Pressurized planar electrochromatography,high-performance thin-layer chromatography and high-performance liquid chromatography—Comparison of performance

Kinetic performance, measured by plate height, of High-Performance Thin-Layer Chromatography (HPTLC), High-Performance Liquid Chromatography (HPLC) and Pressurized Planar Electrochromatography (PPEC) was compared for the systems with adsorbent of the HPTLC RP18W plate from Merck as the stationary phase and the mobile phase composed of acetonitrile and buffer solution. The HPLC column was packed with the adsorbent, which was scrapped from the chromatographic plate mentioned. An additional HPLC column was also packed with adsorbent of 5 μm particle diameter, C18 type silica based (LiChrosorb RP-18 from Merck). The dependence of plate height of both HPLC and PPEC separating systems on flow velocity of the mobile phase and on migration distance of the mobile phase in TLC system was presented applying test solute (prednisolone succinate). The highest performance, amongst systems investigated, was obtained for the PPEC system. The separation efficiency of the systems investigated in the paper was additionally confirmed by the separation of test component mixture composed of six hormones.     

Pressurized planar electrochromatography

Theoretical backgrounds, development, examples of separations, constructional details and principle of action of devices of pressurized planar electrochromatography (PPEC) are presented. Development of the mode is described in respect of operating variables (composition of the mobile phase, pressure exerted on adsorbent layer, mobile phase flow velocity, temperature of separating system, etc.) influencing separation efficiency (kinetic performance, repeatability, separation time). Advantages of PPEC such as high kinetic performance, short separation time and different separation selectivities, especially relative to conventional thin-layer chromatography, are described. Examples of two-dimensional separations are demonstrated to show high separation potential of the mode when combined with conventional thin-layer chromatography (TLC). The PPEC mode is in infancy stage of development, so its challenges are presented as well.     

A Modified Device for Pressurized Planar Electrochromatography and Preliminary Results with On-Line Sample Application

Pressurized planar electrochromatography (PPEC) is a separating technique in which an electric field is applied to force the mobile phase movement through a porous media (electroosmotic effect). High separation efficiency, fast separations and changes in separation selectivity in comparison to liquid chromatography, especially thin layer chromatography (planar chromatography, TLC), are features of this technique. Constructional methodological challenges to PPEC are obstacles to its development and application in laboratory practice. In this article, an attempt to overcome the challenges related to device construction and sample application/injection is described. The introduced device enables both prewetting of the adsorbent layer and electrochromatogram development with a single PPEC device. It also enables simultaneous application/injection of six samples on a chromatographic plate in a stream of the mobile phase (on-line application/injection). In addition, the PPEC chamber was equipped with a thermostat. The device is characterized by an impressive throughput in comparison to the other planar technique, TLC/HPTLC. Although the developed device still needs improvement, it is, in our opinion, a considerable step toward possible automation of this planar separation technique.     

Pressurized planar electrochromatography as a supporting tool for qualitative toxicological chemical analysis with thin-layer chromatography and UV–Vis spectrometry

ABSTRACT

Pressurized planar electrochromatography (PPEC) was applied to support qualitative toxicological chemical analysis performed with thin-layer chromatography (TLC) and UV–Vis spectrometry. Based on retention/migration distance data of substances obtained in TLC and PPEC systems and database of their wavelength maxima of remission UV–Vis spectra as well, a combined fit factor was calculated for substance identification. The involvement of PPEC, TLC, and UV–Vis spectral data together in calculation of the combined fit factor lead to its lower values for substances, which were not identical with reference, in comparison with those when the combined fit factor was calculated using TLC and spectral data only. The results evidence that involvement of PPEC data in qualitative toxicological chemical analysis performed with TLC and UV–Vis spectrometry enhances reliability of it.     

