Analysis of phenolic compounds in Spanish red wines by capillary zone electrophoresis

A simple technique for the routine capillary electrophoretic analysis of phenolic compounds in red wines from different Spanish regions, is described. Separation was achieved using a fused-silica capillary column with a borate buffer at pH 9.5, and UV detection at 280 nm. The compounds identified were: (-)-epicatechin, (+)-catechin, (-)-epigallocatechin, syringic acid, vanillic acid, gallic acid, 3,4 dihydroxybenzoic acid, p-coumaric acid, caffeic acid and the hydroxycinnamic tartaric acid esterscis- andtrans-coumamyl tartaric acid andcis-caffeoyl tartaric acid. Different wines showed very similar electrophoretic profiles, but significant quantitative differences were observed.     

Diversity analysis of lactic acid bacteria in Korean rice wines by culture-independent method using PCR-denaturing gradient gel electrophoresis

A polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was used to determine the presence and diversity of lactic acid bacteria (LAB) in takju, a traditional Korean rice wine. Bacterial DNAs were extracted from 15 commercial rice wines and amplicons of partial 16S rRNA genes were separated by DGGE and intense bands were sequenced. Lactobacillus (Lb.) paracasei, Lb. plantarum, and Leuconostoc pseudomesenteroides were detected in all samples and Lb. harbinensis and Lb. parabuchneri were found with above 80% frequency of occurrence. Unknown species of Lactobacillus were also widely detected. This result revealed that, regardless of products and raw materials, the distribution profiles of LAB in takju products have a common pattern comprising of above predominant species and, furthermore, takju can be regarded as a LAB-rich fermented food providing various probiotics.     

采用毛细管凝胶电泳技术检测蜂王浆新鲜度

新鲜度是评价蜂王浆产品质量一个重要指标。文中采用毛细管凝胶电泳技术对蜂王浆水溶性蛋白质进行了分离测定。结果表明:毛细管电泳技术对蜂王浆水溶性蛋白定量准确、重复性、精密度、稳定性各项指标良好,操作简便,快速,适合于蜂王浆产品的新鲜度的评价。为了筛选新鲜度指示蛋白,将王浆样品在不同温度下放置不同时间,分别进行毛细管电泳分析,最终从蜂王浆19个蛋白峰中筛选到2个蛋白峰,含量随存储温度和时间呈现规律性变化,可以作为蜂王浆新鲜度指示蛋白。     

采用毛细管电泳技术快速检测牛乳中外源蛋白成分

牛乳中非乳源性蛋白成分的检测对于保障牛乳质量安全具有重要意义。介绍了毛细管凝胶电泳技术在牛乳中外源蛋白成分快速检测上的应用。通过优化条件,建立牛乳毛细管凝胶电泳方法,检测到大豆水解蛋白标志峰,分别测定纯大豆水解蛋白溶液及牛乳-大豆水解蛋白混合溶液中的水解蛋白标志峰峰面积-浓度线性关系,并将毛细管电泳与SDS-PAGE垂直板电泳及蛋白芯片电泳进行比较,以验证毛细管电泳结果,并比较不同方法优劣性。     

Quantification of major camel milk proteins by capillary electrophoresis

Proteins from dromedary camel milk (CM) produced in Europe were separated and quantified by capillary electrophoresis (CE). CE analysis showed that camel milk lacks β-lactoglobulin and consists of high concentration of α-lactalbumin (2.01 ± 0.02 mg mL⁻¹), lactoferrin (1.74 ± 0.06 mg mL⁻¹) and serum albumin (0.46 ± 0.01 mg mL⁻¹). Among caseins, the concentration of β-casein (12.78 ± 0.92 mg mL⁻¹) was found the highest followed by α-casein (2.89 ± 0.29 mg mL⁻¹) while κ-casein represented only minor amount (1.67 ± 0.01 mg mL⁻¹). These results were in agreement with sodium dodecyl sulphate-polyacrylamide gel electrophoresis patterns. Overall, CE offers a quick and reliable method for the determination of major CM proteins, which may be responsible for the many nutritional and health properties of CM.     

