On-line Coupling Of Capillary Electrophoresis Research Articles

Comparative study of the vibrating capillary nebulizer (VCN) and commercially available interfaces for on-line coupling of capillary electrophoresis with ICP-MS.

Capillary electrophoresis (CE) is a powerful and sensitive tool for speciation analysis when combined with inductively coupled plasmamass spectrometry (ICP-MS); however, the performance of this technique can be limited by the nature of pneumatic nebulizers. This study compares two commercially available pneumatic nebulizers to a newly introduced vibrating capillary nebulizer (VCN) for on-line coupling of CE with ICP-MS. The VCN is a low-cost, non-pneumatic nebulizer that is based on the design of capillary vibrating sharp-edge spray ionization. As a piezoelectrically driven nebulization source, the VCN creates an aerosol independent of gas flows and does not produce a low-pressure region at the nebulizer orifice. To compare the systems, we performed replicate analyses of sulfate in river water with each nebulizer and the same CE and ICP-MS instruments and determined the figures of merit of each setup. With the CE-VCN-ICP-MS setup, we achieved around 2-4 times lower sensitivity compared to the commercial setups. However, the VCN-based setup provided lower noise levels and better linear correlation from the analysis of calibration standards, which resulted in indistinguishable LOD and LOQ values from the in-house-built VCN-based and commercial setups for CE-ICP-MS analysis. The VCN is found to have the highest baseline stability with a standard deviation of 3500 cts s-1, corresponding to an RSD of 2.7%. High reproducibility is found with the VCN with a peak area RSD of 4.1% between 3 replicate measurements.

On-line coupling of capillary electrophoresis with microdialysis for determining saccharides in dairy products and honey

Free sucrose, lactose, galactose, glucose and fructose were determined in yoghurts, milk and honey using on-line coupling of capillary electrophoresis with microdialysis. The dairy products were diluted 50-fold with 10 mmol/L NaOH and sampled using laboratory-made microdialysis probes. The microdialysate was brought to the entrance of the electrophoretic capillary and the coupling consisted in a polydimethylsiloxane (PDMS) cross connector working in the flow-gating interface regime. The electrophoretic analysis was performed in 50 mmol/L NaOH (pH 12.6) background electrolyte, where baseline separation of the five saccharides was achieved in 3.5 min. The LOQs varied in the range 2.3–7.3 mg/L, the number of separation plates varied between 176,000 plates/m for glucose to 326,000 plates/m for galactose and the relative standard deviation (RSD) for ten consecutive analyses of fruit yoghurt was 0.2% for the migration time and 4.4–7.6% for the peak area.

A simple sheathless CE-MS interface with a sub-micrometer electrical contact fracture for sensitive analysis of peptide and protein samples

Online coupling of capillary electrophoresis (CE) to electrospray ionization mass spectrometry (MS) has shown considerable potential, however, technical challenges have limited its use. In this study, we have developed a simple and sensitive sheathless CE-MS interface based on the novel concept of forming a sub-micrometer fracture directly in the capillary. The simple interface design allowed the generation of a stable ESI spray capable of ionization at low nanoliter flow-rates (45–90 nL/min) for high sensitivity MS analysis of challenging samples like those containing proteins and peptides. By analysis of a model peptide (leucine enkephalin), a limit of detection (LOD) of 0.045 pmol/μL (corresponding to 67 attomol in a sample volume of ∼15 nL) was obtained. The merit of the CE-MS approach was demonstrated by analysis of bovine serum albumin (BSA) tryptic peptides. A well-resolved separation profile was achieved and comparable sequence coverage was obtained by the CE-MS method (73%) compared to a representative UPLC-MS method (77%). The CE-MS interface was subsequently used to analyse a more complex sample of pharmaceutically relevant human proteins including insulin, tissue factor and α-synuclein. Efficient separation and protein ESI mass spectra of adequate quality could be achieved using only a small amount of sample (30 fmol). In addition, analysis of ubiquitin samples under both native and denatured conditions, indicate that the CE-MS setup can facilitate native MS applications to probe the conformational properties of proteins. Thus, the described CE-MS setup should be useful for a wide range of high-sensitivity applications in protein research.

