Simultaneous determination of carbohydrates and organic acids in beer and wine by ion chromatography

The simultaneous determination of mono-organic acids and carbohydrates by ion chromatography with both conductometric and pulsed amperometric detection is described. The carbohydrates, such as mannitol, arabinose, glucose, fructose, lactose, sucrose, raffinose, and maltose, as well as monoorganic acids including acetate, glycolate, formate, pyruvate, and fluoride are separated as anions by ion-exchange chromatography with 0.080 mol/L sodium hydroxide eluent at 1 mL/min within 12 min. Carbohydrates are determined by pulsed amperometric detection and mono-organic acids are determined by suppressed conductivity detection. The species in beverages are determined.     

高效阴离子交换色谱-脉冲安培法检测小鼠尿液中的甘露醇、单糖和乳果糖

采用高效阴离子交换色谱-脉冲安培检测法(HPAEC-PAD)建立了小鼠尿液中甘露醇、单糖(包括半乳糖、葡萄糖、甘露糖和果糖)和乳果糖的分析方法。样品经离心沉淀除去蛋白并过分子膜,以CarboPacTM PA1阴离子交换柱为分离柱,采用NaOH梯度淋洗,脉冲安培四电位检测。结果表明,甘露醇、半乳糖、葡萄糖、甘露糖、果糖和乳果糖在0.1～5.0 mg/L内线性良好,线性相关系数r2为0.988～0.999,样品加标回收率为95.5%～104.2%,检出限为0.0013～0.0048 mg/L。此法准确、快速、简便,能同时对6种糖类化合物进行分析,可以跟踪检测整个糖类代谢过程中甘露醇、单糖和乳果糖之间的代谢关系。     

Ion Chromatographic Analysis of Monosaccharides and Disaccharides in Raw Sugar

A simple and effective method using ion chromatography was developed for the simultaneous determination of five monosaccharides (arabinose, glucose, fructose, xylose, and ribose) and two disaccharides (sucrose and lactose) in raw sugar samples. The separation was performed on a CarboPac PA 10 column using the gradient elution of sodium hydroxide and water as the mobile phase. Monosaccharides and disaccharides were detected by an integrated pulsed amperometric detection (IPAD) using gold working electrode. Acid hydrolysis was used for sample preparation before the analysis of glucose and fructose. All the studied sugars showed good linear ranges within 0.5–100 µg mL⁻¹ with the correlation coefficients higher than 0.997. The limits of detection were all less than 0.5 µg mL⁻¹. The RSDs of the method were less than 10 %. The recoveries of the sugars that spiked in raw sugar samples ranged from 96.1 to 102.4 %. The method was successfully applied for the analysis of sugars in raw sugar samples. Sucrose is the major constituent found in the samples at 97 %.

     

Comparative evaluation of extraction procedures and method validation for determination of carbohydrates in cereals and dairy products by capillary electrophoresis

This work describes the development and application of two optimized electrolytes: 15 mmol/L sorbate, 0.2 mmol/L CTAB, and 35 mmol/L sodium hydroxide for the determination of fructose, glucose, maltose, maltotriose, and sucrose in cereals and 15 mmol/L sorbate, 0.3 mmol/L CTAB, and 55 mmol/L sodium hydroxide for the determination of fructose, glucose, galactose, lactose, and sucrose in dairy products. Both methods were validated with respect to linearity, limits of detection (using both signal-to-noise ratio and the Eurachem approach), recovery tests, and intra- and inter-day precision exhibiting adequate performance. Additionally, statistically similar results were obtained in a comparative study of extraction procedures for carbohydrates using the AOAC protocol, ultrasound extraction, magnetic stirring, and sample dissolution.     

离子色谱脉冲安培法测定蜂蜜中的葡萄糖、果糖、蔗糖

利用离子色谱脉冲安培检测器对蜂蜜中葡萄糖、果糖、蔗糖的测定方法进行了研究。采用CARBOPAC^TM PAl分离柱和脉冲安培检测器,对淋洗分离条件进行了实验,选择50mmol/L NaOH作淋洗液,可使蜂蜜中的葡萄糖、果糖、蔗糖很好地分离。检出限分别为：葡萄糖2μg/kg,果糖2μg/kg,蔗糖5μg/kg。加标回收率为90％－108％。该方法只需简单的前处理、无基体干扰,分离效果好,测定准确率高,适用于蜂蜜中葡萄糖、果糖、蔗糖的测定。     

