Position control and extra-column effects of a microelectrode detector in open-tubular column liquid chromatography

A liquid chromatography system with a potentiometric microelectrode at the exit end of an open-tubular LC column of 3.5m i. d. nearly shows the theoretically [2, 10] predicted performance of 980 000 theoretical plates for non-retained components at a retention time of a few minutes. Possibilities of controlling the position of the microelectrode tip are given. A distance of one column i. d. between detector and column leads to a loss in column performance of about 15%.     

Recent trends in open-tubular capillary electrochromatography

Capillary electrochromatography (CEC), which combines the advantages of the high efficiency of capillary electrophoresis (CE) and the high selectivity of liquid chromatography (LC), has recently received considerable attention. Most CEC experiments have been performed with capillary columns packed with small LC packing materials (1.5–5 μm particle diameter). However, problems such as difficulties in packing the small LC packing materials and fabricating the frits still exist in preparing the CEC column. The use of open-tubular columns in CEC is therefore an alternative approach that can eliminate the problems encountered in packed-column CEC. So far, several types of open-tubular columns have been proposed for CEC separations and in this article recent progress in this area is reviewed.     

Parallel, fluorous open-tubular chromatography using microstructured fibers

Although commonly used in gas chromatography, open-tubular columns for liquid chromatography have seen their development hindered by a number of factors both theoretical and practical. Requiring small diameters, great lengths and specialized detection systems to achieve a proper chromatographic response, columns of this sort have largely been ignored despite the highly desirable column performance an optimized system would provide. Here, we introduce the use of microstructured fibers (MSFs) as a platform for the development of multiplexed open-tubular liquid chromatography (OTLC) columns. The multiple, parallel silica channels presented by the MSF act as a promising substrate for an OTLC column, as they have diameters near the ideal range for interactions (1–3 μm), minimize flow-induced backpressure through their many uniform paths, and increase the loading capacity compared to a single capillary channel of similar size. Additionally, with outer diameters comparable to regular fused silica capillaries, MSFs can easily be employed in conventional chromatographic systems, eliminating the need for specialized equipment. Finally, MSF columns of this type can be functionalized using silane coupling techniques to allow the introduction of a wide variety of stationary phase chemistries. While in this report we explore the potential and limitations of fluorine-functionalized MSFs as OTLC columns, other stationary phase materials could easily be substituted by choosing appropriate silanization reagents. Particular attention here will be paid to the physical and performance characteristics of the fabricated columns, as well as avenues for their improvement and implementation.     

连续二维开管离子交换/反相整体柱毛细管电色谱的构建与评价

在毛细管中原位合成反相整体色谱柱,并在同一根毛细管柱中的其余部分通过在内表面涂覆N-[3-(三甲氧基硅烷)-丙基)]-乙二胺(PEDA)使其具有离子交换功能,制备成连续二维开管离子交换/反相整体柱毛细管电色谱柱.通过对7种有机酸的分离探讨了开管柱中离子交换对分离的影响,进一步以天麻提取物为样品,对二维分离系统加以评价,...     

Combination of electrophoresis,chromatography, and magnetism in a single separation technique: Part 1: a first theoretical evaluation

Magnetic nanoparticles incorporated into the layer of a (polymeric) sorbent, covering the inner surface of a fused silica capillary, can produce—upon applying an electric field across the capillary length—an electromagnetic field that would affect to some extent the separation of charged analytes. A first theoretical assessment of such phenomenon is given here with a view of developing a novel hybrid separation technique based on the principles of electrophoresis, chromatography, and magnetism. Specifically, the effect of built-in magnetic nanoparticles, varying in absolute number, on the strength of axial electric field in an open-tubular column for capillary electrochromatography (CEC) column for CEC—being expressed through the associated changes in near-wall dielectric constant—was analyzed using linearized Poisson–Boltzmann equation.     

高通量高效液相色谱法快速检测西红花中15种非法添加色素

建立高通量的高效液相色谱法同时检测15种西红花中非法添加色素,对于阳性样品采用液相色谱–质谱联用法确证。采用Poroshell 120 EC–C18色谱柱(100 mm×4.6 mm,2.7 μm)分离,以甲醇–0.02 mol/L乙酸铵溶液(用氨水调pH至5.9±0.05)进行梯度洗脱,流量为1.2 mL/min,柱温为35 ℃,以450 nm和500 nm双波长检测。缓冲盐pH值和流动相流量对色谱条件影响大,需严格控制。该方法重现性好,各物质分离度均大于1.5,标准曲线的线性相关系数r为0.998~1.000,样品高、中、低浓度的加标回收率为70.1%~93.1%,测定结果的相对标准偏差为1.2%~6.6%(n=6),检出限为0.01~0.05 mg/kg。该方法精密度良好,分析时间短,分析结果稳定,可用于中药材西红花中15种非法添加色素的快速筛查和含量测定,同时能为相关部门制定监督抽检中药材染色掺伪的补充检验方法。     

