Probucol 4,4′- (Isopropylidenedithio)bis(2,6-di-tert-butylphenol) is a synthetic molecule clinically used for prevention and treatment of hypercholesterolemia and atherosclerosis. Recent studies have shown that the beneficial effects of probucol mainly derive from its anti-inflammatory and antioxidant properties. Gram-negative bacteria are common infectious agents and their wall components, e.g. lipopolysaccharide (LPS), are important elicitors of inflammation. LPS is sensed by tissue resident cells and it triggers a Toll-like receptor 4/MyD88-dependent signaling cascade resulting in endothelial activation, leukocyte recruitment and nociception. Therefore the present study aimed to investigate the anti-inflammatory and analgesic effects of probucol in models of LPS-induced acute inflammation. Probucol at 0.3–30 mg/kg was administrated to male Swiss mice per oral 1 h before intraplantar or intraperitoneal lipopolysaccharide stimulus. Probucol at 3 mg/kg reduced lipopolysaccharide-induced mechanical and thermal hyperalgesia. These effects were accompanied by reduced leukocyte influx and cytokine production in both paw skin and peritoneum exudate. Unexpectedly, probucol did not alter lipopolysaccharide-induced tissue oxidative stress at anti-inflammatory /analgesic dose. On the other hand, probucol inhibited lipopolysaccharide-induced nuclear factor kappa B (NF-κB) activation in paw tissue as well as NF-κB activity in cultured macrophages in vitro, reinforcing the inhibitory effect of probucol over the NF-κB signaling pathway. In this sense, we propose that probucol acts on resident immune cells, such as macrophages, targeting the NF-κB pathway. As a result, it prevents the amplification and persistence of the inflammatory response by attenuating NF-κB-dependent cytokine production and leukocyte recruitment explaining its analgesic effects as well.