Electroosmotic Flow In Capillary Electrophoresis Research Articles

Sulfonic Functionalized Polydopamine Coatings with pH-Independent Surface Charge for Optimizing Capillary Electrophoretic Separations.

Enhancing the pH-independence and controlling the magnitude of electroosmotic flow (EOF) are critical for highly efficient and reproducible capillary electrophoresis (CE) separations. Herein, we present a novel capillary modification method utilizing sulfonated periodate-induced polydopamine (SPD) coating to achieve pH-independent and highly reproducible cathodic EOF in CE. The SPD-coated capillaries were obtained through post-sulfonation treatment of periodate-induced PDA (PDA-SP) coatings adhered on the capillary inner surface. The successful immobilization of the SPD coating and the substantial grafting of sulfonic acid groups were confirmed by a series of characterization techniques. The excellent capability of PDA-SP@capillary in masking silanol groups and maintaining a highly robust EOF mobility was verified. Additionally, the parameters of sulfonation affecting the EOF mobilities were thoroughly examined. The obtained optimum SPD-coated column offered the anticipated highly pH-independent and high-strength cathodic EOF, which is essential for enhancing the CE separation performance and improving analysis efficiency. Consequently, the developed SPD-coated capillaries enabled successful high-efficiency separation of aromatic acids and nucleosides and rapid cyclodextrin-based chiral analysis of racemic drugs. Moreover, the SPD-coated columns exhibited a long lifetime and demonstrated good intra-day, inter-day, and column-to-column repeatability.

Effect of background electrolyte anions on markers of electroosmotic flow in capillary electrophoresis

Effect of background electrolyte anions on markers of electroosmotic flow in capillary electrophoresis

Mobilization of electroosmotic flow markers in capillary zone electrophoresis

UV-absorbing neutral substances are commonly used as markers of mean electroosmotic flow in capillary electrophoresis for their zero electrophoretic mobility in an electric field. However, some of these markers can interact with background electrolyte components and migrate at a different velocity than the electroosmotic flow. Thus, we tested 11 markers primarily varying in their degree of methylation and type of central atom in combination with five background electrolyte cations differing in their ionic radii and surface charge density, measuring the relative electrophoretic mobility using thiourea as a reference marker. Our results from this set of experiments showed some general trends in the mobilization of the markers based on the effects of marker structure and type of background electrolyte cation on the relative electrophoretic mobility. As an example, the effects of an inadequate choice of marker on analyte identification were illustrated in the electrophoretic separation of glucosinolates. Therefore, our findings may help electrophoretists appropriately select electroosmotic flow markers for various electrophoretic systems.

Modulation of Electroosmotic Flow in Open Tubular Capillary Electrochromatography by Chitosan-Assisted Titanium Oxide Nanoparticles Liquid Phase Deposition

Electroosmotic flow is one of the most important driving forces in capillary electrophoresis, which can control the migration velocity and direction of the target substance, thus affecting the separation efficiency and reproducibility of capillary electrophoresis. In this study, a facile method was developed for the preparation of TiO2/chitosan open tubular capillary electrochromatography for modulating electroosmotic flow in capillary electrophoresis. The magnitude and direction of electroosmotic flow were modulated by changing a series of coating preparation parameters, such as deposition time of (NH4)2TiF6 and H3BO3, the concentration of chitosan, and the concentration ratio of (NH4)2TiF6 and chitosan. The results showed that when the concentration of (NH4)2TiF6 was below 2 mg/mL, the surface of the capillary was not modified completely by phosphate ion by Lewis acid–base interaction, resulting in an anodic electroosmotic flow. Furthermore, the feasibility and stability of the column were assessed by separation of nucleotides and alkaloid compounds. With the TiO2/chitosan liquid phase deposition columns with the concentration ratio of 3:7 [(NH4)2TiF6:chitosan], alkaloids were baseline separated within 11 min. The baseline separation of the four nucleotides was achieved using the TiO2/chitosan-coated columns with the concentration ratio of 1.8:7 [(NH4)2TiF6:chitosan]. Furthermore, the relative standard deviations of migration times for inter-day, intra-day, and column-to-column repeatability of the TiO2/chitosan-coated columns with the concentration ratio of 3:7 [(NH4)2TiF6:chitosan] were all less than 5.8%. The strategy of liquid phase deposition of TiO2 nanoparticles provides a promising approach for modulation electroosmotic flow.

