Background: Besides anemia, iron deficiency may cause more subtle symptoms including those of the restless legs syndrome (RLS), the chronic fatigue syndrome (CFS) or sleeping disorders. Objective: The aim of this pre-planned secondary analysis was to compare the frequency and severity of symptoms associated with iron deficiency before and after (intravenous or oral) iron supplementation in iron deficient blood donors. Methods/Design: Prospective, randomized, controlled, single centre trial. (ClinicalTrials.gov: NCT01787526). Setting: Tertiary care center in Graz, Austria Participants: 138 female and 38 male whole blood and platelet apheresis donors aged ≥18 and ≤65 years with iron deficiency (ferritin ≤30ng/ml at the time of blood donation). Interventions: Intravenous iron (1 g ferric carboxymaltose, n=86) or oral iron supplementation (10 g iron fumarate, 100 capsules, n=90). Measurements: Clinical symptoms were evaluated by a survey before iron therapy (visit 0, V0) and after 8-12 weeks (visit 1, V1) including questions about symptoms of RLS, CFS, sleeping disorders, quality of life and symptoms like headaches, dyspnoea, dizziness, palpitations, pica and trophic changes of fingernails or hair. Results: We found a significant improvement in the severity of symptoms for RLS, fatigue and sleep quality (p<0.001). Furthermore, a significant decrease of headaches, dyspnoea, dizziness and palpitations was reported (p<0.05). There was no difference between the type of iron supplementation (intravenous versus oral) and clinical outcome data. Conclusion: Iron supplementation in iron deficient blood donors may be an effective strategy to improve symptoms related to iron deficiency and the wellbeing of blood donors.