Formation of the quaternary ammonium-linked glucuronide of nicotine in human liver microsomes: Identification and stereoselectivity in the kinetics

The formation of the N1-glucuronide metabolite of each nicotine enantiomer was studied in pooled human liver microsomes (n=6). The metabolite formed f rom natural S(-)-nicotine was identified by comparison of the high-pressure liquid chromatography (HPLC) retention time and positive ion electrospray ionization-mass spectral characteristics with a synthetic reference standar d. A radiometric HPLC method was used to quantify the metabolite. The speci ficity of the assay method was demonstrated by experiments in which beta -g lucuronidase treatment of incubated assay samples resulted in elimination o f the peak due to the N1-glucuronide metabolite. The glucuronides of S(-)- and R(+)-nicotine were formed by one-enzyme kinetics, with K-m values of 0. 11 and 0.23 mM and V-max values of 132 and 70 pmol/min/mg of protein, respe ctively. There is marked stereoselectivity in the apparent intrinsic cleara nce values (V-max/K-m) in that the value for S(-)-nicotine is 4 times great er than for the R(+)-isomer (1.2 versus 0.31 mul/min/mg of protein).