NMR studies of oligosaccharides derived from hyaluronate: complete assignment of 1H and 13C NMR spectra of aqueous di- and tetra-saccharides, and comparison of chemical shifts for oligosaccharides of increasing degree of polymerisation

A series of oligosaccharides was prepared from hyaluronate by depolymerisation with bovine testicular hyaluronidase. Complete assignment of the 1 H and 13 C NMR spectra was obtained for the disaccharide, the tetrasaccharide, and the NaBH 4 -treated tetrasaccharide, by using various 1D and 2D NMR methods. The 1 H assignments for the tetrasaccharide differ from the incomplete data reported recently (ref. 11). The 13 C NMR spectra of the aqueous di, tetra-, hexa-, and octa-saccharides of this series show that all resonances, apart from those subject to obvious end effects, have chemical shifts comparable to those of the corresponding resonances of hyaluronate in D 2 O. The observed 13 C chemical shifts suggests that cooperative intramolecular hydrogen bonds probably play a minor role in determining the conformation of hyaluronate in water.