The lithium ion (Li+) shows toxicity against Escherichia coli cells when present in a high concentration in the environment. Since Li+ is extruded from cells via a Na+(Li+)/H+ antiporter, this antiporter must be involved in the detoxification of Li+. Two Na+(Li+)/H+ antiporters (NhaA system and NhaB system) are known to be present in E. coli. We investigated the properties of the antiporters and the participation of these systems in the detoxification of Li+ using mutants lacking one of the antiporters, or lacking both of them. Although the affinity for Li+ of the two systems was almost the same, the Vmax value for Li+ transport of the NhaA system was about 12 times larger than that of the NhaB system. Wild type cells were unable to grow in the presence of 0.7 M LiCl. Although a wild type cell and a mutant lacking the NhaB system grew in the presence of 0.6 M LiCl, a mutant lacking the NhaA system did not. This second mutant grew in the presence of 0.1 to 0.2 M LiCl. A mutant lacking both the NhaA and NhaB systems could not grow in the presence of 30 mM LiCl.