Electrosmotic Flow Research Articles

Enantiomers separation by capillary electrochromatography using polysaccharide-based stationary phases.

The separation of chiral compounds is an interesting and important topic of research because these compounds are involved in some biological processes, fundamentally in human health. Among the various application fields where enantiomers are remarkable, drug analysis has to be considered. Most of the drugs contain enantiomers and very often one of the two isomers could be pharmacologically more active or even dangerous. Therefore, the separation of these compounds is very important. Among the different analytical techniques usually employed, capillary electrochromatography has demonstrated great capability in enantiomers resolution. The great potential of this electromigration technique stands mainly in its high efficiency due to the use of an electrosmotic flow (flat flow profile) and on the high selectivity because of the use of a stationary phase. Chiral separation can be obtained utilizing several chiral stationary phases including a polysaccharide derivative. The aim of this review paper is to summarize the main features of capillary electrochromatography and polysaccharide derivatives of chiral stationary phase. It also report examples of practical applications utilizing this approach.

A simple analytical method for determining inorganic anions and formate in virgin olive oils by capillary electrophoresis with capacitively coupled contactless conductivity detection

This paper describes a novel and simple method for determining chloride, nitrate, sulfate, fluoride, and formate in virgin olive oil (VOO) by capillary electrophoresis with capacitively coupled contactless conductivity detection. The anions were extracted from VOO by a simple ultrasound-assisted aqueous liquid–liquid extraction procedure. The anions were separated in less than 3 min with good peak resolutions using a background electrolyte composed of 15 mmol L−1 histidine solution at pH 4.0, adjusted with lactic acid, and 0.6 mmol L−1 cetyltrimethylammonium hydroxide as electrosmotic flow (EOF) modifier. Tartrate was used as internal standard (I.S.). The proposed method showed good linearity and had coefficients of determination (R2) ranging from 0.9984 to 0.9999. The limits of quantification (LOQ) were between 0.01 and 0.7 mg kg−1. Recovery tests were used to evaluate the efficiency of the extraction procedure; the recovery percentages ranged from 70.2 to 118%, with a relative standard deviation (RSD) varying from 0.7 to 19.4%. The proposed method was applied to the determination of the anions in five different commercial samples of VOO.

Repeatability of Octadecyl Methacrylate-Based Monolithic Columns for Capillary Electrochromatography

Octadecyl methacrylate-based monolithic capillary columns were prepared using octadecyl methacrylate as the monomer, ethylene dimethacrylate as the cross-linking agent, 2-acryloylamido-2-methylpropanesulfonic acid as the ionizable monomer, responsible for a negatively charged stationary phase surface and able to support a cathodic electrosmotic flow, and amyl alcohol and 1,4-butanediol as porogenic solvents. The repeatability of the morphological aspect of the monolithic material was evaluated by scanning electron microscopy (SEM). The uniformity along the same monolithic bed was also investigated by SEM. Replicate determinations of surface area and pore volume were also performed and compared. The repeatability of the obtained electrochromatographic efficiencies for separations of alkylbenzenes using the stationary phases prepared was also evaluated. A column that had a plate height of 31 μm was tested for separation of polycyclic aromatic hydrocarbons. Even though minimal possible variation of experimental parameters was maintained, it was not possible to reproduce the stationary phases of the monolithic columns, since the relative standard deviation for the efficiencies obtained on columns prepared from the same polymerization mixture composition was 62%. The lack of repeatability for the efficiency is thought to be a result of limited homogeneity of the monolithic globular structures along the length of the majority of the prepared columns. The poor precision of preparation was justified by the copolymerization reaction that occurs in the presence of 2, 2′-azobisisobutyronitrile as the initiator agent, whose decomposition starts at the moment that this is added to the reaction mixture.

