Abstract
A method is reported for the separation and recovery of radioactivity in purine compounds, in particular in 3′,5′-cAMP derived from [ 14C]adenine-prelabeled blood platelets. It is based on Randerath's thin-layer ion-exchange chromatography on PEI-cellulose and allows simple, reproducible, and quantitative estimation of radioactivity in most adenine-related purine derivatives. Up to 50 μl of neutralized perchloric acid cell extract (supernatant) can be applied without further treatment to the thin layers. The recovery of added radioactivity as 14C-labeled bases, nucleosides, and nucleotides is 98–100%. Recovery of tritium-labeled compounds appears to vary with their charge and position of label, and is generally not as reproducible. One- and two-dimensional techniques are described. The former resolves: ATP, ADP, AMP, cAMP, IMP, adenosine, adenine, inosine, and hypoxanthine: the latter: ATP, ADP, cAMP, and two additional areas containing, respectively, AMP + IMP and the combined bases and nucleosides.
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