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Interactions between the diketopiperazine cyclo-alanylglycine and four fluorinated alcohols in water–fluoroalcohol mixtures were examined by ¹H{¹⁹F} intermolecular nuclear Overhauser effects (NOE) experiments. The alcohols studied were trifluoroethanol, hexafluoroacetone trihydrate, 1,1,1,3,3,3-hexafluoroisopropanol and perfluoro-t-butanol. The experimental methods used permit detection of solvent–solute NOEs of 0.1% or less. Solute and solvent diffusion coefficients were determined and apparent molecular radii of the fluoroalcohols estimated. Using these data, it was shown that observed ¹H{¹⁹F} intermolecular NOEs are consistent with expectations based on theory. A method for extending conventional theory to take into account the shape of a solute and the exposure of its hydrogens to solvent is described. This approach gives reasonable agreement with experimental results, particularly if it is assumed that solute–solvent interactions take place in such a way that the fluorines of a fluoroalcohol are preferentially oriented toward the solute during solute–solvent encounters. The results support the suggestion that intermolecular ¹H{¹⁹F} NOEs may become a useful tool for studies of peptide and protein conformations in fluoroalcohol–water solvent mixtures. © 2002 Wiley Periodicals, Inc. Biopolymers 64: 227–235, 2002

Uses of pulsed field gradients in the detection of ¹H{¹⁹F} nuclear Overhauser effects in biological systems were examined in experiments wherein gradients were used for coherence selection and recall, as well as simply for the removal of unwanted magnetization components. Strategies for selective detection of these NOEs when the fluorine resonance is broad are proposed. Detection of ¹H{¹⁹F} nuclear Overhauser effects in a carbonic anhydrase-fluorinated inhibitor complex demonstrates the methods considered and shows the superiority of the “inverse” approach to detection of fluorine-proton ¹H{¹⁹} NOEs in macro-molecular systems. Copyright © 1999 John Wiley & Sons, Ltd.

High-resolution and surface coil fluorine-19 NMR spectroscpies have been used to study the metabolism of the liver inducer 3-trifluoromethyl-alpha-ethylbenzhydrol (flumecinol) in rats. Although the fluorine chemical shift range exhibited by metabolites is small, it is possible to identify a number of metabolites or classes of metabolites by direct examination of urine samples. 3-Trifluoromethyl-4'-hydroxy-alpha-ethylbenzhydrol was confirmed as the primary metabolite at low doses (42 mg/kg). At higher doses other metabolites characterized by ring and side chain hydroxylation or further oxidation of the parent compound become apparent. NMR evidence for two previously unrecognized metabolites is described but these structures have not been identified. Studies of liver induction are reported that show that a single large dose of flumecinol exerts a substantial inducing effect on the number and types of metabolites formed after several hours postingestion. Examination of liver metabolism of the drug in vivo showed a single broad resonance, and it was not possible to obtained resolved signals for individual metabolites under the conditions of these experiments. However, nuclear Overhauser experiments showed that the materials detected in the in vivo experiments do not interact significantly with the macromolecular components of the liver.

Fluorine and nitrogen-15 NMR experiments are described which show that 3,5-difluorobenzenesulfonamide binds to human carbonic anhydrase I in a 1:1 complex, that the bound inhibitor is present as an anion and that the aromatic ring of the inhibitor undergoes rapid rotation in the complex in spite of likely interactions with the aromatic ring of Phe-91 which are strong enough to give an appreciable ¹⁹F{¹H} NOE. A previously predicted enhancement of binding by fluorination of the inhibitor was confirmed. © 1998 John Wiley & Sons, Ltd.

Powers and co-workers have provided evidence that thiobenzyl N-heptafluorobutyrylanthranilate (I) is an extremely potent inhibitor of serine proteases, especially alpha-chymotrypsin (Teshima, T., Griffin, J. C., and Powers, J. C. (1982) J. Biol. Chem. 257, 5085-5091). We have prepared additional derivatives of this structure in which fluorine substitutions have been made on the aromatic rings and have attempted to carry out fluorine NMR studies of the interaction of Powers' compound and these new derivatives with chymotrypsin. The solubility of all inhibitors examined in solvent systems compatible with the retention of native enzyme structure is extremely low. While some nmr evidence for complex formation could be obtained, preparations of the complexes examined were metastable and precipitation of the inhibitor eventually limits the amount of complex that can be present in solution to such low levels that nmr experiments are impractical. An unusual effect of solvent composition on fluorine chemical shifts suggests that the conformation of the inhibitors in aqueous solution and when bound to the enzyme is different from that in organic solvents.