Counter-electroosmotic Flow Research Articles

Capillary Zone Electrophoresis Determination of Five Trace Food Additives in Beverage Samples Using Counterflow Transient Isotachophoresis

Transient isotachophoresis (tITP) is a universal online sample preconcentration technology. It uses ITP to focus electrically charged analytes during the initial stage of capillary electrophoresis (CE) analysis, which can effectively improve the sensitivity of CE. In this work, an approach of tITP under counter-electroosmotic flow was developed for capillary zone electrophoresis (CZE) determination of five trace food additives which include benzoic acid, sorbic acid, sunset yellow, allura red, and amaranth in beverage samples. The parameters that affect the preconcentration effect of the system, such as the type of terminating electrolyte and its injection time, the preconcentration time of ITP, and the sample injection time, have been investigated in detail. The mechanism of counterflow-tITP was explored preliminarily. Under optimal conditions, the sensitivity of five food additives was improved 11.9, 11.5, 13.04, 10.05, and 15.3 times. Their limits of detection (LODs, S/N = 3) reached 0.15, 0.10, 0.38, 0.33, and 0.11 μg/ml, with good repeatability (peak area RSDs ≤ 6.9%) and calibration graph linearity (r ≥ 0.9992). Recoveries of five food additives in spiked beverage samples ranged from 92.0 to 100.8% with RSDs between 1.38 and 2.95%. This method is simple, rapid, and sensitive. It has been successfully applied to the separation and detection of trace food additives in cocktails, orange juice, and carbonated beverages. The feasibility of determination of trace additives in beverages by tITP-CZE under the condition of counterflow has been confirmed.

Effect of the flow profile on separation efficiency in pressure-assisted reversed-polarity capillary zone electrophoresis of anions: Simulation and experimental evaluation.

Capillary electrophoresis connected to electrospray ionization mass spectrometry is a promising combination to analyze complex biological samples. The use of sheathless electrospray ionization interfaces, such as a porous nanoelectrospray capillary emitter, requires the application of forward flow (either by pressure or electroosmosis) to maintain the electrospray process. The analysis of solute molecules with strong negative charges (e.g., aminopyrenetrisulfonate labeled glycans) necessitates a reversed-polarity capillary electrophoresis separation mode, in which case the electroosmotic flow is counter current, thus pressure assistance is necessary. In this study, we compared the effect of forced convection with and without counter electroosmotic flow on the resulting separation efficiency in capillary electrophoresis based on flow profile simulations by computational fluid dynamics technique and by actual experiments. The efficiencies of the detected peaks were calculated from the resulting electropherograms and found approximately 950 000 plates/m for electrophoresis with counter electroosmotic flow, 20000 plates/m with pressure only (such as would be in open tubular liquid chromatography), and 480 000 plates/m for electrophoresis with simultaneous counter electroosmotic flow and forward pressure assistance, which validates the simulation data.

Determination of neutral diols and carboxylic acids formed during glycerol electrooxidation by capillary electrophoresis with dual C4D

Methods for determination of glycerol and its electrooxidation products (neutral diols and carboxylates) by capillary electrophoresis (CE) with dual capacitively coupled contactless conductivity detectors (C4D) are presented. Glycerol, dihydroxyacetone and glyceraldehyde were detected as anionic borate complexes in less than 3min under counter Electroosmotic Flow (EOF) mode (resolution of the critical pair of 1.8). Limits of detection (LODs) of 15, 15 and 10µmolL−1 were obtained for glycerol, dihydroxyacetone and glyceraldehyde, respectively. Two methods of separation were used for the separation of carboxylates. The first one used the same Back Ground Electrolyte (BGE) containing borate, and the second used a BGE (pH 6.1) composed by 2-(N-morpholino)ethanesulfonic acid (MES), L-Histidine and a flow modifier. Better separation and LODs for carboxylates were obtained using Mes/Histidine as BGE. However, along with the non-applicability of this BGE to the determination of neutral diols, observation of the C4D signals at two different points of the capillary (10 and 50cm apart from the injection tip) revealed interaction of the flow modifier with some species (mesoxalate and glyoxylate). The electrooxidation of a glycerol sample in alkaline media on an 8cm2 gold working electrode was evaluated by the developed methods. After 16h of electrolysis, 87% of the glycerol had been oxidized and formate, glycolate, hydroxypyruvate and glycerate were detected as the main products.