Physical and electrophoretic characterization of octadecyl methacrylate-based monolithic columns for use in capillary electrochromatography

Organic monolithic columns with high efficiency were obtained for use in capillary electrochromatography (CEC) from a mixture of monomers (constant concentrations) with different proportions of porogenic solvents (1,4-butanediol with isoamyl alcohol, amyl alcohol or cyclohexanol in the presence or absence of water). The stationary phases were prepared from the precursor monomer octadecyl methacrylate (ODMA), the cross-linking agent ethylene dimethacrylate (EDMA) and the ionizable monomer 2-acryloylamido-2-methylpropanesulfonic acid (AMPS), the latter being necessary to make the stationary phase negatively charged, assuring electrosmotic flow (EOF) during analysis. The monolithic columns synthesized were physically characterized by porosimetry and scanning electron microscopy (SEM), and electrochromatographically characterized by calculation of chromatographic parameters. The most efficient column was prepared using the porogenic solvent amyl alcohol:1,4-butanediol in a 65:35 (v/v) ratio and showed a plate height of 38 µm. These columns were applied for the separation of alkyl parabens and polycyclic aromatic hydrocarbons (PAH).

Physical and Electrophoretic Characterization of Octadecyl Methacrylate-Based Monolithic Columns for Use in Capillary Electrochromatography

Organic monolithic columns with high efficiency were obtained for use in capillary electrochromatography (CEC) from a mixture of monomers (constant concentrations) with different proportions of porogenic solvents (1,4-butanediol with isoamyl alcohol, amyl alcohol or cyclohexanol in the presence or absence of water). The stationary phases were prepared from the precursor monomer octadecyl methacrylate (ODMA), the cross-linking agent ethylene dimethacrylate (EDMA) and the ionizable monomer 2-acryloylamido-2-methylpropanesulfonic acid (AMPS), the latter being necessary to make the stationary phase negatively charged, assuring electrosmotic flow (EOF) during analysis. The monolithic columns synthesized were physically characterized by porosimetry and scanning electron microscopy (SEM), and electrochromatographically characterized by calculation of chromatographic parameters. The most efficient column was prepared using the porogenic solvent amyl alcohol:1,4-butanediol in a 65:35 (v/v) ratio and showed a plate height of 38 µm. These columns were applied for the separation of alkyl parabens and polycyclic aromatic hydrocarbons (PAH).

Electrically assisted liquid hydrocarbon transport in porous media

Electrically assisted mass transport of hydrocarbon compounds in natural porous media can be used to recover residual oil from clay rich rock formations, for which other means of recovery is infeasible. Momentum transfer from electrosmotic flow of water achieved on the water wet pore surfaces, and increased permeability due to enlargement of capillaries by colloidal dislodging are thought to be some of the direct mechanisms responsible for oil transport under an applied electric field. In this paper, the efficacy of electrically assisted hydrocarbon recovery in laboratory and field cores of hydrocarbon bearing formations is investigated. In all the laboratory experiments, the mass transport was induced under a constant applied potential gradient, without supplemental water flooding under pressure. It was found that oil recovery was maximized for surrogate cores containing 10–15% clay by mass. At higher clay contents the recovery was limited by low charge efficiency, while at lower clay contents the recovery appeared to be influenced by colloidal transport. The electroosmotic pumping effect was investigated and found to potentially contribute to the oil transport to the cathode reservoir. Low permeability sandstone cores showed increased permeability under applied electric field. Larger oil recovery was observed in the cathode reservoir than in the anode reservoir.

Optimization of an Alternative Methodology for Simultaneous Analysis of Nitrite and Nitrate in Water from Urban Stream by Capillary Electrophoresis under Direct UV Detection

An alternative methodology for simultaneous determination of nitrite and nitrate by capillary zone electrophoresis using direct detection UV at 210 nm under reverse electrosmotic flow is proposed. The choice of the electrolyte composition has taken account: the mobility of the anion buffer and of the solutes; the low absorbance of the buffer in 210 nm; high base line stability and analysis time. The electrolyte optimized has consisted of 100 mmol.L–1 TRIS/HCl buffer and 0.15 mmol.L–1 CTAB at pH 8.2. The proposed method was applied successfully in the analysis of nitrite and nitrate in samples from urban stream in the absence of usual sample pretreatment.

