Electroosmotic flow reversal for the determination of inorganic anions by capillary electrophoresis with methanol-water buffers

Manipulation of the electroosmotic flow (EOF) is essential for achieving optimized separations of small anions by capillary electrophoresis (CE). In this work, efficient suppression or reversal of EOF is achieved upon addition of small amounts of the cationic surfactants, cetyltrimethylammonium bromide (CTAB) or didodecyldimethylammonium bromide (DDAB) to the electrophoretic buffer. Highly stable and reversed EOF are achieved using the surfactants in the presence of up to 50% MeOH. In aqueous and low methanol containing solutions (up to 30%, v/v) surface aggregation of the surfactants at the capillary wall occurs at a concentration below the critical micelle concentration (CMC). The impact of MeOH on reversed EOF is predominantly a function of the diminished zeta potential of the silica, and to a lesser extent on the CMC in the bulk solution of the surfactant. Fast baseline separation and selectivity changes for small inorganic anions are observed when mixed aqueous-organic buffers are employed. Changes in EOF, micellar properties of the surfactant and selectivity for inorganic anions upon addition of various percent of methanol are also discussed.     

Cationic double-chained surfactant as pseudostationary phase in micellar electrokinetic capillary chromatography for drug separations

The cationic double-chained surfactant didodecyldimethylammonium bromide (DDAB) was used as pseudostationary phase (PSP) in micellar electrokinetic capillary chromatography (MEKC). Its performance on the three kinds of drugs, i.e., basic, acidic, and neutral drugs, was systematically investigated. Nicotine, cotinine, caffeine, lidocaine, and procaine were selected as the model basic drugs. Good baseline separation and high efficiency were obtained under the optimal separation condition that consisted of 50mM phosphate (pH 4.0) and 0.08 mM DDAB. Three basic phenylenediamine isomers can also be well separated with DDAB in buffer. In addition, DDAB can form cationic bilayer on the capillary wall, thus the wall adsorption of basic analytes was greatly suppressed. Compared with commonly used CTAB, the separation of basic drugs was significantly improved with a much lower amount of DDAB present in the buffer. The DDAB-involved MEKC also showed superiority to CTAB upon the separation of acidic drugs, amoxicillin and ampicillin. In the case of neutral compounds, a good separation of resorcinol, 1-naphthol and 2-naphthol was achieved with 0.1mM DDAB and 30% (v/v) acetonitrile (ACN) present in buffer. Hence, it was concluded that the double-chained cationic surfactant DDAB can be a good substitute for traditional single-chained surfactant CTAB in MEKC.     

Enhanced stability of surfactant-based semipermanent wall coatings in capillary electrophoresis using oppositely charged surfactant

Semipermanent surfactant coatings are effective for the prevention of wall adsorption of proteins in CE. However, they often suffer from their unsatisfactory coating stability as they essentially degrade from the capillary walls after the surfactants are removed from the buffer. In this paper, we proposed a facile and universal method to improve the stability of semipermanent surfactant coatings based on addition of an oppositely charged surfactant into the coating. Didodecyldimethylammonium bromide (DDAB) and a gemini surfactant, 18-6-18, were used as the model semipermanent coatings, and sodium dodecyl sulfate (SDS) was chosen as their oppositely charged surfactant. SDS can strongly alter the packing parameter P of the cationic surfactants, and consequently mediates the coating stability. With the increase of SDS concentration in coating, the coating stability first dramatically increases due to the enlarged P, and then decreases due to the weakness of electrostatic interaction between the capillary wall and surfactant coating. At the proper SDS concentration, very stable coatings can be obtained that, even after rinsing under 138 kPa for 60 min, the reversed electroosmotic flow (EOF) only decreases by 3.6%. These SDS-enhanced coatings show excellent stability and reproducibility in protein separation (RSD of migration time <1.1% for run-to-run assay, n=9). Also, the high separation efficiency (>500,000 plates/m) and fine recovery of tested proteins indicate that these coatings are powerful in wall adsorption suppression. Finally, we found that the separation efficiency of protein was a more exact indicator for the coating stability than the traditional EOF magnitude.     

