This study aimed to investigate the structural characterization of polysaccharide from Sambucus williamsii Hance roots (SWH1-1) and its effects on inflammation and acute lung injury (ALI). Results showed that SWH1-1 had a molecular weight of 10,040 Da and primarily consisted of arabinose, galactose, glucose and mannose. Its main backbone was composed of →6)-α-D-Glcp-(1→, →6)-α-D-Man-(1→, and →6)-β-D-Galp-(1→. Besides, SWH1-1 could alleviate nitric oxide (NO), tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) contents in lipopolysaccharide (LPS)-treated RAW 264.7 cells. In mice experiment, SWH1-1 alleviated LPS-induced lung injury by reversing lung histological changes, inhibiting apoptosis, and reducing myeloperoxidase (MPO) levels. SWH1-1 also reduced TNF-α, IL-6 and IL-1β values in vivo. SWH1-1 reduced oxidative stress by upregulating superoxide dismutase (SOD), glutathione (GSH) and downregulating malondialdehyde (MDA) contents. Moreover, SWH1-1 inhibited the activation of the Toll-like receptor 4 (TLR4)/nuclear factor κB (NF-κB)/mitogen-activated protein kinase (MAPK) pathway in LPS-injured lung tissues. The results of RAW264.7 cell experiments further verified that SWH1-1 inhibited TLR4/NF-κB/MAPK pathway. Our research displayed that SWH1-1 improved LPS-induced ALI by alleviating inflammation, oxidative stress and TLR4/NF-κB/MAPK pathway. These results suggest that SWH1-1 is a potential anti-inflammatory candidate for the treatment of LPS-induced acute lung injury.