恩施鱼塘坝玉米硒富集及玉米肽抗氧化活性研究

为了探究恩施鱼塘坝地区硒资源的分布特点,土壤因素对玉米中硒含量的影响以及硒含量和玉米肽(CPs)抗氧化活性的关系,本文采用玉米良种(湘玉十号),在鱼塘坝附近选取16个不同地点种植,并分两季采收,利用原子荧光法对玉米中蛋白质、多糖中硒含量进行分析,同时对种植地土壤样本的硒含量和p H值进行了测定。通过Hep G2人体肝癌细胞模型对高硒CPs以及低硒CPs的抗氧化活性进行研究。结果表明:所有玉米种植地土壤均达到富硒土壤标准(0.4 mg/kg),但其生长的玉米中硒含量存在较大差异,玉米对硒的富集受到土壤硒含量、种植时间以及p H值的共同影响;玉米中蛋白和多糖具有很高的硒含量。高硒玉米和低硒玉米制成的玉米肽均表现出较好的抗氧化能力,但前者显著高于后者(P0.05),在Hep G2细胞抗氧化模型中,500μg/m L浓度为玉米肽最佳抗氧化浓度;与较低硒玉米肽处理组相比,经高硒玉米肽预处理的细胞存活率增加了24.16%。综上所述,p H低于5的土壤对玉米吸收硒的能力有严重抑制作用,弱碱性土壤更适合玉米对于硒的利用,且种植时间较长的老玉米硒含量更高;富硒玉米肽的抗氧化能力显著高于低硒玉米肽。     

富硒玉米肽对四氯化碳致小鼠急性肝损伤的保护作用

研究富硒玉米肽对四氯化碳(Carbon tetrachloride,CCl4)诱导的小鼠肝损伤的保护作用。将健康的雄性昆明小鼠70只,随机分成7组:正常对照组、CCl4模型组、富硒玉米肽低、中、高剂量组(100、200、400mg/kg·bw)、普通玉米肽组(200 mg/kg·bw)、水飞蓟素阳性对照组(50 mg/kg·bw),每组10只。测定比较血清中谷丙转氨酶(ALT)活力、谷胱甘肽(GSH)、三酰甘油(TG)含量,同时测定肝组织中丙二醛(MDA)、谷胱甘肽(GSH)的含量及超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)活力,肝脏指数,并观察肝组织病理学变化。结果显示富硒玉米肽低、中、高剂量组均能极显著抑制CCl4致肝损伤小鼠血清ALT活性和TG含量的升高、GSH含量的降低(P0.01);均能极显著提高肝组织中GSH含量和SOD、GSH-Px活力,并降低肝组织中MDA的含量(P0.01),极显著降低由于肝损伤引起的肝脏肿大(P0.01),改善肝组织损伤程度,并存在剂量效应关系。富硒玉米肽高剂量组的护肝效果与50 mg/kg·bw的水飞蓟素相当,相同剂量的富硒玉米肽的保肝作用极显著优于普通玉米肽(P0.01)。结果表明富硒玉米肽对CCl4所致小鼠急性肝损伤有显著的保护作用,且与普通玉米肽相比,富硒玉米肽的有效保肝剂量显著降低。     

富硒大米肽体内抗氧化活性研究

目的:比较普通大米肽、富硒大米肽和硒代蛋氨酸的体内抗氧化活性。方法:将60只老龄(12月龄)雄性昆明小鼠和10只少龄(1月龄)雄性昆明小鼠分成7组(每组10只):老龄模型组(M组)、少龄对照组(YC组)、普通大米肽组(400 mg/kg·bw,Ⅰ组)、富硒大米肽组(400 mg/kg·bw,Ⅱ组)、硒代蛋氨酸组(2.5μg/kg·bw,Ⅲ组)、富硒大米肽+硒代蛋氨酸组(200 mg/kg·bw+1.225μg/kg·bw,Ⅳ组)和白藜芦醇组(10 mg/kg.bw,PC组),其中富硒大米肽组、硒代蛋氨酸组以及富硒大米肽+硒代蛋氨酸组中的硒含量一致,连续灌胃30 d。结果显示:与M组相比,YC组、Ⅱ组、Ⅲ组、Ⅳ组、PC组小鼠血清中超氧化物歧化酶(SOD)活性极显著提高(p0.01);YC组、Ⅰ组、Ⅳ组的还原型谷胱甘肽(GSH)含量显著增加(p0.05),Ⅱ组和PC组GSH含量极显著增加(p0.01);YC组、Ⅱ组、PC组的过氧化氢酶(CAT)活性显著增加(p0.05);Ⅲ组、Ⅳ组的肝匀浆中谷胱甘肽过氧化物酶(GSH-Px)活性显著提高(p0.05),Ⅱ组的GSH-Px活性极显著增加(p0.01);Ⅱ组、Ⅳ组、PC组的T-AOC显著增加(p0.05);YC组、Ⅱ组的丙二醛(MDA)显著减少(p0.05),PC组的MDA极显著减少(p0.01);YC组、PC组的GSH显著增加(p0.05),Ⅱ组的GSH极显著增加(p0.01),以及胸腺、脾脏指数等指标均有不同程度地改善。其中所有指标以富硒大米肽组效果最好,抗氧化活性显著高于同剂量的普通大米肽,大多数指标也显著优于同等硒含量的硒代蛋氨酸组。结论:富硒大米肽中的硒与肽对增强抗氧化能力有一定的增效作用。     