高效阴离子交换色谱-脉冲安培检测法测定烤烟中的水溶性葡萄糖、果糖和蔗糖

用高效阴离子交换色谱-脉冲安培检测法（HPAEC-PAD）测定了烤烟中的水溶性葡萄糖、果糖和蔗糖。采用水浸取及膜过滤法处理烤烟样品,以Dionex CarboPac PA-1阴离子交换柱为色谱柱,0.2 mol/L NaOH水溶液为淋洗液进行分离测定。葡萄糖、果糖和蔗糖的含量与其峰面积的线性关系良好,回收率均在97%以上。方法简便易行,灵敏度高,重现性良好,可以实现对烟草中单糖的快速分离和测定。     

离子色谱中的安培检测方法及其应用

介绍了离子色谱中的安培检测方法(包括恒电位安培检测法、脉冲安培检测法和积分脉冲安培检测法)的原理和应用。脉冲安培检测法与高效阴离子交换色谱结合(HPAEC-PAD)是一种新的分析糖类化合物的方法;积分脉冲安培检测法与高效阴离子交换色谱结合(HPAEC-IPAD)是一种新的氨基酸分析方法。     

烟叶和烟草料液中氨基酸的直接检测及碳水化合物的去除

建立了高效阴离子交换色谱-积分脉冲安培法(HPAEC-IPAD)直接检测烟叶和烟草料液中氨基酸的方法。利用离线除糖的方法,去除烟叶和烟草料液中大量干扰糖类,包括葡萄糖、果糖和蔗糖及麦芽低聚糖等。采用AminoPac PA10阴离子交换柱,以NaOH和NaAc的强碱性溶液为淋洗液,采用梯度洗脱,流速为0.25 mL/min;积分脉冲安培法对氨基酸进行检测,回收率可达76%~105%。此方法可以有效的解决烟叶、烟草料液等含糖量高的样品中糖类化合物的干扰,对氨基酸实现灵敏、准确的定量分析。     

Enzymatic-spectrophotometric determination of sucrose in coffee beans

A spectrophotometric method for determining sucrose is proposed. Sucrose is hydrolyzed by invertase into glucose and fructose. Then, glucose is oxidized in presence of glucose oxidase and the produced hydrogen peroxide reacts with phenol-4-sulfonic acid sodium salt and 4-aminoantipyrine in presence of peroxidase, yielding a pink dye with an absorption maximum at 505 nm. This method was validated following the EURACHEM and VAM project guidelines for method validation. Trueness, precision, robustness, sensitivity and linearity were considered. The method was applied to the determination of sucrose in green and roasted coffee beans. A comparison with the HPLC method with pulsed amperometric detection was carried out.     

Determination of the active ingredients in Gastrodia rhizoma by capillary electrophoresis with electrochemical detection

A simple, reliable and reproducible method, based on capillary electrophoresis (CE) with electrochemical detection (ED), for the determination of five active ingredients and three carbohydrates in extracts of Gastrodia rhizoma is described in this work. The main active ingredients are gastrodin, 4-hydroxybenzyl alcohol, vanillyl alcohol, 4-hydroxybenzylaldehyde and vanillin. Operated in a wall-jet configuration, a 300 microm diameter carbon disc electrode was used as a working electrode, with a good response at +1000 mV (vs. SCE) for 4-hydroxybenzyl alcohol, vanillyl alcohol, 4-hydroxybenzylaldehyde and vanillin; a 300 microm diameter copper disc electrode exhibits a good response at +650 mV (vs. SCE) for gastrodin, sucrose, glucose and fructose. Under optimum conditions, 4-hydroxybenzyl alcohol, vanillyl alcohol, 4-hydroxybenzylaldehyde and vanillin in 100 mmol l(-1) borate buffer (pH 9.2) and gastrodin, sucrose, glucose and fructose in 50 mmol l(-1) sodium hydroxide buffer were baseline separated within 18 min. The response was linear over two orders of magnitude with a detection limit (S/N = 3) in the range 3 x 10(-7)-1.8 x 10(-6) mol l(-1) for all eight analytes. This method was successfully used in the analysis of traditional Chinese medicine, and the assay results were satisfactory.     

离子色谱-柱切换技术同步测定饮料中的三氯蔗糖、葡萄糖、果糖和蔗糖

建立了一种离子色谱-柱切换-安培检测技术同步测定饮料中三氯蔗糖、葡萄糖、果糖和蔗糖的方法。以CarboPac PA10阴离子交换保护柱和分析柱进行分离,用水和250 mmol/L NaOH梯度淋洗,脉冲安培检测。弱保留糖类在25 mmol/L NaOH流动相淋洗下分离检测,通过柱切换将保护柱切换至分析柱后,同时切换至250 mmol/L NaOH淋洗,三氯蔗糖仅通过较短的保护柱分离,4种糖类能够得到同步检测。4种糖类在0.01~20 mg/L范围内有良好的线性关系和较低的检出限,重复性好,样品测定的回收率分别为90.38%~102.88%（三氯蔗糖）、95.56%~102.75%（葡萄糖）、101.66%~114.33%（果糖）和105.03%~106.49%（蔗糖）。该方法可广泛应用于食品中强保留物质的测定。     

