The prognosis for recipients of small liver grafts is poor. The aim of this study was to determine the impact of venous systemic oxygen persufflation (VSOP) with nitric oxide (NO) gas for 30% partial liver preservation and transplantation in rats. After we determined optimal NO concentration as 40 ppm in vitro with the isolated perfused rat liver model, we assessed liver injury and regeneration in vivo at 1, 3, 24 and 168 h after transplantation in the following three groups after 3 h-cold storage (n = 20 per group): control group = static storage; VSOP group = oxygen persufflation and VSOP+NO group = oxygen with NO persufflation. The liver graft persufflation was achieved with medical gas via the suprahepatic vena cava; In comparison with control group after transplantation, VSOP+NO preservation (1) increased portal circulation, (2) reduced AST and ALT release, (3) upregulated hepatic endothelial NO synthase, (4) reduced hepatocyte and bileductule damage and (5) improved liver regeneration. These results suggest that gaseous oxygen with NO persufflation is a novel and safe preservation method for small partial liver grafts, not only alleviating graft injury but also improve liver regeneration after transplantation.