Studies on the analytical performance of a non-covalent coating with N,N-didodecyl-N,N-dimethylammonium bromide for separation of basic proteins by capillary electrophoresis in acidic buffers in 25-and 50-μm capillaries

Capillaries (25-and 50-μm inner diameter) coated with a double-alkyl-chain cationic surfactant N,N-didodecyl-N,N-dimethylammonium bromide (DDAB) were used for the separation of four basic standard proteins in buffers of pH 4 at various ionic strengths. The choice of buffer is critical for the analytical performance. Ammonium ions must be avoided in the buffer used in the non-covalent coating procedure owing to competition with the surfactant. Phosphate buffer gave a better separation performance than some volatile buffers; the reason seems to be a complex formation between the proteins and dihydrogenphosphate ions, which decreases tendencies for adsorption to the capillary surface. The DDAB coating was easy to produce and stable enough to permit, without recoating, consecutive separations of the proteins for up to 100 min with good precision in migration times and peak areas. A strong electroosmotic flow gives rapid separations, which is of special importance when commercial instruments are used, since the choice of the length of the capillary is restricted. Figure EOF stability in 25 micrometer i.d. capillaries. Consecutive injections of mesityloxide performed after an initial coating with 1.0     

Study of the selectivity of inorganic anions in hydro-organic solvents using indirect capillary electrophoresis

In capillary electrophoresis (CE) analysis of small inorganic anions, the ability to control the electroosmotic flow (EOF) and the ability to alter the electrophoretic mobility of the ions are essential to improve resolution and separation speed. In this work, a CE method for separation of small inorganic anions using indirect detection in mixed methanol/water buffers is presented. The suitability of different UV absorbing probes commonly used for indirect detection including chromate, iodide, phthalate, benzoate, trimellitate, and pyromellitate, in mixed methanol/water buffers is examined. The effect of the electrolyte buffer system, including the pH, buffer concentration and the organic solvent on the electrophoretic mobility of the probes and analytes are also investigated. The EOF was reversed using cationic surfactant, cetyltrimethylammonium bromide (CTAB) so ions were separated under co-EOF mode. The organic solvent alters the electrophoretic mobility of the probes and the analytes differently and hence choice of the appropriate probe is essential to achieve high degree of detection sensitivity. Separations of six anions in less than 2.5 min were accomplished in buffers containing up to 30% MeOH. Adjustment of the methanol content helps to improve the selectivity and resolution of inorganic anions. Limit of detection, reproducibility and application of the method for quantification of anions in water samples will also be discussed.     

Nonionic surfactant enhanced semipermanent coatings for capillary electrophoresis of inorganic anions without use of organic additives

Separation of inorganic anions by capillary electrophoresis (CE) is usually conducted in co-electroosmotic mode due to the large electrophoretic mobilities of inorganic anions. Semipermanent surfactant coatings have been shown to be effective for CE of inorganic anions due to their strong capability of electroosmotic flow (EOF) manipulation. However, semipermanent coatings often suffer from their unsatisfactory stability. In addition, organic solvent additives are usually required to adjust the selectivity, which also aggravate the degradation of coating. In this work, a novel semipermanent coating consisting of cationic Gemini surfactant 18-10-18 and nonionic surfactant Tween 20 was developed to separate inorganic anions in CE. This coating is easy to prepare and more stable than pure Gemini coating. The introduction of nonionic surfactant in the coating not only suppresses the reversed EOF but can also adjust the selectivity of separation. Good separations of six model anions were achieved, the separation efficiency was as high as 65040-169700 plates/m and the RSDs of the migration times were less than 0.5 and 2.5% for run-to-run and day-to-day assays, respectively. Calibration curves were linear in the range of 0.05-5.0 mM; the detection limits ranged from 20 to 50 μM. More importantly, no organic solvents are required in the background buffer to achieve the satisfactory separations. This guarantees the coating stability and makes the method greener than most of other methods for CE of inorganic anions.     

