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1.
Summary Although capillary electrophoresis (CE) methods have been included in some regulatory submissions, the formal requirements
for validation of such methods are not well-defined or universally understood. This paper provides a general overview of the
aspects of method performance which need to be considered during the validation of CE methods particularly emphasising areas
of similarity with, and differences between, HPLC and CE. The individual aspects of method performance are reviewed and guidance
given, with illustrative literature examples where possible, on how to carry out suitable validation studies. Performance
criteria are also provided, wherever possible. In this way, a suggested validation approach for CE methods is presented, reflecting
the similarity with that generally adopted for HPLC methods, but accommodating those aspects which are different for, or peculiar
to, CE. 相似文献
2.
M. Sc. Christian Benz Dr. Michael Boomhoff M. Sc. Johannes Appun Prof. Dr. Christoph Schneider Prof. Dr. Detlev Belder 《Angewandte Chemie (International ed. in English)》2015,54(9):2766-2770
Free‐flow electrophoresis is an ideal tool for preparative separations in continuous microflow. With the approach presented herein for coupling free‐flow electrophoresis and mass spectrometry it is now also possible to trace non‐fluorescent compounds and identify them by means of mass spectrometry. The functionality of the method and its potential as an integrated separation unit for microflow synthesis is demonstrated by application to a multicomponent [3+2]‐cycloannulation. 相似文献
3.
It is common to operate equilibrium-based separation methods, such as distillation and extraction, as multistage unit operations, in which equilibrium is presumably achieved within each stage. Two rate-based separation processes, free electrophoresis and magnetic particle separation, have now been operated in multistage mode. Preparative free electrophoresis of particles and solutes has resisted scale-up and is confined to a narrow range of ionic compositions. Natural convection induced in electrophoresis buffers by Ohmic heating has been a strong deterrent and has led to such measures as radial electrophoresis in Couette flow, free-flow electrophoresis, low-gravity electrophoresis, density gradient electrophoresis, and reorienting density gradient electrophoresis, to name a few. The short vertical electrophoresis path exploited in the last-mentioned forms the basis for multistage electrophoresis. A thin-layer countercurrent distribution apparatus was designed and constructed so that up to 20 fractions could be collected on the basis of electrophoretic mobility by applying an electric field. The mixture to be separated starts in a bottom cavity, and successive top cavities collect fractions as separand particles or molecules are electrophoresed upward out of the bottom cavity. Mathematical models of this process were developed, and experiments were performed to verify the predictions of the models by collecting and counting particles in each cavity after fractionation. 相似文献
4.
Nonlinear and nonequilibrium electrophoresis of spherical particles of radius a is shown to be possible when the solid surface allows field or current penetration. At low particle Peclet numbers, transient capacitative charging occurs until the surface polarization completely screens the external field. For a DC applied field [see text], the resulting electrokinetic velocity reaches Dukhin's maximum value of [formula: see text], where [see text] and mu are the liquid permittivity and viscosity. At high Peclet numbers, electroosmotic convection of the electroneutral bulk stops the transient charging before complete field-line exclusion. For an ion-selective and conducting spherical granule, the polarization is then determined by the steady-state Ohmic current driven by the penetrated external field. The high-Peclet electrokinetic velocity is lower, diffusivity-dependent and scales as [see text]. 相似文献
5.
Miniaturization in carbohydrate analysis 总被引:1,自引:0,他引:1
Recent progress of microchip electrophoresis (ME) of carbohydrates is overviewed. Carbohydrate analysis by ME encounters difficulties such as lack of electric charge and deficiency of a chromophore/fluorophore in analyte molecules, however, it benefits from the accumulated knowledge of capillary electrophoresis (CE) and rapid separation of simple sugars also by ME, with high column efficiency comparable to CE, has become possible. Analysis at high pH, with electrochemical detection, is a promising approach because carbohydrates can be ionized by weak dissociation of the hydroxyl groups and the in situ formed ionic species can be effectively separated by the zone electrophoresis mode. The separated species can be sensitively monitored by electrochemical detection on a gold or copper electrode. Ionization as borate complexes and refractometric detection is also possible, though sensitivity is lower. Introduction of UV-absorbing or fluorescent tags is potentially useful but the time-consuming derivatization processes sacrifice the rapidity of ME. Examples of ME of carbohydrates as 1-phenyl-3-methyl-5-pyrazolone (PMP; for simple mono- and oligosaccharides with UV detection), 8-aminopyrene-1,3,6-trisulfonate (APTS; for oligosaccharides ladders with LIF detection), and 4-nitro-2,1,3-benzoxadiazole (NBD-F; for amino sugars and aminoalditols with LIF detection) derivatives are presented, with details of the analytical conditions. Since ME in a short separation channel enables rapid analysis within 1 min, it presents an ideal tool for clinical analysis, as shown in a few papers reporting protocols for specific blood glucose assay. Finally, the usefulness of microfluidic reactors and microarrays for enzyme-assisted carbohydrate analysis as well as glycan profiling is pointed out. 相似文献
6.