Xanthate degradation occurring in flotation process waters of a gold concentrator plant

Xanthates enhance the separation of valuable metals from minerals in hydrometallurgical processes. They degrade to structurally related decomposition products under the chemical and physical conditions in flotation processes, which minimizes the extraction recovery of metals. To monitor low quantities of the decomposition products, the flotation process requires sensitive, qualitative, or even quantitative controlling.The goal of the present research was to develop a sensitive separation method for on-line monitoring of the degradation of collector chemicals in the waters of a gold concentrator plant. To that end, an on-line coupling of capillary electrophoresis with a pressure-assisted field-amplified sample injection (PA-FASI) was developed. The purpose of the overall system set-up was to fortify concentrations during sample introduction in order to study xanthates and thiocarbonate directly in flotation waters during the process. The system developed from these methods was reliable and reproducible, enabling detection of below 10mgL−1 quantities for ethyl and isobutyl xanthates as well as for isobutyl thiocarbonate.

Online Coupling of Capillary Electrophoresis with Direct Analysis in Real Time Mass Spectrometry

The online coupling of capillary electrophoresis with ambient direct analysis in real time mass spectrometry (DART-MS) was realized by a coaxial tip interface. The analytes eluted from capillary electrophoresis (CE) were directly ionized by the metastable helium flux produced by DART and transferred into MS for the detection, with which the online separation and simultaneous detection were achieved. The CE-DART-MS can tolerate higher concentrations of detergents and salts than traditional CE-electrospray ionization (ESI)-MS and avoided the difficulties of collecting CE effluent and cleaning the interface, which simplified the experimental procedures and shortened the analysis time. The performance of the technique was successfully verified by capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) using a mixture of 4-aminoantipyrine, zolmitriptan, and quinine. This online technique showed good repeatability with the relative standard deviations (RSDs; n = 5) of 0.56-1.23% for the retention times and 2.01-7.41% for the peak areas. The quantitative analysis of 4-aminoantipyrine was accomplished in the range of 0.01-0.50 mg/mL with the linear correlation coefficient of 0.9995 and limit of detection of 14.7 fmol. Compared with CE-ESI-MS, the ion suppression effects of nonvolatile salts and detergents were efficiently minimized. The signal intensity remained constant when the concentrations reached 100 mM for sodium borate and 30 mM for SDS (in 30 mM sodium borate buffer). In addition, the proposed method was successfully applied to the detection of the endogenous caffeine in Chinese white tea.

Speciation of the oxidation states of plutonium in aqueous solutions by UV/Vis spectroscopy, CE-ICP-MS and CE-RIMS

For the speciation of the plutonium oxidation states in aqueous solutions, the online coupling of capillary electrophoresis (CE) with inductively coupled plasma mass spectrometry (ICP-MS) has been developed. Depending on the radius/electrical charge ratio, the oxidation states III, IV, V, and VI of plutonium are separated by CE, based on the different migration times through the capillary and are detected by ICP-MS. The detection limit is 20 ppb,i.e.109–1010atoms (10-12–10-13g) for one oxidation state with an uncertainty of the reproducibility of the migration times of ≤1% and ≤5% for the peak area. The redox kinetics of the different plutonium oxidation states in the presence of humic substances (humic and fulvic acid) have been studied. A relatively rapid reduction of Pu(VI) (10 to 1000 h) in contact with Gorleben fulvic or Aldrich humic acid could be observed, depending on the pH of the solution. Furthermore, at pH=1, a reduction to Pu(III) and Pu(IV) in a mixture of all four oxidation states in contact with Gorleben fulvic acid after one month has been observed. In order to improve the sensitivity of the CE method, the offline coupling of CE to resonance ionization mass spectrometry (RIMS) has been explored. First applications of this new speciation method are presented.

Analysis of Hop Acids and Their Oxidized Derivatives and Iso-α-acids in Beer by Capillary Electrophoresis−Electrospray Ionization Mass Spectrometry

This study investigates the applicability of on-line coupling of capillary electrophoresis with electrospray ionization tandem mass spectrometry (CZE-ESI-MS) for the separation and characterization of alpha- and beta-acids and oxidized hop acids from crude extracts of different hop varieties. CZE-ESI-MS with negative-ion electrospray ionization proved to be a suitable technique for the determination of these types of natural compounds and their oxidized derivatives. The CZE parameters (pH, concentration, and buffer type) and ESI-MS parameters (nature and flow rate of the sheath liquid, nebulizer pressure, drying gas flow rate, temperature, and compound stability) were optimized. The optimized method provides the potential for a fast qualitative determination of hop acids and their oxidation compounds. The method was also applied to the determination of iso-alpha-acids in beer.

Online coupling of capillary electrophoresis with mass spectrometry for the identification of biomarkers for clinical diagnosis

Proteomic screening of complex biologic samples is of increasing importance in clinical research and diagnosis. In the postgenomic area it is evident that changes of the composition of body fluids, as well as post-translational modifications of proteins and peptides, provide more information than genetic typing. The study of these changes allows the state of health or disease of particular organs, and consequently, the whole organism, to be described. This review describes the application of capillary electrophoresis coupled online to an electrospray ionization time-of-flight mass spectrometer to the analysis of body fluids obtained from patients for the identification of biomarkers for diagnostic purposes.

