Fructose-1,6-diphosphate (FDP), a glycolytic metabolite, has been reported to protect susceptible organs during hypoxia or ischemia. However, there is paucity of human data on its pharmacokinetics after being exogenously administered. In the current study, the preliminary pharmacokinetics of FDP given orally to humans was investigated, and no typical peak was observed in the serum drug-time curve. Then, the pharmacokinetic studies were performed following multiple doses of FDP in rats, and the Caco-2 monolayer model was used to study the absorption of FDP in vitro. The results suggested that plasma FDP concentration was significantly increased after oral multiple doses of 180 mg kg-1 but not 90 mg kg-1 of FDP, and FDP was partly depleted during the absorption, which was supposed to be consumed by the intestinal epithelium cells. Thus, we conclude that a high dose of FDP should be orally administered in order to get an effective plasma level.