Biomedical subjects

David D Y Chen

Publications and source records attributed to David D Y Chen.

17 recordsLinked to original sources

PubMed2006

Behavior of interacting species in capillary electrophoresis described by mass transfer equation.

Affinity capillary electrophoresis (ACE) has been used to estimate thermodynamic constants of binding interactions with linear or nonlinear regression methods. The accuracy of this approach relies heavily on the binding interaction mechanism, which is controlled by both the nature of the interaction and the experimental conditions. The development of a highly efficient computer-simulated ACE system makes it possible to demonstrate the detailed behavior of any interacting species of a given interaction under any conditions. The order of the mobilities of the complex and the two binding species in their free forms is a key factor to determine what molecules in what locations of the column are involved in the interaction, and the peak shape resulting from such interactions, of a given ACE experiment. In this paper and the supporting materials, 18 scenarios in 6 different combinations of migration orders of the free analyte, free additive, and complex formed are studied by a computer simulation program based on the mass transfer equation. From the study of these situations, we conclude high additive concentration (ensuring high capacity factor) and low analyte concentration (ensuring fast fill-in of the free additive in the analyte plug) are crucial for obtaining accurate results when using the regression methods. On the other hand, the approach to estimate binding constants with computer simulation can be much more accurate as long as accurate and efficient simulation models can be developed, especially when the ratio of the additive and analyte concentrations is not large enough.

Computer Simulation↗

PubMed2006

Analysis of amphetamine, methamphetamine and methylenedioxy-methamphetamine by micellar capillary electrophoresis using cation-selective exhaustive injection.

Cation-selective exhaustive injection (CSEI) is used as an on-line concentration method for the high-sensitivity analysis of illicit amphetamines using CE. Optimum conditions for the determination of amphetamine, methamphetamine and methylenedioxy-methamphetamine were investigated. Sodium dodecyl sulfate (25 mM) in 100 mM phosphate buffer (pH 2.9) with 20% methanol as organic additive was used as the background electrolyte for CE separation. The LOD, based on an S/N of 3:1, was about 0.01 microg/mL using normal capillary micellar electrokinetic chromatography, while by using CSEI in combination with micellar sweeping the sensitivity increased up to 1000-fold with the LOD lower than 50 pg/mL. The reproducibility of CSEI combined with micellar sweeping for analyzing amphetamines was satisfactory (relative standard deviation around 10% by using area ratios against an internal standard). This method is highly sensitive and can be used to analyze trace amount amphetamines in human hair.

Amphetamine↗

PubMed2005

Enumeration algorithm for determination of binding constants in capillary electrophoresis.

With more accurate simulation models and more efficient algorithms becoming available, the binding constants of an affinity interaction can be obtained from much simpler experiments using capillary electrophoresis. With the enumeration algorithm, all possible combinations of the binding constant and the complex mobility in certain ranges that could result in the experimental migration time of an injected analyte are extracted from a 3-D surface, which depicts the migration times resulting from different values of the binding constant and the mobility of the complex formed between the interacting pair, to form a 2-D curve. When the experimental conditions are changed, the analyte migration time will also change. A new 2-D curve can be constructed from another 3-D surface on the basis of the pairs of binding constants and complex mobility values that could result in the new migration time. Because the true binding constant and complex mobility values have to be the same for both experimental conditions under the same temperature, there has to be a point where both 2-D curves will converge. The coordinates of the converging point give the values for a binding constant and a complex mobility that will fit all 2-D curves generated under certain experimental conditions. p-Nitrophenol is used as the analyte, beta-cyclodextrin is used as the additive, and a one-cell model is used to simulate affinity CE. The experimental conditions that can improve the accuracy of the binding constants are discussed.

Algorithms↗

PubMed2005

General approach to high-efficiency simulation of affinity capillary electrophoresis.