富硒大豆低聚肽的抗氧化活性研究

研究了富硒大豆低聚肽在抗氧化方面的作用。将3周龄SD雄性大鼠90只,随机分为6组,即Ⅰ(对照组)、Ⅱ(大豆蛋白组)、Ⅲ(大豆多肽组)、Ⅳ(亚硒酸钠组)、Ⅴ(大豆低聚肽组)、Ⅵ(富硒大豆低聚肽组),在实验环境下,对各组饲喂低硒基础饲料的同时,对第Ⅱ组口腔灌喂大豆蛋白,对第Ⅲ组口腔灌喂大豆多肽,对第Ⅳ组口腔灌喂亚硒酸钠,对第Ⅴ组口腔灌喂大豆低聚肽,对第Ⅵ组口腔灌喂富硒大豆低聚肽,第Ⅰ组口腔灌喂同体积的饮用水。饲喂4周,处死后分别测定各实验大鼠血清和肝脏中的MDA(丙二醛)含量、GSH-Px和SOD活性。结果显示:第Ⅱ、Ⅲ、Ⅴ、Ⅳ、Ⅵ组均能降低血清和肝脏中MDA的含量,提高GSH-Px和SOD活性,但第Ⅱ、Ⅲ组作用不显著,第Ⅴ组作用显著,第Ⅳ、Ⅵ组作用极显著。各组样品对血清和肝脏中MDA含量和GSH-Px活性的影响效果趋于一致,但对血清SOD活性的影响明显强于对肝脏SOD活性的影响,实验结果说明富硒大豆低聚肽具有抗氧化功能,这种功能主要通过降低大鼠血清和肝脏中MDA的含量,提高GSH-Px和SOD活性来实现,富硒大豆低聚肽的抗氧化功能要强于无机硒,明显强于不含硒的大豆低聚肽,推测富硒大豆低聚肽中发挥抗氧化功能的主要是微量元素硒。     

亚硒酸钠和硒代蛋氨酸对人结肠腺癌细胞的毒性作用

探讨两种硒化物对人结肠腺癌Caco-2细胞的毒性作用。将Caco-2细胞暴露在不同浓度的亚硒酸钠(SeIV)和硒代蛋氨酸(SeMet)中培养24 h,用Annexin V-FITC/PI双染色法检测在同样浓度的SeIV和SeMet培养下细胞的凋亡率,并设置不做硒处理的空白对照。用MTT法检测细胞存活率;检测细胞培养液中乳酸脱氢酶(LDH)释放率、细胞内的超氧化物歧化酶SOD活力和谷胱甘肽GSH的含量。相同浓度时(0.8μg Se/mL),与对照相比,SeIV诱导细胞凋亡显著率上升(p0.05),SeMet变化不显著。一定浓度的SeIV(≥0.4μg Se/mL)和SeMet(≥40μg Se/mL)与对照组相比均显著降低细胞的存活率(p0.05);随着Se含量的升高细胞的存活率呈下降趋势,SeIV、SeMet的IC_(50)分别为2.56、215.55μg Se/mL。当SeIV浓度为≥0.8μg Se/mL,SeMet浓度为≥40μg Se/mL时,细胞的LDH释放率均显著高于对照组(14.80%)(p0.05)。当SeIV浓度≥4μg Se/mL,SeMet≥4μg Se/mL时,细胞的SOD活性均显著低于对照组(56.76 U/mg prot)(p0.05)。当SeIV浓度≥4μg Se/mL,SeMet浓度为160μg Se/mL时,细胞的GSH含量均显著低于对照组(61.67μg/mg prot)(p0.05)。一定浓度的SeIV和SeMet会使Caco-2细胞产生氧化应激而导致细胞毒性,诱导细胞凋亡。     

