Characterization of surfactant coatings in capillary electrophoresis by atomic force microscopy.

This paper describes the adsorption mechanisms and aggregation properties of cetyltrimethylammonium bromide (CTAB) and didodecyldimethylammonium bromide (DDAB) surfactants that are used for dynamic coatings in capillary electrophoresis (CE). Atomic force microscopy is used to directly visualize surfactant adsorption on fused silica. It was found that the single-chained surfactant CTAB forms spherical aggregates on silica while the double-chained surfactant DDAB forms a bilayer. Aggregation at the surface occurs at approximately the same surfactant concentration in which EOF reversal is observed in CE. The nearest-neighbor distance between CTAB aggregates varies inversely with buffer pH and becomes constant at the point when the silanol groups are fully ionized. DDAB forms a flat, uniform coating independent of pH. Increasing the buffer ionic strength changes the morphology of the CTAB aggregates from spherical to cylindrical. The change in morphology can alter the surface coverage, which is related to the "normalized" EOF measured in identical buffers. The morphology of a surfactant coating is also shown to affect its ability to inhibit protein adsorption to the capillary wall. Specifically, the full surface coverage provided by DDAB proved superior in a head-to-head comparison with CTAB.

Dielectric friction in capillary electrophoresis: mobility of organic anions in mixed methanol-water media.

The mobilities of a series of organic carboxylates and sulfonates, ranging in charge from -1 to -4, were investigated by capillary electrophoresis using buffers containing 0 to 75% (v/v) methanol. Effective mobilities were measured at a series of ionic strengths, and were extrapolated to zero ionic strength using Pitts' equation to yield absolute mobilities. Generally, higher-charged ions were more strongly influenced by ionic strength, as predicted by the Pitts' equation. Some differences in the ionic strength effects for anions of like charge were observed and were consistent with the relaxation effect. The absolute mobilities of anions were altered by the addition of methanol to the buffer. Analytes with higher charge-to-size ratios were slowed to a greater extent than were ions with lower charge-to-size. As a result, dramatic changes in relative mobility were observed, such as a reversal in migration order between anions of -1 and -4 charge at 75% methanol and 20 mM ionic strength. The mobility changes caused by the addition of methanol are attributed to dielectric friction. Mobilities in the methanol-water solutions were found to depend on analyte charge-to-size and solvent dielectric relaxation time (tau) and were inversely dependent upon solvent dielectric constant (epsilon), as predicted by the Hubbard-Onsager mobility model.

Chemiluminescence nitrogen detection in ion chromatography for the determination of nitrogen-containing anions.

Chemiluminescence nitrogen detection (CLND) provides equimolar response for nitrogen-containing ions such as nitrate, nitrite, cyanide, ammonium and tetradecyltrimethylammonium. Only azide yields a lower response. Nitrite, azide and nitrate are separated on a Dionex AS11 column using 5 nM NaOH as eluent with a 3 microM (1 ng N) limit of detection. Matrices, such as 1:10 diluted seawater, do not degrade these detection limits. CLND also provides equally sensitive (limit of detection 3 microM, 78 ppb) detection of weak acids such, as cyanide, which yield poor sensitivity with suppressed conductivity detection.

Effect of temperature on retention and selectivity in ion chromatography of anions.

The temperature dependence of retention of a wide range of inorganic anions is studied on two commercially available ion exchangers (Dionex AS11 and AS14 columns). Anion retention exhibited both exothermic and endothermic behavior, such that varying the temperature from ambient to 60 degrees C produced selectivity changes. The anions displayed three groupings of temperature dependence: weakly retained singly charged anions (e.g., iodate, bromate, nitrite, bromide and nitrate); multiply charged anions (sulfate, oxalate, phosphate and thiosulfate); and strongly retained singly charged anions (iodide, thiocyanate and perchlorate). Temperature was ineffective at changing the selectivity of retention between anions of the same grouping. However, significant selectivity changes, including elution order reversal, could be achieved between anions from different groupings.

High-sensitivity determination of the degradation products of chemical warfare agents by capillary electrophoresis-indirect UV absorbance detection.

