Development of NMR methods to facilitate drug discovery and formulation
Abstract The goals of this thesis are to apply NMR experiments to problems important in drug discovery and development to (1) probe interactions between small drug molecules and proteins, (2) characterize interactions between drug-like molecules and excipients, and (3) study aspects of surfactant chemistry.
The first of these goals is addressed by applying the epitope mapping experiments, NOESY, STD, and BPPSTE, to map the interactions between the R and S enantiomers of propranolol and α1 acid glycoprotein (AGP). In addition to mapping the interactions between the propranolol enantiomers and AGP, these experiments allowed the comparison of optimal experimental parameters for each type of experiment.
The second goal of this thesis is addressed by applying a newly developed PFG diffusion experiment, the gradient phase encoded spin-lock (GraPES), and ROESY experiments to study the binding of drug-like molecules to an important class of pharmaceutical excipients, cyclodextrins. The GraPES experiment is useful for determining contacts between cyclodextrins and simple small molecules of moderate binding strength. The ROESY experiment can be used to determine the contacts with cyclodextrins for more complex molecules. In addition the GraPES experiment can be applied to ligand-protein binding systems to suppress the protein background. The results of the GraPES experiment further led us to examine the techniques used to analyze diffusion binding interaction or epitope mapping results.
The third goal of this thesis is addressed by (1) following the degradation of polysorbate 20 under accelerated degradation conditions and (2) characterizing monorhamnolipid mixtures. A calibration curve was developed to allow the quantitation of the extent of degradation of polysorbate 20. In addition, this method allows identification of the degradation products. The structures of several new monorhamnolipids were elucidated using a combination of NMR and mass spectrometry. Also, the average pKa of a mixture of monorhamnolipids was determined to be 4.39 ± 0.06 by NMR pH titration. The results of this pH titration indicated that the previous literature average pKa corresponded to a change in rhamnolipid supramolecular structure and not the pKa of the carboxylic acid moiety.