Choice of Experimental Parameters in IPC

Large changes in retention of ionic analytes may occur when the pH, the concentration of organic

Figure 7 Retention factor for the weak base adenine (zB =+ 1) as a function of pH in the mobile phase in the absence (■) and presence (•) of the pairing ion sodium octylsulfonate. The theoretical retention plots in the absence of the pairing ion (dashed line, Y0 = 0) and in its presence (solid line) are calculated from eqn [9] with the following parameters; pK = 4.12, kuB = 8.2, kjB = 2.07, zBFxV0/RT= - 2.398. (Reproduced with permission from Bartha A, Stahlberg J and Szokoli F (1991) Journal ofChromatography 552: 13.)

Figure 7 Retention factor for the weak base adenine (zB =+ 1) as a function of pH in the mobile phase in the absence (■) and presence (•) of the pairing ion sodium octylsulfonate. The theoretical retention plots in the absence of the pairing ion (dashed line, Y0 = 0) and in its presence (solid line) are calculated from eqn [9] with the following parameters; pK = 4.12, kuB = 8.2, kjB = 2.07, zBFxV0/RT= - 2.398. (Reproduced with permission from Bartha A, Stahlberg J and Szokoli F (1991) Journal ofChromatography 552: 13.)

Figure 8 (A) Adsorption isotherm ofoctylsulfonateto a C18-RP-column with the concentration of MeOH in the mobile phase as parameter. (B) Recommended alkylsulfonate type and concentration for different MeOH concentrations in the mobile phase, and (C) recommended tetraalkylammonium ion type and concentration for different MeOH concentrations in the mobile phase. (Reproduced with permission from Bartha A, Vigh Gy and Varga-Puchony Z (1990) JournalofChromatography 499: 423.)

Figure 8 (A) Adsorption isotherm ofoctylsulfonateto a C18-RP-column with the concentration of MeOH in the mobile phase as parameter. (B) Recommended alkylsulfonate type and concentration for different MeOH concentrations in the mobile phase, and (C) recommended tetraalkylammonium ion type and concentration for different MeOH concentrations in the mobile phase. (Reproduced with permission from Bartha A, Vigh Gy and Varga-Puchony Z (1990) JournalofChromatography 499: 423.)

This means that method development in this area usually is successful but requires a basic understanding of the physical processes that determine the retention.

In the development of an IPC method, the first point to consider is the relationship between the pKa value of the analyte ions and the pH value of the mobile phase. To obtain a robust system the mobile phase pH should deviate from the pKa value with more than two units. The pH is controlled by using a 10-50 mm salt buffer in the mobile phase. The buffer salt not only adjusts the pH but also controls the strength of electrostatic interaction between the ionic analytes and the ion pair reagent.

For changing the retention of an ionic analyte there are mainly three parameters to be varied: the type and mobile phase concentration of the ion pair reagent and the concentration of the organic modifier. The physically important parameter that is changed when these three parameters vary is the surface concentra tion of the ion pair reagent, and the corresponding electrostatic surface potential. A guideline for choosing type and concentration of positively and negatively charged ion pair reagents, respectively, as a function of the concentration of MeOH in the mobile phase is shown in Figure 8. The general rules for converting from MeOH to other common types of organic modifiers also apply in this case, e.g. 20% MeOH in the mobile phase corresponds to approximately 8% acetonitrile.

See also: N/Chromatography: Liquid: Mechanisms: Ion Chromatography; Mechanisms: Reversed Phases. Chromatography: Thin-Layer (Planar): Ion Pair Thin-Layer (Planar) Chromatography. III/Chiral Separations: Ion-Pair Chromatography. Peptides and Proteins: Liquid Chromatography.

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