Experimental study on the optimization of general conditions for a free-flow electrophoresis device with a thermoelectric cooler.

With a given free-flow electrophoresis device, reasonable conditions (electric field strength, carrier buffer conductivity, and flow rate) are crucial for an optimized separation. However, there has been no experimental study on how to choose reasonable general conditions for a free-flow electrophoresis device with a thermoelectric cooler in view of Joule heat generation. Herein, comparative experiments were carried out to propose the selection procedure of general conditions in this study. The experimental results demonstrated that appropriate conditions were (i) <67 V/cm electric field strength; (ii) lower than 1.3 mS/cm carrier buffer conductivity (Tris-HCl: 20 mM Tris was titrated by HCl to pH 8.0); and (iii) higher than 3.6 mL/min carrier buffer flow rate. Furthermore, under inappropriate conditions (e.g. 400 V voltage and 40 mM Tris-HCl carrier buffer), the free-flow electrophoresis separation would be destroyed by bubbles caused by more Joule heating. Additionally, a series of applications under the appropriate conditions were performed with samples of model dyes, proteins (bovine serum albumin, myoglobin, and cytochrome c), and cells (Escherichia coli, Streptococcus thermophilus, and Saccharomyces cerevisiae). The separation results showed that under the appropriate conditions, separation efficiency was obviously better than that in the previous experiments with randomly or empirically selected conditions.

A new strategy for highly efficient single-drop microextraction with a liquid-gas compound pendant drop.

Herein, a simple assembly was designed via a capillary and a funnel-like cap to achieve liquid-gas compound pendant drop (CPD) microextraction with great convenience. Due to the increased contact area and adhesion force between the capillary tip and the drop, the proposed method provides considerable flexibility in producing CPDs with different air bubble sizes. Four pesticides were chosen as model analytes to evaluate the proposed method. By using a 1 µL chlorobenzene droplet containing a 1 µL air bubble at a stirring rate of 700 rpm, a 70 to 135-fold enrichment of pesticides was obtained within 3.4 minutes. As compared with a typical SDME, the proposed method showed a 2-fold increase of enrichment factors and a 4-fold decrease of extraction time. Improvement of the extraction efficiency could be ascribed to the increased surface area of the droplet, and the thin film phenomena further improved the extraction kinetics through effective agitation. The results indicate that CPD microextraction could serve as a promising sample pretreatment method for automated high-throughput analyses in a wide variety of research areas.

Negative-pressure-induced collector for a self-balance free-flow electrophoresis device.

Uneven flow in free-flow electrophoresis (FFE) with a gravity-induced fraction collector caused by air bubbles in outlets and/or imbalance of the surface tension of collecting tubes would result in a poor separation. To solve these issues, this work describes a novel collector for FFE. The collector is composed of a self-balance unit, multisoft pipe flow controller, fraction collector, and vacuum pump. A negative pressure induced continuous air flow rapidly flowed through the self-balance unit, taking the background electrolyte and samples into the fraction collector. The developed collector has the following advantages: (i) supplying a stable and harmonious hydrodynamic environment in the separation chamber for FFE separation, (ii) effectively preventing background electrolyte and sample flow-back at the outlet of the chamber and improving the resolution, (iii) increasing the preparative scale of the separation, and (iv) simplifying the operation. In addition, the cost of the FFE device was reduced without using a multichannel peristaltic pump for sample collection. Finally, comparative FFE experiments on dyes, proteins, and cells were carried out. It is evident that the new developed collector could overcome the problems inherent in the previous gravity-induced self-balance collector.

A simple and highly stable free-flow electrophoresis device with thermoelectric cooling system.

Complex assembly, inconvenient operations, poor control of Joule heating and leakage of solution are still fundamental issues greatly hindering application of free-flow electrophoresis (FFE) for preparative purpose in bio-separation. To address these issues, a novel FFE device was developed based on our previous work. Firstly, a new mechanical structure was designed for compact assembly of separation chamber, fast removal of air bubble, and good anti-leakage performance. Secondly, a highly efficient thermoelectric cooling system was used for dispersing Joule heating for the first time. The systemic experiments revealed the three merits: (i) 3min assembly without any liquid leakage, 80 times faster than pervious FFE device designed by us or commercial device (4h); (ii) 5s removing of air bubble in chamber, 1000-fold faster than a normal one (2h or more) and (iii) good control of Joule heating by the cooling system. These merits endowed the device high stable thermo- and hydro-dynamic flow for long-term separation even under high electric field of 63V/cm. Finally, the developed device was used for up to 8h continuous separation of 5mg/mL fuchsin acid and purification of three model proteins of phycocyanin, myoglobin and cytochrome C, demonstrating the applicability of FFE. The developed FFE device has evident significance to the studies on stem cell, cell or organelle proteomics, and protein complex as well as micro- or nano-particles.

Mathematical model and dynamic computer simulation on free flow zone electrophoresis.

In this paper, a mathematical model was proposed for calculating physico-chemical parameters and simulating the separation process of solutes in free-flow zone electrophoresis (FFZE). Computer software was developed and implemented in a Delphi XE2 environment based on the model, which comprises zone electrophoresis, electrolyte solution, hydrodynamics and diffusion as well as conversion equations. The simulation results reveal that (i) the software is capable of simulating a dynamic process of FFZE properly; (ii) the software can simulate operation parameters (e.g., electric field, flow rate and pH value) for experimental optimization; (iii) the simulator is able to display the final electropherogram of numerous analytes in FFZE; and (iv) the electropherogram of FFZE can be automatically converted to that of capillary zone electrophoresis. In order to verify the rationality and reliability of this software, the relevant experiments were conducted and further compared with the simulations. The comparisons demonstrated the high agreement between the simulations and the experiments as well as those cited from the relevant references. In addition, effective mobility, diffusion coefficient and concentration of solutes can be conveniently gained via the software. The developed mathematical model and designed software have evident significance for the optimization of experimental conditions and basic physico-chemical parameters in FFZE.

Experimental investigation of external explosion in the venting process.

Experimental investigations were conducted on the process of combustion and explosion vent in a 200 mm (diameter) x 400 mm (length) vertical cylindrical vessel. When CH(4)-air mixture gases were used and the vent diameter was 55 mm, conditions of Phi (equivalent ratio)=0.8, Phi=1.0 and Phi=1.3 and two ignition positions (at the cylinder center and bottom) were selected. The venting processes and the correlated factors are discussed in this paper.