Enantiomeric Separation by Capillary Electrochromatography on a Sulfated Poly β-Cyclodextrin Modified Silica-based Monolith

A sulfated poly β-cyclodextrin (SPCD) modified silica-based monolithic column was prepared for enantiomeric separation. First, 2-hydroxy-3-allyloxy-propyl-β-cyclodextrin (allyl-β-CD) was bonded onto a bifunctional reagent 3-(methacryloxy)propyltriethoxysilane (γ-MAPS) modified silica-based monolith through radical polymerization; the column was then sulfated with chlorosulfonic acid. The SPCD chiral stationary phase resolved the boring problem associated with desalting when sulfated CDs were synthesized to act as chiral additives. The inorganic salt in the column introduced during the sulfating process could be easily removed by washing the column with water for some time. Chiral compounds investigated were successfully resolved into their enantiomers on the SPCD modified monolith in the capillary electrochromatography (CEC) mode. Due to the existence of the -SO(3)H group, electrosmotic flow (EOF) was obviously increased, and all of the separations could be carried out in 20 min with only a minor loss in the column efficiency and resolution.

Analysis of Broad-Range DNA Fragments with Yttrium Oxide or Ytterbium Oxide Nanoparticle/Polymer Sieving Matrix Using High-Performance Capillary Electrophoresis

plates/m) and the baseline resolution for various DNA standards (100 bp, 500 bp, and 1 kbp DNA ladder) were obtained in 10 min with these matrices. The effects of concentrations of both polyethylene oxide (PEO) and nanoparticles were investigated and the highest performance was obtained at 0.02% PEO with 0.02% YNP or YbNP. Similar sieving power for both YNP and YbNP matrices was observed probably due to the similar sizes of nanoparticles, resulting in the formation of comparable sieving networks for DNA separation. For the reduction of electrosmotic flow, either dynamic or permanent coating of the capillary inner wall was compared and it turned out that PEO was superior to polyvinylpyrrolidone (PVP) or polyacrylamide (PAA) for better separation efficiency.

Surfactant enhanced electrokinetic remediation of DDT from soils

Electrokinetic remediation has been investigated extensively as one of the noble technologies in remediation of metal contaminated soils. However, its applications in remediation of organic contaminants have been limited due to low solubilities of organics in water. In addition, most organic contaminants are non-ionic and therefore, they are not mobile under electrical field. The use of surfactants may increase the remediation efficiency by increasing the solubility of organics. Significant fraction of organics associated with soil, can be transferred to micellar phase, which then can be transported toward either cathode or anode, depending on the ionic group of surfactants. In this study, the removal of hydrophobic organic contaminants from a soil using electrokinetic method was investigated in the presence of surfactants. A nonionic surfactant, Tween 80, and an anionic surfactant, SDBS, were used in the experiments. DDT was chosen as the model organic contaminant. Phase distribution studies and column experiments were conducted. It was found that both Tween 80 and SDBS had similar solubilization potentials for DDT. It was also shown that the aqueous DDT mass could reach from 0.01 to 13% of the total mass in the presence of 7500 mg/L of SDBS. No significant movement of DDT was observed when Tween 80 was used in the column experiments. This was attributed to low rates of electroosmotic flows and strong interaction of Tween 80 with the soil. The amount of surfactant was not enough to mobilize DDT significantly in the column studies. On the other hand, electrokinetic transport with SDBS yielded much better results. DDT transport toward the anode within the negatively charged micelles overcame the opposite electrosmotic flow. This was attributed to the lower degree of interaction between the soil and SDBS, and the electrokinetic transport of negatively charged micelles.

The mechanisms of arsenic removal from soil by electrokinetic process coupled with iron permeable reaction barrier

An innovative remediation system of electrokinetic process coupled with permeable reaction barrier (PRB) was proposed for arsenic removal in soil matrix. Batch tests with PRB media of Fe(0) and FeOOH under potential gradient of 2 V cm(-1) for 5d duration were conducted to evaluate the removal mechanisms of arsenic. Arsenic enhancement of 1.6-2.2 times was achieved when a PRB system was installed in an electrokinetic system. A best performance was found in system with FeOOH layer located in the middle of elctrokinetic cell. This was largely because of higher surface area of FeOOH and the moving of HAsO(4)(2-) to the anode side by electromigration effect was inhibited by the electroosmosis flow. The surface characteristics of PRB media, which were qualified by SEM coupled with energy dispersive spectroscopy (EDS), were clearly confirmed that arsenic was found on the passive layer surface. Results indicated that the removal of As in EK/PRB systems was much more contributed by surface adsorption/precipitation on PRB media than by EK process. Furthermore among the electrical removal mechanisms, electromigration was predominant than electrosmotic flow. Surface adsorption and precipitation were respectively the principal removal mechanism under acid environment, e.g. near anode side, and under basic environment, e.g. near cathode side. The results reported in the present work will be beneficial to optimizing design of batch EK/PRB system and enlarging to the field scale system.