Modulation of electroosmotic flow in capillary electrophoresis by plant polyphenol-inspired gallic acid/polyethyleneimine coatings: Analysis of small molecules

Plant polyphenols can form functional coatings on various materials through self-polymerization. In this paper, a series of modified capillary columns, which possess diversity of charge characteristics for modulating electroosmotic flow (EOF), were prepared by one-step co-deposition of gallic acid (GA), a plant-derived polyphenol monomer, and branched polyethyleneimine (PEI). The physicochemical properties of the prepared columns were characterized by Fourier transform infrared spectroscopy (FT-IR), UV–Vis spectroscopy and scanning electron microscopy (SEM). The magnitude and direction of EOF of GA/PEI co-deposited columns were modulated by changing a series of coating parameters, such as post-incubation of FeCl3, co-deposition time, and deposited amounts of GA and PEI with different relative molecular mass (PEI-600, PEI-1800, PEI-10000, and PEI-70000). Furthermore, the separation efficiencies of the prepared GA/PEI co-deposited columns were evaluated by separations of small molecules, including organic acids, polar nucleotides, phenols, nucleic acid bases and nucleosides. Results indicated that modulating of EOF plays an important role in enhancing the separation performance and reversing the elution order of the analytes. Finally, the developed method was successfully applied to quantitative analysis of acidic compounds in four real samples. The recoveries were in the range of 73.5%–85.8% for citric acid, benzoic acid, sorbic acid, salicylic acid and ascorbic acid in beverage and fruit samples, 101.6%–104.9% for cinnamic acid, vanillic acid, and ferulic acid in Angelica sinensis sample, while 84.6%–97.8% for guanosine-5′-monophosphate, uridine-5′-monophosphate, cytosine-5′- monophosphate and adenosine-5′-monophosphate in Cordyceps samples. These results indicated that the co-deposition of plant polyphenol-inspired GA/PEI coatings can provide new opportunities for EOF modulation of capillary electrophoresis.

β-cyclodextrin-ionic liquid polymer based dynamically coating for simultaneous determination of tetracyclines by capillary electrophoresis.

Tetracyclines are a group of broad spectrum antibiotics widely used in animal husbandry to prevent and treat diseases. However, the improper use of tetracyclines may result in the presence of their residues in animal tissues or waste. Recently, great attention has been drawn towards the green solvents ionic liquids. Ionic liquids have been employed as a coating material to modify the electroosmotic flow in capillary electrophoresis. In this study, a functionalized ionic liquid, mono-6-deoxy-6-(3-methylimidazolium)-β-cyclodextrin tosylate, was synthesized and used for the simultaneous separation and quantification of tetracyclines by capillary electrophoresis. Good separation efficiency could be achieved due to the multiple functions of β-cyclodextrin derived ionic liquid, including the electrostatic interaction, the hydrogen bonding, and the cavity structure in β-cyclodextrin ionic liquid which can entrap the tetracyclines to form inclusion complex. After optimization, baseline separation achieved in 25 min with the running buffer consisted of 10 mmol/L, pH 7.2 phosphate buffer and 20 mmol/L β-cyclodextrin ionic liquid. The satisfied result demonstrated that the β-cyclodextrin ionic liquid is an ideal background electrolyte modifier in the separation of tetracyclines with high stability and good reproducibility. And it is an effective strategy to design and synthesize specific ILs as additive applied in separation.

A facile and versatile approach for controlling electroosmotic flow in capillary electrophoresis via mussel inspired polydopamine/polyethyleneimine co-deposition

Electroosmotic flow (EOF), which reveals the charge property of capillary inner surface, has an important impact on the separation performance and reproducibility of capillary electrophoresis (CE). In this study, a novel, facile and versatile method to achieve diverse and controllable EOF in CE was reported based on the co-deposition of mussel-inspired polydopamine (PDA) and branched polyethyleneimine (PEI) on the capillary inner surface as the hybrid functional coating. After these PDA/PEI co-deposited columns were reinforced by the post-incubation of FeCl3, various magnitude and direction of EOF in CE could be easily achieved by varying a number of preparation parameters, including the mass ratio of DA/PEI and the molecular weight of PEI (including PEI-600, PEI-1800, PEI-10000 and PEI-70000). The separation effectiveness and stability of the hybrid coated columns were verified by the analysis of aromatic acids and aniline derivatives. The results showed that the controllable and diverse EOF was important in enhancing the separation performance of the analytes. The baseline separation of all the five aromatic acids can be achieved in 7min with high separation efficiency by using the PDA/PEI-600 co-deposited column with the mass ratio of 6:1. On the other hand, with the PDA/PEI-70000 co-deposited column with the mass ratio of 6:1, the aniline compounds were easily baseline separated within 10min. By contrast, using the bare and PDA coated columns, the migration of the aromatic acids was very slow and the baseline separation of the aniline compounds cannot be obtained. Moreover, the co-deposited columns showed long lifetime and good stability. The relative standard deviations for intra-day, inter-day and capillary-to-capillary repeatability of the PDA/PEI-600 co-deposited column with the mass ratio of 6:1, which was reinforced by the post-incubation of FeCl3, were all lower than 5%.

