The anti-inflammatory mechanism of Ephedrae Herba was explored in this study. Network pharmacology was used to predict the anti-inflammatory active ingredients and potential targets of Ephedrae Herba and protein-protein interaction network was constructed to screen the core targets. Go and KEGG enrichment analysis for common targets were performed using the DAVID database, and molecular docking was performed to verify the binding ability between the active ingredients and the core targets. Inflammation model of RAW 264.7 cells induced by lipopolysaccharide (LPS) was used to verify the anti-inflammatory effect of Ephedrae Herba. The results of network pharmacology analysis identified nine active components in Ephedrae Herba, 195 potential anti-inflammatory candidate targets, and 2098 inflammatory disease-related targets, among which 102 were characterized as common targets between the active components of Ephedrae Herba and inflammatory disease targets. AKT serine/threonine kinase 1 (AKT1), epidermal growth factor receptor (EGFR), proto-oncogene tyrosine-protein kinase Src (SRC), B-cell lymphoma 2 protein (BCL-2), and prostaglandin-endoperoxide synthase 2 (PTGS2) may serve as key anti-inflammatory core targets, which exhibited favorable molecular docking interactions with active components such as pectolinarigenin, genkwanin, naringenin, luteolin, and diosmetin. GO and KEGG enrichment analyses indicated that the anti-inflammatory effects of Ephedrae Herba may be transduced through signaling pathways including the cancer pathway and phosphoinositide 3-kinase(PI3K)-AKT signaling pathway. In vitro cellular experiments demonstrated that Ephedrae Herba extract significantly reduced LPS-induced nitric oxide (NO) release and the expression levels of pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β) and IL-6 (P<0.05 or P<0.01). Additionally, it remarkably downregulated the expression of key inflammation-activated proteins, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), phosphorylated PI3K (p-PI3K), and phosphorylated AKT(p-AKT) (P<0.05 or P<0.01). This study preliminarily confirmed the anti-inflammatory efficacy of Ephedrae Herba, which exerts its anti-inflammatory effects through a multi-component and multi-target manner. Specifically, it may inhibit the PI3K/AKT signaling pathway, downregulate the expression of iNOS and COX-2, and reduce the synthesis and release of NO and pro-inflammatory cytokines to achieve anti-inflammatory effects.