Determination of intact zeatin nucleotide by direct chemical ionisation mass spectrometry

Abstract

1. INTRODUCTION 9-/3-D-Ribofuranosylzeatin 5 ’ -monophosphate (ZMP) has so far been isolated in crystalline form and identified unambiguously from Zea mays kernels only [l]. Identification was based prin- cipally on enzymic and chemical degradation. However, in a number of plant tissues, nucleotides of zeatin, dihydrozeatin and isopentenyladenine have been identified, by chromatographic methods, as metabolites of exogenous cytokinins, and cytokinin nucleotide formation may be associated with cytokinin uptake [2]. More recent- ly, cytokinin nucleotides have been identified as the primary products of cytokinin biosynthesis [3-51. Indeed, cytokinin nucleotides appear to play a central, and possibly a key, role in cytokinin metabolism [6], However, in the past, most cytokinin isolation and characterisation procedures have tended to concentrate on the cytokinin bases and their glycosides which are normally purified by retention on cation exchange (preferably cellulose phosphate) columns. Surprisingly little effort has been made to identify and quantify cytokinin nucleotides present in the anionic fraction not re- tained by such columns. Nucleotide analyses utilis- ing mass-spectrometry (MS) and gas chromatography (GC)-MS usually require a