Highly efficient sample stacking by enhanced field amplification on a simple paper device.

We present a novel electrokinetic stacking (ES) method based on field amplification on a simple paper device for sample preconcentration. With voltage application, charged probe ions in a solution of lower conductivity stack and form a narrow band at the boundary between the sample and the background electrolyte of higher conductivity. The stacking band appears quickly and stabilizes in a few minutes. With this ES method, three orders of magnitude signal improvement was successfully achieved for both a fluorescein probe and a double-stranded DNA within 300 s. This enhanced stacking efficiency is attributed to a focusing effect due to the balance between electromigration and counter electroosmotic flow. We also applied this ES method to other low-cost fiber substrates such as cloth and thread. Such a simple and highly efficient ES method will find wide applications in the development of sensitive paper-based analytical devices (PADs), especially for low-cost point-of-care testing (POCT).

DISPERSIVE LIQUID-LIQUID MICROEXTRACTION BASED ON SOLIDIFICATION OF FLOATING ORGANIC DROP COMBINED WITH COUNTER-ELECTROOSMOTIC FLOW NORMAL STACKING MODE IN CAPILLARY ELECTROPHORESIS FOR THE DETERMINATION OF BISPHENOL A IN WATER AND URINE SAMPLES

Dispersive liquid–liquid microextraction method based on solidification of floating organic drop (DLLME-SFO) was combined for the first time with counter-electroosmotic flow normal stacking mode (counter-EOF NSM) in capillary electrophoresis (CE) for preconcentration and determination of bisphenol A (BPA) in water and urine samples. Several parameters affecting extraction efficiency, including type and volume of the extraction and disperser solvents, pH, volume of sample and back-extraction solutions, and ionic strength, were systematically studied. In-vial back-extraction of the target analyte from the resulting organic drop into an aqueous phase facilitated the direct application of DLLME-SFO with CE. Under optimum conditions, improvement factors of 1250 (water) and 430 (urine) as compared to conventional capillary zone electrophoresis (CZE) were obtained. Calibration graphs were linear up to 100 µg L−1 with coefficients of determination (R2) ≥ 0.9989 and relative standard deviation (RSD %) ≤ 1.9. Limits of detection (LOD) of 0.8 µg L−1 (water) and 2.5 µg L−1 (urine) were achieved. Because this method required simple and inexpensive devices and very small volumes of nontoxic organic solvents, it is an affordable, efficient, and convenient method for extraction and determination of trace amounts of BPA in water and human urine samples.

Determination of inorganic anions in saliva by electroosmotic flow controlled counterflow isotachophoretic stacking under field-amplified sample injection

Under a strong counter-electroosmotic flow, five salivary inorganic anions, bromide, iodide, nitrite, nitrate and thiocyanate were determined by field-amplified sample injection in combination with isotachophoretic stacking. Separation and concentration conditions were investigated. A terminating electrolyte, 5mM borate, was added in the sample. Under the optimized conditions, Br(-), I(-) and SCN(-) were concentrated online using 150mM HCl-Tris buffer at pH 7.8 in a bare fused capillary, providing more than ten thousand of sensitivity enrichment compared with normal injections. The relative standard deviations (RSDs, n=5) were less than 1% in migration times, 8% in peak areas. Using direct UV detection at 200nm and 226nm, the limits of detection (LODs, S/N=3) were of 0.002-0.01μM. Unfortunately, NO2(-) and NO3(-) could be observed in purified or deionized water. Therefore, a low dilution factor was applied to saliva samples. Due to the matrix effect, samples were injected in a shorter time, and standard addition method was applied to determine all the five inorganic anions in saliva. The RSDs of the migration times and peak areas were in a range of 0.2-0.4% and 3.0-4.0%, respectively. The LODs were 0.2-2.0μM. The salivary levels of the anions obtained were in accord with the reference data. The external standard method can not be adapted to real samples due to biases caused by electrokinetic injection and errors from high dilutions.

