Reversed phase, chromatography systems

To retain solutes selectively by dispersive interactions, the stationary phase must contain no polar or ionic substances, but only hydrocarbon-type materials such as the reverse-bonded phases, now so popular in LC. Reiterating the previous argument, to ensure that dispersive selectivity dominates in the stationary phase, and dispersive interactions in the mobile phase are minimized, the mobile phase must now be strongly polar. Hence the use of methanol-water and acetonitrile-water mixtures as mobile phases in reverse-phase chromatography systems. An example of the separation of some antimicrobial agents on Partisil ODS 3, particle diameter 5p is shown in figure 5. 

Monitoring the Structure, Conformation, and Interactions of Peptides in Reversed-Phase Chromatography Systems by Different Spectroscopic Procedures... 

Vereshchagin, A. G. The partition of polar and non-polar lipids in a reversed-phase chromatography system. J. Chromatogr. [Amsterdam] 14, 184 (1964). 

A. D. Kelmers, H. O. Weeren, J. F. Weiss, P. S. Pearson, M. P. Stulber and G. D. Novelli, Reversed-phase chromatography systems for transfer ribonucliec acid — preparatory scale methods, in Methods in Enzymology (eds., K. Moldave and L. Grossman) Vol. 20, Part C, Academic Press New York, pp 9-34,1971. 

This reversed-phase chromatography method was successfully used in a production-scale system to purify recombinant insulin. The insulin purified by reversed-phase chromatography has a biological potency equal to that obtained from a conventional system employing ion-exchange and size-exclusion chromatographies (14). The reversed-phase separation was, however, followed by a size-exclusion step to remove the acetonitrile eluent from the final product (12,14). 

HPLC requires a mobile phase in which the analytes are soluble. The majority of HPLC separations which are carried out utilize reversed-phase chromatography, i.e. the mobile phase is more polar then the stationary phase. In these systems, the more polar analytes elute more rapidly than the less polar ones. 

If simple sample pretreatment procedures are insufficient to simplify the complex matrix often observed in process mixtures, multidimensional chromatography may be required. Manual fraction collection from one separation mode and re-injection into a second mode are impractical, so automatic collection and reinjection techniques are preferred. For example, a programmed temperature vaporizer has been used to transfer fractions of sterols such as cholesterol and stigmasterol from a reversed phase HPLC system to a gas chromatographic system.11 Interfacing gel permeation HPLC and supercritical fluid chromatography is useful for nonvolatile or thermally unstable analytes and was demonstrated to be extremely useful for separation of compounds such as pentaerythritol tetrastearate and a C36 hydrocarbon standard.12... 

Most small organic molecules are soluble in mixed organic-aqueous solvents and can be easily analyzed using RPLC. However, there are some polar compounds which are not soluble in typical RPLC solvent systems or are unstable in an aqueous mobile phase system. These compounds can be analyzed on an RPLC column with a nonaqueous solvent system. This technique is called "nonaqueous reversed phase chromatography" (NARP).20-21 The NARP technique is primarily used for the separation of lipophilic compounds having low to medium polarity and a molecular weight larger than... 

These systems rely on various combinations of size-exclusion chromatography, reversed-phase chromatography, and zone electrophoresis to characterize amines, peptides, and proteins (Yamamoto etal., 1989 Bushey and Jorgenson 1990 Larmann et al., 1993, Moore and Jorgenson, 1995 Optick and Jorgenson, 1997). Haleem Issaq reviews these separations in Chapter 16 of this book. 

The term chemistry in interphases was first introduced in the field of reverse-phase chromatography [41], In 1995 Lindner et al. transferred the concept to the area of transition metal catalysis [42] and in a recent review the concept is explained in detail [43], The interphase is defined as a region within a system in which the stationary and a mobile component penetrate on a molecular level without the formation of a homogeneous mixture. In these regions the reactive centre on the stationary phase... 

Advanced gradient system For optimum functionality in automated systems designed primarily for reversed-phase chromatography and other gradient techniques, the LKB advanced-gradient system is recommended (Fig 1.2 (c)). Key features include ... 

Reverse phase chromatography describes a system with a non-polar stationary phase and a polar mobile phase. 

The liquid liquid partition chromatography (LLPQ method involves a stationary liquid phase that is more or less immobilized on a solid support, and a mobile liquid phase. The analyte is therefore distributed between the two liquid phases. In conventional LLPC systems, the stationary liquid phase is usually a polar solvent and the mobile liquid phase is an essentially water-immiscible organic solvent. On the other hand, in reversed-phase chromatography (RPQ, the stationary liquid is usually a hydrophobic... 

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