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.     

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.     

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   

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.     

Semi-permanent surfactant coatings for inorganic anion analysis in capillary electrophoresis

Capillary electrophoretic separations of inorganic anions are performed using a capillary coated with a mixture of the cationic surfactant didodecyldimethylammonium bromide (DDAB) and the zwitterionic surfactant 1,2-dilauroyl-sn-phosphatidylcholine (DLPC). These double-chained surfactants form semi-permanent coatings on the capillary wall, which allows the excess surfactant to be removed from the buffer prior to separation. Interactions between surfactant aggregates in the buffer and analyte anions are thus eliminated. The electroosmotic flow (EOF) can be altered from fully reversed (100% DDAB) to near zero (100% DLPC) using different ratios of DDAB and DLPC. Controlling the EOF allows for improved resolution of the anions while maintaining a rapid, co-EOF separation, free from analyte-surfactant additive interactions.     

Dynamic capillary coatings for electroosmotic flow control in capillary electrophoresis

Control of the electroosmotic flow (EOF) is critical for achieving optimal separations by capillary electrophoresis. For instance, manipulation of the EOF can yield either high resolution separations or rapid analyses. Dynamic capillary coatings are a simple and cost-effective approach to altering the EOF. The normal EOF can be slowed using buffer additives such as Mg²⁺ and hexamethonium which ion exchange onto the surface silanols to lower the effective wall charge. Alternatively, cationic polyelectrolytes or cationic surfactants can be used to establish a cationic coating on the capillary wall, which results in a reversed EOF. Practical considerations such as pH stability and reproducibility obtainable with an EOF modifier will be discussed.     

Effect of coating electrolytes on two-tailed surfactant bilayer coatings in capillary electrophoresis

Surfactants such as dioctadecyldimethylammonium bromide (DODAB) form semi-permanent coatings that effectively prevent adsorption of cationic proteins onto the fused silica capillary in capillary electrophoresis (CE). The bilayer coating is generated by flushing the capillary with a 0.1 mM surfactant solution. However, formation of the bilayer is strongly dependent on the coating electrolyte. The effect of counter-ions, electrolyte concentrations and buffer co-ions were monitored based on: the separation of basic model proteins; the adsorption kinetics of DODA⁺ onto fused silica; and dynamic light scattering (DLS) to determine vesicle size. Low concentrations (≤10.0 mM) and/or weakly associating buffers such as phosphate (pH 3.0), acetate (pH 4.0) and chloride should be used for DODAB coating solutions. Dissolving the surfactant in strongly associating electrolyte, such as phosphate at pH 7.0, results in poor coating of the capillary surface. Effective cationic bilayer coatings are formed if the buffer conditions favor formation of vesicles with diameters < 300 nm. Monitoring turbidity at 400 nm provides a convenient means of verifying vesicle diameter variation of <5 nm; that is, that the coating solution is effective.     

Evaluation of poly([2‐(acryloyloxy)ethyl]trimethylammonium chloride) cationic polymer capillary coating for capillary electrophoresis and electrokinetic chromatography separations

Capillary electrophoresis and electrokinetic chromatography are typically carried out in unmodified fused‐silica capillaries under conditions that result in a strong negative zeta potential at the capillary wall and a robust cathodic electroosmotic flow. Modification of the capillary wall to reverse the zeta potential and mask silanol sites can improve separation performance by reducing or eliminating analyte adsorption, and is essential when conducting electrokinetic chromatography separations with cationic latex nanoparticle pseudo‐stationary phases. Semipermanent modification of the capillary walls by coating with cationic polymers has proven to be facile and effective. In this study, poly([2‐(acryloyloxy)ethyl]trimethylammonium chloride) polymers were synthesized by reversible addition‐fragmentation chain transfer polymerization and used as physically adsorbed semipermanent coatings for capillary electrophoresis and electrokinetic chromatography separations. An initial synthesis of poly([2‐(acryloyloxy)ethyl]trimethylammonium chloride) polymer coating produced strong and stable anodic electroosmotic flow of –5.7 to –5.4 × 10⁻⁴ cm²/V⋅s over the pH range of 4–7. Significant differences in the magnitude of the electroosmotic flow and effectiveness were observed between synthetic batches, however. For electrokinetic chromatography separations, the best performing batches of poly([2‐(acryloyloxy)ethyl]trimethylammonium chloride) polymer performed as well as the commercially available cationic polymer polyethyleneimine, whereas polydiallylammonium chloride and hexadimethrine bromide did not perform well.     

