TABLE OF CONTENTS The range of operating modes is much greater in high-performance liquid chromatography (HPLC) because two immiscible phases, stationary and mobile, in contact with one another affect the separation. The variety of phase systems for LC allows for a wide range of selectivities, which can be adjusted to the components of interest in two ways: by varying the stationary phase or by modifying the mobile phase. Chromatographic separation in HPLC is the result of specific interactions of the sample molecules with both the stationary and mobile phases. With an interactive liquid mobile phase, another parameter is available for selectivity in addition to an active stationary phase. In practice, it is preferable to change the selectivity and the capacity by modifying the mobile-phase polarity for different samples while using the same stationary phase.
The operations of sample introduction, chromatographic separation, and detection are performed in exactly the same way in HPLC as in gas chromatography except for those modifications necessary to accommodate a liquid rather than a gas as the mobile phase. That is, the sampling is sequential, the separated components are eluted with the mobile phase, and detection is a dynamic time-dependent process. HPLC can be used whenever the sample can be dissolved in a liquid.
HPLC comprises a number of different LC methods, covering a large number of applications. LC methods can be divided into two forms. The first is retentive LC, in which separation is achieved through interaction of the solutes with the support surface...