Determination of antioxidants in edible grain derivatives from the Canary Islands by capillary electrophoresis

A preliminary study of the antioxidant content of hand-produced grain derivatives from the Canary Islands, named gofio, is developed by means of a capillary electrophoresis (CE) method. Protocatechuic acid, salicylic acid, p-hydroxybenzoic acid, vanillic acid, syringic acid, p-coumaric acid, ferulic acid and sinapic acid were extracted from corn, wheat, barley and pea gofio samples by sonication with methanol and were successfully separated by CE using a polycation, hexadimetrine bromide (HDB), as electrosmotic flow modifier. The separation was achieved using a running buffer consisting of 125 mM boric acid, 49 mM disodium hydrogen phosphate, 0.002% (w/v) HDB and 2.5 mM α-cyclodextrin at pH 7.5. The developed procedure allowed the simultaneous determination of the selected antioxidants in less than 3.5 min. The electrophoretic profile obtained for each kind of flour (corn, wheat, barley and pea) allowed differentiation of the type of gofio. Among the analyzed samples, corn gofio showed the highest antioxidant content. Furthermore, highly roasted corn or wheat gofio samples were also analyzed and their antioxidant contents were lower than those of less roasted samples.

Optimization of capillary coating by hydroxyethyl methacrylate for capillary zone electrophoresis of proteins

This work describes further improvements of coating fused silica capillaries with 2-hydroxyethyl methacrylate (HEMA) by atom transfer radical polymerization (ATRP). First, endcapping with a sterically less bulky silanyl reagent reduces the electrosmotic flow (EOF) by 25% in addition to the 40% EOF reduction caused by HEMA coating compared to a bare fused silica capillary. An additional hydrolysis step was introduced into the preparation of HEMA coated capillaries and leads to better reproducible migration times. The influence of the solvent during ATRP and the resulting polymer coating was investigated by replacement of DMF with water or water–methanol mixtures. The quality of the optimized coating was characterized by protein separations at pH 3. HEMA coated capillaries reveal up to 746 000 plates. The polyvinyl alcohol (PVA) coated capillary provides only half of this efficiency. A long-term test at pH 9 shows good stability of the HEMA coated capillaries in basic medium. Also the numbers of plates in this medium was about 30% higher than for separations with the PVA capillary. In addition, the phosphate buffer was replaced by a volatile ammonium acetate buffer for later use with mass spectrometry (MS).

Capillary electrophoresis of proteins in dextran-coated columns.

A simple coating technique by using uncross-linked dextran has been developed for fused-silica capillaries to be used in capillary electrophoresis of basic proteins. The capillaries were first silanized with a heterobifunctional silane (gamma-aminopropyltriethoxylsilane), which served as a coupling agent between the capillary inner wall and the polysaccharide coating. Dextran of high molecular mass (about 70 kDa) was activated with 1,1'-carbonyldiimidazole. Then the activated dextran was coupled to the primary amino groups that were anchored onto the inner wall of the silanized capillaries. The residual reactive groups on the dextran were further substituted by neutral functions in a coupling reaction with excess ethanolamine. By using dimethyl sulfoxide (DMSO) rather than aqueous buffer as the reaction medium, the extent of substitution was improved by minimizing the residual reactive groups at the surface. Since they are ionogenic, the electrosmotic flow in the system is relatively low. The chemically bound dextran coating showed good reproducibility and stability. In electrophoretic experiments basic proteins were separated with high efficiency by use of the dextran-coated fused-silica capillary columns. The main advantage of the method described here is that both polysaccharide activation and amine-coupling reactions were carried out under mild conditions at room temperature without catalysts. For this reason, the method is recommended to coat the inner wall of microfluidic separation channels which would not tolerate a harsh treatment.