Capillary electrophoresis on-line coupled with hydride generation-atomic fluorescence spectrometry for speciation analysis of selenium

A new method for speciation analysis of two inorganic selenium species was developed by on-line coupling of capillary electrophoresis (CE) with hydride generation-atomic fluorescence spectrometry (HG-AFS) and on-line conversion of Se(VI) to Se(IV). Baseline separation of Se(VI) and Se(IV) was achieved by CE in a 50 cm x 75 microm inside diameter (ID) fused-silica capillary at -20 kV using a mixture of 15 mmol.L(-1) NaH2PO4 and 0.5 mmol.L(-1) cetyltrimethylammonium bromide (pH 7.5) as electrolyte buffer. Se(VI) was on-line reduced to Se(IV) by mixing the CE effluent with concentrated HCl. The precision (relative standard deviation, RSD, n=7) ranged from 0.7 to 1.3% for migration time, 6.4 to 3.7% for peak height response, and 5.9 to 6.1% for peak area for the two selenium species at the 500 microg.L(-1) (as Se) level. The detection limits were 33 and 25 microg.L(-1) (as Se) for Se(VI) and Se(IV), respectively. The recoveries of the two selenium species in five locally collected water samples ranged from 88 to 114%. The developed method was applied to speciation analysis of inorganic selenium species in spiked natural water samples.

On-line capillary electrophoresis-mass spectrometry using dopant-assisted atmospheric pressure photoionization: Setup and system performance

The on-line coupling of capillary electrophoresis (CE) and mass spectrometry (MS) via atmospheric pressure photoionization (APPI) is demonstrated. To achieve CE-APPI-MS, an adapted coaxial sheath-flow interface was combined with an ion-trap mass spectrometer equipped with an APPI source originally designed for liquid chromatography-MS. Effective photoionization of test compounds was accomplished after optimization of several interface and MS parameters, and of the composition and flow rate of the sheath liquid. Further enhancement of the ionization efficiency could be achieved by adding a dopant, such as acetone or toluene, to the sheath liquid to aid indirect ionization. Acetone significantly increased the ionization of the polar test compounds by proton transfer, while toluene was more useful for the enhanced formation of molecular ions from nonpolar compounds. The effect of several common CE background electrolytes (BGEs) on the APPI-MS response of the analytes was also studied. It appeared that in contrast with electrospray ionization, nonvolatile BGEs do not cause suppression of analyte signals using APPI. Therefore, in CE-APPI-MS, a variety of buffers can be chosen, which obviously is a great advantage during method development. Remarkably, also sodium dodecyl sulfate (SDS) did not affect the photoionization of the test compounds, indicating a strong potential of APPI for the on-line coupling of micellar electrokinetic chromatography (MEKC) and MS.

Online CE-MALDI-TOF MS using a rotating ball interface.

We report on the construction and performance of a rotating ball interface for online coupling of capillary electrophoresis (CE) to matrix-assisted laser desorption ionization (MALDI) mass spectrometry with a time-of-flight (TOF) mass analyzer. The interface is based on a rotating stainless steel ball that transports samples from atmospheric pressure to the high vacuum of the mass spectrometer for desorption and ionization. The sample is deposited directly from a 50-microm-i.d. separation capillary onto the 19-mm ball that is rotating at 0.03 to 0.3 rpm. The sample is mixed online with matrix flowing from a separate 50-microm-i.d. capillary. The sample deposit dries before it is rotated past a polymer gasket and into the laser ionization region. Cleaning of the interface is accomplished using solvent-saturated felt, which cleans the ball surface after it rotates out of the ionization chamber. On-line CE-MALDI is demonstrated, and the performance is evaluated with the analysis of a mixture of three peptides: [Lsy8] vasopressin, substance P, and neurotensin. The rotating ball interface to MALDI-TOF MS demonstrated mass detection limit in the high femtomole range. The interface has negligible memory effect and shows no significant electrophoretic peak broadening when operated under optimized conditions.

