Buffer Selection and Preparation

From our discussion in Chapter 2, we know that separation of a mixture of compounds is based on their affinity with the stationary and mobile phases within the system. In reversed phase chromatography, the affinity is based on the polarity of the different components and will vary depending on whether the compounds favour a polar mobile phase (i.e., are more polar in nature) or a nonpolar stationary phase (i.e., are more nonpolar in nature). The separation is measured in terms of retention time and the retention time is heavily dependent on the mobile phase pH. 

When analysing ionic or ionisable compounds, it is recommended that the pH of the mobile phase be controlled. A small change in mobile phase pH can have a significant effect on the chromatography and hence the separation. Therefore, it is necessary to control the pH of the mobile phase in order to maintain and sometimes promote an adequate separation. This can be achieved by the addition of an aqueous buffer to the mobile phase mixture. 

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