Classification and characterization of unknown cytokinins into essential types by in-source collision-induced dissociation electrospray ionization ion trap mass spectrometry

RATIONALE Mass spectrometry is effective for determination of cytokinins, which are bioactive compounds with an adenine-core structure. However, it is difficult to characterize any cytokinin compound without the relevant standard or known molecular structure information. With a limited number of standards, an in-source collision-induced dissociation (CID) method for characterization and classification of unknown cytokinins was described in this study. METHODS Cytokinins were separated by high-performance liquid chromatography and then analyzed by electrospray ionization ion trap mass spectrometry using an in-source CID technique and multiple reaction monitoring (MRM) mode. RESULTS Based on the properties of multi-stage fragmentation in mass spectrometry, naturally occurring cytokinins were classified into four types (zeatin, dihydrogen zeatin, isopentenyl adenine and benzyl adenine) by losing their conjugated sugar, sugar phosphate and other substituents in the source region. Following this technique, seven unknown cytokinins were characterized from roots of maize (Zea mays) without standards and one of them was finally confirmed to be cis-zeatin-riboside. CONCLUSIONS An in-source CID technique combined with MRM mass spectrometry was developed to provide product ion information for identification of cytokinins and to afford guidance for the discovery of unknown cytokinins. Copyright (C) 2012 John Wiley & Sons, Ltd.