© 2022 Elsevier B.V.Inflammation is a critical defense mechanism for body which is crucial for wound healing. However, when it spirals out of control and becomes chronic, it possesses a risk for diseases such as metabolic disorders or cancer. Scutellaria L. species are known with their potent anti-inflammatory effects. We aim to investigate the potential anti-inflammatory effects of Scutellaria brevibracteata Stapf and its active principles with underlying mechanisms using both in vitro and in vivo methods. Aqueous extract of S. brevibracteata (SB) was chromatographed on several columns to get 6 main fractions and 2 pure compounds. The levels of IL-6, NO, and TNF-α were measured in LPS-induced RAW 264.7 cells. In vivo carrageenan-induced paw edema test and immunoblotting in these paws were performed. SB was significantly reduced the inflammation in LPS-induced RAW cells via IL-6 and TNF-α cytokines at 200 μg/mL (P < 0.001) and paw edema via COX-2 and iNOS (P < 0.001). 3-O-methyl kaempferol, isolated from SB, reduced the production of NO, IL-6 and TNF-α in LPS-stimulated macrophages at 50 μM (P < 0.001); it significantly diminished inflammation-induced edema and COX-2, iNOS, and NF-κB protein levels were reduced compared to control group (P < 0.001). It was noticed that the methoxylation of C3 position has no effect on the anti-inflammatory mechanism of action comparing with kaempferol. In vivo studies indicated 3-O-methyl kaempferol was as effective as indomethacin. Considering the side effects caused by indomethacin, this compound can be evaluated as a novel drug candidate in inflammatory diseases with fewer side effects.