Capillary electrophoresis coupled with indirect UV absorbance detection was employed for the determination of the chemical warfare agent degradation products: methylphosphonic acid, ethyl methylphosphonate, isopropyl methylphosphonate, and pinacolyl methylphosphonate. Glutamic acid was used as a buffering agent at its isoelectric point (pH 3.22). In its zwitterionic form, glutamic acid does not act as a competing co-anion in the system, thus providing buffering capacity while maintaining high sensitivity. The indirect probe (phenylphosphonic acid) concentration was lowered to 1 mM from the 10 mM in previous literature studies, further enhancing sensitivity. Detection limits of 2 microM were achieved with hydrodynamic injection and up to 100-fold lower using electrokinetic injection. The increased buffering capacity of this system over previous methods led to migration time reproducibility RSD values of 0.18 to 0.22%. This represents a 10-fold improvement in reproducibility over previous studies with comparable or improved sensitivity.

Indirect laser-induced fluorescence detection for capillary electrophoresis using a violet diode laser.

The violet (415 nm) diode laser is used for indirect laser-induced fluorescence detection in capillary electrophoretic separations of inorganic anions and chemical warfare agent degradation products. Inorganic anions were detected using 8-hydroxypyrene-1,3,6-trisulfonic acid as the indirect probe and achieved submicromolar (40-80 ppb) detection limits in a 2-min separation. The chemical warfare agent degradation products methylphosphonic acid, ethyl methylphosphonate, isopropyl methylphosphonate, and pinacolyl methylphosphonate were detected using the porphyrin tetrakis(4-sulfophenyl)porphine as the indirect probe and achieved detection limits of 0.1 microM (9 ppb), which are 1 order of magnitude better than that achieved using indirect UV detection. Baseline stability achieved with the violet diode laser was excellent, with dynamic reserve (DR) values of > 1000, which are 15 times better than that achieved using an unstabilized HeCd laser.

Prediction of electrophoretic mobilities. 4. Multiply charged aromatic carboxylates in capillary zone electrophoresis.

The electrophoretic mobilites of aromatic carboxylates and sulfonates at zero ionic strength were correlated with models incorporating both hydrodynamic and dielectric friction. The hydrodynamic friction was predicted using either the Hückel spherical ion model or the Perrin ellipsoidal model. Dielectric friction is the charge-induced drag caused by the reorientation of the solvent dipoles in response to the analyte charge. Based on the Hubbard-Onsager and Zwanzig expressions, the dielectric friction is related to z2/V. Expressions incorporating both the hydrodynamic and dielectric frictional terms successfully predicted infinite-dilution mobilities to within 4.4%. The influence of dielectric friction ranged from 3-8% of the overall drag for singly charged analytes to 39% of the total frictional drag for 1,2,4,5-benzenetetracarboxylate.

Altering the selectivity of inorganic anion separations using electrostatic capillary electrophoresis.

A new method for altering the selectivity of inorganic anion separations in capillary electrophoresis is described. Addition of the zwitterionic surfactant 3-(N,N-dimethyldodecylammonio)propane sulfonate (DDAPS) to the background electrolyte modifies the migration order via electrostatic ion chromatography type interactions. Variation of the DDAPS surfactant concentration from 4 to 120 mM monotonically alters the selectivity from electrophoretic mobility based to that of electrostatic ion chromatography, without increasing Joule heating. This technique was applied to the determination of nitrate, nitrite, bromide and iodide in artificial seawater. Detection limits for the anions in 1:5 diluted seawater were 11, 5, 7 and 11 microM, respectively.

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.

Evaluation of column temperature as a means to alter selectivity in the cation exchange separation of alkali metals, alkaline earth metals and amines.

The merits of varying column temperature in a cation exchange separation of alkali metals, alkaline earth metals and amines are considered. Increasing the column temperature (up to 60 degrees C) reduced the retention of all cations, but by varying extents. Consequently, selectivity changes were seen, with reversals in elution order in some cases. To ascertain when temperature is most useful as a separation aid, analytes were classed into three groups according to their temperature behaviour: alkali metals; alkaline earth metals; and amines. Adjusting the column temperature caused selectivity changes between analytes in different groups, but no selectivity changes occurred between analytes in the same group. Further, temperature was compared to the addition of modest amounts of acetonitrile as another means to alter selectivity. The benefits of elevated temperature were not just limited to selectivity changes. Improvements in the efficiencies of all analytes were noted at 60 degrees C. This was especially true for the amines which are severely tailed at ambient temperatures.

