Abstract: AIM: To study the effects of single or combined administration of tetrahydrobiopterin (BH4) and L-2-amino-5-guanidinovaleric acid (L-Arg) on Langendorff immature rabbit heart models. METHODS: Forty-eight immature rabbits were randomly divided into three test groups and one control group (n=12 per group). The isolated immature rabbit hearts were perfused retrogradely in the Langendorff model. The hearts were perfused with 37℃ Krebs-Henseleit buffer (KHB) for 30 min at 70 cm H2O (perfusion pressure). Coronary flow (CF) was recorded before ischemia and 30 min, 60 min and 120 min of reperfusion. The dynamic changes of myocardial NOS activity, NO content, mitochondrial content of malonaldehyde (MDA) and superoxide dismutase (SOD) were measured at different points. The expression level of myocardial HSP70 was also detected. RESULTS: Compared with that in the control group, the production of NO and SOD significantly increased in other groups, and the production of MDA was significantly degraded in other groups. Compared with that in group II and group III, there were no differences in the production of NO and SOD and MDA. Compared with that in group II and group III, the production of NO and SOD and CF significantly increased in group IV, and the production of MDA was significantly degraded. The expression level of myocardial HSP70 was significantly increased. CONCLUSION: Compared with the single administration, the combined administration of BH4 and L-Arg may have better cardioprotective effects by increasing NO and preserving the function of mitochondria.