Electrokinetic effects on fluid flow and particle transport

The effect of counter-electroosmotic flow on the particle trajectories, the particle equilibrium position, and the critical flux was for the first time evaluated in normal flow filtration using numerical solution of the two-dimensional coupled Navier-Stokes, Nernst-Plank, and Poisson equations for a slit pore having a converging entrance. It was shown that the numerical results for the velocity profiles, ion concentrations, and induced streaming potential were in good agreement with analytical expressions obtained for a simple slit shaped. Numerical simulations for particle transport were performed at both constant pressure and constant filtration velocity in the presence of counter-electroosmosis. A significant shift in the particle trajectory and final equilibrium location were shown at constant pressure due to the reduction in the filtrate flux associated with the counter-electroosmotic flow arising from the induced streaming potential. However, simulations conducted at a constant filtration velocity showed only a very small effect of counter-electroosmosis, with the equilibrium position varying by less than 5% for calculations performed in the presence/absence of counter-electroosmosis. This result stems from a very small distortion in the velocity profile in the region above the pore due to the greater contribution from counter-electroosmosis in the region immediately adjacent to the pore wall. This paper will provide a useful framework to evaluate particle transport in the presence of electrokinetic phenomena.

In-line preconcentration of oxidized and reduced glutathione in capillary zone electrophoresis using transient isotachophoresis under strong counter-electroosmotic flow

Transient isotachophoresis (tITP) can improve the sensitivity of capillary electrophoresis (CE). In general, it was carried out under the condition of suppressed electroosmotic flow (EOF). However, some special conditions, such as extreme low pH background electrolyte and coating were needed to achieve the requirements of suppressed EOF. In this work, an approach of tITP under the strong counter-EOF in open system (counter-EOF-tITP) is presented for the rapid and sensitive preconcentrating the reduced glutathione (GSH) and the oxidized glutathione (GSSG) without modifying the capillary and the commercial CE instrument. The parameters of the experimental system, such as the concentration of leading electrolyte, the injected amount of terminating electrolyte and the injected pressure of sample were investigated in detail to understand the mechanism of counter-EOF-tITP. The sensitivity enhancement factors were of 320 for GSH and 280 for GSSG. In addition, the detection limit of 23.4 and 18.0 μg L −1 for GSH and GSSG was achieved, respectively. The method's applicability was demonstrated by determining GSH and GSSG in tomato and human serum.

Co- and Counter-Electroosmotic Flow Capillary Electrophoresis of Anionic Phenols Using UV and MS Detection

Co- and counter-electroosmotic flow (EOF) capillary electrophoresis (CE) of anionic phenols, using UV and mass spectrometry (MS) detection were investigated. The co-EOF CE-UV detection was performed by using hexadimethrine bromide (HDB) as an EOF modifier, which provided the separation in 6.2 min. The counter-EOF CE-MS was obtained without HDB and coupling was achieved by sheath liquid interface. The CE-UV provided the better linearity (r 2 > 0.99) than the CE-MS (r 2 > 0.81), however, both methods gave comparable detection limits (5 µg/mL). CE-UV is efficient for the routine quantitative analysis, whereas CE-MS is useful for identification of the phenols.

High-sensitivity microchip electrophoresis determination of inorganic anions and oxalate in atmospheric aerosols with adjustable selectivity and conductivity detection

A sensitive and selective separation of common anionic constituents of atmospheric aerosols, sulfate, nitrate, chloride, and oxalate, is presented using microchip electrophoresis. The optimized separation is achieved in under 1min and at low background electrolyte ionic strength (2.9mM) by combining a metal-binding electrolyte anion (17mM picolinic acid), a sulfate-binding electrolyte cation (19mM HEPBS), a zwitterionic surfactant with affinity towards weakly solvated anions (19mM N-tetradecyl,N,N-dimethyl-3-ammonio-1-propansulfonate), and operation in counter-electroosmotic flow (EOF) mode. The separation is performed at pH 4.7, permitting pH manipulation of oxalate's mobility. The majority of low-concentration organic acids are not observed at these conditions, allowing for rapid subsequent injections without the presence of interfering peaks. Because the mobilities of sulfate, nitrate, and oxalate are independently controlled, other minor constituents of aerosols can be analyzed, including nitrite, fluoride, and formate if desired using similar separation conditions. Contact conductivity detection is utilized, and the limit of detection for oxalate (S/N=3) is 180nM without stacking. Sensitivity can be increased with field-amplified sample stacking by injecting from dilute electrolyte with a detection limit of 19nM achieved. The high-sensitivity, counter-EOF operation, and short analysis time make this separation well-suited to continuous online monitoring of aerosol composition.

