Reversed-phase chromatography surface chemistry

Obviously, the monolithic material may serve its purpose only if provided with a suitable surface chemistry, which depends on the desired application. For example, hydrophobic moieties are required for reversed phase chromatography, ionizable groups must be present for separation in the ion-exchange mode, and chiral functionalities are the prerequisite for enantioselective separations. Several methods can be used to prepare monolithic columns with a wide variety of surface chemistries. 

The model for ionic retention and ion-pair chromatography that are discussed in Sections 15.2 and 15.3 has been tested and applied to a number of different systems and works very well in most of the cases. From colloid and surface chemistry is known that the model has its limitations, and under certain chromatographic conditions, the presented model will not be valid. The limitations of the model when applied to reversed-phase chromatography of ions still need to be found. Some are self-evident, such as if the pairing-ion concentration is close or above the CMC or when the retention factor is very low so that the accumulation in the double layer is important in comparison to the adsorption, see Ref. [7] for a discussion concerning the accumulation in the double layer. 

An overview and discussion is given of literature methods published after 1989 devoted to the ion-interaction chromatographic determination of inorganic anions. Seventy references are quoted. Ion-interaction chromatography makes use of commercial reversed-phase stationary phase and conventional high-performance liquid chromatography instrumentation. The basis of the technique, the modification of the stationary phase surface, the choice of the ion-interaction reagent as well as the dependence of retention on the different variables involved are discussed. Examples of application in the fields of environmental, clinical and food chemistry are presented. The experimental conditions of stationary phase, of mobile phase composition as well as detection mode, detection limit and application are also summarized in tables. 1997 Elsevier Science B.V. 

The previous chapter discussed the solvent and its interaction with the solute. To complete the chromatographic system the adsorbent has to be selected. As mentioned in Chapter 3.2.1 one has to distinguish between enantioselective and non-enantioselective adsorbents. Both groups of adsorbents are classified into polar, semi-polar and nonpolar adsorbents (see Tab. 4.4). This classification is based on the surface chemistry of the packing material. Interaction between mobile phase and adsorbent characterizes the phase system, which is distinguished between normal phase (NP) chromatography and reversed phase (RP) chromatography. This differentiation is historic and appointed by the ratio of the polarity of the adsorbent and the mobile phase. 

The complex surface chemistry of the metal oxides (section 4.2.1.2) is incompatible with their use in a number of chromatographic techniques. Polymer coated metal oxides are seen as an important approach to extending their scope. Alumina and zirconia particles coated with poly(butadiene), poly(styrene), poly(ethylene oxide), a copolymer of chloromethylstyrene and diethoxymethylvinylsilane and succinylated poly(ethyleneimine), for example, have been prepared for use in reversed-phase, size-exclusion and ion-exchange chromatography [44,46,54,120,134-137]. The methods of preparation are similar to those used for porous silica. 

Reversed-Phase Liquid Chromatography (RPLC) is an important tool in protein chemistry. Examination of sorption isotherms revealed that alcohohc buffers did not desorb proteins near physiological pH in RPLC systems, while buffers containing a poly(ethoxy alcohol) surfactant did not desorb protein at pH 2, but they did at pH 7 with concentrations of surfactant apparently well above the critical micellar concentration (cmc) [2]. It has been proposed that a necessary condition for the desorption of a protein from a surface is that the surface tension of the solvent falls between that of the protein and the surface [6]. This condition is fiilfilled for many proteins with surfactant solutions near conditions of physiological pH and ionic strength. Therefore, it was expected that separations of proteins could be thieved in these conditions. 

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