Polydopamine‐coated open tubular column for the separation of proteins by capillary electrochromatography

The separation and determination of proteins in food is an important aspect in food industry. Inspired by the self‐polymerization of dopamine under alkaline conditions and the natural adhesive properties of polydopamine, in this paper, a simple and economical method was developed for the preparation of polydopamine‐coated open tubular column, in which ammonium persulfate was used as the source of oxygen to induce and facilitate the polymerization of dopamine to form polydopamine. In comparison with a naked fused‐silica capillary, the direction and magnitude of the electro‐osmotic flow of the as‐prepared polydopamine‐coated open tubular column could be manipulated by varying the pH values of background solutions due to the existence of amine and phenolic hydroxyl groups on polydopamine coating. The surface morphology of the polydopamine‐coated open tubular column was studied by scanning electron microscopy, and the thickness of polydopamine coating was 106 nm. The performance of the polydopamine‐coated open tubular column was validated by analysis of proteins. The relative standard deviations of migration times of proteins representing run‐to‐run, day‐to‐day, and column‐to‐column were less than 3.5%. In addition, the feasibility of the polydopamine‐coated open tubular column for real samples was verified by the separation of proteins in chicken egg white and pure milk.     

Unified gas and supercritical fluid chromatography on 50 μm i.d. Columns

A unified approach to sequential gas and supercritical fluid Chromatography using 50 μm i.d. open tubular columns is described. Sample introduction is performed by means of a rotary injection valve. In order that linear velocities can be optimized independently, a second rotary valve in the chromatographic oven is used to direct the flow of column eluate to the flame ionization detector through either fused silica tubing in GC, or a frit restrictor in SFC. Applications of sequential GC-SFC on a 50 μm i.d. open tubular column are demonstrated, and comparisons made between sequential GC-SFC on 50 and 100 μm i.d. columns.     

Fast gas chromatography: packed column solvating gas chromatography versus open tubular column gas chromatography

Packed capillary column solvating gas chromatography (SGC) and open tubular column gas chromatography (GC) were compared with respect to their potentials for fast separations. A recently introduced "universal" peak capacity equation was used to compare the performance of these two methods. The effects of various factors on peak capacity were investigated. Results demonstrate that retention factor and column efficiency are the main factors affecting peak capacity for fast separations. Packed columns produce both high retention factors and high selectivities. While high efficiencies and high peak capacities can be demonstrated by both techniques, open tubular column GC can surpass packed capillary column SGC in both measurements, except for the case of the analysis of simple mixtures in short analysis times, where retention factor and selectivity become important. Practical aspects such as pressure drop and sample capacity are compared for SGC and open tubular column GC. It was found that packed column SGC demonstrates higher sample capacities, but requires much higher column inlet pressures than open tubular column GC. A variety of mobile phases can be used for packed column SGC, which can provide high solvating power for large and polar compounds.     

Cyclic electrophoretic and chromatographic separation methods

A review is given of the application of cyclic analytical methods in capillary electroseparation (CE) and liquid chromatography (LC) systems. Cyclic methods have been used since the early sixties in chromatographic systems to overcome pressure limitations to resolution. From the early nineties on they have also been applied in capillary electroseparation systems to overcome voltage limitations. Some basic theory is given, outlining the temporal development of resolution in cyclic CE and LC systems and calculating the maximal resolution that can be obtained as a function of the operational parameters of pressure and electrical field. Simple equations are given for the temporal change in the peak capacity and the loss of peaks from the systems as it occurs in some cyclic systems. Finally, a circular open tubular chromatographic system is proposed using integrated pumping and continuous detection. The performance of such a system is discussed using magnetohydrodynamic and alternating current electroosmotic pumping as examples of integrated pumps and Shah Convolution Fourier transform detection as an example of a continuous detection method.     

Large diameter open tubular columns in gas chromatographic analysis

Large diameter open tubular columns provide the packed column chromatographer with a simple route to higher resolution gas chromatography. They can be operated in a high-flow (lower resolution) mode that permits their direct substitution for a packed column, or they can be operated in a low-flow (higher resolution) mode that maximizes separation at the cost of longer analysis times. Inlet design and column installation can influence both the chromatographic results and quantiative reliability. Make-up gas is not required in the high-flow mode, and its benefits in the low-flow mode are restricted to enhanced detector sensitivity, provided the outlet end of the column resides in the detector jet (FID). The columns seem fully compatible with all common modes of detection.     

