Structural studies on flexible small molecules based on NMR in oriented mediamethodology and application to natural products

Abstract

This thesis describes the development and application of structural elucidation methodologies based on NMR in aligned media. Nuclear magnetic resonance is arguably the most important technique for the structural analysis of organic molecules in solution. In the last decade, Residual Dipolar Coupling (RDC) analysis emerged as a powerful tool for the determination of the three-dimensional structure of organic molecules in solution, complementing and even outperforming the approach based on the classical NMR observables such as NOE or 3J couplings. While application of RDCs to the structural analysis of proteins developed rapidly, their use with ¿small¿ molecules (typically organic compounds and natural products with MW < 1000 Da) is still scarce. From the spectroscopist point of view, two features of small molecules pose the main obstacles to the application of RDC to their analysis: the scarcity of observable couplings and the complexity stemming from conformational flexibility in solution. Besides, sample preparation with the optimal degree of alignment is still an issue for most classes of compounds. In this thesis, all these topics are addressed and new experimental and computational advancements are presented. 1. Sample preparation. Weak alignment in water and aligning properties of polyacrylamide gels. 2. New observables. Long-range proton¿carbon RDCs. 3. Analysis of flexible organic molecules. Regarding the preparation of weakly oriented samples in solution, technical advancements are presented, involving the development, orientational properties investigation, and quality assessment of new alignment media both in organic solvents and in water, comprising lyotropic liquid crystals and polymeric gels. Conformational flexibility is addressed using the single-tensor approach, that makes possible to fit experimental RDC to mixtures of conformers in fast-exchange equilibrium, facilitating the quantification of the conformational exchange. Bioactive compounds, such as salsolidine (metabolite) and lorcaserin (anti-obesity drug), are analyzed in this way. Long-range RDC are introduced to alleviate the lack of enough one-bond C¿H couplings in molecules with few protons, as is usually the case with many small molecules. A new experimental approach (SJS-HQSC) is presented for the measurement of long-range RDC, that is based on the insertion of the well-known proton-flip element in an HSQC-type experiment. The SJS-HSQC experiment provides very accurate long-range couplings in an experimentally simple and fast way. These methodological advances have been applied to model molecules such as salsolidine, 10-Epi, or the anti-obesity drug lorcaserin as well as to the structural determination of a new natural vinca alkaloid.