Quantitative and mechanistic studies using the oxygen-18 isotope shift in carbon-13 nuclear magnetic resonance spectroscopy
Abstract
The $\sp{18}$O-isotope shift in $\sp{13}$C NMR spectroscopy was used to study the kinetics of oxygen exchange at the anomeric carbon atoms of several ketoses and aldoses. At 25-26$\sp\circ$C and over the pH range from 2 to 10, the relative rates of oxygen exchange for the aldoses studied increased in the following sequence: D-glucose, D-mannose, D-ribose, D-2-deoxyribose. The hydration rates for the open chain forms of the sugars were calculated and the results were analyzed in terms of steric and inductive effects by drawing analogies with the hydration kinetics of simple aldehydes and ketones. Effective molarities associated with ring closure reactions of common monosaccharides are calculated. The position of bond cleavage in the acid-catalyzed hydrolysis of sucrose was elucidated by hydrolyzing the sugar in $\sp{18}$O-water and assaying the incorporation of $\sp{18}$O into the several anomeric products using $\sp{13}$C NMR. Independent supporting experimental evidence was obtained using $\sp1$H NMR. The results clearly indicated fructosyl-oxygen bond cleavage under the reaction conditions employed. A detailed investigation was made of the complex $\sp{13}$C NMR spectrum arising from the various $\sp{18}$O-isotopomers in a solution of $\sp{18}$O-labeled $\rm\lbrack 1,4$-$\sp{13}$C$\sb2\rbrack$ succinic acid. These investigations reveal a way to measure the coupling constant between the carboxyl carbons of succinic acid. This coupling constant was found to increase dramatically with the solution pH. A complex spectrum was also observed in the carbonyl and olefinic regions of $\sp{18}$O-labeled (natural abundance $\sp{13}$C) dimethylmaleic anhydride. The eight signals in each region were interpreted in terms of isotope shifts over one, two, and even three bonds. The quantitative use of the $\sp{18}\rm {O}$ isotope shift in $\sp{13}$C NMR was assessed. The substitution of $\sp{16}$O by $\sp{18}$O in a series of carboxylic acids and ketones did not result in an observable difference in the spin-lattice (T$\sb1$) relaxation behavior of the directly attached $\sp{13}$C nuclei. The use of peak heights to infer the relative proportions of isotopomers (in a sample of 90 spectra in which the relative intensity of one peak with respect to an adjacent peak in no case fell below 25%) gave the same results (within $\pm$4%) as the use of peak areas.
Degree
Ph.D.
Advisors
Etten, Purdue University.
Subject Area
Biochemistry
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