From batch to flow synthesis of 6-substituted purine ribonucleosides by enzymatic transglycosylation

Introduction Purine nucleoside phosphorylases (PNPs, EC 2.4.2.1) catalyze the reversible phosphorolysis of the glycosydic bond of purine nucleosides and, upon addition of a second nucleobase, may transfer the glycosyl moiety to stereoselectively form a new nucleoside (transglycosylation). Results and Discussion Because of its wide substrate specificity,[1] a PNP from Aeromonas hydrophila (AhPNP) was exploited to catalyze a “one-pot, one-enzyme” batch transglycosylation, resulting in a moderate to high conversion (18-65%) of 6-substituted purine bases into a 23-compound library of 6-substituted purine-9-ribosides.[2] Afterwards, AhPNP was covalently immobilized (50% yield) in a pre-packed stainless steel column by reaction with glutaraldehyde and reduction of the imino groups. The resulting AhPNP-IMER (IMmobilized Enzyme Reactor) was coupled to a HPLC apparatus containing an analytical or semi-preparative chromatographic column associated with a UV-visible detector. This system (Figure 1) was exploited to perform the flow-synthesis of five 6-substituted purine ribonucleosides at a mg scale by transglycosylation. Conclusion Under optimized (Design of Experiments, DoE) reaction conditions, coupling of transglycosylation with product separation resulted in a fast and efficient process (52-89% conversion) with minimized sample handling.[3] References [1] Ubiali, D. et al. Adv. Synth. Catal. 2012, 354, 96-104. [2] Ubiali, D. et al. Curr. Org. Chem. 2015, 19, 2220-2225. [3] Calleri, E. et al. Adv. Synth. Catal. 2015, 357, 2520-2528.