The differential equation describing electrophoretic migration can be evaluated with various finite difference schemes (FDSs). However, the accuracy and efficiency can be dramatically different depending on the FDS chosen and the way the algorithm is implemented in a computer simulation program. The monotonic transport scheme is used as the algorithm for the hyperbolic part of the differential equation, and the first-order fully explicit scheme is used for the parabolic part of the equation. The combination of these algorithms minimizes the errors and maintains high efficiency. A circular arrangement of the cells in the computer's memory is used in the implementation of the algorithms, and the use of concentration thresholds to enable and disable cells along the capillary makes the new algorithm highly efficient. Either thermodynamic or kinetic constants can be used in this program to simulate binding interactions between two species for equilibrium and nonequilibrium affinity CE. Simulation results with various parameters are presented. The simulated peak with proper parameters for an equilibrium affinity CE experiment has shape and position similar to that of the experimental peak. The simulated electropherograms for a nonequilibrium affinity CE experiment also show characteristics of the experimental electropherograms.

Journal Article↗

PubMed2004

Continuous electrophoretic purification of individual analytes from multicomponent mixtures.

Individual analytes can be isolated from multicomponent mixtures and collected in the outlet vial by carrying out electrophoretic purification through a capillary column. Desired analytes are allowed to migrate continuously through the column under the electric field while undesired analytes are confined to the inlet vial by application of a hydrodynamic counter pressure. Using pressure ramping and buffer replenishment techniques, 18% of the total amount present in a bulk sample can be purified when the resolution to the adjacent peak is approximately 3. With a higher resolution, the yield could be further improved. Additionally, by periodically introducing fresh buffer into the sample, changes in pH and conductivity can be mediated, allowing higher purity (>or=99.5%) to be preserved in the collected fractions. With an additional reversed cycle of flow counterbalanced capillary electrophoresis, any individual component in a sample mixture can be purified providing it can be separated in an electrophoresis system.

Journal Article↗

PubMed2004

Determination of shapes and maximums of analyte peaks based on solute mobilities in capillary electrophoresis.

In capillary electrophoresis, the relative orders of mobilities of analyte, additive, and the complex formed determine the analyte peak shape in a way similar to the way the binding isotherms determine the peak shapes in chromatography. The three mobilities allow six possible orders; each produces a characteristic peak shape in CE. Equations describing the analyte migration in a CE system with the presence of mobility-changing additives can be implemented into computer programs to predict the migration times of the analyte peak maximums, and the predicted migration times agree well with the experimental results.

Journal Article↗

PubMed2003

Complementary on-line preconcentration strategies for steroids by capillary electrophoresis.

Complementary on-line preconcentration strategies are needed when analyzing different classes of solutes in real samples by capillary electrophoresis (CE) with UV detection. The performance of three different on-line preconcentration (focusing) techniques under alkaline conditions was examined in terms of their selectivity and sensitivity enhancement for a group of steroids, including classes of androgens, corticosteroids and estrogens. Electrokinetic focusing of large sample injection plugs (up to 28% of effective capillary length or 22.1 cm) directly on-capillary can be tuned for specific classes of steroids based on changes in their mobility (velocity) using a multi-section electrolyte system in CE. A dynamic pH junction was applied for the selective resolution and focusing of weakly acidic estrogens using borate, pH 11.0 and pH 8.0 in the background electrolyte and the sample, respectively. Sweeping, using an anionic bile acid surfactant and neutral gamma-cyclodextrin (gamma-CD) under alkaline conditions (pH 8), resulted in focusing and separation of the moderately hydrophobic (non-ionic) classes of steroids, such as androgen and corticosteroids. Optimal focusing and resolution of all test steroids under a single buffer condition was realized by a dynamic pH junction-sweeping format using borate, pH 11.0 and bile acid surfactant with gamma-CD in the BGE, whereas the sample is devoid of surfactant at pH 8.0. The design of selective on-line focusing strategies in CE is highlighted by the analysis of microgram amounts of ethynyl estradiol derived from a female contraceptive pill extract using the dynamic pH junction method, which resulted in over a 100-fold enhancement in concentration sensitivity.

Contraceptives, Oral↗

PubMed2003

Optimization of cellular nucleotide extraction and sample preparation for nucleotide pool analyses using capillary electrophoresis.

