Synthesis of 13 C‐labeled 5‐aminoimidazole‐4‐carboxamide‐1‐β‐D‐[ 13 C 5 ] ribofuranosyl 5′‐monophosphate

5‐Aminoimidazole‐4‐carboxamide‐1‐β‐D‐[ 13 C 5 ] ribofuranosyl 5′‐monophosphate ([ 13 C 5 ribose] AICAR‐PO 3 H 2 ) ( 6 ) has been synthesized from [ 13 C 5 ]adenosine. Incorporation of the mass‐label into [ 13 C 5 ribose] AICAR‐PO 3 H 2 provides a useful standard to aid in metabolite identification and quantification in monitoring metabolic pathways. A synthetic route to the 13 C‐labeled compound has not been previously reported. Our method employs a hybrid enzymatic, and chemical synthesis approach that applies an enzymatic conversion from adenosine to inosine followed by a ring‐cleavage of the protected inosine. A direct phosphorylation of the resulting 2′,3′‐isopropylidine acadesine ( 5 ) was developed to yield the title compound in 99% purity following ion exchange chromatography.