GRADIENT ELUTION WITH SHORTER EQUILIBRATION TIMES IN REVERSED-PHASE ION-PAIR CHROMATOGRAPHY

To reduce the re-equilibration time needed between gradient runs in reversed-phase ion-pair chromatography (RP-IPC), various chromatographic conditions were studied. Five pairing ions of different size (and sorption kinetics), viz., trifluoroacetate, heptafluorobutyrate, octylsulphonate, dodecyl sulphate and tetrabutylammonium, each at two different concentrations, and three types of gradient (acetronitrile, salt and mixed gradients) were examined. Some common nucleotides, such as adenosine-5'-monophosphate, adenosine-5'-diphosphate and adenosine-5'-triphosphate, and some primary amines, such as dopamine, alpha-methyldopamine and serotonin, were used as analytes. The "efficiency" of the various gradient systems was tested in terms of the "speed" of re-equilibration and the average decrease in retention time achieved. Without solving the general problem of slow equilibration in ion-pair gradient elution, chromatographic conditions are recommended under which 1-3 column volumes of solvent A can be sufficient for re-equilibration between gradient runs, i.e., for obtaining reproducible retention times in gradient RP-IPC.