26: Introduction to Chromatographic Separations

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In previous chapters we explored the application of spectroscopy and electroanalytical chemistry to the quantitative analysis of an analyte in a sample. Despite the power of these instrumental methods of analysis, their use is often limited if the sample contains species that will interfere with the analysis. A UV/Vis analysis, for example, is easy to complete if the analyte is the only species present that absorbs light at the analytical wavelength. If two species contribute to the overall absorbance, a quantitative analysis is still possible if we can measure the sample's absorbance at two wavelengths. Things become more complex, however, as the number of analytes or interferents increase or if we do not know the identity of an intereferent.

Chromatography provides a solution to the analysis of complex samples by providing a way to separate the individual species in a sample prior to their analysis by a spectroscopic or electroanalytical method of analysis. In this chapter we provide a general introduction to chromatographic separations. In the four chapters that follow, we will consider specific chromatographic methods.

26.1: A General Description of Chromatography
In chromatography we pass a sample-free phase, which we call the mobile phase, over a second sample-free stationary phase that remains fixed in space. We inject the sample into the mobile phase where its components partition between the mobile phase and the stationary phase. The types of mobile phases and stationary phases, how these two phases contact each other, and how the solutes interact with the two phases are useful ways describe a chromatographic method.
26.2: Migration Rates of Solutes
Our ability to separate two solutes depends on the equilibrium interactions of the solute with the stationary phase and the mobile phase, which effects both the time it takes a solute to travel through the column and how the width of the solute's elution profile. In this section we consider the rate at which the solute moves through the column.
26.3: Zone Broadening and Column Efficiency
Suppose we inject a sample that has a single component. At the moment we inject the sample it is a narrow band of finite width. As the sample passes through the column, the width of this band continually increases in a process we call band broadening. Column efficiency is a quantitative measure of the extent of band broadening.
26.4: Optimization and Column Performance
The goal of a chromatographic separation is to take a sample with more than one solute and to separate the solutes such that each solute elutes by itself. Our ability to separate two solutes from each other—to resolve them—is affected by a number of variables; how we can optimize the separation of two solutes, is the subject of this section.
26.5: Summary of Important Relationships for Chromatography
In this chapter we have introduced many chromatographic variables, some directly measured from the chromatogram, provided by the manufacturer, or from the operating conditions, and some derived from these variables. This section summarizes these variables.
26.6: Applications of Chromatography
Although the primary purpose of chromatography is the separation of a complex mixture into its component parts, as outlined here, a chromatographic separation also provides qualitative and quantitative information about our samples. More detailed examples of qualitative and quantitative applications are found in the chapters that follow.