Ultra-rapid analysis of nitrate and nitrite by capillary electrophoresis.

Rapid analysis of nitrate and nitrite by capillary electrophoresis (CE) has been limited by the ions' very similar electrophoretic mobilities. With a pKa of 3.15, the mobility of nitrite can be selectively reduced using a low pH buffer in CE. A much shorter capillary can be used and separation voltages can be increased. With this method, nitrate and nitrite are separated in just over 10 s. This is roughly 20 times faster than current separation methods. Direct UV detection at 214 nm was employed and offered sub microM detection limits. Total analysis time (pre-rinse, injection, and separation) was less than 1 min, making this method ideal for high-throughput analysis.

Simultaneous separation of cationic and anionic proteins using zwitterionic surfactants in capillary electrophoresis.

The zwitterionic surfactant Rewoteric AM CAS U forms a dynamic wall coating that prevents the adsorption of cationic proteins as well as suppresses the electroosmotic flow (EOF). Addition of polarizable anions to buffers containing this zwitterionic surfactant increases the once suppressed EOF to values nearing +3 x 10(-4) cm2/(V s). The retention of the EOF allows for the separation of analytes of widely different mobilities and is demonstrated by the simultaneous separation of cationic and anionic proteins. Using a buffer containing optimal amounts of the polarizable anion perchlorate and surfactant CAS U, the proteins lysozyme, ribonuclease A, alpha-chymotrypsinogen A, and myoglobin are separated in less than 15 min. Efficiencies as high as 1.5 million plates/m and recoveries greater than 91% are observed for proteins injected in distilled water. Migration time reproducibility is approximately 1% RSD within 1 day and approximately 3% RSD from day to day. The anionic and cationic proteins can be separated over a pH range of 5.5-9, all yielding good efficiencies.

Determination of surfactant concentration using micellar enhanced fluorescence and flow injection titration.

Flow injection titration was used for the determination of anionic, cationic, nonionic and zwitterionic surfactants. The procedure was based on the micellar-enhanced fluorescence of 1,8-anilino-naphthalene sulfonate (ANS). Samples were injected into a carrier stream of phosphate buffer and 1.0 mol l(-1) NaCl. The sample then passed through a mixing chamber which generated the exponential peak shape needed for the titration as well as diluted the sample in the carrier stream to control the pH and ionic strength of the sample. The peak width was linearly related to the logarithm of the surfactant concentration. The minimum detectable concentration was governed by the critical micelle concentration for anionic, zwitterionic and nonionic surfactants, but below the critical micelle concentration for cationic surfactants. The linear range extended for approximately 1.5 orders of magnitude. Reproducibility ranged from 12% at the lower end of the calibration range to 1.1% at higher concentrations. For SDS recoveries of 82-108% were achieved in matrices as concentrated as 1 mol l(-1) in NaCl or Na(2)SO(4).

How to succeed in analytical chemistry: a bibliography of resources from the literature.

Technical excellence is necessary to succeed in a career in analytical chemistry. However there are many other skills necessary for success for which analysts receive no formal training. Talanta has had a tradition of publishing articles on practical aspects of our profession, such as how to write a paper in spectrophotometry or on ion selective electrodes. This article culls the literature for advice on how to purchase equipment, how (and when) to write a manuscript, how to review an article, how to give oral, poster and computer presentations, and how to get a job in analytical chemistry.

Ultrahigh-resolution capillary electrophoretic separation with indirect ultraviolet detection: isotopic separation of [14N]- and [15N]ammonium.

