In addition to well-known analgesic action of tramadol, its potential antinflammatory effects have not been thoroughly evaluated. On the other hand, effectiveness of antioxidants is also reported against inflammation. It is known that glyceryl trinitrate, as a nitric oxide donor, enhance the antioxidative and anti-inflammatory effects. In the present study, the efficacy of the tramadol mixtue with glyceryl trinitrate on cytokines, NF-kappa B expression and oxidative stress marker was examined on the formalin-induced inflammation in rats (Tramadol 5, 10 and 30 mg/kg + nitroglycerine 1 mg/kg). Cytokines (TNF-, IL-6 and IL-10) and oxidative/anti-oxidative stress markers (MDA, GSH) were measured in blood samples. NF-kappa B expression was assessed immunohistochemically in spleen and thymus. The results show that tramadol 30 mg/kg has both anti-inflammatory and anti-oxidative effects. Additionally, it was evidenced that glyceryl trinitrate improves the antiinflammatory and anti-oxidative effects of Tramadol (30 mg/kg) on the formalin-induced inflammation in rats. In this framework, the present study provides a unique approach for the analysis of the efficacy of tramadol and additive effects of glyceryl trinitrate on the acute inflammations in rats.