The effects of the column pressure drop on retention and efficiency in packed and open tubular supercritical fluid chromatography

The effects of the pressure drop across the column on retention and efficiency in SFC have been studied. Numerical methods are described which enable the prediction of hold-up time and pressure drop in both packed and open tubular columns. Predictions of both hold-up time and pressure drop are in good agreement with experimental data. The density gradient along the column can be calculated using the numerical methods and a procedure is described which enables the calculation of the overall capacity factors of the solutes from the density profile in the column. Significant variations of the capacity factor are observed along the column. The effect of the density gradient along the column on local diffusivity and dispersion is studied. The column efficiency in systems with significant pressure drops is affected by changes in: the linear velocity of the mobile phase; the diffusion coefficients; and the capacity factors of the solutes along the column. The overall efficiency of the chromatographic system can be calculated if, as is the case for open tubular columns, adequate plate height equations are available.     

Peak tailoring concept in gas chromatographic analysis of volatile organic pollutants in the atmosphere

An automated gas chromatographic system aiming at performing unattended analysis of volatile organic compounds (VOCs) was developed in laboratory. To encompass VOCs of a wide range of volatility, two different designs of enrichment and separation methods were adopted and compared with performance in analyzing ozone precursors of C3-C12. In the dual-trap dual-column design, lower boiling species (C3-C6) are enriched and separated by one set of trap and column (porous layer open tubular (PLOT)), whereas the enrichment and separation for the higher boiling species (C6-C12) are performed by the other set (wall-coated open tubular (WCOT)). Undesired peaks also inevitably appear on both chromatograms often causing annoyances. To reduce complexity of both the apparatus and the resulting chromatograms, the heart-cut technique was adopted as a base for developing a system, which only uses one trap and one flame ionization detector for constructing two-dimensional GC with PLOT and DB-1. Methods were developed to allow the auxiliary flow pressure in the heart-cut device to be programmed to create dual effects, which not only can perform regular heart-cut actions but can also temporally hold up species in the precolumn for prescribed time intervals. Because it is characteristic for PLOT chromatograms to have reproducible blank retention time windows, segments of a DB-1 trace are produced by the auxiliary flow program aligning perfectly in time with the gaps of the PLOT trace. Subsequently, the two column flows are merged and channeled into single flame ionization detector to produce a very condensed "tailored" chromatogram which is equivalent to overlaying a PLOT and a DB-1 chromatogram on top of each other, except that no peaks are overlapped. This innovative "peak tailoring" concept based on the heart-cut technique is simple in design, easy to build, and extremely rugged for long-term continuous operation as fewer moving parts are involved, which is beneficial for deploying in remote monitoring stations.     

Validation of new microvolume Couette flow linear dichroism cells

Long molecules such as fibrous proteins are particularly difficult to characterise structurally. We have recently designed a microvolume Couette flow linear dichroism (LD) cell whose sample volume is only 20-40 microL in contrast to previous cells where the volume of sample required has typically been of the order of 1000-2000 microL. This brings the sample requirements of LD to a level where it can be used for biological samples. Since LD is the difference in absorption of light polarised parallel to an orientation direction and perpendicular to that direction, it is the ideal technique for determining relative orientations of subunits of e.g. fibrous proteins, DNA-drug systems, etc. For solution phase samples, Couette flow orientation, whereby the sample is sandwiched between two cylinders, one of which rotates, has proved to be the optimal technique for LD experiments in many laboratories. Our capillary microvolume LD cell has been designed using extruded quartz rods and capillaries and focusing and collecting lenses. We have developed applications with PCR products, fibrous proteins, liposome-bound membrane proteins, as well as DNA-dye systems. Despite this range of applications, to date there is nothing reported in the literature to enable one to validate the performance of Couette flow LD cells. In this paper we establish validation criteria and show that the data from the microvolume cells are reproducible, vary by less than 1% with sample reloading, follow the Beer-Lambert law, and have signals linear in voltage over a wide voltage range. The microvolume cell data are consistent with those from the large-volume cells for DNA samples. Surprisingly, upon extending the wavelength range by adding the intercalator ethidium bromide, the spectra in the microvolume and large-volume cells differ by a wavelength dependent orientation parameter. This wavelength variation was concluded to be the result of Taylor-vortices in the large-volume cells which have inner rotating cylinders in our laboratory. Thus the microvolume LD cells can be concluded to provide better data than our large-volume LD cells, though the latter are still to be preferred for titration series as it is extremely difficult to add sample to the capillary cells without introducing artefacts.     

