Anisotropic orientation of lactate in skeletal muscle observed by dipolar coupling in (1)H NMR spectroscopy.

Double quantum (DQ), J-resolved (1)H NMR spectra from rat and bovine skeletal muscle showed a splitting frequency ( approximately 24 Hz) for the lactate methyl protons that varied with the orientation of the muscle fibers relative to the magnetic field. In contrast, spectra of lactate in solution consist of a J-coupled methyl doublet and a J-coupled methine quartet (J(HH) = 7 Hz) with no sensitivity to sample orientation. Spectra acquired in magnetic fields of 4.7, 7, and 11 T showed that the splitting was not due to inhomogeneities in magnetic susceptibility within the muscle, because the magnitude of the splitting did not scale with the strength of B(0) fields. Triple quantum coherence (TQC) spectra revealed two distinct transition frequencies on the methyl resonance. These frequencies resulted from intra-methyl and methine-methyl couplings in this four spin system (A(3)X). Decoupling experiments on the triple quantum coherence showed that the observed frequency splitting was due mainly to the dipolar interactions between the methine and methyl protons of the lactate molecule. Thus, all the proton resonances of the lactate molecules in muscle behave anisotropically in the magnetic field. Adequate design and interpretation of spectroscopic experiments to measure lactate in muscle, and possibly in any cell and organ which contain asymmetric structures, require that both the dipolar coupling described here and the well-known scalar coupling be taken into account.