Separation of four mixtures of pesticides by pressurized planar electrochromatography (PPEC)

Examples of separations of four mixtures of pesticides by pressurized planar electrochromatography (PPEC) under different operating conditions are presented. The samples were separated on a prewetted RP-18W chromatographic plate in a system with acetonitrile-buffer as the mobile phase. A potential of 2.3 kV was applied to a 10 cm long plate to create the electric field. Reproducible retention of pesticides was obtained during PPEC in the closed system when the sorbent layer of the plate was prewetted and equilibrated with the mobile phase. The reported separations of pesticides by PPEC are over 10 times faster than the corresponding separations by TLC.     

Linear voltage profiles and flow homogeneity in pressurized planar electrochromatography

Planar electrochromatography (PEC) is an emerging technique for thin-layer chromatography (TLC) where electroosmosis is the driving force for the solvent, not capillary action. This allows for much faster and constant flow rates in turn yielding increased zone capacities and efficiencies. Instrumental designs have changed greatly over the last few years solving many of the initial instrumentation challenges. We have previously shown that low applied pressure (or no applied pressure) PEC instruments do not give linear voltage drops across the separation path length of a TLC plate, which in turn results in non-stable electroosmotic flow (EOF). By the use of our unique reader electrode grid we have the ability to monitor the potential at eight discrete positions throughout the 10-cm separation path length. We now show that high-pressure PEC instruments, most commonly referred to as pressurized planar electrochromatography (PPEC) do show a linear voltage drop and constant EOF. We compare plate equilibration times of PPEC and low-pressure PEC, use of increased field strengths, as well as sample application designs. In addition, we discuss the use of rhodamine B as a visual marker for reproducible migration and calculation of theoretical plates.     

Preliminary results for 2‐D separation with high‐performance thin‐layer chromatography and pressurized planar electrochromatography

Preliminary results of 2‐D separation of test dye mixture using high‐performance thin‐layer chromatography (HPTLC) and pressurized planar electrochromatography (PPEC) are demonstrated. The advantage of 2‐D HPTLC/PPEC separation is based on different separation selectivities obtained in both HPTLC and PPEC systems. HPTLC RP18 W plates of 5×20 cm from Merck were used in the investigations. In the first dimension, a HPTLC process was performed using 5 cm length of the plate and in the second dimension PPEC separation was obtained applying plate of 20 cm length. PPEC process followed prewetting the chromatographic plate with sample zones on it, which were partly separated after first dimensional (HPTLC) separation. In the experiments, the modified version of PPEC device for 20 cm long chromatographic plate and the reservoir for prewetting the adsorbent layer were applied.     

Correlation of Migration Distance of Peptides in High-Performance Thin-Layer Chromatography and Pressurized Planar Electrochromatography Systems

In a series of our previous papers we have investigated the influence of various variables on retention/migration of peptides in various high-performance thin-layer chromatography (HPTLC) and pressurized planar electrochromatography (PPEC) systems. Here we present a correlation of the selectivity of peptide separation in similar, as well as in various HPTLC and PPEC systems investigated before. Our results show that the selectivity in similar HPTLC and PPEC systems is quite different. This results from the share of electrophoresis in separation of solutes by PPEC. The results suggest that combination of HPTLC and PPEC, with properly selected separation conditions (the same, or even better—different for each technique), may be used for efficient two-dimensional separation of peptides. The best separation can be obtained if PPEC is carried out in two opposite directions (toward the cathode and the anode) simultaneously.     

Progress in planar electrochromatography

Developments in planar electrochromatography in open (PEC) and closed (PPEC) systems are reviewed. The discussion focuses on progress in chamber construction for planar electrochromatography, separating system performance, equilibration of the PPEC process, separation time and selectivity, and the general advantages, disadvantages and prospects of this separation mode. Presented at the 11th International Conference on Chemistry and the Environment, 9–12 September 2007, Torum, Poland.     

Reversed-Phase Pressurized Planar Electrochromatography and Planar Chromatography of Acetylsalicylic Acid,Caffeine, and Acetaminophen

JPC – Journal of Planar Chromatography – Modern TLC - We have investigated the use of pressurized planar electrochromatography (PPEC) and planar chromatography (TLC) for reversed-phase...     