A block copolymer covalent coating acting as surfactants in separation of 2-[hydroxy(4-nitrophenyl)methyl]-cyclopent-2-enone and 4-nitrobenzaldehyde by capillary electrophoresis

An innovative block copolymer capillary coating P(MAn-alt-St)₁₂₇-b-PSt₅₉₂, synthesized by maleic anhydride and styrene, was developed as a new kind of coating for capillary electrophoresis. The covalent bond coating was effectively applied in the separation of raw material (4-nitrobenzaldehyde) and production (2-[hydroxy(4-nitrophenyl)methyl]-cyclopent-2-enone) in a Baylis-Hillman reaction using ammonium acetate with 20% tetrahydrofuran (v/v) as the buffer solution. Electroosmotic flow measurement gave an instantly stable value after 70 times continued injection in 5 days and showed that P(MAn-alt-St)₁₂₇-b-PSt₅₉₂ coatings could suppress electroosmotic flow effectively compared with the bare capillary. The effects of tetrahydrofuran and the pH of buffer on the separation were investigated. The characteristics of the coatings to form micelles similar to surfactants were detected by atomic force microscopy. Moreover, the superiority of this coating was further applied in the separation of four aromatic amines.     

Cationic lipid vesicles as coating precursors in capillary electrochromatography: separation of basic proteins and neutral steroids

1,2-Dioleyl-3-trymethylammoniumpropane (DOTAP) lipid vesicles were employed as coating precursors to obtain a semipermanent cationic lipid bilayer in silica capillary. The coating procedure was relatively fast and simple. Reliable results for the separation of four basic proteins (alpha-chymotrypsinogen A, ribonuclease A, cytochrome C, lysozyme) were obtained by using an acetate buffer under acidic conditions. The RSDs of the migration times were not higher than 0.5% run-to-run and about 1% day-to-day (3 days), while the RSDs of the peak areas were within 7% day-to-day (3 days). The day-to-day RSD of the EOF mobility of about 1%, confirmed that the DOTAP coating was stable for the separation of basic proteins, under acidic buffers. In addition to basic proteins the DOTAP coating was found suitable under acidic conditions for the repeatable separation of neutral steroids. The potential of DOTAP as a carrier in background electrolyte solution was studied.     

Adjusting the selectivity of inorganic anion separation by capillary electrophoresis

The separation of the principal inorganic anions (bromide, carbonate, chlorate, chloride, fluoride, nitrate, nitrite, sulfate, phosphate) has been achieved using a capillary electrophoresis system with indirect UV detection at 260 nm. Several types of cationic surfactants (quaternary ammonium, phosphonium or methonium) were tested as electroosmotic flow modifiers and added to a chromatebased buffer prepared from potassium dichromate. The influence of many physicochemical parameters such as nature and concentration of cationic surfactant, buffer pH, dichromate concentration buffer, voltage and temperature upon the migration time of an analyte anion, peak efficiency, asymmetry factor, and finally resolution has been investigated. A linear relationship between the corrected area and the anion concentration in the 2.5–50 ppm range was obtained, thus allowing the quantitative analysis of anions in mineral water. Finally, by increasing the hydrodynamic injection time, the separation of inorganic anions at a low concentration level of 50 ng/ml was achieved without any loss of resolution.     

Dynamic coating of a capillary with room-temperature ionic liquids for the separation of amino acids and acid drugs by capillary electrophoresis

A room-temperature ionic liquid (IL), 1-ethyl-3-methyl-imidazolium tetrafluoroborate (1E-3MI-TFB), used for the coating of a silica capillary enables one to reduce or invert the electroosmotic flow (EOF) in capillary zone electrophoresis. Excellent separations of amino acids and ary lalkanoic acids were obtained. Such separations could not be obtained in a naked capillary in the presence of the cationic surfactants cetyltrimethylammonium bromide (CTAB) or polycationic polymer hexadimethrine bromide (HDB). The results indicate that 1E-3MI-TFB not only modulates the EOF but also acts as a discriminator. Further experiments indicate that the interaction between hydrogen at C-2 carbon of IL and acid drugs plays an important role in the separation. The text was submitted by the authors in English.     

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.     

Separation of the positional isomers of phthalic acids in hydroorganic solvents using capillary electrophoresis

The positional isomers of phthalic acids (ortho-, meta-, and para-) and benzoic acid could be completely separated by capillary electrophoresis (CE). A simple CE method employing direct detection in mixed methanol/water buffers is presented. The effect of the electrolyte buffer system, including pH, buffer concentration, and organic solvent on the electrophoretic mobility of the analytes, is investigated. The electroosmotic flow is reversed using cationic surfactant and cetyltrimethylammonium bromide so that anions are separated under the co-EOF mode. The resolution of the analytes and selectivity could be improved by the adjustment of the methanol content. Ion association with the surfactant in methanol/water buffer is discussed. The validity of the method in terms of sensitivity, reproducibility, and linearity is also reported. The text was submitted by the authors in English.     

Mixed cationic/anionic surfactants for semipermanent wall coatings in capillary electrophoresis

Mixtures of the cationic surfactant cetyltrimethylammonium bromide (CTAB) with the anionic surfactant sodium dodecyl sulfate (SDS) form more stable coatings in fused-silica capillaries than CTAB alone. The reversed electroosmotic flow (EOF) generated by CTAB/SDS mixtures remains stable for over 80 min after removal of the surfactants from the buffer. Enhanced stability (relative to CTAB alone) was found even when the ratio of SDS to CTAB was as low as 1%. This greater coating stability is attributed to the structural transition from adsorbed micelle to bilayer, which is induced by addition of SDS. Separation of a mixture of basic proteins yielded efficiencies of 364 000-562 000 plates/m and recoveries ranging from 85% to 98%. Migration time reproducibility was less than 0.9% relative standard deviation (RSD) from run to run and less than 2.6% RSD from day to day.     