Analysis of Chinese medicine preparations by capillary electrophoresis–mass spectrometry

In Chinese medicines, herbs are usually prepared before use by patients. Since the preparation procedures convert the original component into one or more products, study of the procedures is usually complex and involves several compounds. On-line coupling of capillary electrophoresis (CE) to mass spectrometry (MS) allows both the efficient separation of CE and the specific and sensitive detection of MS to be achieved. In this study, CE–MS was applied to the determination of alkaloids in Maqianzi (the seed of Strychnos pierrian) and Wutou (aconite root, Radix aconiti praeparata) during the preparation procedure. With optimal CE–MS conditions, alkaloids in both prepared and unprepared Maqianzi were determined successfully in the total ion current (TIC) mode. However, single ion monitoring (SIM) had to be applied for the separation of aconitum alkaloids and their hydrolysis products. Quantification data indicated that MS detection under SIM mode is more sensitive than UV detection. Based on the CE–MS method developed, the hydrolysis of aconitum alkaloids in water and methanol was also studied.

Determination of peptides and proteins in human urine with capillary electrophoresis–mass spectrometry, a suitable tool for the establishment of new diagnostic markers

The on-line coupling of capillary electrophoresis (CE) with electrospray-time-of-flight mass spectrometry (MS) has been used to obtain patterns of peptides and proteins present in the urine of healthy human individuals. This led to the establishment of a “normal urine polypeptide pattern”, consisting of 247 polypeptides, each of which was found in more than 50% of healthy individuals. Applying CE–MS to the analysis of urine of patients with kidney disease revealed differences in polypeptide pattern. Twenty-seven polypeptides were exclusively found in samples of patients. Another 13, present in controls, were missing. These data indicate that CE–MS can be applied as powerful tool in clinical diagnostics.

Determination of phosphorus in phosphorylated deoxyribonucleotides using capillary electrophoresis and high performance liquid chromatography hyphenated to inductively coupled plasma mass spectrometry with an octopole reaction cell

The determination of phosphorus in a biologically relevant sample matrix such as DNA is described. The analytical methodology used is based on a robust on-line coupling of capillary electrophoresis (CE) or high performance liquid chromatography (HPLC) with inductively coupled plasma mass spectrometry, equipped with an octopole reaction system (ORS-ICP-MS). Polyatomic ions formed in the plasma and the interface region of the ICP-MS that interfere with the determination of P at mass 31 were minimised by the addition of helium to the collision cell. Instrumental detection limits of 125 ng L−1 for 31P, and in the range of 18 to 49 ng L−1 for the other trace elements, were achieved under the conditions described. In order to demonstrate the high separation and detection efficiency of the system, the method developed has been applied to the element specific detection of phosphorus in monophosphorylated deoxynucleotides and to enzymatically digested calf thymus DNA after CE and HPLC separation, respectively. Both hyphenated techniques provide baseline separation of the four deoxynucleotide monophosphates (dNMP), which are present in the DNA chain. With CE-ICP-MS, detection limits for phosphorus down to 53 µg L−1 (2′-deoxyguanosin-5′-monophosphate, dAMP) (corresponding to 0.6 pg P absolute) have been achieved. RSDs of the migration times were about 5%. Using HPLC-ICP-MS, detection limits down to 3 µg L−1 (2′-deoxythymidine-5′-monophosphate, dTMP) (corresponding to 0.03 ng P absolute) were obtained. The migration/elution order of the deoxynucleotides investigated was obtained by comparison with the migration/retention times of commercially available dNMP's. Furthermore it was checked if HPLC-ESI-MS can be operated under the same chromatographic conditions for the confirmation of the elution order and for the characterisation of unknown peaks. Finally, flow injection analysis (FIA) was used for the quantification of the phosphorus content in the dNMP samples. Using FIA, detection limits for phosphorus of 2.5 µg L−1 corresponding to 25 pg phosphorus absolute were achieved.

On-line coupling of capillary electrophoresis to hydride generation atomic fluorescence spectrometry for arsenic speciation analysis.

A novel hyphenated technique, on-line coupling of capillary electrophoresis (CE) to atomic fluorescence spectrometry (AFS), was developed for speciation analysis of four environmentally significant and toxic forms of arsenic: arsenite, arsenate, monomethylarsenic acid, and dimethylarsenic acid. Baseline separation of the four arsenic species was achieved by capillary electrophoresis in a 50 cm x 100 microm i.d. fused-silica capillary at 20 kV and using a 20 mmol L(-1) phosphate buffer (pH 6.5). A hydride generation (HG) technique was employed to convert the arsenic species from the CE effluent into their respective hydrides. The CE-AFS interface was constructed on the basis of a cross design for introducing a sheath flow around the CE capillary and a Pt electrode, which provided an electrical connection for stable electrophoretic separations and allowed on-line volatile hydride formation. A laboratory-made gas-liquid separator was used to isolate the generated volatile species from the reaction mixture solution, and an argon flow was used to transport the volatile hydrides into the atomizer of AFS for on-line detection. The precision (RSD, n = 7) ranged from 2.1 to 3.1% for migration time, from 2.8 to 4.2% for peak area response, and from 2.0 to 4.1% for peak height response for the arsenic species at the 1 mg L(-1) (as As) level. The detection limits were in the range of 9-18 microg L(-1) (as As). The recoveries of the four arsenic species in locally collected water samples and urine sample ranged from 91 to 115%. The developed technique was successfully applied to the speciation of the water-methanol extractable arsenic in a sediment sample.

