750 MHz HPLC-NMR spectroscopic identification of rat microsomal metabolites of phenoxypyridines.
Directly coupled 750 MHz HPLC-1H NMR spectroscopy has been applied to the characterisation of low level metabolites of 3-amino-2-(2-fluorophenoxy)pyridine (AP) and 3-nitro-2-(2-fluorophenoxy)pyridine (NP) in rat microsomes. In stop-flow HPLC-NMR mode, the direct injection of microsomal extracts enabled the separation and characterisation of minor metabolites. NP is converted into AP to an extent of 93.4% and this is further metabolised to 4- and 6-hydroxy-AP (6 and 0.6% respectively). Unequivocal identification of these metabolites was achieved without the use of a radiolabel or synthetic standards and thus demonstrates the applicability of directly coupled HPLC-NMR to metabolite identification in in vitro systems. The potential exists for HPLC-NMR and HPLC-NMR-MS to provide rapid metabolic information within the timescale of high throughput lead optimisation exercises in drug discovery.
Corcoran
O
O
Department of Chemistry, Birkbeck College, University of London, UK.
Spraul
M
M
Hofmann
M
M
Ismail
I M
IM
Lindon
J C
JC
Nicholson
J K
JK
D000818:Animals; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D066298:In Vitro Techniques; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D011725:Pyridines / Q000032:analysis / Q000493:pharmacokinetics*; D051381:Rats; D013056:Spectrophotometry, Ultraviolet
D011725:Pyridines