Cell extraction and further sample preparation for nucleotide pool analysis using capillary electrophoresis was faster and simpler using volatile extraction solvents (e.g. organic solvents and de-ionized water) compared to the commonly applied acids dissolved in water (e.g. perchloric acid and trichloracetic acid). Temperature had to be controlled during the whole sample preparation process to prevent degradation, and extracts had to be cleaned from proteins and other large molecules prior to capillary electrophoretic analysis to improve reproducibility. Capillary electrophoresis using borate and cyclodextrins in the background electrolyte was used for determining 11 cellular nucleotides simultaneously. In order to optimize the assay, 0-100% acetonitrile, 0-100% ethanol, and 0-100% methanol in de-ionized water were applied to extract nucleotides from mouse lymphoma cells, and nucleotide yields, recovery, and reproducibility were compared. The assay met the commonly accepted validation limits for biological fluids, if 20-80% acetonitrile in water and 40-60% ethanol in water were used as extraction solvents.

Acyclovir↗

PubMed2003

A quantitative study of continuous flow-counterbalanced capillary electrophoresis for sample purification.

A systematic evaluation of continuous flow-counterbalanced capillary electrophoresis for microscale preparative applications is presented. The success of the technique was found to depend on the specific nature of the analyte and background electrolyte and was most heavily influenced by factors such as solution conductivity and rate of buffer depletion. The performance of the technique for the system studied was ultimately limited by contamination arising from changes in analyte mobilities during extended run times. Marked improvements in both purification rate and yield were observed using the zwitterionic buffer 2-(N-cyclohexylamino)ethanesulfonic acid when compared to similar runs carried out using a borate background electrolyte. A quantitative evaluation of the technique was performed for the analytes studied and the performance of the technique has been compared to fraction collection methods. The highest recovery achieved was 5.2% of the fastest migrating component present in the original sample with a resolution of 9 to the adjacent peak and required 100 min under optimized conditions. Recovery could be further increased to 9.0% in 120 min for the same analyte using pressure-ramped flow-counterbalanced capillary electrophoresis.

Amino Acids↗

PubMed2003

Molecular interaction in capillary electrophoresis.

At the present time, affinity capillary electrophoresis is a well-established method for the measurement of binding constants involving 1:1 interactions. Many recent applications of this technique have illustrated its versatility. If affinity capillary electrophoresis is to be widely applied to the study of all types of interactions it is imperative that we consider that higher order interactions may be present. Applying proper equations or developing new models appropriate to the description of such systems is essential to obtaining reliable binding information.

Artifacts↗

PubMed2002

Proton and hydroxide ion mobility in capillary electrophoresis.

A capillary electrophoresis method for measuring the effective mobilities of proton and hydroxide ions was developed. Photodiode array detection coupled with pH-indicating dyes enabled the detection of proton and hydroxide ions using a conventional CE instrument. The effects of ionic strength and pH of the background electrolyte on the measurement of effective mobility of the proton were first investigated. Mobility measurements were found to be independent of applied voltage. Hydroxide ion mobility was also measured and found to be in agreement with literature values. The effects of pH indicator and proton source on the ion mobility were determined and are described herein.

Journal Article↗

PubMed2002

Affinity capillary electrophoresis using a low-concentration additive with the consideration of relative mobilities.

Most affinity studies in capillary electrophoresis assume that the analyte concentration is much smaller than the additive concentration so that the migration of the analyte has no effect on the concentration of the additive in the capillary. However, in most medium- to high-affinity interactions, the additive concentration has to be kept rather low to observe the changes in analyte mobility before saturation is reached. In this paper, a mathematical model is developed to describe the migration behavior of the analyte in a system where the complex formed becomes concentrated to levels much greater than the original concentration of the additive due to the differences in the mobilities of the analyte, additive, and complex. The analyte is flurbiprofen, the additive is transthyretin, and the stoichiometry of the reaction between the two is 1:2. This study also provides a new algorithm to determine medium- to high-affinity binding interaction constants by CE.

Electrophoresis, Capillary↗

PubMed2002

Applications of on-line weak affinity interactions in free solution capillary electrophoresis.

The impressive selectivity offered by capillary electrophoresis can in some cases be further increased when ligands or additives that engage in weak affinity interactions with one or more of the separated analytes are added to the electrophoresis buffer. This on-line affinity capillary electrophoresis approach is feasible when the migration of complexed molecules is different from the migration of free molecules and when separation conditions are nondenaturing. In this review, we focus on applying weak interactions as tools to enhance the separation of closely related molecules, e.g., drug enantiomers and on using capillary electrophoresis to characterize such interactions quantitatively. We describe the equations for binding isotherms, illustrate how selectivity can be manipulated by varying the additive concentrations, and show how the methods may be used to estimate binding constants. On-line affinity capillary electrophoresis methods are especially valuable for enantiomeric separations and for functional characterization of the contents of biological samples that are only available in minute quantities.