Schultz J Gottlieb DM Petersen M Nesic L Jacobsen S Søndergaard I 《Electrophoresis》2004,25(3):502-511
Methods for classification of two-dimensional (2-DE) electrophoresis gels based on multivariate data analysis are demonstrated. Two-dimensional gels of ten wheat varieties are analyzed and it is demonstrated how to classify the wheat varieties in two qualities and a method for initial screening of gels is presented. First, an approach is demonstrated in which no prior knowledge of the separated proteins is used. Alignment of the gels followed by a simple transformation of data makes it possible to analyze the gels in an automated explorative manner by principal component analysis, to determine if the gels should be further analyzed. A more detailed approach is done by analyzing spot volume lists by principal components analysis and partial least square regression. The use of spot volume data offers a mean to investigate the spot pattern and link the classified protein patterns to distinct spots on the gels for further investigation. The explorative approach in analysis of 2-D gels makes it possible, in a fast and convenient way, to screen many gels in order to determine the protein patterns that form clusters and could be selected for further examination. 相似文献
7.
Collision efficiencies are considered for colloidal suspensions of solid spheres moving in a viscous fluid under the influence of electrophoresis or gravity, Brownian motion, and electrostatic and van der Waals forces. The results are compared to those for convection (electrophoresis or gravity) and diffusion (Brownian motion) acting independently. The collision efficiency increases by many orders of magnitude over that predicted by simply adding diffusive and convective efficiencies in a specific parameter regime. This regime occurs when there is a large energy barrier in the interparticle potential, causing a stable region of parameter space if there is no diffusion. Brownian motion alone will only cause small amounts of aggregation under these conditions. However, for electric fields or buoyancy effects which are only slightly too weak to allow particles to overcome the potential barrier, the addition of weak Brownian motion to a system with convection can cause significant numbers of particles to overcome the energy barrier and aggregate. Copyright 2000 Academic Press. 相似文献
8.
Heinz Engelhardt Wolfgang Beck Jrg Kohr Thomas Schmitt 《Angewandte Chemie (International ed. in English)》1993,32(5):629-649
Capillary electrophoresis is a new analytical technique that has seen a great upswing in recent years, because this adaptation of electrophoresis may be automated and simple, direct quantification is possible. As in conventional electrophoresis, samples with a large range of molecular weights, from inorganic ions to biopolymers such as DNA and proteins, can be separated. Uncharged molecules can also be separated when micelle-forming detergents are added to the buffer; the distribution mechanism that then comes into play increases the separation efficiency of this micellar electrokinetic chromatography. Chiral additives such as cyclodextrins make the separation of enantiomers possible. The rapid increase in the popularity of capillary electrophoresis is reflected in instrument sales and the growing number of scientific publications dealing with the method, and it seems that the future of this technique is assured. 相似文献
9.
Continuous flow zone electrophoresis is an efficient method for the non-destructive separation of biological materials in a flowing film of buffer solution in which the constituents of a sample are separated according to their electrophoretic mobilities, under the influence of an electric field. This paper deals with modeling of flow structure of the buffer solution taking into account the effect of electroosmosis, Joule heating and thermal free convection and the forced convection of axial flow. Modeling of diffusion is examined in a second part. The modeling equations and the corresponding boundary conditions are solved by finite difference methods. The various parameters affecting the quality of the fractionation are analyzed. 相似文献
10.
E. De Lorenzi G. Massolini D. K. Lloyd H. L. Monaco C. Galbusera G. Caccialanza 《Journal of chromatography. A》1997,790(1-2):47-64
A new chiral stationary phase for high-performance liquid chromatography of quail egg white riboflavin binding protein is presented. Several chiral acidic, basic and uncharged drugs were analysed and the influence of the mobile phase's parameters on the retention times and enantioselectivity was evaluated. On the basis of the results obtained, the same protein was studied as a background electrolyte additive in free solution capillary electrophoresis, in order to evaluate if capillary electrophoresis (CE) could be used as a rapid scouting technique for screening the enantioselectivity of novel proteins without immobilisation on a solid support. To investigate if it is possible to directly compare the results obtained by each technique, the CE experiments were planned on the basis of both the findings and ideas originated in liquid chromatography. 相似文献
11.