Pressurized planar electrochromatography of DNS amino acids derivatives in silica gel and silanized silica gel systems with formic acid addition to the water mobile phase

JPC – Journal of Planar Chromatography – Modern TLC - Pressurized planar electrochromatography (PPEC) of dansyl (DNS) derivatives of amino acids in normal- and reversed-phase systems is...     

Influence of sample application mode on performance of pressurized planar electrochromatography in completely closed system

Three modes of sample application on the chromatographic plate are applied at present investigations of pressurized planar electrochromatography (PPEC) systems taking into special attention their influence on performance of the separating system. These modes are as follows: application of the sample solution directly on the chromatographic plate with microsyringe, deposition of sample solution on scrap of adsorbent layer followed by location oft this scrap on the chromatographic plate, application of the sample solution with commercially available aerosol applicator. These modes were combined with prewetting procedures of the chromatographic plates which lead to an accomplishment of equilibration of the stationary phase-mobile phase system. The plots of plate height versus linear flow rate of the mobile phase are presented for PPEC systems for the first time. The best separation performance has been obtained in PPEC system when prewetting of the chromatographic plate followed the sample application with commercially available aerosol applicator. The higher repeatability of migration distance of the solute bands has been obtained in PPEC experiments when the sample application was followed by prewetting the chromatographic plate in comparison to the experiments when these operations were performed in reversed order.     

侧链含氨基的聚(对亚苯基亚乙炔基-对亚苯基-咔唑)三元共聚物的合成及其聚集态下荧光特性

通过Sonogashira偶联反应制备了含有4-[2-(对苯胺)乙烯基]苯(M1)、2,5-二戊烷氧基-1,4-二乙炔基苯(M2)和9-辛基咔唑(M3)3种结构单元的三元共轭聚合物PPEC,并通过核磁共振氢谱确定了3种结构单元的比例为0.54∶1.00∶0.46(M1∶M2∶M3).由于在PPEC的侧链中含有氨基基团,通过在PPEC的THF溶液中分别加入水、甲醇和正己烷,诱导其产生聚集,其聚集后的荧光性质表现出明显的不同,在THF与水的混合溶剂中,PPEC发光强度会随着水含量的增加先是急剧降低,而后在高水含量时发光强度又显著增强;在THF与甲醇的混合溶剂中,PPEC发光强度随着甲醇的加入只是逐渐降低;在THF与正己烷的混合溶剂中,PPEC的发光强度则会随着正己烷的增加而增强.该结果表明氨基通过与水,或者自身所形成氢键作用,改变了PPEC分子链之间的聚集态结构,降低了分子内旋转非辐射能量效率,从而有效改善共轭聚合物的主链发光性质,这为设计聚集态下(或固态下)高性能的荧光共轭聚合物提供了一种新思路.     

Thin-layer chromatographic method for separation and determination of papaverine and its oxidation products

A method is presented for the separation of papaverine, papaverinol and papaveraldine by TLC on silica gel with chloroform saturated with ammonia as the solvent system. The spots are extracted with absolute methanol and each compound determined spectrophotometrically in 1 M hydrochloric acid. Papaveraldine is also determined polarographically in Britton-Robinson buffer, pH 4.1, after its separation and elution. The method is rapid and sensitive and permits the determination of papaverine, after its extraction from drugs, without interference by papaverinol and papaveraldine.     

Studies on Hydrated Stannic Oxide. Part VI. Thin-Layer Chromatography of Some Anions on Hydrated Stannic Oxide: Quantitative Separation of Cr(VI) from Some Ores and Alloys

Abstract

The analytical applications of hydrated stannic oxide as an ion-exchanger have been investigated by thin-layer chromatographic (TLC) technique. Binder-free thin-layers of hydrated stannic oxide are useful for some binary separations. Quantitative separation of Cr(VI) and Mo(VI) from some binary mixtures have been achieved. The method is applied to separate microgram-quantities of Cr(VI) quantitatively from several ores and alloys in ammonia buffer (pH 10) system.     

