Hydrogen-bonding and conformation of agarose in methyl sulfoxide and aqueous solutions investigated by 1H and 13C NMR spectroscopy

The conformation of agarose in dimethyl sulfoxide (Me2SO) or in aqueous solutions has been studied by 1H and 13C NMR spectroscopy. Partial assignment of the 1H NMR spectrum of agarose in Me2SO-d6 has been obtained using 1D and 2D techniques. Coupling constant, chemical shift temperature dependence and exchange rate data for the hydroxyl protons have been used to establish the presence of intramolecular hydrogen bonds. The reported data show that the protons of HO-2 in the 3,6-anhydro-α-l-Galp residue (A) and HO-4 in the β-d-Galp residue (G) have a less rotational freedom and a higher thermal stability than the other hydroxyl protons. On the basis of the exchange rate data, obtained from 1D NOESY experiments with selective excitation of the residual water signal, an inter-residue hydrogen bond is proposed only for G HO-4 in Me2SO, while the involvement of A HO-2 in the agarose-Me2SO complex, recently reported, is suggested. The latter hypothesis is supported by the 13C NMR data obtained in Me2SO-d6, in a Me2SO-d6-D2O mixture and in D2O, which also show that a relatively small conformational rearrangement of the chain segments occurs in aqueous solution. Deuterium isotope effect data indicate the persistence in water of the inter-residue G HO-4 mediated hydrogen bond with the additional involvement of G HO-6. In addition, the negligible isotope effect on the C-2 atom of A would indicate that the corresponding OH group, although free from the agarose-Me2SO complex, is not available for further strong interactions.The hydrogen bonding and conformation of agarose in methyl sulfoxide or in aqueous solutions were studied by NMR spectroscopy. Based on 1H NMR data, an inter-residue H-bond is proposed for HO-4 of β-d-Gal p in Me2SO, while the involvement of HO-2 of 3,6-anhydro-α-l-Gal p in an agarose-Me2SO complex is suggested. These hypotheses are supported by the 13C NMR data.