A new and selective capillary column coated with cationic diazacrown ether for separation of organic and inorganic anions by capillary electrophoresis

A new coated capillary has been introduced for capillary electrophoretic separation of anions by using a positively charged diazacrown ether with a 12-membered ring. A positive charge spread over the inner capillary surface led to a substantial anodic electroosmotic flow (EOF) over the range of migrating buffer of pH 2-11. Under the optimum conditions of 25 mM phosphate buffer at pH 7, the diazacrown-coated capillary showed a successful simultaneous separation of 7 inorganic anions and 13 aromatic anions (including positional isomers) in less than 15 min. The migration times of the sample anions and EOF marker for consecutive runs on a single column were highly reproducible, giving a relative standard deviation of 1%. Theoretical treatment of the migration behavior clearly demonstrated that ion association between the diazacrown and analyte anions is strongly dependent on the nature of the functional groups of anions (e.g., sulfonate groups > carboxyl groups) and the number of negative charges (e.g., trivalent anions > divalent anions > monovalent anions) on the analyte.     

Applications of a sulfonated-polymer wall-modified open-tubular fused-silica capillary in capillary zone electrophoretic separations

A fused-silica capillary that is wall-modified via chemically bonding a sulfonated polymer to the capillary wall has a uniform negative charge density on its surface and produces an electroosmotic flow (EOF) greater than 4 x 10(-4) cm2 V(-1) s(-1) The EOF is nearly independent of buffer pH over the pH range of 2 to 10 and is lower than the EOF obtained for the bare fused-silica capillary at the more basic pH but is higher at the more acidic buffer pH. Optimization of buffer pH can be based on analyte pKa values to improve the overall quality of the capillary zone electrophoresis (CZE) separation of complex mixtures of weak acid and base analytes. Because of the high EOF in an acidic buffer, the capillary is useful for the separation of weak organic bases which are in their cation forms in the acidic buffer. EOF for the sulfonic acid bonded phase capillary can be adjusted via buffer additives such as organic solvent, tetraalkylammonium salts, multivalent cations and alkylsulfonic acids. The advantages of utilizing buffer pH and the EOF buffer modifiers to enhance migration time, selectivity, and resolution in CZE separations with this capillary are illustrated using a series of test analyte mixtures of inorganic anions, carboxylic acids, alkylsulfonic acids, benzenesulfonic acids, sulfas, pyridines, anilines or small-chain peptides.     

Application of dodecyldimethyl (2-hydroxy-3-sulfopropyl) ammonium in wall modification for capillary electrophoresis separation of proteins

A zwitterionic surfactant, dodecyldimethyl (2-hydroxy-3-sulfopropyl) ammonium (C12H25N+(CH3)2CH2CHOHCH2SO3-), named dodecyl sulfobetaine (DSB), was used as a novel modifier to coat dynamically capillary walls for capillary electrophoresis separation of basic proteins. The DSB coating suppressed the electroosmotic flow (EOF) in the pH range of 3-12. At high DSB concentration, the EOF was suppressed by more than 8.8 times. The DSB coating also prevented successfully the adsorption of cationic proteins on the capillary wall. Anions, such as Cl-, Br-, I-, SO4(2-), CO3(2-), and ClO4-, could be used as running buffer modifiers to adjust the EOF for better separation of analytes. Using this dynamically coated capillary, a mixture of eight inorganic anions achieved complete separation within 4.2 min with the efficiencies from 24,000 to 1,310,000 plates/m. In the presence of ClO4- as EOF adjustor, the separation of a mixture containing four basic proteins (lysozyme, cytochrome c, alpha-chymotrypsinogen A, and myoglobin) yielded efficiencies of 204,000-896,000 plates/m and recoveries of 88%-98%. Migration time reproducibility of these proteins was less than 0.5% relative standard deviation (RSD) from run to run and less than 3.1% RSD from day to day, showing promising application of this novel modifier in protein separation.     

