Extracorporeal membrane oxygenation (ECMO) can trigger a systemic inflammatory response syndrome (SIRS) with activation of inflammatory, endothelial, and coagulation system pathways. We assessed in a randomized trial whether a) sweep flow nitric oxide (sNO) reduces these responses, b) what the temporal trajectory of these markers post-ECMO commencement is, and c) how serum marker concentrations are associated with survival. Fifty-three patients were randomized (25 sNO, 28 control). Serum levels of inflammatory cytokines (IL-1, IL-6, IL-8, IL-10, TNF-α), complement (C1q, C3, C4, C5a, C9, factor H, factor B), immune regulators (CD40L, P-Selectin), and coagulation factors (PF4, MBL, ADAMTS13) were measured at (t0) and 1, 12, 24 hours post (t1–t3) ECMO initiation. Sweep flow nitric oxide did not alter serum biomarker trajectories. Across sNO and control groups, serum marker concentrations declined from t0 to t1 with variable patterns thereafter, but no consistent ECMO-associated elevation. Elevated IL-6 and IL-8 before and after ECMO commencement were associated with mortality, whereas higher P-Selectin, CD40 Ligand, and PF4 correlated with survival. Sweep flow nitric oxide did not modify markers of inflammation, endothelial, or coagulation activation during ECMO. Evidence of an ECMO-induced cytokine storm was absent, but serum biomarkers predicted mortality in ECMO-supported neonates and children.