Biopolymer-coated fused silica capillaries for high magnitude cathodic or anodic electro-osmotic flows in capillary electrophoresis

The manipulation of electroosmotic flow in capillary electrophoresis was achieved by coating the inner wall of a fused silica capillary with the biopolymers α-chymotrypsinogen A and dextran sulfate. Simple coating procedures were based on flushing the fused silica capillary with α-chymotrypsinogen A solution to obtain a α-chymotrypsinogen A coating, or to additionally coat with dextran sulfate solution to obtain a α-chymotrypsinogen A-dextran sulfate coating. The biopolymers α-chymotrypsinogen A coated capillary exhibited strong reversed (anodic) electrosmotic flow values as high as −81.7×10−9 m2V−1s−1 at pH 2.0. The α-chymotrypsinogen A-dextran sulfate coated capillary exhibited a cathodic EOF of 62.2×10−9m2V−1 s−1 which remained virtually unaltered over the pH range 2–9. Both coatings showed high stability, as demonstrated by electroosmotic flow reproducibility of 1.0% and 0.7% RSD (n=50), respectively. The α-chymotrypsinogen A coating was found to be tolerant to 0.1 M HCl, whilst the α-chymotrypsinogen A-dextran sulfate coating was tolerant to 1 M NaOH, CH3OH and CH3CN. The coating-to-coating repeatability for the two coatings, as determined by the RSD of the resultant electroosmotic flow values, were 2.25% and 1.85% (n=4), respectively. Four anions and five cations were used as test substances to examine the separation performance of the α-chymotrypsinogen A and α-chymotrypsinogen A-dextran sulfate coatings and high efficiencies (80,000 to 200,000 theoretical plates) and rapid separations were obtained. The separation of isomers of chloroaniline was carried out using the α-chymotrypsinogen A-dextran sulfate coating and a pH 2.5 electrolyte in about one third of the time needed when using a FS capillary. A α-chymotrypsinogen A coated capillary was used for ultra-rapid separation of nitrate and nitrite at acidic pH using a co-electroosmotic flow mode. The separation was completed in less than 10s with a migration time reproducibility of 0.3% RSD (n=10) and sub-μM detection limits.

Assessment of silica-based reversed-phase materials for the analysis of a range of basic analytes by capillary electrochromatography

A range of C18 bonded silica stationary phases, two specifically designed for capillary electrochromatography (CEC) use, have been evaluated for their performance in the CEC analysis of basic analytes of differing physico-chemical properties. The performance of the phases towards basic analytes will be discussed in terms of various CEC parameters and by Principal Component Analysis (PCA) of five LC chromatographic parameters – hydrophobic and steric selectivity, hydrogen bonding and ion-exchange capacities at pH 7.6 and 2.7. The CEC phases, which the PCA highlighted as possessing high non-acidic silanol activity, were shown to give substantially enhanced EOF at pH 7.8 and did not exhibit the phenomenon of “drying-out” at pH values below 6.1 which was pronounced with hydrophobic LC phases. The Hypersil CEC Basic C18 phase was found to provide superior performance in analysing the basic analytes compared to the Hypersil CEC C18 phase, this was attributed by PCA to its non-acidic base silica. Through the use of experimentally derived electrophoretic mobility values, LC retention factor, and electrosmotic flow values in CEC, it was possible to predict the CEC retention behaviour of ten basic analytes to an accuracy of ± 6%.

Protein analysis with large volume sample stacking with an electrosmotic flow pump: a potential approach for proteomics

Large volume sample stacking using electrosmotic flow pump (LVSEP) in capillary electrophoresis is a useful tool for on-line concentration of low-abundant protein samples for proteomics research. While the stacking of only small anions with high p K a values has been reported at low pH buffers (pH<4), protein samples (trypsin inhibitor and α-lactalbumin) with high p I values (p I>4) were stacked successfully in our system using LVSEP. Initially, the whole column of the SIL, or LPA-coated capillary was filled with protein samples (∼1 μg/ml), then a high voltage was applied without polarity switching with high pH buffer (pH 8.5). An enhancement factor of more than 100 times was achieved compared to the conventional pressure injection method.

