Electrokinetic injection electrophoresis

Injecting the Sample The mechanism by which samples are introduced in capillary electrophoresis is quite different from that used in GC or HPLC. Two types of injection are commonly used hydrodynamic injection and electrokinetic injection. In both cases the capillary tube is filled with buffer solution. One end of the capillary tube is placed in the destination reservoir, and the other is placed in the sample vial. 

Fig. 3.172. Non-aqueous capillary electrophoresis with electrochemical detection of a dye mixture containing (a) 1.7 jUg/ml malachite green, (b) 0.70 jug/ml crystal violet, (c) 4.3 /ig/ml rhodamine B, and (d) 9.1 X 10-6 M ferrocene. Experimental conditions capillary dimensions, 95 cm X 75 pm i.d. running electrolyte, acetonitrile containing 1 M HAc and 10 mM NaAc electrokinetic injection, 20 s 5 kV separation voltage 20 kV applied detection potential, 1.55 V. Reprinted with permission from F.-M. Matysik [206].

J. Palmer, N. J. Munro, and J. P. Landers, A Universal Concept for Stacking Neutral Analytes in Micellar Capillary Electrophoresis, Anal. Chem. 1999, 71, 1679 J. Palmer, D. S. Burji, N. J. Munro, and J. P. Landers, Electrokinetic Injection for Stacking Neutral Analytes in Capillary and Microchip Electrophoresis, Anal. Chem. 2001, 73, 725 J. P. Quirino, S. Terabe, and... 

Figure 9.7 Schematic illustration of the flow-gating interface. A channeled Teflon gasket was sandwiched between two stainless steel plates to allow for flow into the electrophoresis capillary, either from the flush buffer reservoir or from the LC microcolumn during an electrokinetic injection.

Eberle et al. [134] separated the enantiomers of omeprazole and structurally related drugs by capillary zone electrophoresis with bovine serum albumin as chiral selector. The separations were carried out on a fused silica column (60 cm x 50 pm, 50 cm to detector) with a buffer consisting of 100-/zM-bovine serum albumin and 7% 1-propanol in 10 mM potassium phosphate pH 7.4. Electrokinetic injection was at 5-8 kV for 7 s. An applied voltage of 300 V/cm was used. Detection was at 290 nm. Detection limits were 0.04 mg/ml for the analytes studied. 

Palmer, J., Burgi, D.S., Munro, N.J., Landers, J.P., Electrokinetic injection for stacking neutral analytes in capillary and microchip electrophoresis. Anal. Chem. 

Capillary electrophoresis separations are dependent on the relative mobilities of analytes under the influence of an electric field and do not depend on mobile phase/stationary phase interactions. A fused silica capillary is filled with a buffer and both ends submerged into two reservoirs of the buffer. A platinum electrode is immersed in each reservoir and a potential difference (5-30 kV) is applied across the electrode. An aliquot of sample of a few nanoliters is injected onto the capillary by either hydrostatic or electrokinetic injection, and the components migrate to the negative electrode. Separations of analytes arise from differences in the electrophoretic mobilities, which are dependent on the mass-to-charge ratio of the components, physical size of the analyte, and buffer/analyte interactions. An electro-osmotic flow (EOF) of the buffer occurs in the capillary and arises as a result of interactions of the buffer with dissociated functional groups on the surface of the capillary. Positive ions from the buffer solution are attracted to negative ions... 

Kappes et al. evaluated the potentiometric detection of acetylcholine and other neurotransmitters through capillary electrophoresis [209]. Experiments were performed on an in-house capillary electrophoresis instrument that made use of detection at a platinum wire, dip-coated in 3.4% potassium tetrakis (4-chlorophenyl) borate/64.4% o-nitrohenyl octyl ether/32.2% PVC in THF. The results were compared to those obtained using capillary electrophoresis with amperometric detection at a graphite electrode. Samples prepared in the capillary electrophoresis buffer were electrokinetically injected (7 s at 5 kV) into an untreated fused silica capillary (88 cm x 25 pm i.d.) and separated with 20mM tartaric acid adjusted to pH 3 with MgO as the running buffer. The system used an applied potential of 30 kV, and detection versus the capillary electrophoresis ground electrode. 

There are two modes of injection in capillary electrophoresis hydrodynamic injection and electroki-netic injection. In hydrodynamic injection, pressure or vacuum are plaeed on the inlet sample vial or the outlet waste vial, respeetively. For electrokinetic injection, the voltage is activated for a short time with the capillary and electrode immersed in the sample. 

Capillary electrophoresis is conducted in capillaries filled with an electrolyte solution. Buffered electrolytes are generally used, since biomolecule mobilities and electroosmotic flow (EOF) are sensitive to pH. The ends of the filled capillaries are placed in electrolyte reservoirs that contain electrodes, and the electrodes are positioned so that electrolysis products do not enter the capillary. A small plug of solution containing the analytes to be separated is pressure- or electrokinetically-injected into one end of the capillary, and a voltage difference is applied to the electrodes such that the analytes of interest migrate toward the other end of the capillary, where they are detected. Analytes with different electrophoretic mobilities migrate at different speeds and become separated as they transverse the capillary. 

For electrokinetic injection a low voltage is applied for a brief time with the inlet of the separation column located in the sample vial. Sample enters the column by the combined effect of electroosmosis and electrophoresis. If it is assumed that the conductivity of the sample solution and background electrolyte is equal, then the length of the injected zone, Ljnj, is given by... 

Stuart, J.N. et al.. Spurious serotonin dimer formation using electrokinetic injection in capillary electrophoresis from small volume biological samples. Analyst, 130, 147, 2005. 

Li and coworkers described the separation of FITC and FllC-labeled antihuman IgG by zone electrophoresis in a glass microchip. The two compounds were electrokinetically injected into the separation channel for 2 s and separated in less than 12 s. A comparison between the separation performance in capillary and microchip showed that higher efficiency was achieved in the microchip format due to the shorter separation length and the higher electric field that were applied in microfluidic structures. 

Hirokawa, T., Okamoto, H., and Gas, B., High-sensitive capillary zone electrophoresis analysis by electrokinetic injection with transient isotachophoreticpreconcentration Electrokinetic supercharging. Electrophoresis, 24,498, 2003. 

Marsh, D.B., and Nutall, K.L. Serum methylmalonic acid by capillary zone electrophoresis using electrokinetic injection and indirect photometric detection. J. Capillary Electmphor, 2, 63,1995. 

A. Caspar and E. Dudds, Application of internal universal cahbration for determination of fuUy dissociated species in capillary electrophoresis using indirect UV detection and electrokinetic injection,... 

Xu, Z. Q., Hirowaka, T., Nishine, T., and Arai, A., High-sensitivity capillary gel electrophoretic analysis of DNAfragments on an electrophoresis microchip using electrokinetic injection with transient isotachophoretic preconcentration. Journal of Chromatography, 990, 53-61, 2003. 

Tsai CH, Yang RJ, Tai CH, Fu LM (2005) Numerical simulation of electrokinetic injection techniques in capillary electrophoresis microchips. Electrophoresis 26 674-686... 

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