Capillary zone electrophoretic separation of peptides and proteins using low pH buffers in modified silica capillaries.

Microfabricated electrophoretic separation devices have been produced by an injection-molding process. The strategy for producing the devices involved solution-phase etching of a master template on a silicon wafer, followed by electroforming more durable injection-molding masters in nickel from the silicon master. One of the nickel electroforms was then used to prepare an injection mold insert, from which microchannel chips in an acrylic substrate were mass-produced. The microchannel devices were used to demonstrate high-resolution separations of double-stranded DNA fragments with total run times of less than 3 min. Run-to-run and chip-to-chip reproducibility was good, with relative standard deviation values below 1% for the run-to-run data and in the range of 2-3% for the chip-to-chip comparisons. Such devices could lead to the production of low-cost, single-use electrophoretic chips suitable for a variety of separation applications, including DNA sizing, DNA sequencing, random primary library screening, and rapid immunoassay testing.

Microchannel electrophoretic separations of DNA in injection-molded plastic substrates

In electrochromatography, electrolyte is driven along a narrow bore chromatographic column at typical HPLC linear velocities by applying a potential gradient of around 50,000 V m−1. Contrasting with pressure driven LC, the tube bore is limited to not more than 200 µm by self heating. Thus miniaturisation is mandatory in electrochromatography whereas it is optional in pressure driven LC unless very small particles (e.g. 1 µm diameter) are used. Because the velocity profile in open tubular electrochromatography is close to that of perfect plug flow, contrasting with the parabolic flow profile in pressure driven LC, open tube electrochromatography provides plate efficiencies limited only by axial diffusion, and comparable to those obtainable in open tubular gas chromatography. Contrasting with pressure driven open tubular LC there is no requirement for the tube bore to be very small (e.g. <10 µm). Although pure electrophoresis is applicable only to ionised solutes, and is not strictly a chromatographic method, neutral species can be chromatographed by partitioning them between two phases which move at different rates, for example by using micellar solutions or colloidal sols as the moving fluid, thereby retaining the high plate efficiency of electrophoresis. With packed tubes electrochromatographic separations of high efficiency can be obtained with 5 µm particles and extremely high efficiencies should be obtainable without sacrifice of eluent velocity by using submicron particles. In principle, electrochromatographic methods can provide a wide range of partitioning methods of extremely high plate efficiency, and will, in future, undoubtedly rival conventional HPLC.

Miniaturisation in pressure and electroendosmotically driven liquid chromatography: Some theoretical considerations

Capillary zone electrophoresis (CZE) is attracting much attention (1-4) as a new separation technique that can complement high-performance liquid chromatography (HPLC). In CZE a migration channel of capillary dimensions is filled with electrolyte; a sample to be analyzed is injected at one end of the channel; and a high voltage is applied across the channel. When the electrolyte contacts the walls of the capillary, the inner surface of the capillary becomes charged, either through the ionization of surface groups on the capillary walls or through the adsorption of charged species from the electrolyte onto the inner surface. In either case, the electrolyte inside the capillary is no longer electroneutral but acquires a net charge, which may be positive or negative. Under the action of the applied electric field, the electrolyte moves toward one end of the capillary, and this movement is referred to as electroosmotic (electroendosmotic) flow. In addition to the bulk flow of the electrolyte, electrophoresis also takes place; that is, the applied electric field exerts a force on positively charged species to cause them to move to the negatively charged electrode and on negatively charged species to cause them to move to the positively charged electrode. As a result, the components in the injected sample separate into distinct zones, based on their different mobilities. However, in many cases, the rate of electrophoretic flow is typically less than the rate of electroosmotic flow. Consequently, species in the injected sample move in one direction-the direction of the electroosmotic flow-and thus the different species can be detected as each zone passes through some suitable detector located downstream from the capillary inlet. Clearly, a precise characterization of the electroosmotic flow is highly desirable not only for understanding CZE but also for optimizing the operation of CZE in analyzing a given sample. One way to measure the electroosmotic flow rate is to record the elution time of an injected uncharged marker solute, which will be carried through the capillary under the action of only electroosmotic flow (4-6). For this purpose it is necessary that the marker solute be truly neutral, that it have negligible interaction with the capillary walls, and that it be readily detected. Another way is to weigh the mass of electrolyte transferred from the capillary inlet to the capillary outlet over a timed interval (7). For this purpose, losses caused by evaporation must be eliminated and the use of a digital balance appears to be recommended. Both of these procedures have been demonstrated to give reliable measurements of the electroosmotic flow rate, provided that some care is taken. We describe here what we believe might be a simpler procedure for measuring the electroosmotic flow rate. I t is based on recording the time history of the current during CZE operation. Thus the new method requires no special type of injected solute or detector, and it can be used by anyone carrying out CZE separations.

Current-monitoring method for measuring the electroosmotic flow rate in capillary zone electrophoresis

The biological inorganic chemistry of zinc ions.

Open-tubular microcapillary liquid chromatography with electro-osmosis flow using a UV detector

Separation of organic and metal ions by high-voltage capillary electrophoresis

In the determination of stability constants of metal complexes, usually the measurements of the activities of free metal ions at very low levels are required. The electrode response of an Orion Model 94–29 copper(II) ion selective electrode at very low activity levels was studied in the various metal buffers composed of copper(II) ions and an excess of ligand. Usually the linear calibration range of the copper(II) ion selective electrode was extended to very low activity levels in the presence of a strong complexing agent. The stability constants of copper(II) complexes with acetate, ammonia, ethylenediamine, glycine, iminodiacetic acid, 1,10-phenanthroline, etc. were determined successfully. In the cases of EDTA and NTA, however, the lower limit of the linear calibration range was abnormally high and the stability constant of copper–EDTA or –NTA chelate could not be obtained. Some experimental results on the potentiometric titration of copper(II) ion with EDTA were also presented.

Genkichi Nakagawa

Papers

Open-tubular microcapillary liquid chromatography with electro-osmosis flow using a UV detector

Separation of organic and metal ions by high-voltage capillary electrophoresis

Use of the Copper(II)-Selective Electrode for the Determination of the Stability Constants of Copper(II) Complexes

Sample dispersion with chemical reaction in a flow-injection system

Open-tubular microcapillary liquid chromatography with 30–40 μm I.D. columns