Increasing flavonoids have been reported to possess anti‐angiogenic effects. Inhibition of angiogenesis plays a critical role in the treatment of cancer, especially in advanced metastatic cancer. In this study, we assessed the effect of Oroxylin A‐7‐glucuronide (Oroxyloside), a main metabolite of Oroxylin A, on angiogenesis in human endothelial cell‐like EA.hy926 cells. Oroxyloside suppressed the migration and tube formation of EA.hy926 cells. Meanwhile, microvessels sprouting from aortic rings and new blood vessels on the chicken chorioallantoic membrane (CAM) were also inhibited. Mechanism studies showed that Oroxyloside reduced the autophosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2/Flk‐1) while it up‐regulated the expression of R‐Ras and VE‐cadherin. In consequence, Oroxyloside inhibited the downstream Akt/MAPK/NF‐κB pathways and then decreased the nuclear translocation and DNA binding ability of NF‐κB. Furthermore, in‐vivo study showed that Oroxyloside exhibited a potential anti‐angiogenic effect in Matrigel plug assay and inhibited growth of xenografted tumors with low systemic toxicity, which could be ascribed to the inhibition of VEGFR2 internalization. Taken together, these results suggested that Oroxyloside could inhibit angiogenesis in vitro and in vivo via suppressing the internalization of VEGFR2 and might serve as a potential antitumor agent. This article is protected by copyright. All rights reserved