《Journal of membrane science》1998,142(2):205-212
A model of pressure-driven membrane process of electrolyte separation is presented. The electric field potential assumed as being known, exact solution for permeate composition is readily obtained. All species are assumed to have the same convection velocity. Local electroneutrality condition is not used. The electric potential has been taken into account under high temperature approximation, thus reducing the problem to algebraic equation in exp(Ψ), where Ψ is dimensionless flow potential, and making it possible to calculate concentrations of ions in permeate. Negative retention is shown to be possible for one-component electrolyte solution. For electrolyte mixtures, concentration of ion with high charge is shown to “govern” the membrane selectivity in respect to low-charge ions. Results obtained are in qualitative accordance with the earlier experimental data on membrane separation of reaction mixtures in homogeneous catalysis. 相似文献
12.
An exact analysis is presented for the diffusiophoresis and electrophoresis of a rigid elliptic cylindrical particle in a uniform applied field oriented arbitrarily with respect to its axis. The range of the interaction between the solute species and the particle surface is assumed to be small relative to the minimum dimension of the particle, but the effect of polarization of the diffuse species in the thin particle-solute interaction layer is incorporated. To solve the conservative equations governing the system, a slip velocity of fluid and normal fluxes of solute species at the outer edge of the thin diffuse layer which balance convection and diffusion of the solute species along the particle surface are used as the boundary conditions for the fluid domain outside the diffuse layer. Expressions for the migration velocity of the particle are obtained in closed forms for the cases of diffusiophoresis in a nonionic solute concentration gradient, diffusiophoresis in a concentration gradient of symmetric electrolyte, and electrophoresis in an external electric field. An interesting feature is found that the diffusiophoretic or electrophoretic velocity of the particle decreases with the reduction of the maximum length of the particle in the direction of migration. Also, the average migration velocity for an ensemble of identical, noninteracting elliptic cylinders with random orientation distribution is obtained for each case considered. 相似文献
13.
《Journal of separation science》2018,41(1):323-335
Enzymes play an essential role in many aspects of pharmaceutical research as drug targets, drug metabolizers, enzyme drugs and more. In this specific field, enzyme assays are required to meet a number of specific requirements, such as low cost, easy automation, and high reliability. The integration of an immobilized‐enzyme reactor to capillary electrophoresis represents a unique approach to fulfilling these criteria by combining the benefits of enzyme immobilization, that is, increased stability and repeated use, as well as the minute sample consumption, short analysis time, and efficient analysis provided by capillary electrophoresis. In this review, we summarize, analyze, and discuss published works where pharmaceutically relevant enzymes were used to prepare capillary electrophoresis‐integrated immobilized‐enzyme reactors in an online manner. The presented assays are divided into three distinct groups based on the drug–enzyme relationship. The first, more extensively studied group employs enzymes that are considered to be therapeutic targets, the second group of assays present tools to assess drug metabolism and the third group assesses enzyme drugs. Furthermore, we examine various methods of enzyme immobilization and their implications for assay properties. 相似文献
14.
On the Criteria of Instability for Electrochemical Systems 总被引:7,自引:0,他引:7
Both cyclic-voltammetry-based and impedance-based experimental criteria that have been developed recently for the oscillatory electrochemical systems are critically appraised with two typical categories of oscillators.Consistent conclusions can be drawn by the two criteria for the category of oscillators that involve the coupling of charge transfer mainly with surface steps(e.g.ad-and desorption)such as in the electrooxidation of C1 organic molecules.Whereas,impedance-based criterion is not applicable to the category of oscillators that involve the coupling of charge transfer mainly with mass transfer(e.g.diffusion and convection) such as in the Fe(CN)6^3- reduction accompanying periodic hydrogen evolution.The reason is that the negative impedance cannot include the feedback information of convection mass transfer induced by the hydrogen evolution.However,both positive and negative nonlinear feedbacks,i.e., the diffusion-limited depletion and convection-enhanced replenishment of the Fe(CN)6^3- surface concentration,that coexist between the bistability,i.e.,Fe(CN)6^3- reduction with and without hydrogen evolution at lower and higher potential sides respectively,are all reflected in the crossed cyclic voltammogram(CCV).It can be concluded that the voltammetry-based criterion(in time domain)is more intuitive,less time-consuming and has a wider range of applications than the impedancebased one (in frequency domain). 相似文献
15.
An analytical study is presented for the diffusiophoresis and electrophoresis of a rigid nonconducting spheroid in a uniform applied field that is oriented arbitrarily with respect to its axis of revolution. The range of the interaction between the solute species and the particle surface is assumed to be small relative to the dimension of the particle, but the effect of polarization of the diffuse species in the thin solute-particle interaction layer is incorporated. To solve the conservative equations governing the system, a slip velocity of fluid and normal fluxes of solute species at the outer edge of the thin diffuse layer which balance convection and diffusion of the solute species along the particle surface are used as the boundary conditions for the fluid domain outside the diffuse layer. Explicit expressions for the migration velocity of the spheroidal particle are obtained for the cases of diffusiophoresis in a nonionic solute concentration gradient, diffusiophoresis in a concentration gradient of symmetric electrolyte, and electrophoresis in an external electric field. An interesting feature is found that the diffusiophoretic or electrophoretic velocity of the particle decreases with the reduction of the maximum length of the particle in the direction of migration. Also, the average migration velocity for an ensemble of identical, non-interacting spheroids with random orientation distribution is obtained for each case considered. 相似文献
16.
