厚叶海带岩藻聚糖硫酸酯分离纯化及其体外肿瘤抑制活性研究

本研究以日本厚叶海带(Kjellmaniella crassifolia)中提取的岩藻聚糖硫酸酯(Fuciodan)为原料,采用DEAESepharose Fast Flow弱阴离子交换柱层析分离纯化,通过苯酚-硫酸法和氯化钡-明胶法检测各组分中多糖及硫酸根(SO_4~2)含量,MTT法分析50、200、800μg/m L的日本厚叶海带岩藻聚糖硫酸酯粗品(F)及分离组分对肝癌细胞Hep G2、肺癌细胞A549和结肠癌细胞HT-29细胞存活率影响。研究结果表明,DEAE-Sepharose Fast Flow弱阴离子交换柱对粗品F的分离纯化效果良好,可以得到5个组分(F-0、F-1、F-2、F-3和F-4);组分F-4多糖与SO_4~(2-)含量最高,分别为64.25%和26.32%。MTT实验结果显示,岩藻聚糖硫酸酯粗品及各分离组分对三种肿瘤细胞均具有肿瘤抑制活性,不同组分对HepG2、A549、HT-29的最低细胞存活率分别为67.69%、56.03%和27.56%。基于以上研究结果,表明日本厚叶海带岩藻聚糖硫酸酯具有很大的发展潜力,开发成为一种有效的天然抗肿瘤药物。     

海带岩藻聚糖硫酸酯降解及基本结构分析

用离子交换色谱法对海带岩藻聚糖硫酸酯粗糖进行分离纯化,然后采用铜-过氧化氢氧化降解法获得低分子量多糖,对降解条件和产物进行研究。得出结论:离子交换色谱中加入一定浓度氯化钠,可以获得高硫酸根和岩藻糖含量的岩藻聚糖硫酸酯。红外光谱和核磁共振分析确定硫酸根的位置在C2或C3上,糖构型属于α-D-吡喃糖。研究表明自由基氧化降解海带岩藻聚糖硫酸酯的降解条件与其硫酸根和岩藻糖含量有关,硫酸根含量越高,多糖越难以降解。最优降解条件:60℃下,500μL 20mg/mL糖溶液,加入0.0267mol/L乙酸铜,F3、F4加10μL 15%H2O2,F5加50μL15%H2O2,反应4h。采用PAGE凝胶电泳方法可以很好的分析氧化降解产物的分子量分布。     

不同分子质量马尾藻岩藻聚糖硫酸酯的制备和化学组成分析

研究H2SO4降解法制备不同分子质量岩藻聚糖硫酸酯的降解条件及其产物的化学组成。依次采用DEAE-52纤维素离子交换层析法、H2SO4降解法、Sephacryl S-200凝胶色谱,制备不同分子质量的岩藻聚糖硫酸酯。运用高效凝胶色谱测定其相对分子质量,并借助衍生化气相色谱和红外光谱对其单糖的组成和结构进行解析。结果表明:在60℃下,含有1%岩藻聚糖硫酸酯的0.05 mol/L H2SO4溶液反应4 h和8 h时后,可以有效可控地将围氏马尾藻岩藻聚糖硫酸酯降解,经纯化后得到MMWF1、MMWF2、LMWF1、LMWF2四组分,其分子质量分别为16.32、15.49、7.65和7.21 k D,总糖含量分别为57.89%、55.45%、56.18%和52.64%,硫酸基含量分别为18.89%、25.11%、14.42%和19.76%。另外,4组分均由鼠李糖、岩藻糖、木糖、甘露糖、葡萄糖和半乳糖组成,但各组分的单糖含量不同,红外光谱测定结果显示其均含有多糖的特征吸收峰和硫酸基的吸收峰。     

超声波辅助酸液浸提海带岩藻聚糖硫酸酯工艺优化

以海带加工下脚料为原材料,通过单因素试验和正交试验,重点考察了对岩藻聚糖硫酸酯提取率的影响较大的几个因素。试验表明,海带原料中岩藻聚糖硫酸酯浸提最佳工艺参数为:在超声波功率150 W下浸提2次,pH1.00、液料比值30 mL/g、浸提温度70℃、浸提时间75 min。在此条件下,浸提得到的岩藻聚糖硫酸酯的平均提取率为35.06%。     

基于铜藻岩藻聚糖硫酸酯的负载岩藻黄质纳米粒子制备与分析

为防止岩藻黄质进入肠道前在胃中分解,以岩藻聚糖硫酸酯和壳聚糖为原料,构建pH值敏感性纳米载体,用于运载岩藻黄质,以实现岩藻黄质在胃肠道环境中的高效递送。本实验从铜藻中提取岩藻黄质、岩藻聚糖硫酸酯,采用高效液相色谱法测定岩藻黄质纯度,采用高效凝胶渗透色谱法、化学法、高效液相色谱法测定岩藻聚糖硫酸酯的分子质量、理化性质、单糖组成。以岩藻聚糖硫酸酯和壳聚糖为原料,采用聚电解质自组装法按照岩藻聚糖硫酸酯-壳聚糖质量比0.6∶1、0.8∶1、1∶1制备负载岩藻黄质的纳米粒子。测定纳米粒子红外光谱评价岩藻聚糖硫酸酯和壳聚糖的结合状态;测定负载岩藻黄质纳米粒子的理化性质及在模拟胃、肠液中粒径、岩藻聚糖硫酸酯复合率、岩藻黄质负载率的变化,评价其在胃肠道中的稳定性,获得在模拟胃环境中稳定、模拟肠环境中裂解且可实现岩藻黄质高效递送的纳米粒子。结果表明,铜藻岩藻黄质的纯度为13.57%;岩藻聚糖硫酸酯的分子质量为342 kDa,总糖质量分数为52.45%,蛋白质量分数为7.96%,糖醛酸质量分数为9.25%,硫酸基质量分数为19.26%;由岩藻糖、半乳糖、甘露糖、葡萄糖醛酸和木糖5种单糖组成,其中岩藻糖和半...     

