Based on recent substrate specificity studies, a series of ribonucleotide based esters and carbamates were synthesized and screened as inhibitors of the phosphoramidases and acyl-AMP hydrolases, Escherichia coli Histidine Triad Nucleotide Binding Protein (ecHinT) and human Histidine Triad Nucleotide Binding Protein 1 (hHint1). Using our established phosphoramidase assay, K(i) values were determined. All compounds exhibited non-competitive inhibition profiles. The carbamate based inhibitors were shown to successfully suppress the Hint1-associated phenotype in E. coli, suggesting that they are permeable intracellular inhibitors of ecHinT.