Novel Mechanism for Dehalogenation and Glutathione Conjugation of Dihalogenated Anilines in Human Liver Microsomes: Evidence for <i>ipso</i> Glutathione Addition ZhangChenghong KennyJane R. LeHoa DeeseAlan FordKevin A. LightningLuke K. FanPeter W. DriscollJames P. HalladayJason S. E. C. A. HopCornelis KhojastehS. Cyrus 2011 The objective of the present study was to investigate the influence of halogen position on the formation of reactive metabolites from dihalogenated anilines. Herein we report on a proposed mechanism for dehalogenation and glutathione (GSH) conjugation of a series of <i>ortho</i>-, <i>meta-,</i> and <i>para</i>-dihalogenated anilines observed in human liver microsomes. Of particular interest were conjugates formed in which one of the halogens on the aniline was replaced by GSH. We present evidence that a (4-iminocyclohexa-2,5-dienylidene)halogenium reactive intermediate (QX) was formed after oxidation, followed by <i>ipso</i> addition of GSH at the imine moiety. The <i>ipso</i> GSH thiol attacks at the <i>ortho-</i>carbon and eventually leads to a loss of a halogen and GSH replacement. The initial step of GSH addition at the <i>ipso</i> position is also supported by density functional theory, which suggests that the <i>ipso</i> carbon of the chloro, bromo, and iodo (but not fluoro) containing 2-fluoro-4-haloanilines is the most positive carbon and that these molecules have the favorable highest occupied molecular orbital of the aniline and the lowest unoccupied orbital from GSH. The <i>para</i>-substituted halogen (chloro, bromo, or iodo but not fluoro) played a pivotal role in the formation of the QX, which required a delocalization of the positive charge on the <i>para</i>-halogen after oxidation. This mechanism was supported by structure–metabolism relationship analysis of a series of dihalogenated and monohalogenated aniline analogues.