Gradient elution reversed-phase

P. W. High speed gradient elution reversed-phase liquid chromatography. 

Stoll, D.R., Cohen, J.D., Carr, P.W. (2006). Fast, comprehensive online two-dimensional high performance liquid chromatography through the use of high temperature ultra-fast gradient elution reversed-phase hquid chromatography. J. Chromatogr. A 1122 (1-2), 123-137. 

RETENTION TIMES, TR (MIN) IN GRADIENT ELUTION REVERSED-PHASE HPLC OF SULPHONATED DYES3... 

FIGURE 5.11 Gradient-elution reversed-phase separation of alkylbenzenes on a Purospher Star RP-18e, 3 am, column (30x4mm i.d.). (A) Non-adjusted linear gradients, 50%-100% acetonitrile in 3min at ImL/ min and at 3mL/min. (B) Adjusted linear gradients, 50%-100% acetonitrile in 3 min at ImL/min and 50%-100% acetonitrile in Imin at 3mL/min. Conditions 40°C, detection UV, 254 nm sample B—benzene, MB—toluene, EB—ethylbenzene, PB—propylbenzene, BB—butylbenzene, AB—amylbenzene, and HB—hexylbenzene. 

Farina, A. et al., HPTLC and reflectance mode densitometry of anthocyanins in Malva Silvestris L. a comparison with gradient-elution reversed-phase HPLC, J. Pharm. Biomed. Anal, 14, 203, 1995. 

D. L. Warner and J. G. Dorsey, Reduction of Total Analysis Time in Gradient Elution, Reversed-Phase Liquid Chromatography, LCGC 1997,15, 254. 

He et al. (1998) developed a gradient elution reversed-phase HPLC technique for separation of pungent principles. HPLC-UV-electrospray MS was used successfully to identify the individual pungent constituents in the chromatogram of ginger extract. Seven compounds were identified positively as the major pungent constituents of... 

Perhaps the worst problem of gradient elution separations is the need to reequilibrate the column with the initial solvent before a second sample can be run. An often-quoted rule of thumb is that up to 20 column volumes of the initial solvent may be necessary for this reequilibration process. The best test of reequilibration is the elution time of a weakly retained solute. These solutes will be greatly affected by an incompletely equilibrated stationary phase, and the retention time will vary. Cole and Dorsey have described a simple and convenient method for the reduction of column reequilibration time following gradient elution reversed-phase chromatography (119). Their method utilizes the addition of a constant 3% 1-propanol to the mobile phase throughout the solvent gradient to provide consistent solvation of the stationary phase. They noted reductions in reequilibration times of up to 78% ... 

Details of numerous HPLC methods that can be used for the purification of plant hormones are presented in Rivier and Crozier [1]. With some tissues, partitioning, cartridge systems and/or immunoaffinity chromatography can provide adequate sample purification prior to ABA and lAA analysis. However, HPLC fractionation is almost always required before GC-SIM analysis of individual cytokinins and GAs. As illustrated in Figs 2 and 3, good separations of free GAs and cytokinins of wide ranging polarity can be obtained by gradient elution, reverse phase HPLC. 

Various chromatographic consequences of protein conformational interconversion have been reported, most of which have been dted for reversed-phase HPLC. The gradient-elution reversed-phase sqiaration of several acid-stable proteins has been shown to give two bands, corresponding to native and denatured forms (Fig. 13 and related discussion). In this case, some of the protein molecules tqtparently denature vriiile retained at the column inlet. Native and denatur protdn are then eluted by the gradient. 

Schellinger, A.P. Stoll, D.R. Carr, P.W. High-speed gradient elution reversed-phase liquid chromatography of bases in buffered eluents Part I. Retention repeatability and column re-equilibration. J. Chromatogr. A, 2008, 1192, 41-53. 

McHowat J, Jones JH, Creer MH. Gradient elution reversed-phase chromatographic isolation of individual glycerophospholipid molecular species. J Chromatogr B 1997 702 21-32. 

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