Cellulose acetate as stationary phase in microcolumn LC

The retention characteristics of alkylbenzenes and polycyclic aromatic hydrocarbons (PAHs) have been examined in liquid chromatography on a microcapillary column packed with cellulose acetate. Particulate and fibrous cellulose acetate were used as the stationary phase. Fibrous cellulose diacetate was found to be of use as an alternative stationary phase for liquid chromatography, although the separation efficiency was low. The retention characteristics for planar PAHs were linearly correlated with the size of the molecules, but the retention behavior for alkylbenzenes and nonplanar PAHs was affected by exclusion. The retention order was strongly influenced by the addition of dimethylformamide or water to methanol in the mobile phase. The results suggested that the retention behavior is influenced by a slight change in the polymer matrix of cellulose acetate.     

Hydrocarbon group-type analysis with microcapillary-column liquid chromatography/conventional infrared spectrometry

Summary Slurry-packed microcapillary column liquid chromatography with conventional infrared detection is described for the group-type analysis of gasolines. The results indicate that the three eluted groups, saturates, olefins and aromatics, from the microcapillary column are easily detected by conventional microcomputerized infrared spectrometer coupled with a Teflon flow-cell with a good signal-to-noise ratio. This study shows the feasibility of microcapillary column liquid chromatography combined with a low-cost, conventional infrared spectrometer as a routine tool which allows the analyst to use an LC-IR system effectively and economically.     

On-line multidimensional chromatography using packed capillary liquid chromatography and capillary gas chromatography

The introduction of selected fractions from a liquid chromatograph into a gas chromatograph has been described; however, analyses were performed by off-line experiments requiring collection and reinjection of the separate fractions or by on-line procedures where disadvantageously, only a fraction of the separated peak or a well resolved component in a mixture could be introduced into a gas chromatograph. This disadvantage is overcome by the apparatus and method described in this paper, which utilizes a multidimensional chromatography system employing a high efficiency, packed capillary LC column coupled on-line to a capillary gas chromatograph. The liquid chromatograph (so designed) can act as a highly efficient clean-up or chemical class fractionation step prior to introduction into the gas chromatograph, significantly reducing sample preparation times in many applications. Thus minor components in a complex matrix can be determined without prior sample clean-up, an example of which is the determination of polychlorinated biphenyls in a complex hydrocarbon matrix.     

Practical limitations of capillary gas chromatography

Glass capillary gas chromatography is a high resolution separation method which allows the qualitative and quantitative analysis of even complex mixtures, which may contain many components–also isomeric–in a wide range of volatilities, polarities and concentrations. The principal limitation of gas chromatographic application is given by an insufficient volatility of the species to be separated. Elevated temperatures have to be applied if the application range is to be extended and to achieve steep peak profiles, i.e. low detection limits at high resolution. The use of elevated temperatures is limited, of course, by the temperature stability of both the solvent (stationary liquid and support) and the solutes. The problems of trace analysis for low volatility compounds at high resolution and its limitational parameters regarding sampling, separation and detection are discussed. The applicability of glass capillary columns in this field is influenced by the following parameters: tailing behaviour; irreversible adsorption of polar and decomposition of unstable solutes; thermal stability of stationary liquid (including the support deactivation); separation efficiency and sample capacity (film thickness). Multidimensional gas chromatography using capillary columns coupled either with a packed or another capilllary column for preseparations may be applied with advantage in the analysis of complex mixtures.     

The effect and potential of using a temperature controlled separation column with ultra-high pressure microcapillary liquid chromatography/tandem mass spectrometry on proteomic analysis

The microcapillary liquid chromatography (μLC)/tandem mass spectrometry (MS/MS) system has become a prevailing analytical platform in proteomics. Typical proteomic studies aimed at proteome-wide identification of peptides and proteins rely heavily on producing an accurate and reproducible solvent-composition gradient throughout microcapillary separation columns to improve LC separation. With the recent advent of targeted proteomic approaches utilizing the LC retention time as a physicochemical parameter for peptides, high reproducibility of LC separation additionally becomes an important factor. In this study, column temperature elevation is utilized to improve reproducibility and separation efficiency of the μLC-MS/MS system. The simple incorporation of a semi-rigid gas line heater allowed precise control of the temperature of microcapillary columns longer than 70 cm, up to 60 °C. Tryptic enolase peptides were used as a standard sample to evaluate the effect of the controlled temperature elevation on the peptide separation efficiency and reproducibility. In addition to the increased reproducibility in peptide elution time due to the controlled column temperature, the temperature elevation resulted in a decrease in the column operation pressure, which, in turn, allowed a higher solvent flow-rate to be employed using the same LC pumps, leading to further improvements in the performance of μLC systems.     

