Solving a 3D Structural Puzzle

Nuclear magnetic resonance (NMR) spectroscopy is a versatile tool in analytical chemistry, highly suitable for structural elucidation of organic molecules—as well as multiple other areas of research. The subjects covered within this thesis all concern methods which allow a shift from covalent to spatial structural information using NMR spectroscopy.

Experimental distances from nuclear Overhauser effect (NOE) correlations, and dihedral angles from 3JHH-coupling constants, were used to obtain 3D structural information for several natural and synthetic compounds. The stereochemistry of novel natural compounds was determined, including that of a bicyclic non-ribosomal peptide (a novel structural motif), a steroid, and several polyketides.

Structural insights were gained for potential anti-cancer agents; the azumamides, including synthetic analogues. Differences in the conformational space of solution state compounds were identified experimentally between structural analogues and compared to the in vitro potency of the compounds. The structures of two peptides that exhibited a high degree of molecular recognition were investigated, resulting in the elucidation of a possible mode of interaction.

Also, a major assumption in the calculation of distances from NOEs, the assumption of equal rotational correlation times between proton pairs, was investigated for molecules in organic solvents. Two spin-state selective (S3) HMBC experiments were developed for measurements of homonuclear and heteronuclear long-range coupling constants, respectively.

The new NMR experiments were based on two existing experiments, the multiplicity-edited HMBC and the HAT HMBC, which were combined to obtain S3 editing of long-range homonuclear coupling constants. The output of the first S3 HMBC experiment was HMBC-type spectra with nJCH correlated cross-peaks, from which n+1JHH-coupling constants were sign-selectively determined with high accuracy.

Very small coupling constants, including previously unreported coupling constants from strychnine, were extracted, with all experimental values correlating very well to theoretical coupling constants from DFT calculation. A pulse segment was developed to change the polarization of the CH-H pairs in the homonuclear S3 HMBC, to gain S3 edited nJCH-coupling constants in the cross-peaks.

While only determining coupling constants to methine carbons, the extracted experimental coupling constants correlated very well to theoretical coupling constants, thus extending the S3 HMBC methodology to include both n+1JHH- and nJCH-coupling constants. Residual dipolar couplings (RDCs) are a relatively late addition to the small molecular NMR community, where alignment media are used to obtain anisotropic samples, which allow for RDCs to be extracted.

The number of inter-nuclear vectors for the correlation of RDCs to 3D structures is often limited for small molecules. Homonuclear RDCs were extracted by use of the homonuclear S3 HMBC that correlated well to alignment tensors from 1DCH-coupling constants, thus increasing the number of internuclear vectors.

The topic of enantiodiscrimination by RDC measurements of rigid organic molecules was also investigated, and new alignment media were developed to allow slight discrimination of enantiomers by stretched polymers. Finally, a new method of back-calculation of RDCs from 3D structures was developed and tested, which copes better with multiple conformers than the commonly used SVD methodology.

The approach thus resulted in good conformer populations for several small molecules, including multiple cinchona alkaloids.