Column chromatography microscale methods

Chromatographic and electrophoretic separations are truly orthogonal, which makes them excellent techniques to couple in a multidimensional system. Capillary electrophoresis separates analytes based on differences in the electrophoretic mobilities of analytes, while chromatographic separations discriminate based on differences in partition function, adsorption, or other properties unrelated to charge (with some clear exceptions). Typically in multidimensional techniques, the more orthogonal two methods are, then the more difficult it is to interface them. Microscale liquid chromatography (p.LC) has been comparatively easy to couple to capillary electrophoresis due to the fact that both techniques involve narrow-bore columns and liquid-phase eluents. 

The field of liquid chromatography is well established, and reliable methods have been developed for analytical and preparative separations. Column miniaturization improves performance for analytical separations. Numerous stationary phases have been developed to separate analytes based on a wide variety of molecular properties including hydrophobicity, ionic interactions, and molecular size. Mobile-phase modifiers can be used to aid in the niinumzation of unwanted interactions with the solid support. Although the field is well established, current research continues to improve separations for both microscale analytical and larger preparative separations. Recent publications will be highlighted that demonstrate the developments toward integrating HPLC components and separation techniques onto microfabricated devices. 

For microscale applications, select a Pasteur pipette (5%-inch) and clamp it upright (vertically). To reduce the amount of solvent needed to fill the column, break off most of the tip of the pipette. Place a small ball of cotton in the pipette and tamp it into position using a glass rod or a piece of wire. Take care not to plug the column totally by tamping the cotton too hard. The correct position of the cotton is shown in Figure 19.7. A microscale chromatography column is packed by one of the dry pack methods described in Part B of this section. 

For gas chromatography with a thermal conductivity detector, it is possible to collect samples that have passed through the column. One method uses a gas-collection tube (see Figure 22.10), which is included in most microscale glassware kits. A collection tube is joined to the exit port of the column by inserting the fS/S inner joint into a metal adapter, which is connected to the exit port. When a sample is eluted from the column in the vapor state, it is cooled by the connecting adapter and the gas-collection tube and condenses in the collection tube. The gas-collection tube is removed from the adapter when the recorder indicates that the desired sample has completely passed through the column. After the first sample has been collected, the process can be repeated with another gas-collection tube. 

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