The Facile Synthesis of 5′-Nucleotides by the Selective Phosphorylation of a Primary Hydroxyl Group of Nucleosides with Phosphoryl Chloride

Abstract 5′-Nucleotides were directly prepared by selective phosphorylation with phosphoryl chloride in the presence of water and pyridine in acetonitrile. The rate of phosphorylation with phosphoryl chloride was markedly improved by the addition of suitable amounts of water and pyridine; the maximal formation of 5′-nucleotides was obtained by mixing these three reactants phosphoryl chloride, water, and pyridine in a molar ratio of 2 : 1 : 2 at a low temperature, followed by adding nucleosides corresponding to a one-fourth or one-fifth of the phosphoryl chloride. By mixing phosphoryl chloride, water, and pyridine in acetonitrile under controlled conditions, a hygroscopic complex (Complex 1), an active phosphorylating agent in the present phosphorylation of nucleosides, was isolated. It was identified as an adduct composed of tetrachloropyrophosphate and pyridinium chloride by means of its infrared spectra and by its chromatographic behavior. One of the proposed structures of the adduct is trichloropyrophosphopyridinium chloride. Both the selectivity in the 5′-hydroxyl group of the nucleosides and the yield of 5′-nucleotides increase upon the addition of pyridinium chloride in the phosphorylation of the unprotected nucleosides with tetrachloropyrophosphate. In the phosphorylation with tetrachloropyrophosphate, the H+ derived from pyridinium chloride in acetonitrile is considered to increase the selectivity for the 5′-hydroxyl group by decreasing the reactivity of the hydroxyl groups in the ribose moiety of the nucleosides with the adduct. Similar results were obtained in the phosphorylation with phosphoryl chloride in the presence of pyridinium chloride in acetonitrile. Thus, several procedures for the selective phosphorylation of unprotected nucleosides to 5′-nucleotides are available.