Modulation of electroosmotic flow in capillary electrophoresis using functional polymer coatings

Capillary electrophoresis (CE) is one of the most powerful techniques for the separation of biomolecules. However, the separation efficiency of proteins in CE is often compromised by their tendency to interact with the silanol groups on the surface of the inner capillary and by an uncontrolled electroosmotic flow. Herein, we report on the synthesis of novel hydrophilic polymeric coatings that can modulate the properties of the capillary walls. The novelty of these poly(N,N-dimethylacrylamide)-based copolymers relies on the simultaneous presence of chemically reactive groups (N-acryloyloxysuccinimide and glycidyl methacrylate) and silane groups in the backbone, which results in highly stable films due to the covalent reaction between the polymer and the glass silanols. Although the functional monomers are reactive towards nucleophilic groups in proteins, they can be effectively blocked in the presence of amino modified agents. In addition, after a careful optimization of monomer concentration, it is possible to confer anti-fouling properties to the polymer coatings, and thus allow for highly efficient acidic and alkaline protein separations. Furthermore, the presence of these monomers makes it possible to modulate the electroosmotic flow from negligible to reduced values, depending on the desired application.

Hydrodynamic suppression of the electroosmotic flow in capillary electrophoresis with indirect spectrophotometric detection

A version of capillary electrophoresis with indirect spectrophotometric detection and the hydrodynamic suppression of electroosmotic flow is studied. It is shown that, to improve the reliability of ion identification, one should calculate electrophoretic mobilities of ions or migration times corrected with regard to the electroosmotic flow rate. Correlations between electrophoretic peak areas of ions and their electrophoretic mobilities are derived. In the studied version of capillary electrophoresis, the accuracy of measuring anion concentrations can be improved using the internal standard method.

Using of S-( −) -2-hydroxymethyl-1,1-dimethylpyrrolidinium tetrafluoroborate as additive to the background electrolyte in capillary electrophoresis

We synthesised and used new type of quaternary ammonium salt [ S-(−)-2-hydroxymethyl-1,1-dimethylpyrrolidinium tetrafluoroborate] as effective additive to acidic background electrolytes. We used this quaternary ammonium salt as effective agent for capillary zone electrophoresis separation of model mixture of five tricyclic antidepressants (amitriptyline, nortriptyline, imipramine, desipramine and clomipramine) as model analytes. We observed that addition of S-(−)-2-hydroxymethyl-1,1-dimethylpyrrolidinium tetrafluoroborate ([HMDP] + [BF 4] −) to acidic background electrolytes leads to suppression of magnitude of electroosmotic flow (EOF) and gradually change the direction of the EOF. Baseline separation of five TAs was achieved by using of 91.1 mmol L −1 (20 g L −1) of [HMDP] + [BF 4] − in 25 mmol L −1 sodium phosphate pH 2.5, where electroosmotic mobility was −11.3 × 10 −9 m 2 V −1 s −1. We achieved baseline separation of five TAs with using of [HMDP] + [BF 4] − as water solution too. We observed that [HMDP] + [BF 4] − can be used as buffer additive, which offers relatively smaller anodic electroosmotic flow instead of cationic surfactants that are mostly used for genarating of anodic electroosmotic flow in capillary electrophoresis.

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.

Moderation of the electroosmotic flow in capillary electrophoresis by chemical modification of the capillary surface with tentacle-like oligourethanes

The surface chemistry of the inner wall of fused-silica capillaries is one important means to control the magnitude as well as the direction of the electroosmotic flow and the adsorption activity. A method was developed to change the surface characteristics of fused-silica capillaries by binding tentacle-like oligourethane groups onto the inner surface. The electroosmotic flow at a buffer pH of 6–9 was reduced by 15 to 40% compared to that in a bare fused-silica tubing, dependent on the type of coating. Sample adsorption is diminished at the same time resulting in a separation of proteins with higher resolution and good migration time precision. At a pH below 4.5 the electroosmotic flow is reversed into the anodic direction, which offers further possibilities for the separation of positively charged analytes as demonstrated for the separation of aromatic and biogenic amines.

