A new quantitative thermospray LC-MS method for nicotine and its metabolites in biological fluids

A rapid thermospray liquid chromatography-mass spectrometry (TSP LC-MS) method is described for the simultaneous determination of nicotine and 17 of its metabolites. Chemical ionization of nicotine and its metabolites separated by reversed-phase HPLC is achieved by postcolumn addition of ammonium acetate buffer with the filament of the ion source turned off. Quantification is accomplished by selectively monitoring the unique protonated molecular ion of each metabolite. Trideuterated cotinine serves as an internal standard. Linear responses for cotinine, demethylcotinine, and trans-3'-hydroxycotinine were observed over a concentration range of 20-8000 ng/mL, and 80-8000 ng/ml for nicotine and nicotine-1'-N-oxide. Of the 17 metabolites examined, only nicotine, cotinine, demethylcotinine, and trans-3'-hydroxycotinine were detected in smokers' urine.