Endotoxins exist anywhere including in water pools, dust, humidifier systems, and machining fluids. The major causal factor is endotoxins in many serious diseases, such as fever, sepsis, multi-organ failure, meningococcemia, and severe morbidities like neurologic disability, or hearing loss. Endotoxins are also called lipopolysaccharide (LPS) and are important pathogens of acute lung injury (ALI). Rutin has potential beneficial effects including anti-inflammation, antioxidation, anti-hyperlipidemia, and anti-platelet aggregation. Pre-treatment with rutin inhibited LPS-induced neutrophil infiltration in the lungs. LPS-induced expression of vascular cell adhesion molecule (VCAM)-1 and inducible nitric oxide synthase (iNOS) was suppressed by rutin, but there was no influence on expression of intercellular adhesion molecule-1 and cyclooxygenase-2. In addition, activation of the nuclear factor (NF)κB was reduced by rutin. Furthermore, we found that the inhibitory concentration of rutin on expression of VCAM-1 and iNOS was similar to NFκB activation. In conclusion, rutin is a potential protective agent for ALI via inhibition of neutrophil infiltration, expression of VCAM-1 and iNOS, and NFκB activation. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 185–191, 2016.