The separated components are visualized as a plot of the volume of the mobile phase eluted through the column versus the detector signal. This plot, called a chromatogram, shows the location of the individual peaks and the quality of resolution of these peaks. In Figure 1, there are two well-separated peaks corresponding to the two components A and B in the sample. Component A red requires solvent of volume V e A to elute at its maximum concentration. Component B green has near complete access to the pores of the matrix.
Therefore, its flow through the column is retarded. It elutes just before one column volume worth of the solvent can pass through the column i. The location of the peaks, their heights, and widths convey a great deal of information about the sample components as well as the column. Importantly, you can use peak locations to determine the molecular weights of the sample components.
The distribution of an analyte between the stationary and mobile phases depends on its size. It is described by a parameter called its equilibrium distribution coefficient K d. K d is the ratio of the concentrations of the component in the stationary and the mobile phases. For component B, K d can be written as. So, K d depends on particle size; therefore, it is essential to determine K d to estimate particle size. It is difficult to determine the concentrations of an analyte in the stationary and mobile phases.
Therefore, you express K d in terms of the easily measurable parameters, V e and V o. We know that the flow of component B through the column is retarded and it elutes at volume V e B. V T is the total solvent accessible volume, a parameter that is difficult to measure. Recent citations: Sneha Munshi et al. Related articles Based on techniques. MacKinnon , , Springer Protocols less.
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Chapter 5 Gel permeation chromatography.
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