Use of phosphorus-32 labeled polyphosphoric acid for synthesis of labeled purine and pyrimidine 5'-ribonucleotides

Phosphorus-32 labeled polyphosphoric acid was used to phosphorylate the 2′,3′-O-isopropylidene derivaties of purine and pyrimidine ribonucleosides. The reaction mixture, which contained isopropylidene ribonucleoside 5′-phosphates, was subjected to selective hydrolysis, which removed the isopropylidene protecting group. The polyphosphate chains remained intact. Chromatography revealed the presence of ribonucleoside 5′-mono-, di-, tri-, tetra-, and higher phosphates. The distribution of radioactive label was established by liquid scintillation counting and enzyme assay. The triply labeled adenosine triphosphate prepared in this manner proved to be enzymically active and gave a positive firefly bioluminescence response.