Analysis of carbohydrates in plants by high-performance anion-exchange chromatography coupled with electrospray mass spectrometry

A mass spectrometer was coupled to high-performance anion-exchange chromatography (HPAEC) with the help of electrochemical neutralization of the eluent and post-column addition of lithium chloride for carbohydrate analysis. Parallel selective channels (single ion monitoring) were used to decrease the detection limits and separate unresolved peaks. The mass specific detection allowed the simultaneous analysis of a wide range of sugar alcohols, mono-, di- and oligosaccharides. Carbohydrates extracted from leaves of poplar submitted to drought stress were analyzed using pulsed amperometric detection (PAD), then mass spectrometry. It allowed the confirmation of peak attribution and the identification of salicin as a major compound in the extracts. Different responses to water deficit and re-hydration were obtained for several carbohydrates, suggesting different roles in osmoprotection processes.     

高效阴离子交换色谱-脉冲安培检测法分析啤酒和麦汁中的糖

建立了高效阴离子交换色谱-脉冲安培检测(HPAEC-PAD)同时测定单糖、二糖和多种低聚糖的方法。采用水、0.25 mmol/L NaOH溶液和1 mol/L NaAc溶液三元梯度淋洗,在CarboPac PA-100色谱柱上,11种糖在40 min内达到良好分离;采用积分脉冲安培检测方法,无需对样品进行复杂的前处理或衍生反应便可直接检测。11种糖的检出限(S/N=3)在13～88 pg范围内。将该方法用于啤酒和麦汁样品中单糖、二糖及低聚糖的分析取得了很好的结果,样品中的加标回收率为81%～107%。     

Fast determination of sugars in Coke and Diet Coke by miniaturized capillary electrophoresis with amperometric detection

The fast separation capability of a novel miniaturized capillary electrophoresis with amperometric detection (CE-AD) system was demonstrated by determining sugar contents in Coke and diet Coke with an estimated separation efficiency of 60,000 TP/m. Factors influencing the separation and detection processes were examined and optimized. The end-capillary 300 microm Cu wire amperometric detector offers favorable signal-to-noise characteristics at a relatively low potential (+0.50 V vs. Ag/AgCl) for detecting sugars. Three sugars (sucrose, glucose, and fructose) have been separated within 330 s in a 8.5 cm length capillary at a separation voltage of 1000 V using a 50 mM NaOH running buffer (pH 12.7). Highly linear response is obtained for the above compounds over the range of 5.0 to 2.0 x 10(2) microg/mL with low detection limit, down to 0.8 microg/mL for glucose (S/N = 3). The injection-to-injection repeatability for analytes in peak current (RSD < 3.6%) and for migration times (RSD < 1.4%) was excellent. The new miniaturized CE-AD system should find a wide range of analytical applications involving assays of carbohydrates as an alternative to conventional CE and micro-CE.     

Preliminary investigation of the simultaneous detection of sugars,ascorbic acid,citric acid,and sodium benzoate in non-alcoholic beverages by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was used for the simultaneous detection of sugars, ascorbic acid, citric acid, sodium and/or potassium benzoate in non-alcoholic beverages, with meso-tetrakis(pentafluorophenyl)porphyrin (MW 974) as a matrix. Using potassium hydroxide as dopant, fructose/glucose was detected as the potassiated molecule at m/z 219, whereas potassiated sucrose, [Sucrose. K](+), was detected at m/z 381. Using sodium hydroxide as dopant, the fructose and sucrose ions were detected at m/z 203 and 365, respectively. Citric acid generated multiple ions at m/z 269, 307, and 345, which were assigned to [Citricbond;H+2K](+), [Citricbond;2H+3K](+), and [Citricbond;3H+4K](+), respectively. However, a stored methanolic solution of citric acid produced additional ions at m/z 283, 297, and 321, which were attributed to [Citricbond;2H+CH(3)+2K](+), [Citricbond;3H+2CH(3)+2K](+), and [Citricbond;3H+CH(3)+3K](+), respectively, due to esterification that took place during storage. The limits of detection in water were: ascorbic acid, 0.30 wt%; citric acid, 0.5 wt%; and sodium benzoate, 0.001 wt%. In the beverage formulations, the limits of detection were: ascorbic acid 0.3 wt%, citric acid 0.3 wt%, and sodium benzoate 0.02 wt%. Spiking a water or beverage solution that contained ascorbic and/or citric acid with less than 0.6 wt% of tartaric acid lowered the detection limits of ascorbic and citric acids to 0.2 wt%. This study demonstrates the potential for using MALDI-TOFMS in the quality control analyses of non-alcoholic beverages, particularly with regard to the detection of low molecular weight organic acids in commercial beverage formulations.     

