The importance of chromatographic separation in LC/MS/MS quantitation of drugs in biological fluids: detection, characterization, and synthesis of a previously unknown low-level nitrone metabolite of a substance P antagonist.
The observation of an interference peak in plasma samples from dogs dosed with compound I led to the discovery of an unidentified metabolite. The unknown metabolite had the same molecular weight as the parent drug, and their fragmentation profiles were also quite similar. LC/MS/ MS analysis of the plasma extracts of dogs and rats dosed with I and its deuterium-labeled analogue suggested a nitrone structure for the unknown metabolite. Synthesized nitrone matched the unknown metabolite with identical retention time and nearly identical fragmentation profile. The nitrone slowly decomposed in acidic aqueous solution at ambient temperature and also underwent in-source, thermal-induced hydrolysis during electrospray ionization mass spectrometric analysis. The reaction of the nitrone with diethyl acetylenedicarboxylate readily generated a [2 + 3] cycloaddition product. The example shown here clearly demonstrates that precautions must be taken when LC/MS/MS quantitation is conducted in the selected reaction monitoring mode.
Wu
Yunhui
Y
Department of Pharmaceutical Research & Development, Merck Research Laboratories, P.O. Box 4, West Point, Pennsylvania 19486, USA. yunhui_wu@merck.com
Farrell
James T
JT
Lynn
Kari
K
Euler
Danielle
D
Kwei
Gloria
G
Hwang
Tsang-Lin
TL
Qin
Xue-Zhi
XZ
D000818:Animals; D001826:Body Fluids; D002853:Chromatography, Liquid; D004285:Dogs; D009589:Nitrogen Oxides / Q000032:analysis; D004364:Pharmaceutical Preparations / Q000097:blood; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization; D013373:Substance P / Q000037:antagonists & inhibitors* / Q000097:blood* / Q000378:metabolism
D009589:Nitrogen Oxides; D004364:Pharmaceutical Preparations; C015388:nitrones; D013373:Substance P