Reversed-phase stationary phases

Optimized HPLC separation allows most betaxanthins to be separated on a Cl8 reversed phase stationary phase according to their respective polarities. - Considerable progress was achieved by the introduction of a highly polar silica-based column, which allowed major improvement of peak resolution, especially at early... 

HPLC), an advanced, accurate, and well-developed technique for both qualitative and quantitative measurements based on reverse phase stationary phases, or ion pairs applied for analytical or preparative purposes. 

Not only in HPLC, but also in modem thin-layer chromatography, the application of reversed-phase stationary phases becomes increasingly important. The advantage of the hydrophobic layers in comparison with the polar, surface-active stationary phases is the additional selectivity and a reduced hkehhood of decomposition of sensitive substances. 

Zarzycki and coworkers [77] studied the influence of temperature on the separation of cholesterol and bile acids using reversed-phase stationary phases. The best chromatographic conditions for the separation of mnlticomponent samples of steroids were chosen. Experiments were performed on wettable plates with RP-18W and at the temperatnres of 5, 10, 20, 30, 40, 50, and 60°C. The studies showed (Figure 9.9) that the degree of separation in the high-temperature region can be increased by an improvement of the efficiency of the chromatographic system. However, a relatively weak retention-temperatnre response for the studied steroids was observed. 

O.A.G.J. van der Houwen, R.H.A. Sorel, A. Hulshoff, J. Teeuwsen, and A.W.M. Indemans, Ion-exchange phenomena and concomitant pH shifts on the equilibration of reversed-phase stationary phase materials with ion-pairing reagents , J. Chromatogr., 1981,209, 393. 

KayiUo, S., Deimis, G.R., and ShaUiker, R.A., An assessment of the retention behaviour of polycyclic aromatic hydrocarbons on reversed phase stationary phases selectivity and retention on C18 and phenyl-type surfaces, J. Chromatogr. A, 1126, 283, 2006. 

Wheeler, J.F., Beck, T.L., Klatte, S.J., Cole, L.A., and Dorsey, J.G, Phase-transitions of reversed-phase stationary phases—cause and effects in the mechanism of retention, J. Chromatogr. A, 656, 317, 1993. 

FIGURE 8.6 SEM of an organo-silica hybrid monolith prepared with the precursor A-octadecyldimethyl[3-(trimethoxysilyl)propyl] ammonium chloride to provide reversed phase stationary phase and anodic EOF. (a) Cross-sectional view (magnification 1800 times) and (b) longitudinal view (magnification 7000 times). (Reprinted from J. D. Hayes, A. Malik, Anal. Chem., 72 4090 (2000). With permission. Copyright American Chemical Society 2000.)... 

Doyle, C. A. and Dorsey, J. G., Reversed-Phase HPLC Preparation and Characterization of Reversed-Phase Stationary Phases, In Handbook of HPLC, Katz, E., Eksteen, R., Schoenmakers, P. and Miller, N., Eds., Marcel Dekker, New York, pp. 293-323, 1998. 

Refluxing benzene solutions of Cjq in the presence of a 20-fold excess of BujSnH leads to hydrostannylation (Scheme 6.15) [73]. Multiple additions can also take place. To maximize the yield of the monoadduct CgoHSnBuj (24), the time dependence of the reaction was followed quantitatively by HPLC. After about 4 h, the concentration of the monoadduct 24 reaches its maximum. Compound 24 can be isolated by preparative HPLC on a Cjg-reversed-phase stationary phase with CHCI3-CH3CN (60 40, v/v) as eluent. The structure of C5oHSnBu3 (24) was determined by NMR spectroscopy and other methods, showing that a 1,2-addition takes place regio-selectively (Scheme 6.15) [73]. 

From the previous discussion, it is clear that when an ionic solute adsorbs to the reversed-phase stationary phase, it will create an electrostatic surface potential that will repel ions with the same charge from the surface and that the magnitude of the repulsion is represented by Equation 15.11. With the help of Equations 15.11 and 15.12, the adsorption isotherm for an ionic solute can be derived in the following way ... 

Figure 5.1 Scheme illustrating potential interactions between methylimidazolium cation and phenyl-based reversed-phase stationary phase. Arrows indicate hydrogen bonding, hydrophobic, and tt-tt interactions. Anion is not shown for clarity. (Adapted from Stepnowski, R, Nichthauser, J., Mrozik, W., and Buszewski, B., Anal. Bioanal. Chem., 385, 1483-1491, 2006.)... 

Surface-confined ionic liquids as reversed-phase stationary phases in liquid chromatography... 

This protocol focuses on the analysis of chlorophyll a and b, and the more nonpolar derivatives, including pheophytins and pyropheophytins. An octadecyl-bonded, reversed-phase stationary phase is used with a methanol/water mixture and ethyl acetate mobile phases in a gradient elution to provide rapid and complete separation of the major chlorophyll derivatives in 25 to 30 min. This is coupled with traditional UV/visible spectrophotometric detection at 654 nm to selectively screen these photosynthetic pigments in food and plant tissues. 

Alkylpenicillamine/Cu(II) complex dynamically ( ) adsorbed to reversed-phase stationary phase... 

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. 

Modification of the reversed-phase stationary phase choice of the ion-interaction reagent detection. 190... 

H. Richardson and B. A. Bildenmeyer, Bare silica as a reversed-phase stationary phase liquid chromatographic separation of antihistamines with buffered aqueous organic mobile phases, J. Pharm. Sci., 75 1480(1984). 

Figure 2.3 Reaction of silica gel with a functional group to produce a reversed-phase stationary phase.

A few polymeric reversed-phase stationary phases are available which provide the advantage that they can be operated over a wider pH range than the silica-based columns. Polymeric columns, however, tend to be less efficient than silica-based ones and are often less retentive. 

The popularity of reversed-phase chromatography can be explained by its unmatched simplicity, versatility, and scope.12 Although reversed-phase chromatography is used routinely for separating non-polar, non-ionic compounds, it is also possible and practical to separate ionic compounds on standard reversed-phase stationary-phase materials by using secondary equilibria, such as ion suppression, ion-pair formation, metal complexation, and micelle formation. To take advantage of these secondary equilibria,... 

It is well known from ion pairing chromatography that surfactants adsorb onto reversed phase stationary phases. Knowledge of the lengthy equilibrations necessary before ion pairing separations leads to an intuitive belief that gradient elution micellar... 

Since the discovery of IPC, this separation strategy has been devoted to increasing the mediocre retention of ionized samples on reversed phase stationary phases via ion-pair formation with a suitable IPR to increase the analyte hydrophobicity and, in turn, its retention. It is therefore clear that most IPC separations are performed under reversed phase conditions. Even if normal phase chromatography exploits polar... 

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