Column properties and flow profiles of a flat, wide column for high-pressure liquid chromatography

The design and the construction of a pressurized, flat, wide column for high-performance liquid chromatography (HPLC) are described. This apparatus, which is derived from instruments that implement over-pressured thin layer chromatography, can carry out only uni-dimensional chromatographic separations. However, it is intended to be the first step in the development of more powerful instruments that will be able to carry out two-dimensional chromatographic separations, in which case, the first separation would be a space-based separation, LC(x), taking place along one side of the bed and the second separation would be a time-based separation, LC(t), as in classical HPLC but proceeding along the flat column, not along a tube. The apparatus described consists of a pressurization chamber made of a Plexiglas block and a column chamber made of stainless steel. These two chambers are separated by a thin Mylar membrane. The column chamber is a cavity which is filled with a thick layer (ca. 1mm) of the stationary phase. Suitable solvent inlet and outlet ports are located on two opposite sides of the sorbent layer. The design allows the preparation of a homogenous sorbent layer suitable to be used as a chromatographic column, the achievement of effective seals of the stationary phase layer against the chamber edges, and the homogenous flow of the mobile phase along the chamber. The entire width of the sorbent layer area can be used to develop separations or elute samples. The reproducible performance of the apparatus is demonstrated by the chromatographic separations of different dyes. This instrument is essentially designed for testing detector arrays to be used in a two-dimensional LC(x) x LC(t) instrument. The further development of two-dimension separation chromatographs based on the apparatus described is sketched.     

Diffusioosmosis and electroosmosis in a capillary slit with surface charge layers

This study analytically examines the steady diffusioosmotic and electroosmotic flows of an electrolyte solution in a fine capillary slit with each of its inside walls covered by a layer of adsorbed polyelectrolytes. In this solvent-permeable and ion-penetrable surface charge layer, idealized polyelectrolyte segments are assumed to distribute at a uniform density. The electric double layer and the surface charge layer may have arbitrary thicknesses relative to the gap width between the slit walls. The electrostatic potential distribution on a cross section of the slit is obtained by solving the linearized Poisson–Boltzmann equation, which applies to the case of low potentials or low fixed-charge densities. Explicit formulas for the fluid velocity profile due to the imposed electrolyte concentration gradient or electric field through the slit are derived as the solution of a modified Navier–Stokes/Brinkman equation. The results demonstrate that the structure of the surface charge layer can lead to an augmented or a diminished electrokinetic flow (even a reversal in direction of the flow) relative to that in a capillary with bare walls, depending on the characteristics of the capillary, of the surface charge layer, and of the electrolyte solution. For the diffusioosmotic flow with an induced electric field, competition between electroosmosis and chemiosmosis can result in more than one reversal in direction of the flow over a range of the Donnan potential of the adsorbed polyelectrolyte in the capillary.     

毛细管电色谱法的研究进展

比较全面地综述了毛细管电色谱法的发展及研究状况。     

Study of electro-optical properties of liquid crystal/reactive mesogen-coated liquid crystal display fabricated by slit coater

We have recently proposed a slit coater method for printable liquid crystal (LC) devices. In this method, the LC/reactive mesogen (RM) mixture was directly coated uniaxially onto the substrate surface by shear flow between the slit nozzle and the substrate, and the liquid crystalline molecules were anchored by in situ photopolymerisation of RM monomers. In this study, the dependence of electro-optical properties and device parameters of LC sample cells fabricated by the slit coater on ultraviolet (UV) exposure dosage was investigated. Moreover, effects of UV exposure dosage and thermal annealing on the azimuthal anchoring energy of LC sample cells fabricated by the slit coater are presented.     

