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To determine the natural course of zinc protoporphyrin/heme ratio (ZnPP/H) and its role in the detection of iron deficiency (ID) and iron-deficiency anemia (IDA) in the first 4 months of life in moderately preterm infants.ZnPP/H was measured at 1 week, 6 weeks and 4 months postnatal age in a prospective cohort of 161 Dutch infants born at a gestational age of 32+0 to 36+6 weeks who did not receive an erythrocyte transfusion or iron supplementation.ZnPP/H levels decreased in the first 6 weeks and increased thereafter. At 4 months postnatal age, ZnPP/H was higher in the 11 (8.5%) infants with IDA (mean (s.d.): 260.8 (16.1)) but not in the 27 (21.3%) infants with ID (mean (s.d.): 177.0 (15.1)) compared with normal infants (mean (s.d.): 157.3 (12.5)).In moderately preterm infants, ZnPP/H can be of additional value to detect infants at risk for IDA due to iron-deficient erythropoiesis at 4 months of age.