1. The chromatographic elution patterns on Sepharose 6B of the supernatant from mucosal homogenates prepared 10 min after administration of copper into duodenal segments in vivo, indicate that copper is bound preferentially in the fraction of mucosal transferrin. 2. In iron deficiency the amount of 64Cu-copper taken up into the duodenal mucosa is more than two times higher and the amount bound to proteins of the supernatant is also increased to approximately the same degree whereas the amount transferred into the body is diminished to one fourth. 3. In the iron deficient group 64Cu-copper was also bound to a fraction which contains probably metallothionein. 4. The distribution of copper in the supernatant was changed due to a simultaneous administration of iron; the amount of copper bound in the transferrin fraction decreased in favor of the metallothionein fraction and another copper binding fraction was eluted between the transferrin and the metallothionein fraction. 5. Copper in a tenfold molar excess inhibited the iron absorption; simultaneously, the iron bound in the iron binding fractions of the supernatant was remarkably diminished. 6. The results suggest that the affinity of copper to two mucosal iron binding proteins, transferrin and metallothionein, is at least partly responsible for the inhibitory effect of copper on iron absorption in iron deficiency.