Characteristics of electroosmotic flow in capillary electrophoresis in water and in organic solvents without added ionic species

Although proper buffering is essential for repeatable separations in capillary electrophoresis, electroosmotic flow can be developed in many solvents without the addition of buffer components. In this study we detected electroosmotic flow in fused silica capillary in the following solvents without the addition of ionic species: water, deuterium oxide, acetonitrile, acetone, 2-butanone, formamide, N-methylformamide, N,N-dimethylformamide, methanol, ethanol, 1-propanol, dimethyl sulfoxide, ethyl acetate, tetrahydrofuran, and morpholine. The electroosmotic flow was in the range of 3.4×10−9–1.8×10−m2/V s. About 100 times weaker electroosmosis was detected in glacial acetic acid. We also detected electroosmotic flow in N-methylacetamide, which is solid at room temperature (melting range 28–30°C). No electroosmotic flow was found in inert solvents such as n-hexane and chloroform. The zeta potential of the fused silica capillary was calculated for the solvents where electroosmotic flow was found. The zeta potential of the capillary wall was also studied in water–methanol mixtures, where it has a maximum at about 40% methanol concentration. ©1999 John Wiley & Sons, Inc. J Micro Sep 11: 199–208, 1999

Separation of chemical warfare agent degradation products by the reversal of electroosmotic flow in capillary electrophoresis.

We report the development of analyses for nerve agent degradation products or related species by the reversal of electroosmotic flow in capillary electrophoresis (CE). The developed methods were used in this laboratory for analysis of samples in the second and third official proficiency tests (International Round-Robins) for the Provisional Technical Secretariat/Preparatory Commission for the Organization for the Prohibition of Chemical Weapons, and those results are reported here. Analytes studied include methylphosphonic acid (a dibasic acid), the monoisopropyl ester of ethylphosphonic acid, and the monoalkyl esters of methylphosphonic acid (R = ethyl, isopropyl, isobutyl, pinacolyl (3,3-dimethyl-2-butyl), cyclohexyl, and 2-ethylhexyl). The cationic surfactants used here for the reversal of electroosmotic flow are didodecyldimethylammonium hydroxide and cetyltrimethylammonium hydroxide. CE methods using conductivity or indirect UV detection provide a good separation efficiency and very high sensitivity for the analysis of such compounds. The detection limits for these species were about 75 micrograms/L when using conductivity detection and about 100 micrograms/L when using indirect UV detection. Because pH plays an important role in the CE separation of the alkylphosphonic acids and their monoesters, the influence of pH on these separation systems was investigated. Electrolytes were stable for at least 3 months. Excellent separation efficiency and freedom from interference due to common anions were obtained in the developed methods which typically achieved complete separations in less than 3 min. The method was applied to aqueous leachates of soil, wipes of surfaces, and vegetation sampled from a field site known to have been exposed to nerve agents and subsequently cleaned up. The data from these environmental samples indicated that the method can be expected to be useful for environmental monitoring.

Characterization of the Cationic Surfactant Induced Reversal of Electroosmotic Flow in Capillary Electrophoresis

Reversal of electroosmotic flow in capillary electrophoresis can be achieved by the addition of cationic surfactants to the electrophoretic buffer. This reversal of flow is caused by the formation of a bilayer or hemimicelle at the walls of the capillary, effectively making the wall charge positive. The bilayer can be formed either by adsorption of monomeric surfactants or by admicelles (surfactant pairs), depending on the ionic strength of the buffer and the surface charge at the capillary walls. At concentrations of cationic surfactant sufficient to form the bilayer (i.e., greater than the critical micelle concentration) the electroosmotic flow velocity is independent of pH above a pH of 4. Adjustment of the ionic strength causes minor variations in the reversed electroosmotic flow rate due to the opposing forces of increased concentration of adsorbed surfactant and decreased double layer thickness. The nature of the anionic counterion in solution has a strong effect on the magnitude of the reversed ele...

Modeling of the Impact of Ionic Strength on the Electroosmotic Flow in Capillary Electrophoresis with Uniform and Discontinuous Buffer Systems

A new dynamic computer model permitting the combined simulation of the temporal behavior of electroosmosis and electrophoresis under constant voltage or current conditions and in a capillary which exhibits a pH-dependent surface charge has been constructed and applied to the description of capillary zone electrophoresis, isotachophoresis, and isoelectric focusing with electroosmotic zone displacement. Electroosmosis is calculated via use of a normalized wall titration curve (mobility vs pH). Two approaches employed for normalization of the experimentally determined wall titration data are discussed, one that considers the electroosmotic mobility to be inversely proportional to the square root of the ionic strength (method based on the Gouy-Chapman theory with the counterion layer thickness being equal to the Debye-Hückel length) and one that assumes the double-layer thickness to be the sum of a compact layer of fixed charges and the Debye-Hückel thickness and the existence of a wall adsorption equilibrium of the buffer cation other than the proton (method described by Salomon, K.; et al. J. Chromatogr. 1991, 559, 69). The first approach is shown to overestimate the magnitude of electroosmosis, whereas, with the more complex dependence between the electroosmotic mobility and ionic strength, qualitative agreement between experimental and simulation data is obtained. Using one set of electroosmosis input data, the new model is shown to provide detailed insight into the dynamics of electroosmosis in typical discontinuous buffer systems employed in capillary zone electrophoresis (in which the sample matrix provides the discontinuity), in capillary isotachophoresis, and in capillary isoelectric focusing.