HPLC-ICP-MS在线联用分析食品中无机硒和硒氨基酸的形态

以亚硒酸盐(Se(Ⅳ))、硒酸盐(Se(Ⅵ))、硒胱氨酸(Se Cys_2)、硒蛋氨酸(Se Met)、甲基硒代半胱氨酸(Me Se Cys)和硒代乙硫氨酸(Se Et)为硒形态的目标分析物,采用Dionex Ion Pac AS11色谱柱(250 mm×4.0 mm)为分离柱,通过优化流动相的p H、浓度、甲醇含量、流速和色谱柱的柱温等因素对六种目标硒形态分离及不同提取方法对目标分析物提取效率的影响,建立了高效液相色谱-电感耦合等离子体质谱(HPLC-ICP-MS)在线联用同时测定食品中无机硒和硒氨基酸的分析方法。Me Se Cys、Se Met、Se Et、Se(Ⅳ)、Se Cys_2和Se(Ⅵ)的检出限分别为0.25、0.20、0.35、0.15、0.30、0.15μg Se/L;Me Se Cys和Se Cys_2的线性范围为2.0~2500μg Se/L,Se(Ⅳ)和Se(Ⅵ)线性范围为1.2~1200μg Se/L,Se Met和Se Et的线性范围分别是1.5~1500μg Se/L和2.2~2200μg Se/L,各硒形态的线性相关系数均不少于0.9997。将该方法应用于食品中硒形态的分析,其加标回收的回收率为80.8%~106.7%,相对标准偏差(RSDs,n=3)为4.7%~9.6%。方法具有操作简单、方便快速、灵敏度高和环境友好等优点,可满足食品中硒形态定量分析。     

螺旋藻富硒转化含硒蛋白质的鉴定及其抗氧化活性研究

研究富硒螺旋藻(Se SP)培养生物转化含硒蛋白质(Se P)中硒的含量及分布,观察Se P在体外对氧自由基的清除效应。藻体总蛋白(TP)用SDS-PAGE电泳分离后,用电感偶合等离子体质谱技术(ICP-MS)对不同分子量范围蛋白质中的硒含量进行精确定量;通过激光烧蚀(LA)-ICP-MS对纯化含硒藻蓝蛋白(Se PC)硒含量在电泳胶上进行原位检测;用制备SDS-PAGE凝胶电泳分离纯化不同分子量的Se P组份,化学发光方法检测Se P在体外对超氧自由基和羟自由基的清除作用。结果发现,与未富硒培养的螺旋藻(SP)相比,Se SP总蛋白中硒含量提高了32倍,可达805.48μg/g,其中75.81%分布于25 ku以下小分子量含硒蛋白质(LMWSe P)中,LA-ICP-MS证明纯化Se PC亚基含有稳定共价结合活性硒元素,LMWSe P对自由基的直接最大清除率在70%以上。结果提示,利用Se SP培养可转化生产活性Se P,Se SP中低分子量Se P组份是一种具有较高抗氧化活性的天然活性硒资源,其中硒的存在形态、合成转化机制和体内生物活性有待深入研究。     

富硒牡蛎肽的制备及其抗氧化和血管紧张素转化酶抑制活性研究

为开发兼具抗氧化和血管紧张素转化酶(angiotensin converting enzyme, ACE)抑制活性的富硒牡蛎肽，该研究以富硒牡蛎蛋白为原料优化制备富硒牡蛎肽，并对其硒含量、氨基酸组成、抗氧化活性和ACE抑制活性进行评价。结果表明，富硒牡蛎肽的最佳酶解条件为时间4 h、温度37℃、酶底比0.5%、pH 1.0、底物质量浓度7 g/100mL。该条件下制备的富硒牡蛎肽富含与抗氧化和ACE抑制活性相关的疏水性氨基酸和酸性氨基酸，其清除DPPH自由基和ABTS阳离子自由基的EC₅₀值分别为1.365、1.074 mg/mL,并呈现出良好的细胞抗氧化活性(EC50:1.114μg/mL),对HepG2细胞的氧化损伤具有显著的保护作用，且呈量效关系。此外，富硒牡蛎肽的ACE抑制活性较强，这可能与其良好的抗氧化活性具有内在关联性。富硒牡蛎肽具有良好的抗氧化和降血压活性，该研究结果为牡蛎源富硒肽研究奠定了前期基础，为天然富硒营养健康食品的开发提供了理论依据。     

