Applications

Thin layer electrochromatography can be divided into three main forms depending on the major factor governing the separation. While not mutually exclusive, since most separations include some element of the other modes, these broadly arise from electrophoretic solute migration, electroosmotic solvent flow and the natural spin-off from the heating effects arising from the applied potential, electrothermal elution.

The most commonly encountered examples of TLE are based around electrophoretic separations in aqueous solvent. Not surprisingly, given the historical success of paper electrophoresis, several workers have used thin layers of microcrystalline cellulose. In addition, cellulose acetate and silica have been used for the separation of proteins. Other applications have included the separation of starches, amino acids (and various derivatives) (Figure 4), organometallic compounds (Figure 5) and transition metal ions.

Electrophoretic separations are not limited to aqueous solvent systems and the higher resistance of nonaqueous solvents gives the advantage of lower currents and reduced heating effects. The separation

Figure 4 The separation of amino acids by two-dimensional thin layer electrophoresis-thin-layer chromatography with an aqueous electrolyte. The chromatographic media was plastic-backed cellulose layer. Electrophoresis in the first dimension using a 4%(v/v) aqueous formic acid electrolyte was followed by chromatographic elution in the second dimension with butanol-0.4%pyridineacetic acid (22:10:10, v/v/v). Adapted from E. McEvoy-Bowe (1985) Journal ofChromatography 347: 199-208, with permission.

Figure 4 The separation of amino acids by two-dimensional thin layer electrophoresis-thin-layer chromatography with an aqueous electrolyte. The chromatographic media was plastic-backed cellulose layer. Electrophoresis in the first dimension using a 4%(v/v) aqueous formic acid electrolyte was followed by chromatographic elution in the second dimension with butanol-0.4%pyridineacetic acid (22:10:10, v/v/v). Adapted from E. McEvoy-Bowe (1985) Journal ofChromatography 347: 199-208, with permission.

of a number of dyes using ethanol as the solvent is shown in Figure 6. In this separation electro-osmotic flow effects were suppressed to reveal the electrophoretic migration of the charged dyes, resulting in completely different elution orders.

In TLE, solvent migration from EOF is easily confused with capillary-induced flow resulting from localized solvent evaporation. Broadly speaking, EOF is to be expected from wet polar solvents, protic solvents or from those that are capable of autoprotolysis.

With vertical tank systems, and particularly those employing nonpolar solvents, there must remain some uncertainty over whether thermal effects have been responsible for any solvent migration observed. This is the case in the pioneering planar systems studied by Pretorious (Figure 7), in which nonpolar solvents such as benzene were allegedly used. Our attempts to reproduce this work with a vertical tank system resulted in an electrically driven solvent

Figure 5 A thin-layer electropherogram of platinum chloro-amine complexes. The chromatographic media was microcrystal-line cellulose thin layers and electrolyte was 0.1 M NaClO4, at

Figure 6 Nonaqueous thin-layer electrochromatography (TLE) and conventional thin-layer chromatography (TLC) of a dye mixture: (a) Oil Blue, (b) Rhodamine B, (c) Neutral Red, (d) Diazine Green, (e) Brilliant Green. The chromatography media was silica (electroosmotic flow suppressed) and the solvent was ethanol.

Figure 6 Nonaqueous thin-layer electrochromatography (TLE) and conventional thin-layer chromatography (TLC) of a dye mixture: (a) Oil Blue, (b) Rhodamine B, (c) Neutral Red, (d) Diazine Green, (e) Brilliant Green. The chromatography media was silica (electroosmotic flow suppressed) and the solvent was ethanol.

Figure 7 An early thin-layer electrochromatography (TLE) separation of nonionic compounds. The chromatography media was dichlorodimethylsilane-treated silica and the solvent was unspeci-

Figure 5 A thin-layer electropherogram of platinum chloro-amine complexes. The chromatographic media was microcrystal-line cellulose thin layers and electrolyte was 0.1 M NaClO4, at

Figure 7 An early thin-layer electrochromatography (TLE) separation of nonionic compounds. The chromatography media was dichlorodimethylsilane-treated silica and the solvent was unspeci-

500 V for 5 min. Adapted from M Lederer and E Leipzig-Pagani fied. Adapted from V. Pretorius etal. (1974) Journal of Chromato-

(1998) Analytica ChimicaActa 358: 61-68, with permission.

graphy 99: 23-30, with permission.

migration and chromatographic separation resulting largely from thermal effects and not EOF. By changing to more polar solvents, in a horizontal tank, we have shown that true electroosmotic flow could be achieved. The separation of a number of pyrimidines on silica eluted with ethanol showed elution characteristics similar to those obtained by conventional TLC, but with higher separation efficiency and in one tenth of the time (Figure 8).

More recently, high-voltage nonaqueous TLE employing electroosmosis as the main driving force has been applied to the separation of a wide range of acidic, basic and neutral organic compounds, with considerable success.

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