The metabolism of 1,2-dihaloethanes (DHEs) to glutathione-containing metabolites by freshly isolated rat hepatocytes was investigated. 1,2-Dichloroethane (DCE), 1,2-dibromoethane (DBE), and 1-bromo-2-chloroethane (BCE) were metabolized to S-(2-hydroxyethyl)glutathione (HEG), S-(carboxymethyl)glutathione (CMG), and S,S'-(1,2-ethanediyl)bis(glutathione) (GEG). The formation of these glutathione-containing metabolites was concomitant with the depletion of intracellular glutathione (GSH) and accounted for 58%, 84%, and 71% of the DCE-, BCE-, and DBE-induced loss of intracellular GSH, respectively. The covalent binding of [14C]DBE to hepatocyte protein reached 18.7 nmol/mL of cell suspension (7.8 nmol/mg of protein) within 2.0 h of incubation. Half of this covalent binding occurred within 0.5 h of incubation (4.0 nmol/mg of protein) in the presence of high levels of intracellular GSH (30% of initial GSH level at 0.5 h). Hepatocyte metabolism of 2-chloroacetic acid produced only CMG. 2-Chloroethanol metabolism gave rise to CMG and HEG in a 11.5:1.0 ratio; 2-chloroacetaldehyde produced almost equal amounts of CMG and HEG. GEG formation was increased significantly for DBE and BCE when GSH was added to the medium during treatment, suggesting that the GSH conjugates S-(2-haloethyl)glutathione are exported from the hepatocytes. These results indicate that the glutathione S-transferase-catalyzed conjugation of GSH with the DHEs is responsible for the majority of the DHE-induced GSH depletion. The S-(2-haloethyl)glutathione conjugates appear responsible for the extensive covalent binding to protein observed during [14C]DBE metabolism.