The effects of adding increasing concentrations of magnesium (Mg), zinc (Zn), and iron (Fe) on calcium (Ca) uptake into rat brush border membrane vesicles was measured. At ratios of 1:1, based on the RDA, none of these minerals were found to significantly decrease calcium uptake. Mg, however, inhibited Ca uptake at an RDA ratio of 3:1 (Mg:Ca). At higher RDA ratios (Zn:Ca 10:1), Zn also decreased Ca uptake. In a comparison of the molar ratios of the above cations, Zn inhibited Ca uptake the most (0.12:1), possibly by competing directly at high-affinity Ca binding sites. Fe did not decrease Ca uptake but rather demonstrated an enhancing effect at high concentrations (20:1). When mixtures of the above divalent cations were studied, the effects on Ca uptake could largely be explained by the actions of the ions individually.