Hesperetin, a major bioflavonoid in sweetoranges and lemons, exerts an anti-inflammatory effect inpulmonary diseases; however, its effect on lipopolysaccharide(LPS)-induced acute lung injury is unclear. This studyinvestigated the effect of hesperetin on LPS-induced lunginflammatory response. Mice were intratracheally instilledwith 5 mg/kg body weight LPS, and then were given hesperetinorally (10, 20, and 30 mg/kg body weight) 1 h later. Hesperetin dramatically suppressed the levels of interleukin-6 and tumor necrosis factor-α, as well as the number ofinflammatory cells in bronchoalveolar lavage fluid. Besides,it reduced lung injury, wet weight/dry weight ratio, andmyeloperoxidase and lactate dehydrogenase activities, andenhanced superoxide dismutase activity. In addition, hesperetinsignificantly downregulated the Toll-like receptor4 (TLR4) and myeloid differentiation factor 88 (MyD88)protein expression and suppressed nuclear factor-kappa B(NF-κB) activation in lung tissue. Together, these resultsindicated that the anti-inflammatory effect of hesperetin isassociated with the TLR4–MyD88–NF-κB pathway, andthat hesperetin shows therapeutic potential for LPS-inducedacute lung injury.