Basic Chromatographic Theory

These basic parameters, retention time and peak width, can be used to derive a number of other parameters that express the quality of the achieved chromatographic separation. In the following paragraphs, a brief summary of the most important parameters of chromatographic theory are discussed. 

The capacity factor k (equation 2.1) describes the velocity of the analyte relative to the velocity of the mobile phase. Each compound spends a different amount of time interacting with the mobile and stationary phase. The average velocity of a sample compound is dependent on how much time it spends in the mobile phase. If k is much smaller than 1, then the analyte moves too quickly and the elution time is so short that an exact determination of is difficult. If the sample moves too slowly, the separation time is very high. A good value for k would be between about 1 and 5. The selectivity factor a (equation 2.2) describes the relative velocities of the analytes with respect to each other. The selectivity describes how well a chromatographic method can distinguish between two analytes. 

The parameters that influence band broadening can be approximated by the van Deemter equation (equation 2.5) which is valid for gas and liquid chromatography as well as capillary electrophoresis (see chapter 3.2). 

The ultimate goal of a separation is to achieve a high resolution, R, (equations 2.6 and 2.7). If Rs = 1.5, then peaks of identical area overlap by only 

an /fi = 1 equals a peak overlap of 4 %. Peak resolution can be optimised by increasing the selectivity and minimising band broadening. 

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Basic chromatographic theory relates the retention factor of a solute to the equilibrium constant for adsorption, K, according to... 

Optimization of the separation of these samples is much more challenging than samples of homologues or oligomers. Basic chromatographic theory (equations 1-4 and related text) provides little direction for these separations, due to peak reversals that occur almost universally when conditions are changed, particularly temperature or composition. Historically, researchers have generally focused on only one or, at most, two experimental variables at a time, and have chiefly used trial-and-error as their optimization "strategy". 

Knox and Hartwick [1] included counter ions to ensure electrical neutrality in the adsorption process. We submit that their approach is not acceptable on the basis of the extensive studies that demonstrate the development of surface potential as a consequence of the stronger adsorbophilic attitudes of the IPR compared to that of its counter ion [31]. From basic chromatographic theory, if ion-pairing equilibria are taken into account, the retention factor of the analyte, k, is given by ... 

J. A. Jonsson, Chromatographic theory and basic principles. Marcel Dekker Inc., New York 1987. 

J0NSSON, J. A., in Chromatographic Theory and Basic Principles (Jonsson, J. A. ed.), Chapter 3, Dispersion and Peak Shapes in Chromatography (Marcel Dekker, 1987). 

Chromatographic Theory and Basic Principles, edited by Jan Ake Jdnsson... 

It is illustrated in Figure 5.1a. Clearly, both Rr and Rr can be easily calculated from their respective chromatograms, making them very useful measures for describing chromatographic results. Recall also that Rr = 1/(1 + k) and that k is proportional to the partition coefficient K, the basic thermodynamic variable in chromatographic theory. 

Isocratic separation of test compounds is a useful way to demonstrate the performance of a system. Basic chromatographic characteristics, such as theoretical plates, are easily measured and can be compared to what is expected from theory and to performance of other chromatographic systems. Figure 17-4 is a UHPLC chromatogram obtained under isocratic conditions on a 43-cm-long capillary column packed with 1.0-pm nonporous Cl 8 particles (Eichrom... 

Ion Chromatography, edited by James G. Tarter 38. Chromatographic Theory and Basic Principles, edited by Jan Ake Jonsson 39. Field-Flow Fractionation Analysis of Macromolecules and Particles, Josef Janca 40. Chromatographic Chiral Separations, edited by Morris Zief and Laura J. Crane 41. Quantitative Analysis by Gas Chromatography, Second Edition, Revised and Expanded, Josef... 

General references on chromatography include P. Sewell and B. Clarke. Chromatographic Separations. New York Wiley, 1988 Chromatographic Theory and Basic Principles, J. A. Jonsson, Ed. New York Marcel Dekker, 1987 A. Braithwaite and F. 1. Smith, Chromatographic Methods, 5th ed. London Blackie, 1996. 

As a rule, chromatographic optimizations are based on the trial-and-error approach, relying on experimentation, and the basic relationships of chromatographic theory. In HPLC [58-62] is used the well-known expression for the resolution... 

Before considering chromatographic theory in more detail it is important to understand the basic parameters. Fig. 2.2 shows a typical chromatogram, which represents the concentration of solute eluting from a column (as determined by the response of a sample detector) plotted against either time or volume. 

The theory of chromatography, especially of the hydrodynamics of chromatography, is a subject that chemists may tend to shy away from. Nevertheless, a basic awareness column dimensions or the best particle size can have a significant impact on the throughput or the sensitivity of a method. Therefore a fundamental grasp of chromatographic theory is important. 

Gazes, J., and R. Scott. 2002. Chromatography theory. Boca Raton, FL CRC Press. Heftmann, E. 2004. Chromatography Fundamentals and techniques. New York Elsevier. Jonsson, J. A. 1987. Chromatographic theory and basic principles. Boca Raton, FL CRC Press. 

Jonsson, J.A. (ed.) Chromatographic Theory and Basic Principles. Marcel Dekker, Inc. New York, 1987. Miller, J.M. Chromatography Concepts and Contrasts.WA y. New York, 1987. 

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