Glycoscreening by on-line sheathless capillary electrophoresis/electrospray ionization-quadrupole time of flight-tandem mass spectrometry.

An analytical approach based on sheathless on-line coupling of capillary electrophoresis (CE) and electrospray ionization (ESI) quadrupole time-of-flight (QTOF) mass spectrometry (MS) has been developed for providing new insight into the characterization of carbohydrate mixtures. The home-built sheathless CE/

On-Line Capillary Electrophoresis-Electrospray Mass Spectrometry for the Analysis of Pharmaceuticals

The on-line coupling of capillary electrophoresis (CE) with mass spectrometry using the electrospray-ionization mode (ESI-MS) is a promising combination of two analytical techniques. While CE provides high separation efficiency per unit of time, mass spectrometry affords high sensitivity and selectivity as well as molecular structural information. Three different projects are under way in our laboratory and are presented to illustrate the potential of CE-ESI-MS in pharmaceutical analysis. First, the determination of Ecstasy and other amphetamine derivatives in urine samples is reported. Second, the analysis of secondary metabolites, present in plant extracts such as tropane alkaloids, is depicted. Furthermore, the enantioselective analysis of pharmaceutical drugs and metabolites in human plasma is also described.

Capillary electrophoresis/electrospray ionization mass spectrometry (CE/ESI-MS) as a powerful tool for trace element speciation

The on-line coupling of capillary electrophoresis (CE) and electrospray ionization mass spectrometry (ESI-MS) for speciation analysis is entailed with a number of difficulties due to technical and methodical reasons. The optimization of several parameters in order to provide stable operating conditions are illustrated in detail: the positioning of the CE capillary, the CE buffer system, the concentration sensitivity and the detection mode. Additionally, off-line ESI-MS investigations of metal species with different stabilities were carried out in order to assess the influence of the ESI-process on the analyzed samples. It turned out, that metal ions and weak metal complexes may undergo gas-phase ligand replacement and intramolecular charge transfer reactions. Therefore, the structure of species will be altered. Covalent organometallic compounds and strong metal complexes, on the other hand, will not be altered and will be detected as charged molecular ions and/or as charged ion-solvent clusters. Examples of different compounds illustrate this fact.

On-line capillary electropho resis-Mass spectrometry

A brief review of interfaces for on-line coupling of capillary electrophoresis (CE) and mass spectrometry (MS) is presented. To enable the CE-MS system, electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) were introduced as MS interfaces for connecting with CE instruments, while the partial-filling method was developed to suppress the introduction of non-volatile components or additives in CE buffers as well as ionic species such as ionic micelles into the MS interface. Strategy for the coupling of micellar electrokinetic chromatography (MEKC) and MS is also discussed. Examples of CE-MS are briefly shown.

Application of capillary electrophoresis-electrospray ionisation mass spectrometry to arsenic speciation

The on-line coupling of capillary electrophoresis (CE) and electrospray ionisation mass spectrometry (ESI-MS) is a very useful tool for speciation analysis. This hyphenated technique provides elemental (isotopic pattern, if the element is not monoisotopic) as well as structural (molecular mass and/or fragmentation) information on an unknown species. Owing to several properties (high separation efficiency, low ‘flow rates’), CE is best suited as the separation device for this coupling. The geometrical dimensions of both systems require the use of rather long CE capillaries (up to 100 cm), which leads to long total analysis times. The application of pressure along the capillary during or after the CE separation shortens the total analysis time dramatically. The effects of this ‘pressure mobilisation’ on detection limits, peak shape and resolution are discussed in detail. The technique was applied to the speciation of arsenic. A CE method was developed, providing the separation of six arsenic species of interest in a single run {arsenite [As(III)], arsenate [As(v)], methylarsonic acid (MMA), dimethylarsinic acid (DMA) arsenobetaine (AsB) and arsenocholine (AsC)}. The method used an acidic electrolyte system (ammonium acetate-acetic acid) for pH stacking. With the exception of As(III) and MMA, the arsenic species were baseline-separated from each other. Detection limits were calculated as 60-480 µg L –1 for the arsenic species. The only exception was arsenite, As(III), with a detection limit of 50 mg L –1 . The method was applied to standard mixtures and urine samples.