Streaming potential in open and packed fused-silica capillaries

The surface properties of novel stationary phases in packed and open tubular columns for capillary electrochromatography (CEC) were examined by measuring the streaming potential in a home made apparatus. The surfaces investigated include materials such as porous styrenic sorbents and octadecyl-silica as well as fused-silica tubing, in both raw and surface modified forms. Functionalization of the surface was carried out, for instance, by reductive amination or organosilane grafting on to capillary inner wall. The dependence of the streaming potential on pH was examined with aqueous solutions in the pH range from 2.5 to 9.0. Electrokinetic properties of 50 μm I.D. fused-silica capillaries have been determined by both streaming potential and electrosmotic flow measurements. Both methods gave similar pH profiles of the ζ-potential and the isoelectric points. This confirms the viability of our approach to evaluate the specific charged groups of the packing which is one of the important factors influencing electrosmotic flow (EOF) velocity and protein adsorption during a chromatographic run. In addition to bare silica capillaries, styrenic monolithic columns with different surface functionalities, which have been extensively used in our laboratory for CEC separation of peptides and proteins, were employed for comparison of two methods. Plots of zeta potential as a function of percent ACN show a complex behavior, indicating that zeta potential cannot be predicted simply from binary mixture solvent properties. It is demonstrated that the evaluation of the zeta potential by the streaming potential method is nondestructive, relatively fast, without untoward effects introduced by Joule heating and yet another means for the characterization of the surfaces under conditions employed in CEC.

Electrophoretic behaviour of oligonucleotides and mono-, di- and triphosphate nucleotides by capillary zone electrophoresis.

A systematic investigation has been made into the mechanisms of the capillary zone electrophoresis (CZE) separation of 12 common nucleotides (mono-, di- and triphosphorylated) and polydeoxythymidylic acid oligonucleotides (pd(T)5-18) using electrophoretic mobility values calculated from migration time data. Relationships between electrophoretic mobility and the physicochemical characteristics of the analytes (charge, dissociation constants, charge-to-mass ratio) and the background electrolyte conditions (buffer strength, percentage organic modifier and buffer pH) were characterised. Nucleotide migration was dominated by the negatively charged phosphate groups. Additionally, there were important contributions to migration behaviour from the ionised amide groups of the nucleobases guanine and uracil at higher buffer pH values or with the presence of methanol in the electrolyte. Calculated electrophoretic mobility values for the nucleotides showed a substantially improved (5-fold) inter-run repeatability compared with migration time data. These studies show the value of representing nucleotide migration data as electrophoretic mobility in CZE for obtaining a more thorough analysis of separation mechanisms and to compensate for variation in migration time data caused by small changes in electrosmotic flow. Oligonucleotides pd(T)5-11 could be adequately resolved from their nearest neighbour, but the limit of single-base separation was pd(T)10 from pd(T)11 under the conditions used. It was calculated that a difference in charge-to-mass ratio of 2.64 x 10(-5) was required for resolution under the CZE conditions used.

Dynamics of capillary electrochromatography: II. Comparison of column efficiency parameters in microscale high-performance liquid chromatography and capillary electrochromatography

In capillary electrochromatography (CEC) the flow of the mobile phase is generated by electrosmotic means in high electric field. This work compares band spreading measured experimentally in several packed capillaries with electrosmotic flow (EOF) and viscous flow under otherwise identical conditions. The data were fitted to the simplified van Deemter equation for the theoretical plate height, H= A+ B/ u+ Cu, in order to evaluate parameters A and C in each mode of flow in the different columns. The ratio of these two parameters obtained with the same column in microscale HPLC (μ-HPLC) and CEC was used to quantify the attenuation of their contribution to band spreading upon changing from viscous flow (in μ-HPLC) to electrosmotic flow (in CEC). The capillary columns used in this study were packed with stationary phases of different pore sizes as well as retentive properties and measurements were carried out under different mobile phase conditions to examine the effects of the retention factor and buffer concentration. In the CEC mode, the value of both column parameters A and C was invariably by a factor of two to four lower than in the μ-HPLC mode. This effect may be attributed to the peculiarities of the EOF flow profile in the interstitial space and to the generation of intraparticle EOF inside the porous particles of the column packing. Thus, band spreading due to flow maldistribution and mass transfer resistances is significantly lower when the mobile phase flow is driven by voltage as in CEC, rather than by pressure as in μ-HPLC.