Egg protein ovotransferrin-derived peptides (IRW and IQW) can attenuate tumor necrosis factor (TNF)-induced inflammatory responses and oxidative stress in endothelial cells. The present study investigates the structural requirements and the molecular mechanisms underlying these events. While IRW significantly inhibited TNF-induced up-regulation of intercellular cell adhesion molecule-I (ICAM-1) and vascular cell adhesion molecule-I (VCAM-1), IQW could only inhibit the up-regulation of ICAM-1.The anti-inflammatory effects of these peptides appeared to be mediated by the nuclear factor-κB (NF-κB) pathway, which was differentially regulated by IRW and IQW. Both IRW and IQW exhibited anti-oxidant effect as shown by reduction of TNF-induced superoxide generation. The structural integrity of these peptides was essential for their activities, since dipeptides or the combination of constituent amino acids did not exhibit the same effect. Our study demonstrated the significance of the structural integrity of these two tri-peptides in attenuating endothelial inflammation and oxidative stress, indicating their potential as nutraceuticals.