Fully electrochemical hyphenated flow system for preconcentration,cleanup, stripping,capillary electrophoresis with stacking and contactless conductivity detection of trace heavy metals

Successful coupling of electrochemical preconcentration (EPC) to capillary electrophoresis (CE) with contactless conductivity detection (C⁴D) is reported for the first time. The EPC–CE interface comprises a dual glassy carbon electrode (GCE) block, a spacer and an upper block with flow inlet and outlet, pseudo-reference electrode and a fitting for the CE silica column, consisting of an orifice perpendicular to the surface of a glassy carbon electrode with a bushing inside to ensure a tight press fit. The end of the capillary in contact with the GCE is slant polished, thus defining a reproducible distance from the electrode surface to the column bore. First results with EPC–CE–C⁴D are very promising, as revealed by enrichment factors of two orders of magnitude for Tl, Cu, Pb and Cd ion peak area signals. Detection limits for 10 min deposition time fall around 20 nmol L^?1 with linear calibration curves over a wide range. Besides preconcentration, easy matrix exchange between accumulation and stripping/injection favors procedures like sample cleanup and optimization of pH, ionic strength and complexing power. This was demonstrated for highly saline samples by using a low conductivity buffer for stripping/injection to improve separation and promote field-enhanced sample stacking during electromigration along the capillary.     

Analysis of inorganic cations in biological samples by the combination of micro-electrodialysis and capillary electrophoresis with capacitively coupled contactless conductivity detection

Micro-electrodialysis (μED) and CE were combined for rapid pretreatment and subsequent determination of inorganic cations in biological samples. Combination of μED with CE greatly improved the analytical performance of the latter as the adsorption of high molecular weight compounds present in real samples on the inner capillary wall was eliminated. Fifty microliter of 80-fold diluted human body fluids such as plasma, serum and whole blood was used in the donor compartment of the μED system requiring less than 1?μL of the original body fluid per analysis. Inorganic cations that migrated through a cellulose acetate dialysis membrane with molecular weight cut-off value of 500?Da were collected in the acceptor solution and were then analyzed using CE-C?D. Baseline separation of inorganic cations was achieved in a BGE solution consisting of 12.5?mM maleic acid, 15?mM L-arginine and 3?mM 18-crown-6 at pH 5.5. Repeatability of the CE-C?D method was better than 0.5% and 2.5% for migration times and peak areas, respectively; limits of detection of all inorganic cations in the presence of 2?mM excess of Na(+) were around 1?μM and calibration curves were linear with correlation coefficients better than 0.998. Repeatability of the sample pretreatment procedure was calculated for six independent electrodialysis runs of artificial and real samples and was better than 11.8%. Recovery values between 96.3 and 110% were achieved for optimized electrodialysis conditions of standard solutions and real samples; lifetime of the dialysis membranes for pretreatment of real samples was estimated to 100 runs.     

Preparation and characterisation of dual-layer latex-coated columns for open-tubular capillary electrochromatographic preconcentration of cations combined in-line with their separation by capillary electrophoresis

A dual-layer ion-exchange latex-coated column was prepared and characterised for on-capillary preconcentration of cations using an open-tubular ion-exchange CEC format. After preconcentration, the analyte cations were eluted with a transient isotachophoretic gradient and separated by CE. The latex double layer was established by first coating the negatively charged wall of the capillary with a layer of cationic quaternary ammonium anion-exchange Dionex AS5A latex particles (60 nm diameter), and then coating a layer of anionic sulphonated cation-exchange Dionex CS3 latex particles (300 nm diameter) onto the underlying AS5A layer. The adhesion of layers is based on electrostatic attractions. Several dual-layer capillaries were characterised for their EOF and ion-exchange capacity and this showed that coatings could be prepared reproducibly by a simple flushing procedure. The dual-layer columns exhibited a moderate, pH-independent EOF (ca. 26 x 10(-9 )m2V(-1)s(-1)) and an ion-exchange capacity of 57 microequiv./g (or 2.69 nequiv./column). Using an 8 cm length of coated capillary combined with a 72 cm length of untreated capillary, a method for on-line preconcentration and separation of monovalent organic bases, alkali metal ions and alkaline earth metal ions by CE was developed. Recoveries for the preconcentration step were 48% for 4-methylbenzylammonium, 43% for benzylammonium, 30-32% for alkali metal ions and 71-75% for alkaline earth cations. In all cases, recoveries were reproducible with RSDs being less than 6.2%. The influences of the ion-exchange selectivity coefficient of the analyte and the sample-loading rate on analyte recovery were also examined. The proposed method was utilised for the determination of alkaline earth cations and low microM detection limits were obtained.     

