯化虾蛋白进行检测分析。患者识别的南美白对虾致敏原的分子质量依次约为200、175、116、85和36kDa,通过硫酸铵盐析及等电点沉淀等方法可以得到电泳纯的虾主要过敏蛋白。免疫印迹结果证实纯化的虾蛋白是具有过敏原性的原肌球蛋白,在此基础上,建立了虾原肌球蛋白的酶联免疫吸附测定方法。     

凡纳对虾原肌球蛋白硫酸铵沉淀分离纯化方法的优化

虾原肌球蛋白(tropomyosin,TM)食物过敏是一种全球性的食品安全问题,研究发现TM的分离纯化对于系统准确地鉴定和控制过敏具有重要的意义,鉴于此,本研究以南美白对虾(Litopenaeus vannamei)为对象,改进了TM分离纯化的方法。结果表明,抽提液p H 6.5~7.5、硫酸铵饱和度30%时,可以有效分离纯化TM;经喹啉酸法测定其质量浓度最高可达43μg/μL。该方法省去了高效液相色谱、层析技术等对TM的进一步纯化,减少了纯化步骤,避免了在这些纯化步骤中出现的蛋白损失及蛋白稀释;高质量浓度的TM溶液可无须经过冷冻干燥,直接用于对其理化性质及过敏原性的研究。     

菲律宾蛤仔过敏原原肌球蛋白的鉴定与分子克隆

目的了解菲律宾蛤仔中过敏原的情况,对其主要过敏原进行鉴定和分子克隆。方法采用聚丙烯酰胺凝胶电泳和免疫印迹验证原肌球蛋白,双向电泳对蛋白等电点进一步确定。利用差示扫描量热法对蛋白的热性能测定以及蛋白克隆和测序来分析原肌球蛋白。结果菲律宾蛤仔原肌球蛋白的分子量在37 k Da左右,等电点为5.1,热稳定性较强。原肌球蛋白的基因序列全长为855 bp,编码284个氨基酸,对序列进行同源对比,相似性较高。结论本实验证实了原肌球蛋白为菲律宾蛤仔的过敏原,为认识菲律宾蛤仔过敏原提供基础数据。     

原肌球蛋白过敏原的研究进展

原肌球蛋白(Tropomyosin)是一种具有重要调节功能的蛋白质,在很多动物中起着重要作用,但大多数原肌球蛋白都能引发食物过敏反应。本文综述了原肌球蛋白性质功能、原肌球蛋白与疾病和健康的关系以及原肌球蛋白过敏性的消减研究,为进一步对原肌球蛋白过敏原的研究提供一定的参考。     

牡蛎过敏原Crag 1的结构与功能

Crag 1是牡蛎中主要过敏原,属于一种原肌球蛋白。原肌球蛋白是存在于肌肉和非肌肉细胞中高度保守的、分子量大约为34~38 kDa的一种结合蛋白。目前已经明确Crag 1的一级结构是由284个氨基酸构成,二级结构以α-螺旋为主,软体类主要过敏原原肌球蛋白均没有复杂的三、四级空间结构。Crag 1的过敏原性不是来自构象、非连续性表位而是来自于线性、连续性表位。Crag 1在92~105区段(IQLLEEDMERSEER)中具有IgE抗原结合表位,其他的IgE抗原结合表位有待进一步试验确认。Crag 1作为一种原肌球蛋白,对肌肉有一定的调节作用。不同物种原肌球蛋白的序列同源性较高并且存在一定的交叉反应。综述介绍了Crag 1的基本结构性质与功能,系统地描述了Crag 1,讨论了现阶段研究牡蛎过敏的意义及存在的问题,为以后治疗软体动物、甲壳类动物食品的过敏性疾病,尤其是治疗牡蛎引起的过敏疾病,提供一定的参考依据。     

甲壳类水产品过敏原及其致敏性消减技术研究进展

甲壳类水产品味道鲜美,营养丰富,广受消费者喜爱,但可诱发机体产生严重过敏反应,甚至危及生命,已成为全球范围内日益严重的食品安全问题。概述了目前已鉴定的甲壳类水产品过敏原的结构和免疫性质,及其致敏性消减技术原理和研究进展;已报道的甲壳类水产品过敏原有原肌球蛋白、精氨酸激酶、肌质钙结合蛋白、肌球蛋白轻链、磷酸丙糖异构酶和血蓝蛋白等,其中原肌球蛋白为甲壳类水产品的主要过敏原,可与72%~98%的甲壳类食品过敏患者血清产生特异性IgE反应。利用物理加工消减甲壳类水产品过敏原致敏性,主要通过传统热处理、微波、超高压、低温等离子体和辐照等物理作用力诱导蛋白质变性,进而破坏蛋白质的致敏性表位;酸处理和糖基化等化学修饰消减技术可以通过改变过敏原结构、形成新化学键等方式掩盖或直接破坏致敏性表位;酶处理和发酵处理等生物修饰消减技术则直接降解过敏原致敏性表位。未来仍需要通过过敏表位的靶向消减、多种消减技术协同、动物与人体试验开展,探究过敏原结构和表位修饰的影响机制,推进过敏原消减技术的实际应用,为低敏甚至脱敏甲壳类食品的研发提供参考。     

