Abstract

The formation of the N1-glucuronide metabolite of each nicotine enantiomer was studied in pooled human liver microsomes (n = 6). The metabolite formed from 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 standard. A radiometric HPLC method was used to quantify the metabolite. The specificity of the assay method was demonstrated by experiments in which β-glucuronidase treatment of incubated assay samples resulted in elimination of the peak due to theN1-glucuronide metabolite. The glucuronides ofS(−)- 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, respectively. There is marked stereoselectivity in the apparent intrinsic clearance values (V_max/K_m) in that the value for S(−)-nicotine is 4 times greater than for the R(+)-isomer (1.2 versus 0.31 μl/min/mg of protein).