Recent developments in electrokinetically driven analysis on microfabricated devices

This review is devoted to the rapid developments in the field of microfluidic separation devices in which the flow is electrokinetically driven, and where the separation element forms the heart of the system, in order to give an overview of the trends of the last three years. Examples of microchip layouts that were designed for various application areas are given. Optimization of mixing and injection strategies, designs for the handling of multiple samples, and capillary array systems show the enormous progress made since the first proof-of-concept papers about lab-on-a-chip devices. Examples of functional elements for on-chip preconcentration, filtering, DNA amplification and on-chip detection indicate that the real integration of various analytical tasks on a single microchip is coming into reach. The use of materials other than glass, such as poly(dimethylsiloxane) and polymethylmethacrylate, for chip fabrication and detection methods other than laser-induced fluorescence (LIF) detection, such as mass spectrometry and electrochemical detection, are described. Furthermore, it can be observed that the separation modes known from capillary electrophoresis (CE) in fused-silica capillaries can be easily transferred to the microchip platform. The review concludes with an overview of applications of microchip CE and with a brief outlook.     

Cover Picture: Electrophoresis 2'2011

Issue no. 2 is a regular issue assembled of 16 solid and original research articles distributed over 3 distinct parts. Part I is on novel trends in fundamentals and methodologies including theoretical models for selectivity of charged solutes in MEKC, system peaks in indirect detection, measuring epimerization constants by MEEKC, bundled CE using micro‐structured fibers, 2‐D separations by coupling CIEF and CEC, high speed DNA CE, MCE of N‐glycans and mucin expression in a microfluidic gradient device. Part II is concerned with detection, sensitivity enhancement, on‐column preconcentration and microdialysis sampling involving the design of continuous full filling CEC‐ESI‐MS using nanoparticles, CE‐fluorescence using tapered optical fiber, CZE separation of pesticide residues in water samples with acid‐assisted on‐column preconcentration and CE‐LIF to detect neurotransmitter amino acids and carbamathione in brain microdialysis samples. Novel methods for the separation and profiling of various proteins and large nucleic fragments are described in 4 consecutive papers grouped in part III. Featured articles include: Theoretical models of separation selectivity for charged compounds in micellar electrokinetic chromatography (( 10.1002/elps.201000405 )) Bundled capillary electrophoresis using microstructured fibres ( 10.1002/elps.201000442 )) Two‐dimensional separation system by on‐line hyphenation of capillary isoelectric focusing with pressurized capillary electrochromatography for peptide and protein mapping ( 10.1002/elps.201000419 )) Microchip electrophoresis of N‐glycans on serpentine separation channels with asymmetrically tapered turns ( 10.1002/elps.201000461 ))     

Single-drop microextraction as a powerful pretreatment tool for capillary electrophoresis: A review

Single drop microextraction (SDME) is a convenient and powerful preconcentration and sample cleanup method for capillary electrophoresis (CE). In SDME, analytes are typically extracted from a sample donor solution into an acceptor drop hanging at the inlet tip of a capillary. The enriched drop is then introduced to the capillary for CE analysis. Since the volume of the acceptor drop can be as small as a few nanoliters, the consumption of solvents can be minimized and the preconcentration effect is enhanced. In addition, by covering the acceptor phase with an organic layer or by using an organic acceptor phase, inorganic ions such as salts in the sample solution can be blocked from entering the acceptor phase, providing desalting effects. Here, we describe the basic principles and instrumentation for SDME and its coupling with CE. We also review recent developments and applications of SDME-CE.     

Rapid and simple pretreatment of human body fluids using electromembrane extraction across supported liquid membrane for capillary electrophoretic determination of lithium

Electromembrane extraction was used for simultaneous sample cleanup and preconcentration of lithium from untreated human body fluids. The sample of a body fluid was diluted 100 times with 0.5 mM Tris solution and lithium was extracted by electromigration through a supported liquid membrane composed of 1-octanol into 100 mM acetic acid acceptor solution. Matrix compounds, such as proteins, red blood cells, and other high-molecular-weight compounds were efficiently retained on the supported liquid membrane. The liquid membrane was anchored in pores of a short segment of a polypropylene hollow fiber, which represented a low cost, single use, disposable extraction unit and was discarded after each use. Acceptor solutions were analyzed using capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C(4) D) and baseline separation of lithium was achieved in a background electrolyte solution consisting of 18 mM L-histidine and 40 mM acetic acid at pH 4.6. Repeatability of the electromembrane extraction-CE-C(4) D method was evaluated for the determination of lithium in standard solutions and real samples and was better than 0.6 and 8.2% for migration times and peak areas, respectively. The concentration limit of detection of 9 nM was achieved. The developed method was applied to the determination of lithium in urine, blood serum, blood plasma, and whole blood at both endogenous and therapeutic concentration levels.     