Reliable identification of grapevine rootstock varieties using RAPD PCR on woody samples

Since grapevine ( Vitis spp .) rootstock material is being traded increasingly as disbudded woody material a lack of distinctive morphological features on such material necessitates an alternative and reliable means of identification. Methods described here were developed for rapid and efficient extraction of DNA from woody samples rich in phenolic compounds and polysaccharides, and for subsequent identification of varieties by RAPD PCR. Using these methods, and with the application of only one selected RAPD primer, we were able to differentiate sixteen rootstock varieties, including the seven varieties most commonly used in Germany. Problems commonly encountered with reproducibility of RAPD patterns were avoided by choosing primers with a dinucleotide sequence and a high G/C content that allowed a rather high annealing temperature of 45°C. Methods described here should also be useful for other horticultural crops, especially those with woody tissues rich in phenolic compounds and polysaccharides.     

Chlorohydrins of bisphenol A diglycidyl ether (BADGE) and of bisphenol F diglycidyl ether (BFDGE) in canned foods and ready-to-drink coffees from the Japanese market

BADGE.2HCl and BFDGE.2HCl were determined in 28 samples of ready-to-drink canned coffee and 18 samples of canned vegetables (10 corn, 5 tomatoes and 3 others), all from the Japanese market. HPLC was used as the principal analytical method and GCMS for confirmation of relevant LC fractions. BADGE.2HCl was found to be present in one canned coffee and five samples of corn, BFDGE.2HCl in four samples of canned tomatoes and in one canned corn. No sample was found which exceeded the 1mg/kg limit of the EU for the BADGE chlorohydrins. However the highest concentration was found for the sum of BFDGE.2HCl and BFDGE.HCl.H₂O at a level of 1.5mg/kg. A Beilstein test confirmed that all cans containing foods contaminated with BADGE.2HCl or BFDGE.2HCl had at lest one part coated with a PVC organosol.     

European priorities for research to support legislation in the area of food contact materials and articles

A strong science base is required to underpin the planning and decision-making process involved in determining future European community legislation on materials and articles in contact with food. Significant progress has been made in the past 5 years in European funded work in this area, with many developments contributing to a much better understanding of the migration process, and better and simpler approaches to food control. In this paper this progress is reviewed against previously identified work-areas (identified in 1994) and conclusions are reached about future requirements for R&D to support legislation on food contact materials and articles over the next 5 or so years.     

Aroma characterization of various apricot varieties using headspace–solid phase microextraction combined with gas chromatography–mass spectrometry and gas chromatography–olfactometry

The characterization of the aromatic profile of several apricot cultivars with molecular tracers in order to obtain objective data concerning the aromatic quality of this fruit was undertaken using headspace–solid phase microextraction (HS–SPME). Six apricot cultivars were selected according to their organoleptic characteristics: Iranien, Orangered, Goldrich, Hargrand, Rouge du Roussillon and A4025. The aromatic intensity of these varieties measured by HS–SPME–Olfactometry were defined and classified according to the presence and the intensity of grassy, fruity and apricot like notes. In the six varieties, 23 common volatile compounds were identified by HS–SPME–GC–MS. Finally, 10 compounds, ethyl acetate, hexyl acetate, limonene, β-cyclocitral, γ-decalactone, 6-methyl-5-hepten-2-one, linalool, β-ionone, menthone and (E)-hexen-2-al were recognized by HS–SPME–GC–O as responsible of the aromatic notes involved in apricot aroma and considered as molecular tracers of apricot aromatic quality which could be utilized to discriminate apricot varieties.     

Textural Properties of Cold-set Gels Induced from Heat-denatured Whey Protein Isolates

A 9% whey protein (WP) isolate solution at pH 7.0 was heat-denatured at 80°C for 30 min. Size-exclusion HPLC showed that native WP formed soluble aggregates after heat-treatment. Additions of CaCl2 (10–40 mM), NaCl (50–400 mM) or glucono-delta-lactone (GDL, 0.4–2.0%, w/v) or hydrolysis by a protease from Bacillus licheniformis caused gelation of the denatured solution at 45°C. Textural parameters, hardness, adhesiveness, and cohesiveness of the gels so formed changed markedly with concentration of added salts or pH by added GDL. Maximum gel hardness occurred at 200 mM NaCl or pH 4.7. Increasing CaCl2 concentration continuously increased gel hardness. Generally, GDL-induced gels were harder than salt-induced gels, and much harder than the protease-induced gel.     