北大西洋瓜参岩藻聚糖硫酸酯分离纯化及结构分析

目的:从北大西洋瓜参(Cucumaria frondosa)体壁中提取纯化得到一种北大西洋瓜参岩藻聚糖硫酸酯(CF-FUC),并对其结构特征进行研究。方法:利用阴离子交换层析和凝胶过滤层析纯化出CF-FUC组分,以高效凝胶过滤色谱法测定分子质量,以离子色谱法测定硫酸根含量,以柱前衍生高效液相色谱法测定单糖组成,并通过核磁共振技术推测其精细结构。结果:CF-FUC的分子质量为360kD,硫酸根含量为(29.8±3.2)%,单糖组成为岩藻糖、半乳糖、氨基葡萄糖、氨基半乳糖物质的量比为1:0.22:0.07:0.07,岩藻糖存在两位硫酸酯化α型吡喃岩藻糖及非硫酸酯化α型吡喃岩藻糖两种形式。结论:CF-FUC结构有别于其他海参岩藻聚糖硫酸酯(SC-FUC)。     

响应面优化海带岩藻聚糖硫酸酯的提取工艺研究

以海带加工下脚料为试验材料,运用响应面分析,优化超声波辅助酸液提取岩藻聚糖硫酸酯工艺条件。单因素试验显示,在超声波功率为150 W,浸提2次时能达到较好稳定效果。在上述基础上,选取的自变量为浸提液pH值、液料比(mL/g)、浸提温度(℃)和超声波时间(min),响应值为岩藻聚糖硫酸酯提取率(%),结合Box-Behnken试验设计,探讨岩藻聚糖硫酸酯提取率受各因素及其交互作用的影响。结果表明,最佳工艺参数:浸提液为p H1.60、液料比为37∶1(mL/g)、浸提温度为78℃和超声波处理时间为71 min。经试验验证得到,33.86%的岩藻聚糖硫酸酯提取率与34.30%的理论预测值相比,其相对误差为1.28%。由此可见,经优化的工艺参数可靠,回归模型预测效果较好。研究结果可为从海带加工下脚料中岩藻聚糖硫酸酯的提取提供技术参考。     

马尾藻岩藻聚糖硫酸酯纯化及降血脂功能研究

以亨氏马尾藻为原料,运用超声波浸提工艺提取得到的岩藻聚糖硫酸酯(SPF),采用乙醇分级沉淀法纯化后经脱脂、脱蛋白得到初级纯化的岩藻聚糖硫酸酯,喂食高血脂症的小鼠,探讨其降血脂的功能。结果表明:亨氏马尾藻岩藻聚糖硫酸酯高、中、低3个剂量组的降血脂效果都优于药品对照脂必妥的降血脂效果,并且以高剂量的降血脂效果最佳,能够极显著(P0.01)的降低总胆固醇、低密度脂蛋白、甘油三酯的含量,能显著(P0.05)提高高密度脂蛋白的含量,且高于空白组;能显著降低动脉硬化指数,动脉粥样硬化指数AI1和AI2都有极显著影响(P0.01)。说明亨氏马尾藻初级岩藻聚糖硫酸酯多糖具有明显的降血脂功能。     

半叶马尾藻中岩藻聚糖硫酸酯的提取纯化及抗氧化研究

从半叶马尾藻中提取得到的岩藻聚糖硫酸酯（SPF）,采用十六烷基三甲基溴化铵（CTAB）沉淀法纯化得SPF1,然后经DEAEC-52柱层析得F1和F2两个级分。化学组成分析表明,这4个级分均含有岩藻糖、硫酸基、糖醛酸以及少量的蛋白质。采用比色法研究岩藻聚糖硫酸酯清除羟基自由基（·OH）及超氧阴离子自由基（O2^-·）的作用,结果显示,4种岩藻聚糖硫酸酯级分均有清除活性氧自由基的能力,并且随着纯度的提高其抗氧化能力越强。     

海参岩藻聚糖硫酸酯的超滤纯化工艺研究

采用超滤膜分离技术对海参岩藻聚糖硫酸酯提取液进行纯化,研究膜截留分子量、跨膜压力等对膜通量及纯化效果的影响,并测定了膜污染后的清洗效果。结果发现:截留分子量200 k Da的UV200膜最适于海参岩藻聚糖硫酸酯的分离纯化,所得固形物产品中总糖含量高达56.12%,蛋白质含量仅为6.37%,岩藻糖含量为36.34%,硫酸根含量为10.75%;超滤工艺优化结果显示,跨膜压强以0.2 MPa为宜,蛋白质脱除率高达91.09%,色素脱除率可达65.30%;膜污染后,采用含酶强碱性清洗剂可有效去除膜面污染物。     

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.