On-line cation-exchange preconcentration and capillary electrophoresis coupled by tee joint interface

An on-line preconcentration method based on ion exchange solid phase extraction was developed for the determination of cationic analytes in capillary electrophoresis (CE). The preconcentration-separation system consisted of a preconcentration capillary bonded with carboxyl cation-exchange stationary phase, a separation capillary for zone electrophoresis and a tee joint interface of the capillaries. Two capillaries were connected closely inside a 0.3 mm i.d. polytetrafluoroethylene tube with a side opening and fixed together by the interface. The preparations of the preconcentration capillaries and interface were described in detail in this paper. The on-line preconcentration and separation procedure of the analysis system included washing and conditioning the capillaries, loading analytes, filling with buffer solution, eluting analytes and separating by capillary zone electrophoresis (CZE). Several analysis parameters, including sample loading flow rate and time, eluting solution and volume, inner diameter and length of preconcentration capillary etc., were investigated. The proposed method enhanced the detection sensitivity of CE-UV about 5000 times for propranolol and metoprolol compared with normally electrokinetic injection. The detection limits of propranolol and metoprolol were 0.02 and 0.1 microg/L with the proposed method respectively, whereas those were 0.1 and 0.5 mg/L with conventional electrokinetic injection. The experiment results demonstrate that the proposed technique can increase the preconcentration factor evidently.     

毛细管电泳-非接触式电导法直接测定偏硅酸

采用毛细管电泳-电容耦合非接触式电导检测(CE-C<'4>D),以2.4mmol/L KOH+1.6mmol/L K<,2>HPO<,4>+0.4 mmol/L 十六烷基溴化铵(CTMAB)为电泳运行液,融硅石英毛细管(45cm×50μm,有效长度40 cm),负高压分离(-15 kV),偏硅酸可在6.0 min内实现...     

毛细管电泳多脉冲溶出安培检测方法的研究

 摘要：将用于电化学检测的三电极与驱动电泳分离的电化学系统的接地电极在毛细管出口处的外面作适当的布置,可最大程度地减少高压电场对安培检测的干扰。多阶脉冲溶出安培检测方式提高了电流检测灵敏度,并可在一定程度上通过不同的溶出电位鉴别分离组分。将该方法应用于铜、锌、铅、铊、镉等离子的毛细管电泳分离,得到了较满意的结果。     

Potentiometric stripping analysis for lead in urine

Potentiometric stripping analysis is based on the preconcentration of analytes by means of potentiostatic reduction and amalgamation at a thin-film mercury electrode. After preconcentration, the potentiostatic circuitry is disconnected and the amalgamated metals are oxidized either by mercury(II) ions or by dissolved oxygen. Lead can be determined in acidified urine samples by potentiometric stripping analysis after the addition of Triton X-100. In deaerated samples the detection limit is 1 μg l^?1, and in non-deaerated samples 12 μg l^t-1, the preconcentration time being 16 min.     

A novel technique for on-capillary preconcentration of anionic compounds applied to the trace analysis of rapamycin in human blood by capillary electrophoresis

A capillary electrophoretic (CE) method with UV detection at 278 nm has been developed for analysis of the immunosuppressant rapamycin (sirolimus) in human blood at low microg.L(-1) levels. Separation has been achieved in an acidic carrier electrolyte containing sodium dodecyl sulfate and 20% v/v methanol. For sample cleanup and preconcentration, both an off-line solid-phase extraction step using a silica-based reversed-phase material and a newly developed on-capillary focusing technique have been employed. The subsequent treatment of rapamycin under alkaline conditions leads to a cleavage of the lacton bond of the molecule, generating a negatively charged carboxylic group which allows electrokinetic injection into the CE instrument. During the injection process, the negatively charged analyte migrates into an acidic carrier electrolyte, so that it becomes neutral due to protonation and is focused at the capillary inlet. Injection times of 300 s at -7.5 kV could be applied without band-broadening. Results for real samples indicated that the method is fully suited for routine applications and may be an attractive alternative to established liquid chromatographic techniques.     