Tests of potential functional barriers for laminated multilayer food packages. Part I: Low molecular weight permeants

The advent of the functional barrier concept in food packaging has brought with it a requirement for fast tests of permeation through potential barrier materials. In such tests it would be convenient for both foodstuffs and materials below the functional barrier (sub-barrier materials) to be represented by standard simulants. By means of inverse gas chromatography, liquid paraffin spiked with appropriate permeants was considered as a potential simulant of sub-barrier materials based on polypropylene (PP) or similar polyolefins. Experiments were performed to characterize the kinetics of the permeation of low molecular weight model permeants (octene, toluene and isopropanol) from liquid paraffin, through a surrogate potential functional barrier (25 μm-thick oriented PP) into the food simulants olive oil and 3% (w/v) acetic acid. These permeation results were interpreted in terms of three permeation kinetic models regarding the solubility of a particular model permeant in the post-barrier medium (i.e. the food simulant). The results obtained justify the development and evaluation of liquid sub-barrier simulants that would allow flexible yet rigorous testing of new laminated multilayer packaging materials.     

Occurrence of bisphenol-F-diglycidyl ether (BFDGE) in fish canned in oil

The levels of bisphenol-F-diglycidyl ether (BFDGE) were quantified as part of a European survey on the migration of residues of epoxy resins into oil from canned fish. The contents of BFDGE in cans, lids and fish collected from all 15 Member States of the European Union and Switzerland were analysed in 382 samples. Cans and lids were separately extracted with acetonitrile. The extraction from fish was carried out with hexane followed by re-extraction with acetonitrile. The analysis was performed by reverse phase HPL C with fluorescence detection. BFDGE could be detected in 12% of the fish, 24% of the cans and 18% of the lids. Only 3% of the fish contained BFDGE in concentrations considerably above 1mg/kg. In addition to the presented data, a comparison was made with the levels of BADGE (bisphenol-A-diglycidyl ether)analysed in the same products in the context of a previous study.     

Contribution of European research to risk analysis

The European Commission's, Quality of Life Research Programme, Key Action 1—Health, Food & Nutrition is mission-oriented and aims, amongst other things, at providing a healthy, safe and high-quality food supply leading to reinforced consumer confidence in the safety of European food. Its objectives also include the enhancing of the competitiveness of the European food supply. Key Action 1 is currently supporting a number of different types of European collaborative projects in the area of risk analysis. The objectives of these projects range from the development and validation of prevention strategies including the reduction of consumers risks; development and validation of new modelling approaches; harmonization of risk assessment principles, methodologies, and terminology; standardization of methods and systems used for the safety evaluation of transgenic food; providing of tools for the evaluation of human viral contamination of shellfish and quality control; new methodologies for assessing the potential of unintended effects of genetically modified (genetically modified) foods; development of a risk assessment model for Cryptosporidium parvum related to the food and water industries; to the development of a communication platform for genetically modified organism, producers, retailers, regulatory authorities and consumer groups to improve safety assessment procedures, risk management strategies and risk communication; development and validation of new methods for safety testing of transgenic food; evaluation of the safety and efficacy of iron supplementation in pregnant women; evaluation of the potential cancer-preventing activity of pro- and pre-biotic ('synbiotic') combinations in human volunteers. An overview of these projects is presented here.     

Influence des ions sur le pouvoir hydratant de l'ure e: e tude sur peau de porc ex vivo

This study deals with the influence of ions (NaCl and MgSO4) in a W/O emulsion containing 10% urea. Moisturization kinetics are assessed by corneometry on pig skin ex vivo. The formula's influence on urea penetration is measured by infrared spectrometry with an ATR device and the stripping method. Corneometry and spectroscopy were chosen to record simultaneously the hydratation levels and urea localization into superficial cell layers. Urea crystallization after evaporation of emulsions and aqueous solutions is described. Results show that urea does not hydrate nor penetrate when applied to the skin through an aqueous gel. In a W/O emulsion, sodium chloride increases the ability of urea to moisturize without improving penetration. In vitro urea crystallization is disturbed by sodium chloride or magnesium sulphate for solutions and emulsions. This stabilization by ions is correlated with good moisturization values. The stabilization of urea in the solute state provided by ions increases its water epidermal binding capacity without enhancing penetration.