Modification of the electroosmotic flow and separation selectivity of anions in electrochromatography with pseudo-stationary phases of C14-alkyldimethylammoniopropane sulfonate zwitterionic surfactants by addition of salts to the background electrolyte

The effects of salts (NaCl, NaClO4, MgCl2, CeCl3) added to background electrolyte (BGE) solutions (10 mmol L(-1) sodium phosphate, pH 7.2) on electroosmotic flow (EOF) and the separation selectivity of anions (chloride, bromide, iodide, nitrite, nitrate, chlorate, thiocyanate, iodate, chromate, and molybdate ion) by capillary electrochromatography using the zwitterionic surfactant 3-(N,N-dimethylmyristylammonio)propane sulfonate (C14N3S) as a pseudo-stationary phase were investigated. There are two mechanisms affecting the separations: 1. the cations and anions of the added salts interact with the zwitterionic surfactant to varying degrees, thus changing the overall retention of the analytes; and 2. they change the EOF and the resulting apparent mobilities. It was shown that a BGE containing perchlorate and a low concentration of zwitterionic surfactant (2 mmol L(-1)) gave a stable and reproducible EOF and the concentration of perchlorate could be used to manipulate the separation selectivity for polarizable anions, such as iodide and thiocyanate. These effects are discussed in terms of measured association constants describing the interaction of anions and cations with the zwitterion.     

Use of 1-alkyl-3-methylimidazolium-based ionic liquids as background electrolytes in capillary electrophoresis for the analysis of inorganic anions

Separation of inorganic anions in CE is often a challenging task because the electrophoretic mobilities of inorganic anions are comparable to or even greater than the EOF mobility. In this study, we present the use of ionic liquids (ILs) as background electrolytes (BGEs) in CE of inorganic anions. The 1-alkyl-3-methylimidazolium-based ILs as BGEs dynamically coated the capillary wall and induced a reversed EOF. This allowed the anions to comigrate with the EOF and yielded a rapid separation. Increasing the alkyl chain length of the ILs and BGE concentration can significantly improve the separation resolution. With 40 mM 1-butyl-3-methylimidazolium tetrafluoroborate as BGE, good separations of five model anions (Br-, I-, NO2(-), NO3(-), and SCN-) were achieved in a range of buffer pH values. The separation efficiency was as high as 34 600-155 000, and the RSDs of the migration times were less than 0.8% (n = 5).     

Comparison of the separation of large DNA fragments in the presence and absence of electroosmotic flow at high pH

This paper describes the analysis of large DNA fragments at pH > 10.0 by capillary electrophoresis (CE) in the presence of electroosmotic flow (EOF) using hydroxyethylcellulose (HEC) solution. HEC solution in the anodic reservoir enters the capillaries filled with high-pH buffer by EOF after sample injection. With respect to resolution, sensitivity, and speed, separation conducted under discontinuous conditions (different pH values of HEC solutions and buffer filling the capillary) is appropriate. Using HEC solution at concentrations higher than its entanglement threshold ensures a good separation of large DNA fragments in the presence of EOF at high pH. In addition to pH and HEC, the electrolyte species, dimethylamine, methylamine, and piperidine, play different roles in determining the resolution. The separation of DNA fragments ranging in size from 5 to 40 kilo base pairs was completed in 6 min using 1.5% HEC prepared in 20 mM methylamine-borate, pH 12.0, and the capillary filled with 40 mM dimethylamine-borate, pH 10.0. In comparison, this method allows faster separations of large DNA fragments compared with that conducted in the absence of EOF using dilute HEC solutions.     

Polypyrrole-coated capillaries for capillary zone electrophoresis

Fused-silica capillaries are permanently coated by silanization with 3-{[3-(N-pyrrole)-2-hydroxypropyl]amino}propyltriethoxysilane followed by oxidative polymerization of the pyrrole moieties with iron (III) or peroxodisulfate in the presence of chloride, perchlorate, or dextransulfate as anions. This approach allows to modulate the EOF in its magnitude as well as in its direction. With the small anions chloride and perchlorate, the EOF is reversed below pH 5 while with the large dextransulfate polyanions (DS) the EOF is relatively constant over the pH range from 2.5 to 9.4. This can be of advantage at low pH, at which the EOF of uncoated capillaries is close to zero. Application for separation of some herbicides is shown. The lifetime of PP-modified capillaries is satisfactory: the decrease in EOF is less than 3% during 80 analyses (160 min) and less than 5% over three months of storage. The reproducibility of capillary modification is about 5% (RSD of EOF).     