Separation of isotopically labeled [14N]/[15N] ammonium was performed with capillary electrophoresis. This ultrahigh-resolution separation was based on mobility counterbalance with precise control of the anodic electroosmotic flow. Mixtures of zwitterionic surfactant (Rewoteric AM CAS U) and cationic surfactant (cetyltrimethylammonium bromide) were used as buffer additives to modify the electroosmotic mobility. Indirect ultraviolet detection was used with benzyltributylammonium as the buffer coion. Baseline-resolved peaks of [14N]- and [15N]ammonium were obtained within 11 min. The detection limit was 0.01 mM for both [14N]-and [15N]ammonium. Linear calibration in concentration was observed up to 1.0 mM for [15N]ammonium and 2.0 mM for [14N]ammonium. Calibration of the isotopic ratio, [15N]ammonium concentration to total ([14N] and [15N])ammonium, was valid from 5 to 95%.

Factors affecting selectivity of inorganic anions in capillary electrophoresis.

Capillary zone electrophoretic separations of inorganic anions are largely governed by the intrinsic (infinite dilution) mobility of the anion. This in turn is a function of the hydrodynamic friction caused by the size of the ion and the dielectric friction caused by the charge density of the anion re-orienting the surrounding solvent. The influence of these factors on the mobility of anions is examined in both water and nonaqueous solvents. The influence of other experimental parameters, such as ionic strength, ion association, electroosmotic flow modifier concentration, and the addition of complexing agents such as polymeric cations, cyclodextrins, crown ethers and cryptands are also reviewed. From this discussion, some rules of thumb as to when different approaches will be most effective are drawn.

Prediction of electrophoretic mobilities. 3. Effect of ionic strength in capillary zone electrophoresis.

Plots of mobility versus the square root of ionic strength (I(1/2)) do not show the linear behavior predicted by Kohlrausch's law. Classical electrolyte theory states that such deviations are to be expected due to the finite size of the ions. This paper uses the Pitts equation to account for the effect of ionic size on the ionic strength dependence of mobilities in CZE. Experimental mobilities for carboxylates, phenols, and sulfonates of -1 to -6 charge in aqueous buffers ranging from 0.001 to 0.1 M ionic strength were described by µ(-) = µ(0) - Az (I(1/2)/(1 + 2.4I(1/2))), where the constant in the denominator is empirically determined. Infinite dilution mobilities (µ(0)) determined by extrapolation of mobility data to zero ionic strength based on this expression yielded excellent agreement (100.3 ± 3.3%) with literature values for 14 compounds in a variety of buffers. The Pitts equation provides a reasonable estimate of the constant A for solutes up to a charge of -5. However, this constant also depends on temperature and the nature of the buffer counterion, presumably due to ion association. Thus it is most appropriate to determine the constant A empirically for a given buffer system.

Prediction of electrophoretic mobilities. 1. Monoamines.

The mobility of an ion is of fundamental importance in capillary electrophoresis. The size, shape, and other physicochemical parameters of monoamines are determined using molecular modeling. These parameters are used to generate regression expressions to predict absolute (infinite dilution) mobilities. Molecular volume or mass is the strongest determinant of electrophoretic mobility. However, molecular volumes calculated via molecular modeling varied systematically depending on the software used, and so molecular mass is the favored descriptor. Neither the classical spherical (Hückel) nor ellipsoidal (Perrin) models were reasonable predictors of mobility. In accord with empirical expressions, such as the Wilke-Chang equation for diffusion, the absolute mobilities correlate with mass (or volume) to a much greater power than predicted by Stokes's law. Incorporation of the effects of hydration using the McGowan waters of hydration increments further improved the predictions. The best equation for predicting absolute mobilities of monoamines is µ(0) = [(5.55 ± 0.73) × 10(-)(3)]/[W((0.579)(±)(0.026)) + (0.171 ± 0.054)H] where W is the molecular weight and H is the mean waters of hydration calculated using the McGowan increments. The uncertainties are the standard deviations of the parameters. This equation yielded an average prediction error of 4.1% for the data set used to generate the expression (literature absolute mobilities for 34 monoamines possessing no other functional groups), 7.2% for an independent data set from the literature (absolute mobilities for seven monoamines possessing other functional groups), and 3.3% for an experimentally determined data set (13 monoamines determined using capillary electrophoresis).