Excretion of copper from copper pre-loaded rat liver parenchymal cells was investigated. The copper pre-loaded cells were isolated from the liver of copper-administered rats and were maintained for 4 days in modified Williams medium E; sodium pyruvate was increased while calcium was reduced in concentration, and 10% fetal bovine serum and insulin were added. For the first 30 hr after plating, loss of copper was slow, with a half-life of more than 100 hr: subsequently the rate of loss was faster, with a half-life of 51 +/- 9 hr. Copper pre-loaded in the cells was mostly bound to metallothionein and the amount of copper-metallothionein decreased with time of culture. Furthermore, the relative ratio of the two iso-metallothioneins changed with length of culture.