Fast method to determine furosine in breakfast cereals by capillary zone electrophoresis

Furosine is a well-know marker of the extent of the thermal treatment applied to foodstuffs. A fast, reproducible and accurate capillary zone electrophoresis (CZE) method to analyse furosine focused on complex matrices, such as cocoa-added breakfast cereals, is presented. A representative group of 17 commercial cocoa-added breakfast cereals were determined by the reference method (HPLC) and by CZE. In addition, eight ready-to-eat breakfast cereals were tested to point out the application of the procedure to a simpler matrix. Data from both methodologies are compared by F-test and unpaired Student's t-test, showing no significant differences among them and an adequate correlation between methods was found (r ²=0.9852; p=0.000). The CZE method provides a valuable analytical tool for determination of furosine in cocoa-added breakfast cereals and is suitable for routine analysis where presence of highly retained material could limit the resolution and shelf-life of the analytical columns applied for HPLC. The analysis is carried out in 3 min. The stability of the migration time and the good correlation between HPLC–CZE results support the effectiveness of the method.     

Determination of trans-Resveratrol in wines,herbs and health food by capillary electrophoresis with electrochemical detection

A method based on capillary electrophoresis with electrochemical detection (CE–ED) was developed for the determination of trans-Resveratrol in wines, Chinese medicinal herb Polygonum cuspidatum Sied. et Zucc., and Zijin capsule. The effects of some important factors such as injection time, and applied potential to working electrode were investigated. Operated in a wall-jet configuration, a 300 μm diameter carbon-disk electrode was used as the working electrode, which exhibits good response at +0.85 V (vs. SCE) for trans-Resveratrol. Linearity was obtained in the concentration range from 1.0×10⁻⁴ to 5.0×10⁻⁷ g/ml. The detection limit (S/N=3) was 5.96×10⁻⁸ g/ml. This proposed method has been successfully applied to analyze several actual samples with satisfactory assay results.     

Analysis of fish and squid myofibrillar proteins by capillary sodium dodecyl sulfate gel electrophoresis: actin and myosin quantification

 A capillary electrophoretic method for the separation and quantification of fish and squid myofibrillar proteins was developed. The method uses sodium dodecyl sulfate and β-mercaptoethanol for solubilization and analysis of myofibrillar protein subunits. The separation of the different polypeptides is achieved by the sieving effect of the gel inside the capillary. A calibration curve for myosin heavy chain and actin UV absorbance quantification of these proteins was developed. Received: 2 November 1999 / Revised version: 16 February 2000     

The interaction of proteins during gel electrophoresis

Reversible polymerisation and A + B ? C type reactions among proteins during zone electrophoresis may give rise to tailing, extra bands and mobility changes. One or more of these signs are likely to appear with the milder type of interaction, i.e. equilibrium constant in the range 10²–10⁸, whether the time to equilibrium is measured in hours or less. Two-dimensional electrophoresis is more useful for diagnosing interactions. No definite evidence has been obtained for protein-protein interactions among the slow-moving gliadins of Bison and Cappelle varieties and the total number of bands probably represents the minimum number of molecular species present. The electrophoretic pattern of a given variety does not appear to alter qualitatively when the year or place of growth is changed. It is estimated that a wheat protein concentration needs to be above ～0·1 mg/ml to be observable on a starch gel.     

A robust capillary electrophoresis method for the determination of organic acids in wines

A robust capillary electrophoresis (CE) method for the determination of the six main organic acids in wines is described. The CE method is based on the use of 2,6- pyridinedicarboxylic acid (PDC) as background electrolyte (BGE) for the indirect UV detection of tartaric, malic, citric, succinic, acetic and lactic acids. Ethylenediaminetetracarboxylic acid (EDTA) was added to the BGE to eliminate interferences from trace metals on the determination of citric acid. Problems related to linearity reported in the literature using the same BGE were shown to be the result of the sample pH being lower than that of the buffer. Although this could be overcome simply by adjusting the pH of the sample, electrokinetic injection was found to be a better alternative, improving at the same time the sensitivity of the determination. Linearity over the range 100–5,000 mg/l (ppm) for all acids was achieved, requiring only 80× dilution of standards and samples prior to injection. Within day repeatabilitys (n=7) of migration times (RSD%<0.15) and peak areas (RSD%<5) as determined for a wine sample were found to be excellent. The method was applied to 20 South African wine samples, and the results are compared to data obtained by ion-exclusion liquid chromatography. Succinic acid was consistently overestimated using the latter technique, while for the other acids good agreement between the methods was noted.     