Theoretical aspects on the regulation of electroosmotic flow in capillary electrophoresis by adding charged amphiphiles

The regulation of electroosmotic flow (EOF) in capillary electrophoresis is of great importance in many analytical applications. In this paper, the variation of EOF with the concentration of amphiphile is studied by adding a charged amphiphile to the background electrolyte (BGE) when using a C 8-coated capillary. A theory relating the change in concentration of amphiphile in the BGE to the variation in observed EOF is presented. The basis of the theory is that the adsorbed amphiphile creates an electrostatic surface potential that is calculated from the linearised Poisson-Boltzmann equation. The adsorption isotherm of the amphiphile to the capillary surface is described by an electrostatic surface potential modified linear isotherm. By assuming that the ζ-potential in the Smoluchowski equation is proportional to the electrostatic surface potential, an equation describing the EOF as a function of amphiphile concentration is obtained. The proposed theory is experimentally tested by adding octane sulfonate, tetrabutylammonium or tetrapentylammonium ion to the BGE. It is shown that, in these cases, the proposed theory describes the EOF as a function of amphiphile concentration well and that, as expected, the proportionality constant between the electrostatic surface potential and the ζ-potential is less than one. Some practical aspects and problems that occur with this type of measurement are also discussed.

Method for measuring very weak, residual electroosmotic flow in coated capillaries

Many different methods exist for measuring the velocity of electroosmotic flow in capillary electrophoresis (CE). The easiest and the most common one is photometric detection of a neutral marker. This method can be used on any CE unit, while other methods may require some modifications or special equipment. However, this method is not very suitable for measuring weak electroosmosis, since very long runs are necessary for the neutral marker to arrive at the detector. For this case, an alternatice method is proposed here. It assumes that the neutral marker is injected by means of electroosmosis, then rinsed by applying a low pressure and recorded. The resulting peak is quantified and, according to the amount of sample injected, the velocity of electroosmosis can be estimated. The proposed method is much faster than the traditional one; its duration is determined mainly by the time of electroosmotic injection. It can be used for estimation of the quality of capillary coatings, for which the residual electroosmotic flow is one of the characteristics.

Hydrolytically stable amino-silica glass coating material for manipulation of the electroosmotic flow in capillary electrophoresis

A hydrolytically stable amino-silica glass coating material was fabricated inside fused-silica capillaries, using sol-gel technology. Aminopropyltriethoxysilane was used as the precursor in the glass formation process. The net charge at the surface of the coating material depends on the degree of protonation of the amino groups and the degree of ionization of the silanol groups, thus enabling manipulation of the magnitude and direction of the electroosmotic flow (EOF). At low pH (<6.0), the coating bears a net positive charge, which results in an electroosmotic flow from the cathode toward the anode and minimizes the wall-solute interactions of basic species. At high pH (>6.5), the coating surface bears a net negative charge and the coated capillary behaves like an uncoated one, having an EOF in the cathodic direction. The amino-silica glass coating has also been shown to be extremely stable under both acidic and basic conditions. The reproducibility of the electroosmotic mobility of five different capillaries was found to be 7% R.S.D. The utility of the material is shown with the separation of basic proteins, peptides and basic compounds.

Theoretical Description of the Influence of External Radial Fields on the Electroosmotic Flow in Capillary Electrophoresis

The effect of applying a radial voltage on the electroosmotic flow in capillary electrophoresis has been studied from a theoretical point of view. Based on Stern's model for the electric double layer on the surface of a fused silica capillary and on the Gouy-Chapman theory for the diffuse layer, equations describing the relation between the electroosmotic mobility and the radial electric field were derived. The thickness of the stagnant solution layer on the surface of the capillary, an important parameter in the calculations, was estimated from the electroosmotic mobility found in high-pH solutions. The theory developed predicts the experimental findings that the effect of the radial field levels off at high applied voltages and that it is smaller when the electroosmotic mobility without radial field is already high. The theoretical results were compared with experimental data taken from the literature. A good quantitative agreement was found.