Nuclear magnetic resonance spectroscopy of peracetylated oligosaccharides having C-13-labeled carbonyl groups in lieu of permethylation analysis for establishing linkage substitutions of sugars

Peracetylation of free hydroxyl groups in model saccharides with [C-13-carb onyl]acetic anhydride resulted in additional splittings of sugar ring proto n signals in NMR spectra, due to 3-bond J couplings between each acetyl car bonyl carbon and a sugar ring proton at that position. Quantification of 14 4 of these 3-bond coupling constants in different saccharide structures sho wed a range between 2.5 and 4.7 Hz, whereas ail possible 4-bond couplings b etween sugar ring protons and acetyl carbonyl carbons were within linewidth (<0.5 Hz). Therefore, further splitting of sugar ring proton signals in th e range of 2.5-4.7 Hz upon acetylation with a [C-13-carbonyl]acetyl group i dentifies that position as (formerly) having a free hydroxyl group. This pe rforms the same basic function as pennethylation analysis, but does not req uire hydrolysis of glycosidic linkages. Additionally, proton-detected 2D he teronuclear multiple bond correlation (HMBC) experiments or proton-detected heteronuclear correlation spectroscopy (hetCOSY) enabled ring proton-carbo nyl-C-13 3-bond J connectivities to be correlated with high sensitivity. Mo dified NMR pulse sequences are reported that include frequency selective de coupling schemes to enable coupling constants to be determined from 2D data . The tailored pulse sequences resulted in higher spectral resolution and s ensitivity for [C-13-carbonyl]-ring proton correlations. (C) 1999 Elsevier Science Ltd. All rights reserved.