Electromigration methods in the determination of synthetic food dyes

Conditions were optimized for separating synthetic food dyes E 102, E 110, E 122, E 124, E 128, E 129, and E 133 by two-dimensional thin-layer chromatography (TLC) on Sorbfil plates using the mobile phase methanol-2-propanol-ethyl acetate-water (1: 1: 2: 2) in both the first (high-performance TLC) and second (electroosmotic TLC) directions. Conditions were selected for separating E 122, E 124, and E 133 by capillary zone electrophoresis; a borate buffer solution with pH 9.2 containing 10 vol % of acetonitrile was used. The time of separation was 16 min.     

高速逆流色谱法分离茶叶中的生物碱

以氯仿－甲醇－磷酸二氢钠缓冲液（２３ｍｍｏｌ／Ｌ,ｐＨ５．６）（体积比４∶３∶２）为溶剂系统,上相为固定相,下相为移动相,从茶叶的总生物碱中分离出三个成分,其中一个是咖啡碱,一个是茶碱,另一个待定。将分离结果与ＴＬＣ分离结果相比较,证实了高速逆流色谱法的有效及实用性。     

Indirect resolution of enantiomers of penicillamine by TLC and HPLC using Marfey's reagent and its variants

TLC and HPLC methods were developed for indirect chiral separation of penicillamine (3,3-dimethylcysteine) enantiomers after derivatization with Marfey's reagent (FDNP-Ala-NH(2)) and two of its structural variants, FDNP-Phe-NH(2) and FDNP-Val-NH(2). The binary mobile phase of phenol-water (3:1 v/v) and solvent combinations of acetonitrile and triethylamine phosphate buffer were found to give the best separation in normal and reversed-phase TLC, respectively. The diastereomers were also resolved on a reversed-phase C18 HPLC column with gradient elution of acetonitrile and 0.01 m trifluoroacetic acid. The results due to these three reagents were compared. The method was successful for checking the enantiomeric impurity of l-penicillamine in d-penicillamine and to check the enantiomeric purity of pharmaceutical formulations of d-penicillamine. The method was validated for linearity, repeatability, limit of detection and limit of quantification.     

Behavior of peptides and computer-assisted optimization of peptides separations in a normal-phase thin-layer chromatography system with and without the addition of ionic liquid in the eluent

The addition of an ionic liquid into the mobile phase appeared to be useful in optimization of chromatographic separation of peptides. Different behavior of peptides in thin-layer chromatography (TLC) was observed after addition of 1-ethyl-3-methylimidazolium tetra fluoroborate to the eluent in comparison to the system without the ionic liquid. Nonlinear dependence of the retention coefficient, R(M), of peptides on the volume percentage of acetonitrile in the eluent was found in normal-phase TLC with and without immidazolium tetra fluoroborate in the mobile phase. In general, R(M) increased with increasing concentration of acetonitrile. In TLC systems without the ionic liquid, R(M) can be described well with a quadratic function. On the other hand, in a TLC system with an ionic liquid as the additive to the mobile phase, the retention behavior is better described with a third-degree polynomial function. The potential usefulness of ionic liquids for optimization of separation of peptides was demonstrated. Optimization of the separation conditions was supported by a commercially available computer program.     

Role of buffer concentration and applied voltage in obtaining a good separation in planar electrochromatography

Planar electrochromatography is performed by applying an electric field across a thin layer chromatography (TLC) plate. In addition to electroosmotic flow in the axial direction, there is also flow to the surface of the TLC layer, and this can substantially degrade the quality of separation. This effect is offset by Joule heating which causes evaporation of liquid from the layer surface, and which under some conditions causes degradation of separation quality by excessive drying of the layer. It is shown that pH, buffer concentration, and applied voltage control the balance between liquid being driven to the surface and liquid evaporating from the surface due to Joule heating. Conditions are discussed which result in good separation quality, or in separations degraded by either excessive wetting or drying of the layer. The above separations were performed at constant voltage. A chromatogram is presented that shows that a good separation is also obtained at constant power, i.e. under conditions where there is a constant amount of Joule heating.