Aquatic weeds are widely used as animal feed in developing countries. However, information concerning element bioavailability from these plants is lacking. A combination of an in vitro method (PBET) and an in vivo feeding trial was conducted in the present study in order to investigate potential element bioaccessibility and real bioavailability from Pistia stratiotes (PS). Cu, Fe, Mn, Zn and Pb concentrations in PS biomass, artificial gastrointestinal fluids and rat tissues were determined using ET-AAS and ICP-OES. PS exhibited elevated Fe, Mn and Pb levels. The PBET revealed high bioaccessibility of all monitored elements from PS biomass. The results of the in vivo trial were inconsistent with those of the PBET, because animals fed PS exhibited low levels of essential elements in the tissues. The consumption of a PS supplemented diet significantly decreased total Fe levels and increased total Pb accumulation in exposed animals. Significantly reduced amounts of essential elements in the intestinal walls indicated a potential disruption in nutrient gastrointestinal absorption in animals fed PS.