Dual column gas chromatographic system for use in mass spectral determination of nitrosamines.

A packed gas chromatographic column and a support coated open tubular (SCOT) column are connected in series. Between the columns are two micro-volume switching valves, one enabling solvent to be vented. Short retention nitrosamines are passed through both columns, whereas longer retention nitrosamines by-pass the SCOT column by means of the other switching valve.     

Two-dimensional symmetry breaking of fluid density distribution in closed nanoslits

Stable and metastable fluid density distributions (FDDs) in a closed nanoslit between two identical parallel solid walls have been identified on the basis of a nonlocal canonical ensemble density functional theory. Similar to Monte Carlo simulations, periodicity of the FDD in one of the lateral (parallel to the walls surfaces) directions, denoted as the x direction, was assumed. In the other lateral direction, y direction, the FDD was considered uniform. It was found that depending on the average fluid density in the slit, both uniform as well as nonuniform FDDs in the x direction can occur. The uniform FDDs are either symmetric or asymmetric about the middle plane between walls; the latter FDD being the consequence of a symmetry breaking across the slit. The nonuniform FDDs in the x direction occur either in the form of a bump on a thin liquid film covering the walls or as a liquid bridge between those walls and provide symmetry breaking in the x direction. For small and large average densities, the stable state is uniform in the x direction and is symmetric about the middle plane between walls. In the intermediate range of the average density and depending on the length L(x) of the FDD period, the stable state can be represented either by a FDD, which is uniform in the x direction and asymmetric about the middle of the slit (small values of L(x)), or by a bump- and bridgelike FDD for intermediate and large values of L(x), respectively. These results are in agreement with the Monte Carlo simulations performed earlier by other authors. Because the free energy of the stable state decreases monotonically with increasing L(x), one can conclude that the real period is very large (infinite) and that for the values of the parameters employed, a single bridge of finite length over the entire slit is generated.     

一种多通道匀浆装填毛细管色谱柱的新装置及应用

针对目前毛细管色谱柱装柱效率低、不同批次装填的毛细管色谱柱之间性能差异大的问题,我们发展了一种多通道匀浆装填毛细管色谱柱的新装置。该装置以液相色谱泵提供压力、采用磁力搅拌保持匀浆液均匀分散,一次可装填多达6根毛细管色谱柱。以牛血清白蛋白(BSA)的胰蛋白酶酶切肽段混合物为样本,选择峰容量、蛋白覆盖率、3个特定离子的保留时间以及毛细管色谱柱柱压为指标,在毛细管液相色谱-质谱联用系统上对装填的反相毛细管色谱柱的性能进行了评价。分别考察了一次装填的6根毛细管色谱柱、两次装填的12根毛细管色谱柱以及一次装填1根与一次装填6根毛细管色谱柱的性能及稳定性。实验结果表明:同一批次装填的6根毛细管色谱柱的性能相近;不同批次装填的12根毛细管色谱柱的峰容量和覆盖率没有明显的区别,但保留时间和毛细管色谱柱柱压的稳定性较差;一次装填1根和一次装填6根毛细管色谱柱柱性能的稳定性与两次分别装填6根毛细管色谱柱的稳定性相近,即采用本装置可显著提高毛细管色谱柱的装填效率且每次装填毛细管色谱柱的数量不会对柱性能产生影响。     

Wall modified photonic crystal fibre capillaries as porous layer open tubular columns for in-capillary micro-extraction and capillary chromatography

Wall modified photonic crystal fibre capillary columns for in-capillary micro-extraction and liquid chromatographic separations is presented. Columns contained 126 internal parallel 4 μm channels, each containing a wall bonded porous monolithic type polystyrene-divinylbenzene layer in open tubular column format (PLOT). Modification longitudinal homogeneity was monitored using scanning contactless conductivity detection and scanning electron microscopy. The multichannel open tubular capillary column showed channel diameter and polymer layer consistency of 4.2 ± 0.1 μm and 0.26 ± 0.02 μm respectively, and modification of 100% of the parallel channels with the monolithic polymer. The modified multi-channel capillaries were applied to the in-capillary micro-extraction of water samples. 500 μL of water samples containing single μg L⁻¹ levels of polyaromatic hydrocarbons were extracted at a flow rate of 10 μL min⁻¹, and eluted in 50 μL of acetonitrile for analysis using HPLC with fluorescence detection. HPLC LODs were 0.08, 0.02 and 0.05 μg L⁻¹ for acenaphthene, anthracene and pyrene, respectively, with extraction recoveries of between 77 and 103%. The modified capillaries were also investigated briefly for direct application to liquid chromatographic separations, with the retention and elution of a standard protein (cytochrome c) under isocratic conditions demonstrated, proving chromatographic potential of the new column format, with run-to-run retention time reproducibility of below 1%.     

