Isolated rat liver mitochondria rapidly accumulate chromate (1.2 microM 51CrO4(2-)) to about 0.25-0.30 nmol Cr/mg protein. The relative uptake decreases with increasing chromate doses. Chromate uptake decreases when pH is raised from 7.0 to 7.5.N-ethylmaleimide (0.25 mM) and butylmalonate (5 mM) inhibit chromate uptake to 70% and 30% of control values, respectively, whereas mersalyl (40 nmol/mg protein) causes an inhibition of greater than 95%. Both sulphate and phosphate decrease mitochondrial chromate uptake, the former being more effective in lower doses (5 mM). These results indicate that transport of chromate is mediated both on the dicarboxylate and the phosphate carrier. The extensive mitochondrial chromium accumulation can be explained by trapping of chromium, probably by reduction of chromate to the trivalent form, within the mitochondria. Release of chromium after chromate loading was seen after 15 min. Added after chromate loading, mersalyl partly prevents this release. Trivalent chromium as 51CrCl3 is taken up to a much lower degree than hexavalent chromium as 51CrO4(2-). The presence of glutathione (5 mM) reduces the uptake both of 51Cr-III and 51Cr-VI, indicating extramitochondrial reduction of Cr-VI to Cr-III and subsequent binding to GSH.