Oxidative and defluorinative metabolism of fludalanine, 2-2H-3-fluoro-D-alanine

The antibacterial agent fludalanine [2-2H-3-fluoro-D-alanine (DFA)] is a potent inhibitor of bacterial alanine racemase, an enzyme required for the generation of D-alanine, an essential component of the bacterial cell wall. Primary metabolism of DFA involves its oxidation to fluoropyruvate (FP); this organic fluoride is then rapidly reduced to fluorolactate (FL) which is the major organic metabolite in laboratory animals. Gas-liquid chromatographic chemical ionization mass spectrometric assays were developed for these two metabolites. FL is the predominant organofluoride metabolite of DFA in the circulation. FP was detected in the urine although recovery was very low. The rapid conversion of FP to FL precludes assay of the former in serum. Maximum serum FL concentrations in the rat appear about 1 hr after the dose of DFA and are relatively constant for several hours thereafter. The peak FL concentration is proportional to the dose of DFA; repeated daily dosing of DFA appears to cause neither saturation nor induction of metabolic pathways. Comparison of FL concentrations determined using the GC/MS assay with those based on an enzymic method specific for L-(+)-FL demonstrated that only the latter isomer is found in the plasma of monkeys dosed with DFA. In vivo exchange studies involving the alpha-proton of FL indicate that a small FP pool exists and is in equilibrium with FL. A crude pyruvate dehydrogenase complex isolated from beef heart mitochondria was shown to produce equimolar quantities of acetate, CO2, and fluoride from FP.