Experiments with capillary columns having unconventional cross-sectional geometry

The potentials of unconventional capillary columns, e.g. flat, crinkled, whisker and helically coiled open-tubular columns (HOT columns) were evaluated. In comparison to circular columns, no improvement in the column performance, when used under normal capillary GC circumstances, could be observed.     

Determination of Total Radiochemical Purity of [18F]FDG and [18F]FET by High-Performance Liquid Chromatography Avoiding TLC Method

The goal of this work was to present two high-performance liquid chromatography (HPLC) method that could be applied for the determination of the total radioactive purity of 2-deoxy-2-[¹⁸F]fluoro-D-glucose ([¹⁸F]FDG) and O-(2-[¹⁸F]fluoroethyl)-L-tyrosine ([¹⁸F]FET). The separation of [¹⁸F]fluoride ions, [¹⁸F]FET and [¹⁸F]FET intermediate was accomplished on LiChrosper RP-18, 250?×?4 mm, 5 µm (Merck) analytical column. For mobile phase 10 mM potassium dihydrogen phosphate buffer at pH7 (A) and acetonitrile (B) was used: 0–2 min: 15% B; 2–12 min: 85% B; 12–15 min: 15% B, respectively. Analysis of [¹⁸F]FDG was performed using LiChrosper 100 NH₂, 250?×?4.5 mm, 5 µm (Merck) analytical column. The initial mobile phase composition was 10 mM KH₂PO₄ buffer (pH7) and acetonitrile (15:85, v/v) and the acetonitrile ratio was decreased to 15% at 2 min after the sample injection and held for 5 min. Complete elution of [¹⁸F]fluoride ions from stationary phases could be achieved by adding 10 mg/mL K[¹⁹F]F to radioactive samples in a ratio 1:1 during the sample preparation. Recovery of [¹⁸F]fluoride ions ranged from 99.5 to 100.6%. The validation of the developed methods showed good results for linearity (r²?=?0.9981–0.9996), specificity (R_S?=?3.7–10.2), repeatability (%Area RSD_%?=?1.2–4.3%) and limit of quantitation (LOQ?=?1.6–4.5 kBq). During the cross-validation similar radiochemical purity values were obtained by the novel HPLC methods and thin layer chromatography performed according to the recommendations of the Ph. Eur. monographs.

     

Column technology for capillary electrochromatography

Column technologies for capillary electrochromatography (CEC) are reviewed. To achieve high efficiency, the inner diameters of open-tubular and packed columns should be less than 25 and 200 μm, respectively. To obtain acceptable separation speed under typical CEC conditions (e.g. 30 kV, 1 mm s⁻¹ electroosmotic flow velocity, and 2–4×10⁻⁸ m² V⁻¹ s⁻¹ electroosmotic mobility) the column lengths for open-tubular and packed columns should be less than 120 and 60 cm, respectively. Capillary CEC columns are generally classified into three types: packed, open-tubular, and continuous-bed or monolithic. The various column preparation procedures and the advantages and disadvantages of each column type are discussed in detail.     

Perhydroxylcucurbit[6]uril as a highly selective gas chromatographic stationary phase for analytes of wide‐ranging polarity

This work describes the separation performance of a perhydroxylcucurbit[6]uril stationary phase for capillary gas chromatography. The perhydroxylcucurbit[6]uril stationary phase exhibits higher resolving capabilities for critical analytes with better peak shapes than cucurbit[6]uril and conventional stationary phases. The advantageous separation feature of the perhydroxylcucurbit[6]uril stationary phase may originate from its unique structure and favorably balanced interactions with the analytes. Also, the perhydroxylcucurbit[6]uril column shows good separation repeatability with relative standard deviations in the range of 0.01–0.13% for intraday, 0.37–0.82% for interday, and 1.0–4.7% for column‐to‐column repeatability.     

Separation of enantiomers by capillary electrochromatography

The current popularity of capillary electrochromatography (CEC) has led to an increasing number of studies on the development and evaluation of enantioselective CEC systems. These studies clearly demonstrate that the most prominent advantage of electrically driven separation methods, the vastly increased column efficiency as compared to pressure-driven chromatography, can also be experimentally achieved for the separations of enantiomers. In analogy to high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE), several approaches have been used. The addition of a chiral selector to the mobile phase is the simplest method. Less erroneous and more elegant approaches are those that use open-tubular, conventional packed, and monolithic columns containing chiral stationary phases that stereoselectively interact with enantiomers. This review evaluates the new techniques and compares them to enantioselective HPLC and CE. Further, it describes the various concepts of enantioselective CEC and focuses on the current ‘state-of-the-art' column technology.     

