Study design for studies 1-3. (A) Study 1 (n = 25), fasting subjects received 40, 80, 160, and 240 mg Fe at 8:00 am and were randomly allocated to start the study either with single or with consecutive day doses (crossover design). Subjects acted as their own controls. Hepcidin and iron status was assessed at 8:00 am,12:00 pm, and 5:00 pm (days 1-2) and at 8:00 am on days 3, 4, and 5 (single dose schedule) or at 8:00 am,12:00 pm, and 5:00 pm (days 1-3) and 8:00 am on days 4 and 5 (consecutive dose schedule). (B) Study 2 (n = 16) foresaw only 1 week of supplementation and only 2 consecutive 60-mg Fe doses. (C) Study design of study 3 (n = 13) where bi-daily supplementation was tested: the diet of the subjects was controlled between subjects to maintain at least 3 hours of fasting between iron dosages, which were given at 10 hours and at 16 hours after a standardized breakfast and lunch, respectively. A full description and more detailed representation of the study design are available as online supplemental material. Numbers refer to consecutive study days. LFe, labeled iron supplement administration; L, determination of isotopic composition (iron absorption).