Heart‐cutting 2D‐CE with on‐line preconcentration for the chiral analysis of native amino acids

The use of transient moving chemical reaction boundary (tMCRB) was investigated for the on‐line preconcentration of native amino acids in heart‐cutting 2D‐CE with multiple detection points using contactless conductivity detection. The tMCRB focusing was obtained by using ammonium formate (pH 8.56) as sample matrix and acetic acid (pH 2.3) as a BGE in the first dimension of the heart‐cutting 2D‐CE. Different experimental parameters such as the injected volume and the concentration in ammonium formate were optimized for improving the sensitivity of detection. A stacked fraction from the first dimension was selected, isolated in the capillary, and then separated in the second dimension in the presence of a chiral selector ((+)‐(18‐crown‐6)‐2,3,11,12‐tetracarboxylic acid). This on‐line tMCRB preconcentration coupled with heart‐cutting 2D‐CE was applied with success to the chiral separation of D ,L ‐phenylalanine, and D ,L ‐threonine in a mixture of 22 native amino acids. The sample mixture was diluted in 0.8 M of ammonium formate, and injected at a concentration of 2.5 μM for each enantiomer with a volume corresponding to 10% of the total capillary volume. An LOD (S/N=3) of 2 μM was determined for L ‐threonine.     

Coupled affinity-hydrophobic monolithic column for on-line removal of immunoglobulin G, preconcentration of low abundance proteins and separation by capillary zone electrophoresis

A butyl methacrylate-co-ethylene dimethacrylate (BuMA-co-EDMA) monolith was synthesized by UV initiated polymerization at the inlet end of a 75 microm I.D. fused silica capillary that had been previously coated with a protein compatible polymer, poly(vinyl)alcohol. The monolith was used for on-line preconcentration of proteins followed by capillary electrophoresis (CE) separation. For the analysis of standard proteins (cytochrome c, lysozyme and trypsinogen A) this system proved reproducible. The run-to-run %RSD values for migration time and corrected peak area were less than 5%, which is typical of CE. As measured by frontal analysis using lysozyme as solute, saturation of a 1cm monolith was reached after loading 48 ng of protein. Finally, the BuMA-co-EDMA monolithic preconcentrator was coupled to a protein G monolithic column via a zero dead volume union. The coupled system was used for on-line removal of IgG, preconcentration of standard proteins and CE separation. This system could be a valuable sample preparation tool for the analysis of low abundance proteins in complex samples such as human serum, in which high abundance proteins, e.g., human serum albumin (HSA) and immunoglobulin G (IgG), hinder identification and quantification of low abundance proteins.     

毛细管电泳和毛细管电色谱技术在农药残留检测中的应用

由于毛细管电泳（CE）和毛细管电色谱（CEC）具有所需样品体积小、分离效率高等特点,越来越多的学者已将它们应用到农药残留（简称农残）检测中,并将它们同各种不同的检测器以及样品浓缩方法相结合,以提高检测的灵敏度。本文对CE和CEC两种方法中所涉及的常见的样品预浓缩方法进行了简要的介绍。对各种不同类型的检测器（如紫外检测、荧光检测、电化学检测以及质谱检测等）的优缺点及其在农残检测中的应用情况进行了评述;同时对手性农药的CE和CEC分离检测情况进行了特别介绍;并对CE和CEC在农残分析与检测中的应用前景进行了展望。     

Development and validation of a capillary electrophoresis method with capacitively coupled contactless conductivity detection (CE-C(4) D) for the analysis of amikacin and its related substances

Amikacin is a semisynthetic aminoglycoside antibiotic derived from kanamycin A that lacks a strong UV absorbing chromophore or fluorophore. Due to the physicochemical properties of amikacin and its related substances, CE in combination with capacitively coupled contactless conductivity detection (CE-C(4) D) was chosen. The optimized separation method uses a BGE composed of 20 mM MES adjusted to pH 6.6 by l-histidine and 0.3 mM CTAB that was added as flow modifier in a concentration below the CMC. Ammonium acetate 20 mg.L(-1) was used as internal standard. 30 kV was applied in reverse polarity on a fused silica capillary (73/48 cm; 75 μm id). The optimized separation was obtained in less than 6 min with good linearity (R(2) = 0.9996) for amikacin base. It shows a good precision expressed as RSD on relative peak areas equal to 0.1 and 0.7% for intraday and interday, respectively. The LOD and LOQ are 0.5 mg.L(-1) and 1.7 mg.L(-1) , respectively.     