芯片液相色谱技术进展

微型化是现代分析仪器发展的重要趋势。微型化液相色谱仪器在提供与常规尺度液相色谱相同甚至更高分离效率的同时,可以有效减少溶剂和样品的消耗;在液相色谱-质谱联用中,低流速进样可以有效提高质谱离子源的离子化效率,提高质谱检测效率;对于极微量样品的分离,微型化的液相色谱可以有效减少样品稀释;液相色谱的微型化还有利于液相色谱仪器整体的模块化和集成化设计。芯片液相色谱是在微流控芯片上制备色谱柱并集成相应的流体控制系统和检测系统。芯片液相色谱是色谱仪器微型化的一种重要方式,受到学术界和产业界的普遍关注,但是这一方式也充满挑战。液相色谱微流控芯片需要在芯片基底材料、芯片色谱柱的结构设计、微流体控制技术、检测器技术等方面做出创新,使微流控芯片系统适配液相色谱分离技术的需要。目前芯片液相色谱领域面临的主要问题在于芯片基底材料的性质难以满足芯片液相色谱进一步微型化和集成化的需求;因此芯片液相色谱在未来的发展中需要着重关注新型微流控芯片基底材料的开发以及微流控芯片通道结构的统一设计。该文着重介绍了芯片液相色谱技术近年来的研究进展,并简要展示了商品化芯片色谱当前的发展情况。     

The use of high performance liquid chromatography for the analysis and monitoring of iron carbonyl complexes in a reaction mixture

The suitability of high performance liquid chromatography (HPLC) as an analytical and preparative tool for the analysis and isolation of the products resulting from a reaction of diironnonacarbonyl [Fe₂(CO)₉] with di-t-butyl-sulfurdiimine [DBSD] was investigated. Reserved phase chromatography with octyl-modified silica as stationary phase and methanol/water mixtures as mobile phase showed a high selectivity for the separation of the products, and by use of gradient elution the separation of 17 products was completed within 30 min, which meant that the progress of the reaction could be monitored, A semi-preparative column (dimensions 250 X 10 mm) was useful in isolating some of the products; with this column 7 mg of reaction products could be injected without loss in efficiency.     

LC‐MS/MS methods for the determination of edaravone and/or taurine in rat plasma and its application to a pharmacokinetic study

Three liquid chromatography–tandem mass spectrometry (LC‐MS/MS) methods were respectively developed and validated for the simultaneous or independent determination of taurine and edaravone in rat plasma using 3‐methyl‐1‐p‐tolyl‐5‐pyrazolone and sulfanilic acid as the internal standards (IS). Chromatographic separations were achieved on an Agilent Zorbax SB‐Aq (100 × 2.1 mm, 3.5 µm) column. Gradient 0.03% formic acid–methanol, isocratic 0.1% formic acid–methanol (90:10) and 0.02% formic acid–methanol (40:60) were respectively selected as the mobile phase for the simultaneous determination of two analytes, taurine or edaravone alone. The MS acquisition was performed in multiple reaction monitoring mode with a positive and negative electrospray ionization source. The mass transitions monitored were m/z [M + H]⁺ 175.1 → 133.0 and [M + H]⁺ 189.2 → 147.0 for edaravone and its IS, m/z [M ? H]^? 124.1 → 80.0 and [M ? H]^? 172.0 → 80.0 for taurine and its IS, respectively. The validated methods were successfully applied to study the pharmacokinetic interaction of taurine and edaravone in rats after independent intravenous administration and co‐administration with a single dose. Our collective results showed that there were no significant alterations on the main pharmacokinetic parameters (area under concentration–time curve, mean residence time, half‐life and clearance) of taurine and edaravone, implying that the proposed combination therapy was pharmacologically feasible. Copyright © 2014 John Wiley & Sons, Ltd.     

Sample introduction in high speed capillary gas chromatography; input band width and detection limits

The speed of analysis in capillary gas chromatography can be substantially increased by reduction of the column inner diameter. However, special demands are then posed upon instrumental design. In particular, the sampling system is highly critical because it has to be capable of delivering extremely small injection band widths which must be compatible with the column inside diameter. This study focuses on the evaluation of two potentially suitable sample introduction systems with respect to input band width and detection limits and their compatibility with small bore (≦ 100 μm) columns in capillary gas chromatography. One of them allows liquid on-column injection, based on liquid splitting, of only a few nl onto small bore (≦ 100 ?m) fused silica columns. For gases, input band widths as low as 1 ms are obtained with this system. The other one is part of a miniaturized gas chromatograph with extremely low dead volume interfaces and detector volumes. It allows input band widths for gases of a few ms. Without any preconcentration ppm concentrations are measured in gaseous samples with a 80 ?m thick film capillary column. It will be shown that a further reduction of the minimum detectable amount and analysis time is possible with this equipment.