Trace determination of arsenic species by capillary electrophoresis with direct UV detection using sensitivity enhancement by counter- or co-electroosmotic flow stacking and a high-sensitivity cell.

Stacking techniques used independently and also with a high-sensitivity cell (HSC) were employed to optimise sensitivity and detection limits in the direct photometric detection of the following eight arsenic species by capillary zone electrophoresis (CZE): arsenite, arsenate, monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), 4-hydroxy-3-nitrophenylarsonic acid (Roxarsone), p-aminophenylarsonic acid (p-ASA), 4-nitrophenylarsonic acid (4-NPAA), and phenylarsonic acid (PAA) (target analytes). The stacking mechanisms, optimised stacking and separation conditions, and concentration sensitivity enhancement factors were discussed and compared for (i) normal stacking mode (NSM, sometimes also referred to as field-amplified stacking) in an uncoated fused-silica capillary in the counter-electroosmotic flow (EOF) mode, (ii) large-volume sample stacking (LVSS) with polarity switching, and (iii) the less often applied stacking method of co-EOF NSM stacking with EOF reversal using a poly(diallydimethylammonium chloride) (PDDAC)-coated capillary. The optimal injection volumes were 7.4, 60 and 17.2% of the total capillary volume, for the above three methods, respectively. LVSS with polarity switching gave the lowest limit of detection (LOD). The use of the HSC further reduced the LOD of each target analytes by a factor of 5-8 times. By combining LVSS and HSC, LODs of the target analytes could be reduced by a factor of 218-311, to 5.61, 9.15, 11.1, and 17.1 microg/L for As(III), DMA, MMA, and As(V), respectively. The method was demonstrated to be applicable to the determination of the target analytes in tap water and lake water, with recoveries in the range of 89.4-103.3%.

Electrophoretic chiral separation of pharmaceutical compounds with multiple stereogenic centers in charged cyclodextrin media

This paper reports the use of sulfated β-cyclodextrins (S-β-CDs) (degree of substitution (DS) = 7-11 and 15), hepta-6-sulfato-β-CD (HS-β-CD), heptakis (2,3-dimethyl-6-sulfato)-β-CD (HDMS-β-CD), and heptakis (2,3-diacetyl-6-sulfato)-β-CD (HDAS-β-CD) in conjunction with capillary zone electrophoresis (CZE) to separate enantiomers of several commercially available pharmaceutical compounds with multiple stereogenic centers. Compounds studied include eucatropine, fenoterol, nadolol, nafronyl, nylidrin, and pentapiperide. S-β-CD with a relatively high degree of substitution is shown to be effective in separating several of these compounds due to the high selectivity. Resolution of four isomers was achieved for several of the test compounds under counter-electroosmotic flow (EOF) conditions in less than ten minutes. Data illustrating the effects of CD concentration and pH are presented. It is also shown that the migration order of isomers can be manipulated by changing either the CD concentration or buffer pH.

High-speed chiral separations on microchip electrophoresis devices.

High-speed electrophoretic chiral separations have been successfully performed in a microfabricated device by employing cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC). Utilizing short separation channels and relatively high field strengths in combination with small volume-defined injection plugs, and operating in counter-electroosmotic flow conditions, fast and efficient separations of fluorescein insothiocyanate (FITC)-labeled amino acid enantiomers were obtained. Analysis time ranged from 75 s for the most basic amino acids to 160 s for the most acidic ones with associated efficiencies from 7000 up to 28 000 effective plates (100 000 to 395 000 plates/m). Buffer parameters were varied in order to study the effect on chiral resolution. A buffer system consisting of 100 mM borate (pH 9.4), 30 mM of SDS, and 10 mM gamma-CD as chiral selector provided adequate resolution of the majority of FITC-amino acid enantiomers tested.

