HIGH-RESOLUTION DIFFUSION AND RELAXATION EDITED ONE-DIMENSIONAL AND 2-DIMENSIONAL H-1-NMR SPECTROSCOPY OF BIOLOGICAL-FLUIDS

A new approach to the characterization of biomolecules in whole biolog ical fluids is presented based on simplification of H-1 NMR spectra by utilizing differences in molecular diffusion coefficients alone and c ombinations of relaxation and diffusion parameters. New NMR pulse sequ ences incorporating both spectral editing features together with solve nt water resonance elimination are presented. The methods are exemplif ied using whole human blood plasma, and it is shown that it is possibl e to obtain NMR spectra of the slowly diffusing species (generally lar ge molecules) by diffusion editing, the slowly relaxing species (gener ally small molecules) by spin relaxation editing, or spectra showing a ny range of molecular mobility using a combination of the two methods. The diffusion-based editing methods are also applicable to the select ion of resonances in two-dimensional NMR spectroscopy of biofluids, an d we show this for the first time by fire production of H-1-H-1 diffus ion-edited TOCSY spectra of human blood plasma where the resonance int ensities are weighted according to the molecular diffusion coefficient . In this case, by measuring a diffusion-edited H-1-H-1 TOCSY NMR spec trum of plasma, it is possible to obtain signals from only the macromo lecular components, and this may be of benefit in the analysis of bloo d lipoproteins. In complex biofluids, the combination of diffusion and relaxation editing brings about considerable spectral simplification leading to an easier resonance assignment process. We also demonstrate the production of H-1 NMR spectra with intensities corresponding to d iffusion coefficient rather than number of protons, and this operas up new possibilities for pattern recognition classification of samples b ased an altered molecular mobility features of biofluid components.