3种毛细管电泳方法分析牛乳蛋白的比较

对比毛细管凝胶电泳(CGE)、毛细管未涂层管区带电泳(CZE)和涂层管区带电泳(CZE)在牛乳蛋白分离和定量分析,结果表明在分离条件、分离效果和定量特征上,毛细管涂层区带电泳优于其他两种电泳。应用涂层毛细管区带电泳对不同阶段婴幼儿配方乳粉的蛋白组成进行测定,分离效果较好。     

Analysis of non-coloured phenolics in port wines by capillary zone electrophoresis Influence of grape variety and ageing

 Different experimental varietal port wines were analysed in order to determine differences in the concentrations of phenolic compounds between some commonly used grapes varieties (Touriga Francesa, Touriga Nacional, Tinta Roriz and Tinta Barroca). Although all samples had the same qualitative pattern of non-coloured phenolic compounds, Touriga varieties contained more than twice the concentration of these compounds compared to the Tinta varieties. A preliminary study of the effect of ageing on these compounds revealed an overall increase in those components associated with the bitter or astringent characteristics of food and beverages (tyrosol, gallic acid and catechin). Received: 1 August 1997     

Analysis of non-coloured phenolics in port wines by capillary zone electrophoresis Influence of grape variety and ageing

 Different experimental varietal port wines were analysed in order to determine differences in the concentrations of phenolic compounds between some commonly used grapes varieties (Touriga Francesa, Touriga Nacional, Tinta Roriz and Tinta Barroca). Although all samples had the same qualitative pattern of non-coloured phenolic compounds, Touriga varieties contained more than twice the concentration of these compounds compared to the Tinta varieties. A preliminary study of the effect of ageing on these compounds revealed an overall increase in those components associated with the bitter or astringent characteristics of food and beverages (tyrosol, gallic acid and catechin). Received: 1 August 1997     

毛细管电泳在蛋白质及其水解产物分析中的应用及进展

蛋白质是人体所有细胞和组织的重要成分,由氨基酸通过脱水缩合组成的多肽链经过盘曲折叠而构成。由于蛋白种类多且结构复杂,传统方法很难对其进行分离分析,电泳方法由此诞生。毛细管电泳是一种将电泳和色谱有机结合的快速分离技术。本文针对食品和药品领域,对毛细管电泳在蛋白质及其水解产物分析中的最新应用及研究进展进行了综述,主要从毛细管电泳的基本原理及主要类型、毛细管电泳与其他技术的联用及应用现状、毛细管电泳在食品和药品中蛋白质及其水解产物的分析应用方面进行了阐述和总结,并展望了毛细管电泳技术在特殊医学用途配方食品中蛋白质和多肽分子量测定方面的发展前景,以期为特殊医学用途配方食品中整蛋白及多肽分子量分布测定方法的开发提供参考。     

毛细管电泳法对乳及乳制品中乳源蛋白的研究

采用毛细管电泳方法对原料乳、市售鲜奶、不同厂家的巴氏灭菌乳、不同厂家和产地超高温灭菌乳(UHT)、调味乳、乳酸饮料、复原乳、酸奶、奶粉中蛋白成分进行检测。选择聚乙烯醇涂层毛细管,采用柠檬酸缓冲体系,在紫外检测214nm、分离电压20kV条件下对乳及乳制品中的α一乳白蛋白(α-La)、β一乳球蛋白(β-Lg)、α-酪蛋白(α-CN)、β-酪蛋白(β-CN)和k-酪蛋白(k-CN)进行分离测定。结果表明:五种蛋白的含量在原料乳(巴氏灭菌乳、市售鲜奶)、UHT乳、酸奶、调味乳、乳酸饮料、复原乳中依次降低,而UHT乳含量随保质期的增加而减少,奶粉中蛋白质含量因其适应人群而有差异。乳及乳制品中蛋白质的含量与其存在形式、产地及加工工艺相关。     

Two-dimensional gel electrophoresis of proteins and peptides in bovine milk

Proteins and peptides in bovine milk and colostrum were separated by two-dimensional gel electrophoresis (2-DE). The proteins were separated in the first dimension by isoelectric focusing in the pH ranges 4–7 and 6–11, and in the second dimension by sodium dodecyl sulphate polyacrylamide gel electrophoresis on 12.5% T homogeneous gels. The peptides were separated in the pH range 3–10 in the first dimension, while 15% T homogeneous gels were used in the second dimension. 2-DE evaluation of samples with various somatic cell counts showed an increased number of peptides with increased cell count. 2-DE analysis of samples heated at 65 and 85 °C for 30 min showed a decrease in the number of proteins and peptides with increased temperature. Colostrum exhibited a peptide pattern in which the intensity and number of spots decreased with time postpartum. These results show that 2-DE is a useful tool to detect variations of proteins and peptides in milk.     