Separation and determination of the macrolide antibiotics (erythromycin, spiramycin and oleandomycin) by capillary electrophoresis coupled with fast reductive voltammetric detection

Separation and determination of erythromycin, spiramycin and oleandomycin by capillary zone electrophoresis coupled with fast reductive voltammetric detection using an Hg-film electrode was investigated in a simple aqueous phosphate buffer system. The influence of pH, concentration of phosphate, applied voltage, capillary length and dimension on the separation was examined and optimized. The entire separation of erythromycin, spiramycin, and oleandomycin was achieved in a 0.2 mol/L phosphate buffer system without organic modifiers. The electrochemical detection parameters, such as electrode material, applied waveform, scan rate, preconcentration potentials and preconcentration times, were investigated and discussed. This approach provides high separation efficiency and high sensitivity for all compounds, with detection limits (3 x peak-to-peak baseline noise) of 7.5 x 10(-8) mol/L for spiramycin, and 3 x 10(-7) mol/ L for erythromycin and oleandomycin. The calibration plot of peak areas for each separated peak vs. concentration of analyte was found to be linear over three orders of magnitude.     

Determination of Phenols,Inorganic Anions,and Carboxylic Acids in Kraft Black Liquors by Capillary Electrophoresis

Two methods are presented for the quantitative capillary electrophoretic (CE) determination of phenolic lignin degradation compounds as well as of inorganic anions and organic acids in Kraft black liquors. Important phenolic lignin degradation compounds can be rapidly separated by co-electroosmotic CE after acidification of the liquors and subsequent extraction of the compounds with chloroform. A capillary electrophoretic separation of phenolic compounds is performed by using a phosphate/borate electrolyte system and UV detection at 214 nm. In addition, a HPLC method using a gradient with water, methanol, and acetic acid is also developed. Inorganic ions which are of importance to the pulping process can be determined by simply diluting the black liquors after sampling and subsequent analysis with a chromate electrolyte system and indirect UV detection at 185 nm. In addition, the concentration of low molecular aliphatic carboxylic acids can be determined simultaneously within the same run. By method optimization it is possible to separate the anions within one minute and, at the same time, to increase the resolution of the solutes. The electrolyte systems for the CE separations were optimized by varying the pH value and by adding organic solvents. Short separation times are obtained by adding a polycationic EOF modifier (hexadimethrine bromide) to the electrolyte which reverses the electroosmotic flow. A migration of the anionic analytes in the same direction as the electroosmotic flow is thus established.     

Complementary on-line preconcentration strategies for steroids by capillary electrophoresis

Complementary on-line preconcentration strategies are needed when analyzing different classes of solutes in real samples by capillary electrophoresis (CE) with UV detection. The performance of three different on-line preconcentration (focusing) techniques under alkaline conditions was examined in terms of their selectivity and sensitivity enhancement for a group of steroids, including classes of androgens, corticosteroids and estrogens. Electrokinetic focusing of large sample injection plugs (up to 28% of effective capillary length or 22.1 cm) directly on-capillary can be tuned for specific classes of steroids based on changes in their mobility (velocity) using a multi-section electrolyte system in CE. A dynamic pH junction was applied for the selective resolution and focusing of weakly acidic estrogens using borate, pH 11.0 and pH 8.0 in the background electrolyte and the sample, respectively. Sweeping, using an anionic bile acid surfactant and neutral gamma-cyclodextrin (gamma-CD) under alkaline conditions (pH 8), resulted in focusing and separation of the moderately hydrophobic (non-ionic) classes of steroids, such as androgen and corticosteroids. Optimal focusing and resolution of all test steroids under a single buffer condition was realized by a dynamic pH junction-sweeping format using borate, pH 11.0 and bile acid surfactant with gamma-CD in the BGE, whereas the sample is devoid of surfactant at pH 8.0. The design of selective on-line focusing strategies in CE is highlighted by the analysis of microgram amounts of ethynyl estradiol derived from a female contraceptive pill extract using the dynamic pH junction method, which resulted in over a 100-fold enhancement in concentration sensitivity.     

Electrophoretic separations on chips with hydrodynamically closed separation systems

This review focuses on capillary electrophoretic separations performed on capillary electrophoresis chips (CE chips) with hydrodynamically closed separation systems in a context with transport processes (electroosmotic flow (EOF)) and hydrodynamic flow (HDF)) that may accompany the separations in these devices. It also reflects some relevant works dealing with conventional CE operating under such hydrodynamic conditions. The use of zone electrophoresis (ZE), isotachophoresis (ITP) and their on-line combination (ITP-ZE) on the single-column and column-coupling CE chips with the closed separation systems and related problems are key topics of the review. Some attention is paid to sample pretreatment in the separations performed on the CE chips. Here, mainly potentialities of the ITP-ZE combination in trace analysis applications of the miniaturized systems are discussed in a broader extent. Links between the ZE separation and detection provide a frame for the discussion of current status of the detection on the CE chips. Analytical applications illustrate potentialities of the CE chips operating with the closed separation systems (suppressed HDF and EOF) to the determination of small ions present in various matrices by ZE, ITP and ITP-ZE.