Capillary electrophoresis of seed albumins fromVicia species using uncoated and surface-modified fused silica capillaries

Summary Capillary zone electrophoresis has been developed for the separation of seed albumins fromVicia faba using both uncoated and polyoxyethylene ether (Brij-35) coated octadecysilane derivatized capillaries. Optimal separation conditions were found by studying the effect of pH, buffer composition and applied voltage. The nonionic surfactant/C₁₈ coated capillary significantly reduced albumin adsorption and electroosmotic flow (EOF). A gradual washing out of the surfactant from the coated capillary during use altered not only the magnitude of the EOF, but also its reproducibility. The introduction of hydrophilic polymer solutions between analyses for dynamic modification of the Brij/C₁₈ coated capillary surface prevented desorption of coating material, allowed optimization of resolution and ensured stability of the EOF. CE with surface-modified capillaries was then used to compare seed albumin profiles of severalVicia species. This technique appears to provide a powerful tool for use in taxonomic investigations.     

Capillary electrophoresis of seed albumins fromVicia species using uncoated and surface-modified fused silica capillaries

Summary Capillary zone electrophoresis has been developed for the separation of seed albumins fromVicia faba using both uncoated and polyoxyethylene ether (Brij-35) coated octadecylsilane derivatized capillaries. Optimal separation conditions were found by studying the effect of pH, buffer composition and applied voltage. The nonionic surfactant/C₁₈ coated capillary significantly reduced albumin adsorption and electroosmotic flow (EOF). A gradual washing out of the surfactant from the coated capillary during use altered not only the magnitude of the EOF, but also its reproducibility. The introduction of hydrophilic polymer solutions between analyses for dynamic modification of the Brij/C₁₈ coated capillary surface prevented desorption of coating material, allowed optimization of resolution and ensured stability of the EOF. CE with surface-modified capillaries was then used to compare seed albumin profiles of severalVicia species. This technique appears to provide a powerful tool for use in taxonomic investigations.     

Cationic gemini surfactant as dynamic coating in CE for the control of EOF and wall adsorption

The cationic gemini surfactant ethylene bis(1-dodecyldimethylammonium) dibromide was used as a dynamic coating to control EOF and prevent wall adsorption of basic proteins in CE for the first time. This gemini surfactant shows a more powerful capability in EOF reversal than traditional single-chained surfactant. The gemini surfactant reverses the EOF at a concentration level even less than 0.01 mM, and the EOF magnitude is affected by surfactant concentration, pH, ionic strength, and ions added in buffer. Highly efficient and rapid protein separation (N > 300,000) was obtained with buffer containing 2 mM gemini surfactant under pH ranging from 3 to 6. The effects of surfactant and buffer concentration on protein separation were investigated in detail. Under the optimal conditions, good repeatability (RSD of migration time <0.6% for run-to-run and <2.5% for day-to-day assays) and recovery (>90%) of tested proteins were obtained. This new dynamic coating is also suitable for biosample analysis.     

Long-chained gemini surfactants for semipermanent wall coatings in capillary electrophoresis of proteins

