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Instrument Parameters Controlling Retention Precision in Gradient Elution Reversed-phase Liquid Chromatography

NCJ Number
Journal of Chromatography A Volume: 1371 Dated: December 2014 Pages: 90-105
Date Published
December 2014
16 pages

This study examined the effect of temperature, initial and final mobile phase composition, gradient time, and flow rate on the retention time precision under gradient elution conditions for various types of low MW solutes.


The study determined the retention factor in pure water and the solute-dependent solvent strength parameters of Snyder's linear solvent strength theory (LSST) as a function of temperature for three different groups of solutes. The effect of small changes in the chromatographic variables by use of the LSST gradient retention equation were estimated. Peaks at different positions in the chromatogram have different sensitivities to changes in these instrument parameters. In general, absolute fluctuations in retention time are larger at longer gradient times. Drugs showed less sensitivity to changes in temperature compared to relatively less polar solutes, non-ionogenic solutes. Surprisingly, results showed that fluctuations in temperature, mobile phase composition, and flow rate had less effect on retention time under gradient conditions compared to isocratic conditions. Overall temperature and the initial mobile phase composition are the most important variables affecting retention reproducibility in gradient elution chromatography. (Publisher abstract modified)

Date Published: December 1, 2014