UBC Theses and Dissertations

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UBC Theses and Dissertations

Chemical and spectroscopic studies of complex organic molecules Bernstein, Michael Alec


This thesis concerns itself mainly with carbohydrates--particularly their physical properties and structural elucidation by spectroscopic methods. Within this broad area a variety of topics are pursued. The findings are divided into two sections; the first deals with several spectroscopic approaches applicable to the study of carbohydrates attached to proteins (glycoproteins) and the second concentrates on the use of recently devised NMR experiments which assist in the determination of molecular structure and conformation. The studies in Section I depend on methods for "activating" specific loci in glycoprotein glycans. These materials were then "tagged" with nitroxide spin-labels, which reported on the motional flexibility of the glycans, which were found to be relatively mobile, and hydrated. These data were corroborated in an analogous H² NMR relaxation study, and this (superior) method reported very similar motional behavior to that derived from ESR. Finally in this section, the utility of synthetic glycoproteins in the analysis of ¹³C NMR spectra of glycoproteins was studied. First a useful method for neoglycoprotein synthesis was devised, and proteins bearing homogeneous and known sugars were prepared. It was determined that the ¹³C chemical shifts of the pendant sugars are comparable with those of the corresponding glycoside. Enzymic oxidation studies suggested that this approach may also greatly assist with the assignment of such carbohydrate molecules. The second section begins with a detailed survey of recent advances in high-resolution NMR spectroscopy which may be useful in organic chemistry. The experiments are explained in a simple conceptual fashion with illustrative data for trideuteriomethyl 2,3,4,6-tetra-0-(trideuterioacetyl)-α-D-glucopyranoside given throughout. This chapter is intended to be of assistance to the practicing organic chemist who has little experience in the areas of two-dimensional (2D) and other multi-pulse NMR procedures. A limited number of these experiments are used extensively in later chapters. Chapter II.3 details a new procedure whereby an oligosaccharide may be characterized using NMR spectroscopy, only. The method is "de novo" in that little need be known about the molecule prior to analysis; it relies on the characterization of all coupling pathways within each constituent monosaccharide and the determination of linkage order by inter-residue nuclear Overhauser enhancements (n0e's). To increase the chemical shift dispersion, the molecule is derivatized - here as a per-0-acetate. The ¹³C and ¹H NMR spectra of the plant alkaloid, brucine, were then studied. The emphasis here was on the choice and utilization of selected 1- and 2D NMR methods to determine the molecular structure and conformation of a complex organic molecule in the minimum of time. Elucidation of the final conformational detail required a strong emphasis to be placed on n0e experiments. Chapter II.4 details an NMR study on a cardenolide glycoside, digoxin. Using the strategies devised earlier, the molecule was studied at 500 MHz with the aim of complete spectral assignment. The spectrum is complicated by extensive signal overlap and the molecule's self-associative properties. For this reason, the lipophilic (steroidal) moiety displays broader lines which resist analysis by spin-echo experiments. The homonuclear and heteronuclear chemical shift correlation experiments performed well under these conditions, allowing the assignment of all sugar and some steroidal protons/carbons. The genin resonances proved relatively intractable, even with the powerful n0e experiment.

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