NMR STUDIES OF CELLULAR METABOLISM

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Summary of Work: As a consequence of the widespread fluoridation of water and the occasional exposure of some populations to considerably higher fluoride levels, it is important to understand the biochemistry of this element. This study was motivated by the question of whether fluoride undergoes any significant anabolism into fluorinated organic compounds which could be a basis for toxicity. In fact, one such process is known: the pyruvate kinase catalyzed phosphorylation of fluoride. In order to investigate this process further, we looked at several other kinases, and at the in vitro reaction of fluoride with nucleotides in the presence of magnesium. Two of the other kinases studied: glycerokinase and acetate kinase, were also found to exhibit fluorokinase activity, although to a lesser extent than pyruvate kinase. Magnesium ions and other divalent ions were found to catalyze both the fluorokinase reaction as well as the formation of fluoronucleotides, e.g. adenosine 5'-O- fluorophosphate and adenosine 5'-O-(2-fluorodiphosphate). Although these reactions can proceed at 37 C, the concentrations of fluoride and divalent cations required for significant production of these compounds exceed those in biological systems by several orders of magnitude. Hence, these processes would not pose significant toxicity for the cell. In addition to these studies, collaborative studies with Prof. David Thompson on the metabolism and potential toxicity of p-alkylphenols have continued. Previous studies on simple, unhindered alkylphenols have been extended to examine the effects of additional fluorine or other halogen substituents. As in the previous studies, evidence has been derived indicating metabolic transformation via formation of quinone methide intermediates. Additionally, quinone methide metabolic transformation of 4-hydroxyphenylacetone, an analog of acetaminophen, has also been demonstrated, and a glutathione adduct characterized by NMR spectroscopy.