Determination of Cytokinins by Ion Suppression-Reverse Phase High Performance Liquid Chromatography

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Abstract
A method for the separation of cytokinins has been developed by the use of ion suppression-reverse phase high performance liquid chromatography (IS-RP HPLC) with high efficiency, specificity and low sensitivity. Four reverse phase chromatographic columns were compared for their ability to separate and quantify six cytokinins with respect to signal response, resolution (RS), capacity factor (k1) and efficiency, N (number of theoretical plates). The optimal aqueous and organic phase components for the resolution of these cytokinins were determined with respect to pH, ionic strength and the proportion of the organic solvents, methanol and acetonitrile. Heptane sulfonic acid was used as an organic modifier. The optimal HPLC operating conditions were applied to separate and quantify the cytokinins present in a culture supernatant of Azotobacter chroococcum.