Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) has been identified as a critical mediator of cell death (necroptosis and apoptosis) and inflammation. Necrostatin-1 (Nec-1) and 7-Cl-O-Nec-1 (Nec-1s) are widely used as selective small-molecule inhibitors of RIPK1 in various culture cells and disease models. NAD(P)H: quinone oxidoreductase 1 (NQO1) is a ubiquitous flavoenzyme that catalyzes the reduction and detoxification of quinones and other organic compounds. Here, we showed that Nec-1 and Nec-1s could bind and inhibit NQO1 activity. Similar to dicoumarol, the specific inhibitor of NQO1, both Nec-1 and Nec-1s significantly suppress NQO1-dependent cell death. However, dicoumarol failed to reverse necroptosis induced by TNFα/BV6/Z-VAD-FMK (TBZ) in HT29 cells. These findings suggest that besides RIPK1, NQO1 might be another target for Nec-1 and Nec-1s and provide new insights for the interpretation of Nec-1-based experimental results.