Endogenous cytokinins in three genera of microalgae from the chlorophyta

Nine axenic microalgal (Chlorophyta) strains from three genera (Protococcus, Chlorella, and Scenedesmus) were analyzed for endogenous cytokinins. Cytokinin-like activity was detected using the excised cucumber cotyledon bioassay. Five strains showed no cytokinin-like activity and four strains, low cytokinin-like activity. Ethanolic extracts of the microalgae containing a mixture of deuterium-labeled standards were purified using a combined DEAE-Sephadex octadecysilica column and immunoaffinity column based on wide-range specific mon-oclonal antibodies and analyzed by HPLC linked to a micromass single quadrupole mass spectrometer with an electrospray interface and a photodiode array detector. There were similar trends in cytokinin profiles for the nine microalgal strains investigated, although concentrations did vary. Both isopentenyladenine and isopentenyladenosine were detected in all nine strains. cis-Zeatin and cis-zeatin riboside occurred at higher concentrations than the trans isomers, whereas trans-zeatin-O-glucoside and trans-zeatin riboside-O-glucoside were dominant over the cis isomers. Dihydrozeatin and its conjugates were not detected in any significant amounts. The aromatic benzyladenine always occurred at higher concentrations than benzyladenosine. The topolins were well represented with all three isomers (ortho, meta, and para) being detected, with ortho-topolin and ortho-topolin riboside occurring at higher concentrations than the other isomers. However, for the O-glucosides, the meta isomers (meta-topolin-O-glucoside and meta-topolin riboside-O-glucoside) occurred at higher concentrations than the other isomers. No N-glucosides were detected (isopentenyladenine-9-glucoside, zeatin-9-glucoside, dihydrozeatin-9-glucoside, benzyladenine-9-glucoside, ortho-topolin-9-glucoside, and meta-topolin-9-glucoside). Generally, zeatin and topolin conjugates were the dominant forms of isoprenoid and aromatic cytokinins, respectively. There was no distinct trend in the proportions of isoprenoid to aromatic cytokinins.