Background: Iron deficiency anemia is the most prevalent nutrient deficiency during pregnancy, yet there are few data on the effect of prenatal iron supplementation in women in developing countries. Objective: Our objective was to describe the effect of iron supplementation on hematologic changes during pregnancy, and the effect on those changes of adding zinc to the supplements. Design: Pregnant women were enrolled in a randomized, double-masked study conducted at a hospital in a shantytown in Lima, Peru. Women were supplemented daily from 10-24 wk gestation to 4 wk postpartum with 60 mg Fe and 250 μg folic acid with or without 15 mg Zn. Hemoglobin and ferritin concentrations were measured in 645 and 613 women, respectively, at enrollment, at 28-30 and 37-38 wk gestation, and in the cord blood of 545 neonates. Results: No differences in iron status were detected by supplement type, but hematologic changes were related to initial hemoglobin status. Women with anemia (hemoglobin <110 g/L) showed steady increases in hemoglobin concentration throughout pregnancy whereas women with relatively higher initial hemoglobin concentrations had declining values during mid pregnancy, then rising values by 37-38 wk gestation. Women with an initial hemoglobin concentration >95 g/L showed increases in serum ferritin by the end of the pregnancy. Despite supplementation, women with poorer hematologic status; who were younger, single, and multiparous; and who consumed fewer supplements were more likely to have anemia at the end of pregnancy. Conclusions: These hematologic changes are congruent with the effects of iron supplementation reported in placebo-controlled trials and the addition of zinc did not significantly affect them.