Geometry has been considered as one of the important parameters in nanoparticle design because it affects cellular uptake, transport across the physiological barriers, and in vivo distribution. However, only a few studies have been conducted to elucidate the influence of nanoparticle geometry in their in vivo fate after oral administration. This article discloses the effect of nanoparticle shape on transport and absorption in gastrointestinal(GI) tract. Nanorods and nanospheres were prepared and labeled using fluorescence resonance energy transfer molecules to track the in vivo fate of intact nanoparticles accurately. Results demonstrated that nanorods had significantly longer retention time in GI tract compared with nanospheres. Furthermore, nanorods exhibited stronger ability of penetration into space of villi than nanospheres, which is the main reason of longer retention time. In addition, mesenteric lymph transported 1.75% nanorods within 10 h, which was more than that with nanospheres (0.98%). Fluorescent signals arising from nanoparticles were found in the kidney but not in the liver, lung, spleen, or blood, which could be ascribed to low absorption of intact nanoparticles. In conclusion, nanoparticle geometry influences in vivo fate after oral delivery and nanorods should be further investigated for designing oral delivery systems for therapeutic drugs, vaccines, or diagnostic materials.