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When samples contain ionizable compounds, the mobile phase pH can be one of the most important variables in the control of retention in a reversed-phase HPLC (RP-HPLC) separation. However, if it is not controlled properly, pH can be a source of many problems. Since most compounds analyzed by RP-HPLC contain one or more acidic or basic functional groups, most mobile phases require pH control. For this reason, buffers are widely used. This video highlights some of the important aspects of mobile phase pH for the practical chromatographer