Role of Joule heating in dispersive mixing effects in electrophoretic cells: convective-diffusive transport aspects 总被引:1,自引:0,他引:1
This contribution addresses the problem of solute dispersion in a free convection electrophoretic cell for the batch mode of operation, caused by the Joule heating generation. The problem is analyzed by using the two-problem approach originally proposed by Bosse and Arce (Electrophoresis 2000, 21, 1018-1025). The approach identifies the carrier fluid problem and the solute problem. This contribution is focused on the latter. The strategy uses a sequential coupling between the energy, momentum and mass conservation equations and, based on geometrical and physical assumptions for the system, leads to the derivation of analytical temperature and velocity profiles inside the cell. These results are subsequently used in the derivation of the effective dispersion coefficient for the cell by using the method of area averaging. The result shows the first design equation that relates the Joule heating effect directly to the solute dispersion in the cell. Some illustrative results are presented and discussed and their implication to the operation and design of the device is addressed. Due to the assumptions made, the equation may be viewed as an upper boundary for applications such as free flow electrophoresis. 相似文献
17.
Capillary electrophoresis (CE) is a well-established and frequently used technique in the pharmaceutical industry. Therefore an appropriate analytical instrument qualification (AIQ) is required for quality assurance. AIQ forms the basis of a quality management followed by analytical method validation, system suitability tests (SSTs) and quality control checks. Two parts of the AIQ, namely the operational qualification (OQ) and the performance qualification (PQ) are of particular interest in the daily routine of the laboratory. A new concept for OQ and PQ was developed to assure the correct function of a CE system. The significance of each parameter, possible test methods as well as acceptance criteria will be presented and discussed in detail. Especially temperature adjustment by the cooling system and the voltage supply must be tested for accurate and precise operation. The detector noise, wavelength accuracy and detector linearity have to be checked as well. Finally, the injection linearity, accuracy and precision need to be qualified. The proposed set of qualification procedures is easy to implement and was already tested on five CE instruments from three different manufacturers. A time- and cost-saving continuous PQ was derived, using results from method-specific SSTs and some additional experiments. This holistic concept continuously surveys the most relevant parameters, hence assuring the suitability of the used instruments and decreasing their downtimes. 相似文献
18.
《Journal of separation science》2018,41(11):2473-2478
Capillary electrophoresis connected to electrospray ionization mass spectrometry is a promising combination to analyze complex biological samples. The use of sheathless electrospray ionization interfaces, such as a porous nanoelectrospray capillary emitter, requires the application of forward flow (either by pressure or electroosmosis) to maintain the electrospray process. The analysis of solute molecules with strong negative charges (e.g., aminopyrenetrisulfonate labeled glycans) necessitates a reversed‐polarity capillary electrophoresis separation mode, in which case the electroosmotic flow is counter current, thus pressure assistance is necessary. In this study, we compared the effect of forced convection with and without counter electroosmotic flow on the resulting separation efficiency in capillary electrophoresis based on flow profile simulations by computational fluid dynamics technique and by actual experiments. The efficiencies of the detected peaks were calculated from the resulting electropherograms and found approximately 950 000 plates/m for electrophoresis with counter electroosmotic flow, 20 000 plates/m with pressure only (such as would be in open tubular liquid chromatography), and 480 000 plates/m for electrophoresis with simultaneous counter electroosmotic flow and forward pressure assistance, which validates the simulation data. 相似文献
19.
20.
Is capillary electrophoresis on microchip devices able to genotype uridine diphosphate glucuronosyltransferase 1A1 TATA‐box polymorphisms?
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Angelo Minucci Giulia Canu Maria De Bonis Elisabetta Delibato Ettore Capoluongo 《Journal of separation science》2014,37(12):1521-1523
In this commentary, we focused our attention on capillary electrophoresis. It achieves the efficient separation of molecular species by the application of high voltages to samples in solution. Actually, capillary electrophoresis can be performed on microchip devices, based on an automated and miniaturized electrophoresis system, based on lab‐on‐a‐chip technology. By this technology it is possible to separate nucleic acid fragments (DNA or RNA) with respect to sizing accuracy and sizing resolution. Currently, two automated capillary electrophoresis on microchips devices are available: the Agilent 2100 Bioanalyzer and the Experion? Automated Electrophoresis System. In this study, we evaluated if the CE is able to distinguish the three uridine diphosphate glucuronosyltransferase 1A1 TATA‐box genotypes. 相似文献