Introduction

In normal phase chromatography the dominant interactions between the solute and the stationary phase that cause retention and selectivity are polar in nature. If dispersive interactions dominate, then the separation system is called 'reversed phase' chromatography. To comprehend the retention mechanisms involved in normal phase chromatogra-phy, it is necessary to understand the nature of the different interactive forces that are involved between the solutes and the two phases and how they occur. The retention of a solute is directly proportional to the magnitude of its distribution coefficient (K) between the mobile phase and the stationary phase which, in turn, will depend on the relative affinity of the solute for the two phases, i.e. the interactive forces between the solute molecules and the two phases. Consequently, the stronger the forces between the solute molecule and the molecules of the stationary phase, the larger the distribution coefficient and the more the solute is retained. There are only three basic types of molecular force, all of which are electrical in nature. These three forces are called 'dispersion forces', 'polar forces' and 'ionic forces'. Despite there being many different terms used to describe molecular interactions (e.g. hydrophobic forces, n-n interactions, hydrogen bonding, etc.) all interactions between molecules are the result of composites of these three basic molecular forces. Dispersion forces arise from charge fluctuations throughout a molecule resulting from random electron/nuclei vibrations. They are typical of those that occur between hydrocarbons and other substances that have either no permanent dipoles or can have no dipoles induced in them. In biotechnology and biochemistry, dispersive interactions are often referred to as 'hydropho-bic' or 'lyophobic' interactions, apparently because dispersive substance such as the aliphatic hydrocarbons do not dissolve readily in water. Polar interactions arise from electrical forces between localized charges such as permanent or induced dipoles. Polar forces are always accompanied by dispersive interactions and may also be combined with ionic interactions. Polar interactions can be very strong and produce molecular associations that approach, in energy, to that of a weak chemical bond (e.g. 'hydrogen bonding'). Ionic interactions arise from permanent negative or positive charges on the molecule and thus usually occur between ions. Ionic interactions are exploited in ion exchange chromatography where the counter-ions to the ions being separated are suited in the stationary phase. To achieve the necessary retention and selectivity between the solutes for complete resolution, it is necessary to select a phase system that will provide the optimum balance between dispersive, polar and ionic interactions between the solute molecules and the two phases.

To achieve retention, the forces between the solute molecules and the stationary phase must dominate. If the molecules are largely dispersive in character then dispersive interactions must dominate in the stationary phase and, by suitable choice of mobile phase, polar interactions are made to dominate in the mobile phase. Conversely, if the substances to be separated are largely polar or polarizable then polar interactions must dominate in the stationary phase and, by suitable choice of mobile phase, dispersive interactions must be made to dominate in the mobile phase. When polar interactions dominate in the stationary phase, historically, the separation system has been termed 'normal' chromatography. When dispersive interactions dominate in the stationary phase, historically, the separation system is said to be 'reversed phase' chromatography. Thus in normal chromatog-raphy polar interactions dominate in the stationary phase and dispersive interactions are made to dominate in the mobile phase.

The polar forces that retain solutes in normal chromatography vary in strength and somewhat in mechanism. The strongest polar forces arise from dipole-dipole interactions, where charge centres on the solute molecule interact with the opposite charge centres on the stationary phase. Compounds, such as those containing the aromatic nucleus and thus (K) electrons, are said to be 'polarizable'. When such molecules are in close proximity to a molecule with a permanent dipole, the electric field from the dipole induces a counter dipole in the polarizable molecule. This induced dipole acts in the same manner as a permanent dipole and thus polar interactions occur between the molecules. Induced dipole interactions are, as with polar interactions, always accompanied by dispersive interactions. Aromatic hydrocarbons can be retained and separated purely by dispersive interactions when using a dispersive stationary phase (reversed phase chromatography) or they can be retained and separated by combined induced-polar and dispersive interactions by using a polar stationary phase such as silica gel (normal phase chromatography). The strongly polar hydroxyl groups inducing dipoles in the easily polarizable aromatic nucleus. A single molecule can possess different types of polarity; phenylethanol, for example, will possess both a permanent dipole as a result of the hydroxyl group and also be polarizable due to the aromatic ring. More complex molecules can have many different interactive groups.

The most common, and in fact traditional, normal phase system used in liquid chromatography consists of silica gel as the stationary phase and a mobile phase that is predominantly an alkane or a mixture containing a high proportion of an alkane. This system will be used to examine the type of interactive mechanisms that can take place in normal phase chromatography. The second solvent(s) can be more dispersive such as methylene chloride or a more polar such as ethyl acetate, «-propanol or even ethanol. To reduce retention, either the interactive character of the stationary phase must be reduced or the interactive character of the mobile phase increased or both. Both effects can be achieved by modifying the mobile phase. However, in normal phase chromatog-raphy, when employing mixed mobile phases, the interactions on the silica surface can become quite complex and the mechanism of retention needs some discussion. The mechanisms involved in mobile phase interactions with the solute are quite different to those involved with the stationary phase and thus, they will be considered separately.

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Solar Panel Basics

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