Chromatographic separation, modes normal phase chromatography

Matrix The components within a mixture that provide support and structure but are not directly relevant to the analytes of interest. Blood is an example of a matrix in the examination of drugs of abuse Mobile phase The phase that carries the analyte through the stationary phase and is used to influence the chromatographic separation Mode of separation Denotes the mechanism by which the separation takes place. It is characterised by the stationary phase and the solvents used to elute the analytes of interest. It can be classed as reversed phase, normal phase, ion exchange, and chiral chromatography... 

Generally, better chromatographic performance is found with chiral separations in the normal phase for most column manufacturers. It is also likely that the easier solvent removal after collecting the isolated enantiomer, is what drove the industry to normal phase chromatography for chiral applications. It is advantageous to the chiral chromatographer that the majority of the commercially available normal phase LC CSPs and modifiers can be used on both LC and SFC instrumentation. This flexibility allows methods developed using one mode to be transferred to the other... 

It is important for the analyst to be able to select the best stationary phase to use for a particular chromatographic analysis. Silica gel can be used in two modes of chromatographic separations as a stationary phase in normal elution development or as a stationary phase in exclusion chromatography. 

Solvents, Extraction and Separation of Pigments. All solvents were freshly glass distilled and ethers were freed of peroxides over highly activated (0% H2O) basic alumina. All procedures were in dim yellow light and extracts/isolates maintained frozen under N2, whenever possible. Chromatographic separation utilized microcrystalline cellulose, Sephadex LH-20, and silica gel in normal and reverse phase modes. Extraction and chromatography is detailed elsewhere (20). 

The selection of mobile phases to be used in conjunction with these columns is dependent on the mode of chromatography (reversed phase versus normal phase) and on the physicochemical properties of the API (and related substances). However, in all instances, it is important to ensure that the API and all related substances can be monitored and are separated. Accordingly, it is of paramount importance that the initial chromatographic conditions elute all the individual components from the column and that no components are eluted at the solvent front (i.e., all compounds of interest are retained). In practice, this dictates the use of broad (5-90% strong solvent in 60 min) gradients. 

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