高效液相色谱-电感耦合等离子体质谱法检测富硒苹果中5种硒形态

目的建立高效液相色谱-电感耦合等离子体质谱联用技术(high performance liquid chromatographyinductively coupled plasma mass spectrometry,HPLC-ICP-MS)测定富硒苹果中5种硒形态的方法。方法对样品提取方法、流动相的类型、浓度、p H值等条件进行考察及确定,采用柠檬酸超声提取的方式处理样品,流动相为浓度为5 mmol/L、p H=5.0的柠檬酸溶液。选用Hamiltion PRP-X100阴离子分析柱,色谱进样量为100μL,质谱采用碰撞池模式进行测定。结果本方法在15 min内可以完全分离5种硒形态,硒代胱氨酸(Se Cys2)、甲基硒代半胱氨酸(Me Se Cys)、亚硒酸根(Se(Ⅳ))、硒代蛋氨酸(Se Met)、硒酸根(Se(Ⅵ))的检出限分别为0.6、0.7、1、0.9、1μg/L,样品加标回收率范围为82.1%~98.3%,相对标准偏差为1.6%~3.8%。结论本方法可以简单、快速、准确地测定富硒苹果中硒的5种形态。     

过氧化氢诱导H epG 2细胞氧化应激模型的建立

建立过氧化氢(H_2O_2)介导的HepG2细胞氧化应激模型,为筛选抗氧化活性物质以及揭示抗氧化应激机制提供细胞学模型。采用不同浓度H_2O_2处理Hep G2细胞不同时间,噻唑蓝(3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide,MTT)法检测细胞存活率,二氯荧光黄双乙酸盐(2′,7′-Dichlorofluorescin diacetate,DCFH-DA)法检测活性氧(Reactive oxygen species,ROS)水平,分光光度法检测丙二醛(Malondialdehyde,MDA)含量,以及超氧化物歧化酶(Superoxide dismutase,SOD)、过氧化氢酶(Catlase,CAT)和谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-Px)活性。结果表明,随着H_2O_2浓度升高和作用时间延长,Hep G2细胞存活率降低,ROS水平和MDA含量升高,SOD、CAT和GSH-Px活性降低。与对照组相比,200μmol/L H_2O_22处理Hep G2细胞6 h,细胞存活率显著降低(P0.05),仅为63.1%;ROS水平和MDA含量显著升高(P0.05),分别为127.1%和135.1%;SOD、CAT和GSH-Px活性显著降低(P0.05),分别为77.28%、78.56%和77.41%。H_2O_2诱导HepG2细胞建立氧化应激模型的最佳条件为H_2O_2作用浓度200μmol/L,作用时间6 h。     

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.     

An internet compendium of analytical methods and spectroscopic information for monomers and additives used in food packaging plastics

An internet website (http://cpf.jrc.it/smt/) has been produced as a means of dissemination of methods of analysis and supporting spectroscopic information on monomers and additives used for food contact materials (principally packaging). The site which is aimed primarily at assisting food control laboratories in the European Union contains analytical information on monomers, starting substances and additives used in the manufacture of plastics materials. A searchable index is provided giving PM and CAS numbers for each of 255 substances. For each substance a data sheet gives regulatory information, chemical structures, physico-chemical information and background information on the use of the substance in particular plastics, and the food packaging applications. For monomers and starting substances (155 compounds) the infra-red and mass spectra are provided, and for additives (100 compounds); additionally proton NMR are available for about 50% of the entries. Where analytical methods have been developed for determining these substances as residual amounts in plastics or as trace amounts in food simulants these methods are also on the website. All information is provided in portable document file (PDF) format which means that high quality copies can be readily printed, using freely available Adobe Acrobat Reader software. The website will in future be maintained and up-dated by the European Commission's Joint Research Centre (JRC) as new substances are authorized for use by the European Commission (DG-ENTR formerly DGIII). Where analytical laboratories (food control or other) require reference substances these can be obtained free-ofcharge from a reference collection housed at the JRC and maintained in conjunction with this website compendium.     

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.     

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.     

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.     

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.     

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.     

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.     

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.