A semipermanent coating for preventing protein adsorption at physiological pH in kinetic capillary electrophoresis

Protein adsorption to the inner capillary wall hinders the use of kinetic capillary electrophoresis (KCE) when studying noncovalent protein-ligand interactions. Permanent and dynamic capillary coatings have been previously reported to alleviate much of the problems associated with protein adsorption. The characteristic limitations associated with permanent and dynamic coatings motivated us to look at a third type of coating - semipermanent. Here, we demonstrate that a semipermanent capillary coating, designed by Lucy and co-workers, comprised of dioctadecyldimethylammonium bromide (DODAB) and polyoxyethylene (POE) stearate, greatly reduces protein adsorption at physiological pH - a necessary requirement for KCE. The coating (i) does not inhibit protein-DNA complex formation, (ii) prevents the adsorption of the analytes, and (iii) supports an electoosmotic flow required for many applications of KCE. The coating was tested in three physiological buffers using a well-known DNA aptamer and four proteins that severely bind to bare silica capillaries as standards. For every protein, a condition was found under which the semipermanent coating effectively suppresses protein adhesion. While no coating can completely prevent the adsorption of all proteins, our findings suggest that the DODAB/POE stearate coating can have a broad impact on CE at large, as it prevents the absorption of several well studied, highly adhesive proteins at physiological pH.     

LC Separation of Co-Existing Cetylpyridinium Chloride, Tetradecyltrimethylammonium Bromide and Dodecyltrimethylammonium Bromide on Silica TLC Plates with Aqueous-Organic Eluents

Thin-layer chromatography (TLC) of three cationic surfactants was performed on silica TLC plates with various solvent systems. The mutual separation of cetylpyridinium chloride (CPC), tetradecyltrimethylammonium bromide (TTAB) and dodecyltrimethylammonium bromide (DTAB) was achieved on silica TLC plates with ethanol: 1% aqueous ammonium chloride (4:6, v/v) as an eluent. Effects of cations and anions in the mobile phase on mobility and separation of CPC, TTAB and DTAB were examined. The interference due to the presence of metal cations as impurities on the resolution in the mixture of CPC, TTAB and DTAB was also examined. The limits of detection of CPC, TTAB and DTAB estimated were 0.015, 0.031 and 0.062 μg zone⁻¹, respectively. The developed method was utilized to identify these surfactants in different spiked water samples after their preliminary separation.     

表面活性剂在高效毛细管电泳中的作用

表面活性剂作为缓冲液添加剂已广泛用于高效毛细管电泳中,综述了阴离子、阳离子、两性离子、非离子及手性等多种表面活性剂在离子、中性分子、手性化合物、多肽和蛋白质分离等方面的作用,介绍了其作用机理与改善高效毛细管电泳分离的原理。     

Small diamines as modifiers for phosphatidylcholine/phosphatidylserine coatings in capillary electrochromatography

Greater stability of liposome coatings and improved resolution of model steroids in capillary electrochromatography (CEC) were sought by adding small diamines (ethylenediamine, diaminopropane, bis-tris-propane, or N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid, HEPES)) to the liposome solution before coating of fused silica capillaries. The phospholipid coatings consisted of 1 mM of 8:2 mol% phosphatidylcholine (PC)/phosphatidylserine (PS) and 5 mM of modifier in buffer solutions (acetate, phosphate, or Tris) at pH 4.0-7.4. The coating was based on a published procedure, and five steroids were used as neutral model analytes in evaluation of the coating. The results showed that under optimal conditions, the small linear diamines increased the packing density of anionic phospholipids, leading to improved separations. In addition, the choice of buffer for the liposome coating and separation appeared to influence the performance of the coatings. While buffers with amino groups take part in the phospholipid bilayer formation, buffers like phosphate may even have negative effect on coating formation. The factors affecting phospholipid coatings with diamines as modifiers are clarified.     

Nonaqueous affinity capillary electrophoresis investigation of small molecule molecular recognition

The conditional binding constants for a bis-guanidinium-like receptor and a series of dicarboxylate ligands have been determined in two buffer/solvent systems, namely 25 mM ammonium acetate/1% acetic acid in acetonitrile/methanol (7:3 v:v) and 30 mM N-methyl morpholine/15 mM methanesulfonic acid in acetonitrile/methanol (9:1 v:v). The latter buffer has not been applied before in capillary electrophoresis. The binding constants in both solvent systems decrease as the dicarboxylate length increases. The binding constants are larger in the less competitive N-methyl morpholine buffer. The dicarboxylates associate only weakly with a dicationic analog of the receptor, p-xylyl trimethylammonium, which is not a hydrogen bond donor.