New methods for measurement of long-range proton-carbon coupling constants in oligosaccharides

Abstract

The geometry about the glycosidic l inkage plays a crucial role in the conformational analysis of oligosaccharides. Currently, most interglycosidic constraints are obtained from proton-proton NOE measurements. However, as has been pointed out ( 1, 2)) these spectral parameters can often be interpreted in terms of more than one set of conformer populations. Additional experimental constraints are therefore needed in order to allow for a more accurate description of saccharide conformations. The use of interglycosidic heteronuclear coupling constants ( 3Jn,.coc. and 3Jn,,coc,.) to determine the values of torsion angles Cp and + about the glycosidic l inkage is well known (3)) but their measurement (4, 5) has been lim ited by the low natural abundance of 13C and its low sensitivity. The introduction of inverse experiments, with the corresponding gain in sensitivity, has resulted in much wider interest in the measurement and interpretation of “J cn coupling constants (6-13). Most of the proposed experiments are, however, designed for the structural analysis of proteins (8-13) and usually performed on isotopically enriched samples. Recently, two different one-dimensional inverse-detected schemes for the evaluation of nJCH coupling constants of saccharides with natural abundance 13C have been proposed ( 14, 15). In the present Communicat ion we describe a novel two-dimensional inverse-detected NMR technique for the measurement of long-range heteronuclear coupling constants. A convenient strategy for the determination of interglycosidic 3JHcin saccharides has been developed. The methodology is based upon the pulse sequences shown in F ig. 2. The method is illustrated on a mode l disaccharide, methyl-b-xylobioside, 1, the ‘H NMR spectrum of which is shown in F ig. 1. The proposed experiment can be characterized as a heterocorrelated experiment optimized for nJCH in which single-quantum 13C magnetization ( 16) evolves during the evolution period t, of a two-dimensional experiment. The initial building block of the pulse sequence (Fig. 2a) is a refocused INEPT ( 17) optimized for ’ JCH. The 13C magnetization further evolves during the period A2 only under the effect of “JCH.