聚丙烯酰胺凝胶电泳法快速检测植物源性食品中的蛋白成分

目的建立十二烷基磺酸钠-聚丙烯酰胺凝胶电泳法(sodium dodecyl sulfate polyacrylamide gel electrophoresis,SDS-PAGE)鉴定核桃、杏仁、花生、大豆中的蛋白成分,并分析植物蛋白饮料中蛋白质的来源。方法采用一步法植物活性蛋白质提取试剂盒分别提取预处理后的核桃、杏仁、花生、大豆等样品中的蛋白质成分,计算蛋白质的相对分子量,之后分别进行SDS-PAGE检测得到这几种蛋白成分的特征条带,并对不同样品中蛋白成分的电泳图谱进行比对。结果每一物种都有相对应的蛋白指纹图谱,不同种类的核桃含有相同的蛋白质亚基,具有高同源性,但核桃与花生、黄豆和杏仁含有不同种类大小的蛋白,具有高特异性。结论本方法快速、准确、灵敏,可有效分析植物蛋白饮料中各类蛋白质的来源,对食品中植物源性成分的鉴定具有重要意义。     

Detection of Clostridium botulinum neurotoxin coding genes: analysis of PCR products by real time versus capillary gel electrophoresis methods

In this work, two PCR-based methods have been developed for the detection of Clostridium botulinum strains carrying the gene coding for C. botulinum neurotoxin C (BoNTC) responsible for avian botulism. Both methods are based on the same amplification primers designed using multiple sequence alignments between toxin C coding sequences from DNA sequence databases. The first is a real-time PCR method, using a Taqman-MGB probe. The second uses conventional end-point PCR, followed by capillary gel electrophoresis with laser-induced fluorescence detection (CGE-LIF). A comparison between both methods has been established for the individual and simultaneous detection of toxin C (BONTC) or bacterial 16S (BACT) sequences from C. botulinum. The results indicate that, in general, the same sensitivity was achieved by using RT-PCR and PCR-CGE-LIF allowing the detection of both C. botulinum amplicons from concentrations as low as 7 × 10⁻⁵ μg/ml of total genomic DNA. Some other features from RT-PCR and CGE-LIF are also critically discussed in this work, including quantification capability, size determination, analysis speed and identification strategies, to provide enough information to adequately select the best analytical technique in each case. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

烟草叶片蛋白质双向电泳技术体系的建立

以烟草品种K326旺长后期叶片为材料,用IEF/SDS-PAGE双向凝胶电泳技术对5种蛋白质提取方法(TCA/丙酮沉淀法、Mg/NP-40法、苯酚法、TCA-苯酚法和可溶性蛋白质提取法)进行比较,并对自制管胶pH范围进行探索。结果表明:苯酚法图谱中蛋白质条带最多、最清晰,并可获得较高纯蛋白质得率,为0.0420%,且图谱中蛋白质条带数最多、最清晰。自制管胶pH范围优化试验结果表明,以两性电解质pH 3-10与pH 5-8比例为1:3的2-DE图谱质量最佳,蛋白质可得到较好的分离。     

Analysis of malts by capillary electrophoresis

A range of malts, as well as their high‐ and low‐molecular‐mass fractions, has been examined by capillary electrophoresis in phosphate buffer, pH 2.5, and in carbonate buffer, pH 9.5, and the results have been compared with those for roasted barley and for caramels. The malts fall into two categories: (i) the lightly roasted malts, where the high‐molecular‐mass coloured fraction is negatively charged at pH 9.5 and positively charged at pH 2.5; and (ii) the highly roasted malts (and the roasted barley), where the high‐molecular‐mass fraction migrates close to the electro‐osmotic flow at both pH 9.5 and 2.5, implying that it carries little or no charge. The former category shows migration patterns similar to Class III caramels, whereas the latter migrates differently from Class I, III and IV caramels as well as from the former. Capillary electrophoresis therefore has considerable potential for differentiating between malts and between malts and caramels and roasted barley. © 2002 Society of Chemical Industry