XTerra reversed-phase

Gritti, F. and Guiochon, G, Effect of the pH, the concentration and the nature of the buffer on the adsorption mechanism of an ionic compound in reversed-phase liquid chromatography, ii. Analytical and overload band profiles on Symmetry-C-18 and Xterra-C-18, J. Chromatogr. A, 1041, 63, 2004. 

Primary screens were performed in duplicate on mixtures of 80 compounds (1 pM per compound, 5 pM protein), where samples in each mixture were orthogonally pooled so that no two compounds that are in one well are also together in another well. Primary hits were achieved when the same compound in two wells were observed. The GPC spin-column eluates were partially resolved using a reversed-phase C18 HPLC column (Waters Xterra 2.1 x 20.0 mm) with a 1-min ballistic gradient and total cycle time of 2 min. The HPLC eluates were analyzed with a Micromass eight-way MUX interfaced to an LCT Tof ESI-MS system. Achieved throughput was 100 000 compounds per day. 

F. Gritti and G. Guiochon, Effect of the ionic strength of salts on retention and overloading behavior of ionizable compounds in reversed-phase liquid chromatography I. XTerra-Ci8,/. Chromatogr. A 1033 (2004), 43-55. 

PDE5 inhibitor (Fig. 8.17). The anal)d e and internal standard (sildenafil, II) were extracted by liquid-liquid extraction with diethyl ether/dichlor-omethane (70 /30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on reverse phase Xterra MS C18 column with a mobile phase of 10 mM ammonium formate/acetonitrile (10/90, v/v, pH adjusted to 3.0 with formic acid). The protonate of analyte was quantitated in positive ionization by MRM with a mass spectrometer. The mass transitions m/z 390.4 > 268.0 and m/z 475.5 > 58.3 were used to measure I and II, respectively. The assay... 

Reversed-phase LC is ideally suited for the analysis of polar and ionogenic analytes, and as such is ideally suited to be applied in LC-MS. Reversed-phase LC is the most widely used LC method. Probably, over 50% of the analytical applications are preformed by reversed-phase LC. Nonpolar, chemically-modified silica or other nonpolar packing materials, such as styrene-divinylbenzene copolymers (XAD, PRP) or hybrid silicon-carbon particles (XTerra), are used as stationary phases in combination with aqueous-organic solvent mixtures. Silica-based packing materials are used more frequently than polymeric packing materials. 

The current development in reversed-phase chromatography goes in the direction of sterically or chemically protected phases (shield), phases with hydrophilic endcapping (AQ, EPS) and hybrid phases (XTerra). 

For LC separation, various reverse-phase analytical columns can be used (Table 1). Most commonly. Cl8 columns are used. The LC column used for the analytical method described below is an XTerra C MS, 3.5 pm (150x2.1 mm I.D.) reversed-phase column (Waters, Milford, MA). 

Fig. 6. Chromatograms of a mixture of steroid contaminants obtained with reversed-phase XTerra RP18. From top to bottom column temperature 15 °C, gradient run time 30 min and 100 min column temperature 45 °C, gradient run time 30 min and 100 min.

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