Animals↗

PubMed2002

Accurately describing weak analyte-additive interactions by capillary electrophoresis.

When modeling analyte-additive interactions in capillary electrophoresis (CE), it is necessary to correct for all changes in the apparent electrophoretic mobility of an analyte that are not due to specific binding. Current models based on dynamic complexation have corrected for bulk viscosity changes in the background electrolyte (BGE) when additives are used, while assuming negligible changes in the dielectric constant and other physicochemical properties of the solution. In this report, a study of weak interactions between deoxyribonucleotides and hydroxypropyl-beta-cyclodextrin (HP-beta-CD) revealed significant nonideality in binding isotherms. Changes in the dielectric properties of the solution due to the addition of high concentrations of HP-beta-CD to the BGE was observed to alter the electrophoretic mobility of analytes. A relative dielectric correction factor was required to normalize analyte mobilities to a reference state of zero additive concentration. The use of both a relative dielectric factor and a viscosity correction factor was found to increase the accuracy of the model, reflected by a higher degree of correlation between predicted and measured analyte mobilities. This type of correction is particularly relevant when studying weak analyte binding interactions or when using high concentrations of additive in the BGE. This work is vital for accurate determination of weak binding constants and mobility values, as well as providing a deeper understanding of the fundamental parameters influencing a separation in CE.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed2002

Univalent salts as modifiers in micellar capillary electrophoresis.

The influence of three univalent salts (LiCl, NaCl and RbCl) on the separation of amino acids labelled with 3-(4-carboxybenzoyl)-quinoline-2-carboxaldehyde (CBQCA) in micellar capillary electrophoresis has been studied. Capacity factors for a series of eight CBQCA-labelled amino acids in a sodium dodecyl sulfate (SDS) micellar system containing different concentrations of salt were measured and were found to be related to both the hydrodynamic radius of the salt counter-ion (Li(+), Na(+), Rb(+)) and the relative hydrophobicity of the amino acid. Affinities of the analytes for the micelles were generally observed to decrease as the salt concentration in the background electrolyte was increased from 10 to 50 mM. This decrease in affinity was greatest in the presence of the salt counter-ion with the smallest hydrodynamic radius and is primarily due to an increased resistance to mass transfer. Furthermore, interaction of hydrophobic analytes with the micelles is greater than that of hydrophilic analytes at all salt concentrations due to the greater strength of the hydrophobic interactions and this effect is also enhanced in the presence of a smaller counter-ion. No negative effects due to Joule heating or electromigrative dispersion were observed for low to moderate concentrations of salt, which suggests that the use of simple univalent salts to modify analyte/micelle affinities can be a practical method for improving the separation of complex mixtures.

Amino Acids↗

PubMed2002

Multiple sprayer system for high-throughput electrospray ionization mass spectrometry.

A multiple sprayer electrospray ion source for high-throughput analysis is described. The ion source is comprised of multiple electrospray capillaries, each with an ion lens located near the tip. The electric potentials applied to the ion lenses are used to control the sprayers. The use of ion lenses eliminates the need for mechanical blocking devices to selectively enable or disable the sprayers, and results in a less expensive and more reliable set-up. Sprayers can be enabled or disabled within approximately 50-250 ms when the lens potentials are controlled manually. For simultaneous operation of multiple electrospray capillaries, it is advantageous to orient the capillaries so that the spray from each passes directly in front of the entrance aperture of the mass spectrometer.

Journal Article↗

PubMed2002

An atmospheric pressure ion lens that improves nebulizer assisted electrospray ion sources.

An atmospheric pressure ion lens improves the performance and ease of use of a nebulizer assisted electrospray (ion spray) ion source. The lens is comprised of an oblong-shaped stainless steel ring attached to an external high voltage power supply. The lens is located near the tip of the conductive sprayer, and is maintained at a potential less than that of the sprayer. The ion lens improves the shape of the equipotential lines in the vicinity of the sprayer tip. This lens gives approximately a 2-fold reduction in the signal RSD, a 2-fold increase in the ion signal, an increase in the number of multiply charged ions, and a much broader range of usable sprayer positions.

Journal Article↗