Adenosine 3′:5′-cyclic monophosphate in higher plants: Isolation and characterization of adenosine 3′:5′-cyclic monophosphate from Kalanchoe and Agave

1.3′:5′-Cyclic AMP was extensively purified from Kalanchoe daigremontiana and Agave americana by neutral alumina and anion- and cation-exchange column chromatography. Inclusion of 3′:5′-cyclic [8-3H]AMP from the point of tissue extraction permitted calculation of yields. The purification procedure removed contaminating material that was shown to interfere with the 3′:5′-cyclic AMP estimation and characterization procedures. 2. The partially purified 3′:5′-cyclic AMP was quantified by means of a radiochemical saturation assay using an ox heart 3′:5′-cyclic AMP-binding protein and by an assay involving activation of a mammalian protein kinase. 3. The plant 3′:5′-cyclic AMP co-migrated with 3′:5′-cyclic [8-3H]AMP on cellulose chromatography, poly(ethyleneimine)-cellulose chromatography and silica-gel t.l.c. developed with several solvent systems. 4. The plant 3′:5′-cyclic AMP was degraded by ox heart 3′:5′-cyclic nucleotide phosphodiesterase at the same rates as authentic 3′:5′-cyclic AMP. 1-Methyl-3-isobutylxanthine (1 mM), a specific inhibitor of the 3′:5′-cyclic nucleotide phosphodieterase, completely inhibited such degradation. 5. The concentrations of 3′:5′-cyclic AMP satisfying the above criteria in Kalanchoe and Agave were 2-6 and 1 pmol/g fresh wt. respectively. Possible bacterial contribution to these analyses was estimated to be less than 0.002pmol/g fresh wt. Evidence for the occurrence of 3′:5′-cyclic AMP in plants is discussed.