Development of a Capillary Electrophoresis Assay Based on Free Sulfate Determination for the Direct Monitoring of Sulfoesterase Activity

A capillary electrophoresis assay of sulfoesterase activity was developed that overcomes the main drawbacks encountered with the usual methods for sulfate determination in complex biological medium. Conditions are described allowing direct measurement of inorganic sulfate that is enzymatically produced in the reaction mixture. The main features of this method are electrokinetic sample introduction, which allows selective extraction of sulfate from the matrix into the separation capillary, counter-electroosmotic flow migration mode, indirect absorbance detection and use of an internal standard for quantitative performances. Likewise, perfect linearity was obtained for concentrations of sulfate up to 40 ppm. The limits of detection and quantification were 0.2 and 0.6 ppm, respectively. The run-to-run and day-to-day precision are 1 and 4.5%, respectively, for sulfate concentrations varying from 35 ppm down to 1 ppm. The accuracy was established for the synthetic p-nitrocatechol sulfate substrate by comparison with the classical spectrophotometric assay. The method was applied to the kinetic monitoring of the activity of a sulfoesterase extracted from the marine mollusc Pecten maximus on fucoidan, a bioactive sulfated fucose-based polysaccharide derived from brown algae. For the first time, a sulfoesterase activity was shown to be effective on such sulfated polysaccharides.

Capillary electrophoresis chiral separations of basic compounds using cationic cyclodextrin.

Chiral separations of basic enantiomers were carried out by using a cationic cyclodextrin (CD), quaternary ammonium beta-cyclodextrin (QA-beta-CD), under counter-electroosmotic flow (counter-EOF) conditions. The special characteristics of using a cationic CD to separate cationic enantiomers is that the EOF can be reversed and the analyte-CD complexation is reduced. This is especially useful for chiral separation of cationic compounds, which strongly bind with neutral and anionic CDs (such as tricyclic amine compounds). The reduction in the binding constants between the CD and the cationic enantiomers makes it easier to control the optimum CD concentration. The application of the cationic CD also eliminated the peak tailing problem caused by electrodispersion. The effect of pH and the concentration of QA-betaCD on chiral separation has been studied. At pH 3.02, no separation for any of the enantiomeric amines was observed. At pH 8.20, chiral separation of some tricyclic compounds was achieved at very high resolution due to the counter-EOF setup. At pH 11.6, most enantiomers were neutral and chiral separation of some bicyclic compounds can be obtained.

Determination of free fatty acids by capillary zone electrophoresis

Capillary zone electrophoresis was investigated for the separation of free fatty acids as an alternative to well established techniques such as GC or HPLC. The analysis was performed with indirect UV detection in a counterelectroosmotic flow mode using a diethylbarbiturate carrier electrolyte at a pH between 10 and 11 in a mixed aqueous-organic solvent. Separation of saturated and unsaturated fatty acids could be achieved after bromination of the double bonds. Problems with wall adsorption of fatty acids could be overcome by increasing the temperature and using a high concentration of a zwitterionic reagent to inactivate the silica surface. Increased sensitivity could be achieved after preconcentration on alumina. The method was applied to the determination of free fatty acids in dairy products. The advantages compared to traditional methods include short analysis times and simple preparation steps.

Pore size distribution effects on electrokinetic phenomena in semipermeable membranes

Electrokinetic phenomena occurring in charged ultrafiltration membranes can significantly influence the transport characteristics of these membranes. Theoretical calculations have been performed to evaluate the effects of different log-normal pore size distributions on the solvent flow rate, the induced streaming potential, and the membrane zeta potential. The solvent flow rate increased with an increase in the breadth of the pore size distribution, with this effect being much more pronounced for pressure-driven flow than for electrically-driven flow. The streaming potential induced by the convective fluid flow significantly alters the flow profiles through the membrane due to the very different dependence of the pressure-driven and counter-electroosmotic flow on the pore radius. This causes a relatively large negative (reverse) flow to develop through the smallest pores in the distribution. The membrane zeta potential was also a function of the pore size distribution, even for membranes having the same hydraulic permeability and surface charge density. These results provide important insights into the effects of different pore size distributions on membrane transport and on the proper interpretation of these electrokinetic phenomena.