Medicinal plants have long been used as an alternative to traditional drugs for the treatment of inflammatory conditions due to the classical side effects and restricted access of various commercially available drugs, such as steroids (GCs) and nonsteroidal anti-inflammatory drugs (NSAIDs). Sambucus australis is a Brazilian herb that is commonly used to treat inflammatory diseases; however, few studies have examined the use of this species in the treatment of inflammatory conditions. The present study aims to evaluate the potential anti-inflammatory activity of S. australis in vitro. We established spleen cell cultures stimulated with pokeweed mitogen (PWM) to evaluate the production of proinflammatory cytokines, such as IL-4, IL-5, IFN-y, and IL-10 (by ELISA), and the expression of the transcription factor NF-kB (by RT-PCR). In addition, we evaluated the levels of nitric oxide in macrophage cultures and the membrane-stabilizing activity of S. australis methanolic extract (EMSA). Treatment with EMSA at concentrations of 100, 50, 25 and 12.5 µg/ml significantly decreased IL-4 (p<0.001) and IL-5 (p<0.001) levels. Treatment with 100 µg/ml EMSA reduced IFN-у (p<0.001) levels. Moreover, at 100 mg/ml, EMSA also increased IL-10 production and reduced NF-kB expression (p<0.01). In macrophage cultures stimulated with LPS, EMSA decreased nitric oxide levels (p<0.001) at all concentrations tested (100, 50, 25 and 12.5 µg/ml). Additionally, EMSA had a protective effect in the erythrocyte membrane stabilization assay. Taken together, these results suggest that S. australis has anti-inflammatory potential in vitro, characterized by the reduction of both inflammatory cytokines and the expression of NF-kB along with the up-regulation of IL-10.