Alltech集束毛细管柱和常规毛细管柱的比较

对集束毛细管气相色谱柱的色谱性能,如流速对柱效的影响、柱温对柱效的影响、柱容量等进行了考察,并将其对典型火炸药成分ＤＮＴ,ＴＮＴ的分离与常规毛细管气相色谱柱进行了比较。结果表明,集束毛细管气相色谱柱综合了填充柱与石英毛细管气相色谱柱之优点,弥补了二者的不足,是一种柱容量较大且分离效能好、可以在高载气流速下操作的新型气相色谱柱。     

Effects of Monomer Flow Rate,Flow Configuration,and Reactor Geometry on the Rate of Plasma Polymerization

The effects of flow rate on the plasma polymerization of ethylene in an rf discharge were investigated using both a tubular and a bell-jar-type of reactor. Both reactors contained parallel plate internal electrodes. Experiments with the tubular reactor showed that both the total thickness of the deposit and its distribution in the axial direction were strong functions of the flow rate. At low flow rates the polymer thickness decreased in the flow direction, while at high flow rates the polymer thickness increased. Each of these observations is explained by a simple model of plasma polymerization. Using the bell-jar reactor, different monomer flow distribution configurations were tested to determine their effect on the distribution of polymer thickness. It was found that distribution or diffusion of the monomer inflow provided a more uniform film.     

Electric conductivity in a fibrous porous medium with thin but polarized double layers

The electric conduction in the fibrous medium constructed by a homogeneous array of parallel, identical, charged, circular cylinders having an arbitrary zeta potential filled with the solution of a symmetrically charged electrolyte is analytically examined. The thickness of the electric double layers surrounding the dielectric cylinders is assumed to be small relative to the radius of each cylinder and to the gap width between two neighboring cylinders, but the polarization of the mobile ions in the diffuse layers is allowed. The effect of interactions among individual cylinders is taken into explicit account by employing a unit cell model. The appropriate equations of conservation of electrochemical potential energies of ionic species are solved for each cell, in which a cylinder is envisaged to be surrounded by a coaxial cylindrical shell of the fluid solution. Analytical expressions for the effective electric conductivity are obtained in closed forms as functions of the porosity of the fiber matrix and other characteristics of the porous system. Comparisons of the results of the cell model with different conditions at the outer boundary of the cell are made. Under an otherwise identical condition, the electric conductivity in a porous medium composed of an array of parallel cylinders in the transverse direction is smaller than that of a suspension of spheres. The effect of interactions among the cylinders or spheres on the effective conductivity can be quite significant under appropriate conditions.     

Theoretical optimisation of the side-wall of micropillar array columns using computational fluid dynamics

The present study provides an overview of the ideal side-wall position in micro-pillar array columns for the case of semi-embedded side-walls. The position has been determined using computational fluid dynamics simulations of the flow field in flow domains with different side-wall shifts. Optimal side-wall shift values are presented for a wide range of shapes (cylinders, and diamonds and hexagons with different aspect ratios) and packing densities. Simple linear correlations that allow calculating the optimised side-wall geometries for the different considered variety of shapes and packing densities could be established. Interestingly, only two correlations are needed to represent all investigated cases: one correlation for all diamonds, and one correlation for the cylinders and all hexagons. Compared to the case of a flat side-wall, the minimal feature size on the mask can be increased by a factor of 2.5 in the case of cylindrical pillar bed with external porosity ? = 0.4, implying that that much smaller pillar diameters can be used in the bulk of the bed before the minimal feature size on the mask falls below the lithography resolution.