Eight Karplus relationships have been determined for use in conformational studies of saccharide N-acetyl side-chains in solution by NMR spectroscopy. Density functional theory was used to calculate (1)H-(1)H, (13)C-(1)H, and (13)C-(13)C NMR spin-coupling constants in four model compounds that mimic alpha-gluco, beta-gluco, alpha-allo, and beta-allo 2-acetamido-2-deoxy-D-aldohexopyranosyl ring configurations in order to study the effects of C1 and C3 configuration adjacent to the N-acetyl group on coupling behavior. Six vicinal J-couplings sensitive to the C2-N2 torsion angle were parametrized: (3)J(H2,NH), (3)J(H2,CO), (3)J(C1,NH), (3)J(C3,NH), (3)J(C1,CO) and (3)J(C3,CO). Two vicinal J-couplings sensitive to amide bond conformation (cis and trans amide) were also investigated, namely, (3)J(NH,CH(3)) and (3)J(C2,CH(3)). In relevant cases, comparisons were made to analogous coupling pathways found in proteins to evaluate the effects of peptide versus saccharide pathway structure on coupling magnitude. These parametrizations allow multiple, redundant J-couplings within the N-acetyl fragment of saccharides and related structures to be used for more confident assignments of side-chain conformation in biologically important saccharides, especially those where this structural element may play a role in molecular recognition or other biological processes. This application is illustrated in an analysis of experimental J-couplings measured within the N-acetyl side-chain of a (13)C-labeled methyl N-acetyl-alpha-D-glucosaminide.