TY - JOUR T1 - Characterization of Nicotine and Cotinine<em>N</em>-Glucuronidations in Human Liver Microsomes JF - Drug Metabolism and Disposition JO - Drug Metab Dispos SP - 1484 LP - 1490 DO - 10.1124/dmd.30.12.1484 VL - 30 IS - 12 AU - Miki Nakajima AU - Eriko Tanaka AU - Jun-Tack Kwon AU - Tsuyoshi Yokoi Y1 - 2002/12/01 UR - http://dmd.aspetjournals.org/content/30/12/1484.abstract N2 - The nicotine and cotinine N-glucuronidations in human liver microsomes were characterized. The Eadie-Hofstee plots of nicotine N-glucuronidation in human liver microsomes were clearly biphasic, indicating the involvement of multiple enzymes. The apparentKm and Vmaxvalues were 33.1 ± 28.1 μM and 60.0 ± 21.0 pmol/min/mg and 284.7 ± 122.0 μM and 124.0 ± 44.0 pmol/min/mg for the high- and low-affinity components, respectively, in human liver microsomes (n = 4). However, the Eadie-Hofstee plots of cotinine N-glucuronidation in human liver microsomes were monophasic (apparent Km= 1.9 ± 0.3 mM, Vmax = 655.6 ± 312.3 pmol/min/mg). The nicotine and cotinineN-glucuronidations in the recombinant human UDP-glucuronosyltransferases (UGTs) (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A8, UGT1A9, UGT1A10, UGT2B7, and UGT2B15) expressed in baculovirus-infected insect cells or human B-lymphoblastoid cells that are commercially available were determined. However, no recombinant UGT isoforms showed detectable nicotine and cotinineN-glucuronides (the concentrations of nicotine and cotinine were 0.5 and 2 mM, respectively). Nicotine and cotinineN-glucuronidations in pooled human liver microsomes were competitively inhibited by bilirubin as a substrate for UGT1A1 (Ki = 3.9 and 3.3 μM), imipramine as a substrate for UGT1A4 (Ki = 6.1 and 2.7 μM), and propofol as a substrate for UGT1A9 (Ki = 6.0 and 12.0 μM). The nicotineN-glucuronidation (50 μM nicotine) in 14 human liver microsomes was significantly (r = 0.950,P &lt; 0.0001) correlated with the cotinineN-glucuronidation (0.2 mM cotinine), indicating that the same isoform(s) is involved in both glucuronidations. Furthermore, weak correlations between imipramine N-glucuronidation and nicotine N-glucuronidation (r = 0.425) or cotinine N-glucuronidation (r = 0.517) were observed. In conclusion, the involvement of UGT1A1 and UGT1A9 as well as UGT1A4 in nicotine and cotinine N-glucuronidations in human liver microsomes was suggested, although the contributions of each UGT isoform could not be determined conclusively. The American Society for Pharmacology and Experimental Therapeutics ER -