Urinary metabolites of a novel quinoxaline non-nucleoside reverse transcriptase inhibitor in rabbit, mouse and human: identification of fluorine NIH shift metabolites using NMR and tandem MS

1. The urinary metabolites of (S)-2-ethyl-7-fluoro-3-oxo-3,4-dihydro-2H-qui noxaline-carboxylic acid isopropylester (GW420S67X) have been investigated in samples obtained following oral administration to rabbit, mouse and huma n. GW420S67X underwent extensive biotransformation to form hydroxylated met abolites and glucuronide conjugates on the aromatic ring, and on the ethyl and isopropyl side-chains in all species. In rabbit urine, a minor metaboli te was detected and characterized as a cysteine adduct that was not observe d in mouse or man. 2. The hydroxylated metabolites and corresponding glucuronide conjugates we re isolated by semi-preparative HPLC and characterized using NMR, LC-NMIR a nd LC-MS/MS. The relative proportions of fluorine-containing metabolites we re determined in animal species by F-19-NMR signal integration. 3. The fluorine atom of the aromatic ring underwent NIH shift rearrangement in the metabolites isolated and characterized in rabbit, mouse and human u rine. 4. The characterization of the NIH shift metabolites in urine enabled the d etection and confirmation of the presence of these metabolites in human pla sma.