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A simple and convenient mode—moving chemical reaction boundary method–capillary zone electrophoresis (MCRBM–CZE)—was designed for the enhancement of separating efficiency of CZE. In this mode, the transient MCRBM is used for the on-line pre-treatment of sample. By analyses of tryptophan(Trp) and phenylalanine(Phe) as an example, the experiments by MCRBM–CZE were carried out and further compared with those by normal CZE without the transient MCRBM. The results reveal that by carefully selected appropriate electrolytes, a strong condensation effect can be achieved by using MCRBM–CZE; this effect can greatly improve the separation efficiency, resolution and peak height of Trp and Phe in CZE as compared with those of normal CZE of Trp and Phe. Even if the sample comprises high concentrations of salt, such as 80 mM NaCl(concentration of sodium ion up to 145.6 mM), the same condensation effect can also been observed; this implies obvious significance for biological samples like urine and serum. However, if the electrolytes was chosen inappropriately only a poor compression effect of sample was observed in the MCRBM–CZE runs. [Copyright &y& Elsevier]

We investigated several factors, such as temperature, current intensity (i), time (t) and the product (mA min mm−2, viz., C mm−2) of i and t, etc., that obviously affect the moving neutralization reaction boundary method (MNRBM). The results manifest that the temperature and the product ti have a strong influence on the movement rate of the boundary. The data prove that about 0.6 C mm−2 (being equivalent to 10 mA min mm−2) is a critical point. If the product ti is lower than the critical point, a good quantitative agreement exists between the observed and theoretical values, but if it is higher than the critical point, the agreements are poor. The optimized experimental conditions are: (1) 18–20 °C room temperature, (2) 0.6–0.8 mA mm−2, (3) less than 10 mA min mm−2, (4) 1% agarose gel, (5) daily prepared solution and gel containing NaOH. The optimized MNRBM is of benefit for the studies on MNRB itself, isoelectric focusing and capillary zone electrophoresis as will be partially shown in this paper. [Copyright &y& Elsevier]

The traditional recycling free-flow isoelectric focusing (RFFIEF) suffered from complex structure, tedious operations and poor extensibility as well as high cost. To address these issues, a novel reciprocating free-flow isoelectric focusing device (ReFFIEF) was developed for proteins or peptides pre-fractionation. In the new device, a reciprocating background flow was for the first time introduced into free flow electrophoresis (FFE) system. The gas cushion injector (GCI) used in the previous continuous free-flow electrophoresis (CFFE) was redesigned for the reciprocating background flow. With the GCI, the reciprocating background flow could be achieved between the GCI, separation chamber and transient self-balance collector (tSBC). In a run, process fluid flowed to and from, forming a stable reciprocating fluid flow in the separation chamber. A pH gradient was created within the separation chamber, and at the same time proteins were focused repeatedly when passing through the chamber under perpendicular electric field. The ReFFIEF procedure was optimized for fractionations of three model proteins, and the optimized method was further used for pre-fractionation of model human serum samples. As compared with the traditional RFFIEF devices developed about 25 years ago, the new ReFFIEF system showed several merits, such as simple design and structure, user-friendly operation and easy to extend as well as low cost. [ABSTRACT FROM AUTHOR]

Herein, a novel strategy was developed to separate and prepare target protein from complex sample by free-flow electrophoresis (FFE), which mainly based on the charge-to-mass ratio ( C / M ) analysis of proteins. The C / M values of three model proteins, namely Cytochrome C (Cyt C), myoglobin (Mb) and bovine serum albumin (BSA) were analyzed under different pH and the separation of these proteins was predicted by CLC Protein Workbench software. Series of experiments were performed to validate the proposed method. The obtained data showed high accordance with our prediction. In addition, the chamber buffer (CB) of FFE system was optimized to improve the resolution of separation. Meanwhile, in order to evaluate the analytical performance of the proposed method, Cyt C was extracted from swine heart and further separated by FFE based on C / M analysis. Results showed that Cyt C was completely separated from the crude sample and a purity of 96.9% was achieved. The activity of prepared Cyt C was 98.3%, which indicate that the proposed method is promising in a wide variety of research areas where the native properties of proteins should be maintained for downstream analysis. [ABSTRACT FROM AUTHOR]

Highlights: [•] Thermoelectric coolers and ceramic plate combined as temperature control system for stable long-term run of FFE for the first time. [•] Simple fastening structure designed for easy assembly of FFE device with good anti-leakage ability within 3min. [•] Simple and rapid method for exhausting air bubble in FFE within 5s. [•] An electroconductive membrane with about 90% efficiency of voltage usage in FFE. [ABSTRACT FROM AUTHOR]

Abstract: In this paper, the controllable band width, resolution and sample loading were investigated by the injection system of free-flow electrophoresis (FFE) with gratis gravity. Two general injection methods were described herein. The first method was the one in which sample injection fluxes were variable with constant background flux, while the second was the one in which the background fluxes were flexible with stable sample flux. With methyl green and crystal violet as two viewable model compounds, a series of experiments were performed, and the experimental results revealed that (1) the sample band width could be under desiring control through the regulation of ratios between sample and background fluxes, (2) the separative resolution could be also adjusted elaborately via the regulation of flux ratios during the separation of methyl green and crystal violet with only one charge disparity, and (3) the sample loading could be conveniently controlled via the flux ratios and an approximate maximum sample loading could be selected under the condition of just completed separation of two adjoining solutes. In addition, it was observed that the flux ratio had soft influence on the separative resolution of two solutes. These results were of significance to the designs on band width, resolution and sample loading in the newly developed FFE device with gratis gravity. [Copyright &y& Elsevier]

Abstract: This paper introduces a mathematic mode of moving chelation boundary (MCB) for computer simulation of a continuous EDTA-based sample sweeping in capillary electrophoresis (CE). Besides the equations of MCB used herein, the mode also includes electro-neutrality equation, constant current density, jump boundary condition of MCB, Kohlrausch’ regulating function expressed in MCB formulation, product of water, ionic apparent mobility, ionic strength and conductivity of electrolyte as well as simple equilibrium reaction, etc. The simulation software is developed based on the mode. With the software the relevant simulation is carried out, and the corresponding experiments on a MCB are performed. The results on the simulation and experiments demonstrate that (1) the software can simulate a dynamic process, characteristic peak shape and relevant electrophoregram of a MCB; (2) the simulator can quantitatively compute velocities of MCB and complex boundary (CB), all of ionic concentrations (especially the concentration of complex) and sweeping efficiency; (3) these simulation results mentioned above are generally in accordance with the experiments. The simulation software holds evident significances for the study on a MCB and conditional optimization in such an EDTA-based sample sweeping of metal ion in CE. [Copyright &y& Elsevier]