In this paper, we presented the first example of using gemini surfactants as semipermanent coatings in CE for protein separation. These coatings are based on the self-assembly of a series of cationic gemini surfactants, alkanediyl-alpha,omega-bis(dimethylalkylammonium bromide) (m-s-m), on the capillary wall. The coatings can keep stable for a long time without surfactant in the buffer, e.g., after the surfactants were removed from the buffer, the reversed EOF only decreased by 3.6 and 3.9% for 18-2-18 and 16-2-16 coatings over 60 min under continuous electrophoretic conditions. The coating stability increased with the alkyl chain length m. The double long chains of geminis (m > or = 14) yielded a good coating stability; meanwhile, the spacer group acted as an EOF modifier. Thus, this bifunctional surfactant coating provided a new buffer-independent method for EOF control. For 18-s-18 series, the best coating stability and largest EOF were obtained at s = 10. Ranging s from 3 to 10 yielded a linear fine-tuning of EOF and thereby allowed the adjustment of the protein apparent mobility. Highly efficient separation (>500 000 plates/m) was achieved with all the 18-s-18 coatings. Excellent run-to-run and day-to-day reproducibility (RSD of migration time 相似文献   

Polymeric ionic liquid as a background electrolyte modifier enhancing the separation of inorganic anions by capillary electrophoresis

A novel and easy method for the separation of inorganic anions by capillary electrophoresis using a polymeric ionic liquid (PIL), poly(1-vinyl-3-butylimidazolium bromide) as a background electrolyte modifier has been developed. The PIL has been proved to generate a reversed electroosmotic flow which reduces the analysis time and improves the separation significantly. Effects of the PIL concentration and buffer composition (pH and concentration) were evaluated on basis of the resolution and efficiency of the sample. Under optimum conditions, good separation of six model inorganic anions was achieved with high efficiency and excellent reproducibility within 3 min. The results obtained indicate that the combination of reversed EOF and the association between the analytes and the PIL on the capillary wall or BGE play a prominent role in the separation of anions. Therefore, the PIL presents a useful alternative for the BGE modifier in the study of inorganic anions by CE.     

Multiple effect of surfactants used as additives in background electrolytes in capillary zone electrophoresis: cetyltrimethylammonium bromide as example of model surfactant

Surfactants are frequently used in the preparation of background electrolytes (BGEs) in capillary zone elcetrophoresis (CZE) in order to affect and to optimize both the electroosmotic flow (EOF) and the separation process. Their effects are, however, always multiple, the resulting situation may be very complex and the separation process may even be destroyed. We use the surfactant cetyltrimethylammonium bromide (CTAB) as a model example and bring experimental results and related discussion which elucidate the multiple effect of surfactants in an integrated way. It is shown that even at concentration levels lower than 10(-4) M CTAB strongly reduces the cathodic EOF in bare fused-silica capillaries and converts it into anodic EOF. The magnitude and polarity of the EOF depends not only on the concentration of CTAB but also on the composition of BGEs used. The interactions of CTA cations with the bare capillary wall reduce sorption of cationic analytes and enables their analysis. CTA cations at levels below their critical micelles concentration (CMC) already interact with anionic analytes and reduce their mobilities. This association is strong with highly charged anions and by this, the reversal of the EOF, applying BGEs with highly charged anions is less effective. These interactions are competitive and also depend on the composition of the BGE used. At levels above its CMC, CTAB forms micelles and enables the application of the micellar electrokinetic capillary chromatography (MEKC) mode and the analysis of, e.g., neutral components. Simultaneously, it is shown that the presence of CTAB may increase the number of potentially formed system zones.     

pH-independent large-volume sample stacking of positive or negative analytes in capillary electrophoresis

In capillary electrophoresis, the short optical path length associated with on-column UV detection imposes an inherent detection problem. Detection limits can be improved using sample stacking. Recently, large-volume sample stacking (LVSS) without polarity switching was demonstrated to improve detection limits of charged analytes by more than 100-fold. However, this technique requires suppression of the electroosmotic flow (EOF) during the run. This necessitates working at a low pH, which limits using pH to optimize selectivity. We demonstrate that LVSS can be performed at any buffer pH (4.0-10.0) if the zwitterionic surfactant Rewoteric AM CAS U is used to suppress the EOF. Sensitivity enhancements of up to 85-fold are achieved with migration time, corrected area, and peak height reproducibility of 0.8-1.6%, 1.3-3.7%, and 0.8-4.9%, respectively. Further, it is possible to stack either positively or negatively charged analytes using zwitterionic surfactants to suppress the EOF.