Theoretical investigation of low detection sensitivity for underivatized carbohydrates in ESI and MALDI

Electrospray ionization (ESI) and matrix‐assisted laser desorption/ionization (MALDI) mainly generate protonated ions from peptides and proteins but sodiated (or potassiated) ions from carbohydrates. The ion intensities of sodiated (or potassiated) carbohydrates generated by ESI and MALDI are generally lower than those of protonated peptides and proteins. Ab initio calculations and transition state theory were used to investigate the reasons for the low detection sensitivity for underivatized carbohydrates. We used glucose and cellobiose as examples and showed that the low detection sensitivity is partly attributable to the following factors. First, glucose exhibits a low proton affinity. Most protons generated by ESI or MALDI attach to water clusters and matrix molecules. Second, protonated glucose and cellobiose can easily undergo dehydration reactions. Third, the sodiation affinities of glucose and cellobiose are small. Some sodiated glucose and cellobiose dissociate into the sodium cations and neutral carbohydrates during ESI or MALDI process. The increase of detection sensitivity of carbohydrates in mass spectrometry by various methods can be rationalized according to these factors. Copyright © 2016 John Wiley & Sons, Ltd.     

Comparison of Pulsed Electrochemical Detection Modes Coupled with Microchip Capillary Electrophoresis

There is a need to develop analytical methods that are capable of rapidly measuring small biological markers in the field of metabolomics. Among others, carbohydrates play an important role biologically yet are traditionally hard to detect since they have no chromophore or fluorophore. In the present report, the first application of integrated pulsed amperometric detection (iPAD) coupled with microchip electrophoresis to the analysis of glucose, mannose, sucrose, maltose, glucosamine, lactose, maltotriose and galactose is demonstrated. iPAD is an electrochemical detection mode that can be used for direct detection of carbohydrates, amines and sulfur containing compounds. The effect of different solution parameters, including the buffer concentration, pH and the concentration of SDS on both separation and detection response was analyzed. In addition, a comparison study between PAD and iPAD was performed using glucose, glucosamine, sucrose and maltose as model carbohydrates.     

Polysaccharides analysis of sinorhizobial capside by on-line anion exchange chromatography with pulsed amperometric detection and mass spectrometry coupling

High performance anion exchange chromatography (HPAEC)-pulsed amperometric detection (PAD) is a performing technique for carbohydrate analysis, due to the selectivity and sensitivity of the detection. The identification occurs through retention times. In absence of standards, structural characterization of complex polysaccharides requests the coupling of HPAEC-PAD with electrospray ionization (ESI)-MS. This is a technological challenge, due to the non-volatility and high conductance of the eluents. Therefore, a desalting device has been installed on-line between the PAD and the MS. On-line HPAEC-MS has only been rarely described. We report here successful analysis of biological acidic oligosaccharides, allowing for the first time to demonstrate that membrane anchored 3-deoxy-D-manno-2 octulosonic acid (Kdo) homopolymers are consensus sinorhizobial capsular polysaccharide (KPS).     

毛细管区带电泳-间接紫外检测法快速测定食品中乳糖、蔗糖、葡萄糖和果糖

建立了毛细管区带电泳-间接紫外检测快速测定食品中乳糖、蔗糖、葡萄糖和果糖的方法。以水或5 mmol/L醋酸为样品提取液,未涂层熔融石英毛细管(30.2 cm(有效长度20 cm)×50 μm)为分离柱,4 mmol/L山梨酸钾+10 mmol/L磷酸钠+30 mmol/L NaOH(pH 12.56)+0.5 mmol/L十六烷基三甲基溴化铵(CTAB)为分离缓冲液,在-8 kV下分离,于254 nm波长下检测,10 min内实现了食品中上述4种糖的同时分离与测定。乳糖、蔗糖、葡萄糖和果糖的检出限(S/N=3)分别为50、75、25和25 mg/L,定量限(S/N=10)分别为150、225、75和75 mg/L,回收率在87.0%~107.0%之间,相对标准偏差在1.2%~4.7%之间。整个实验过程未使用有机溶剂。用该法测定了9种食品样品及1个质控样品,结果表明该法简单、快速、准确,适用于食品中乳糖、蔗糖、葡萄糖和果糖的日常测定。