Background: The effect of vitamin D replacement on hemoglobin (Hb) concentration in subjects with concurrent deficiencies of vitamin D and iron is not known. Methods: We report on an investigator-initiated, randomized, single-blinded, placebo-controlled, 12-week interventional trial. Thirty subjects with iron-deficiency anemia (serum ferritin <15 µg/l) were randomized to an intervention arm (cholecalciferol, i.e. vitamin D3, 0.6 million units i.m. once) or placebo. In all subjects, iron deficiency was corrected with parental iron. Other causes of anemia were excluded with appropriate investigation. The primary end point was a rise in Hb concentration. Results: Baseline parameters of age, BMI, hemogram values and levels of serum ferritin, 25-hydroxyvitamin D [25(OH)D] and parathyroid hormone (PTH) were similar in the 2 arms. Twelve weeks after vitamin D replacement, there was a significant increase in 25 (OH)D levels (57.7 ± 20.5 vs. 14.1 ± 6.2 ng/ml, p < 0.0001) and a decrease in PTH levels (32.4 ± 16.4 vs. 52.9 ± 18.4 pg/ml, p = 0.003) in subjects in the intervention arm when compared to the placebo arm. However, the increments in serum ferritin and Hb concentration in the intervention and placebo arm did not differ. Conclusion: Vitamin D replacement in subjects with iron-deficiency anemia after iron correction does not improve Hb concentration further.