Our previous studies have shown that the baicalin could blocked infection of chlamydia trachomatis (C. trachomatis)-infected cells in vitro. Toll-like receptor 2 and 4 (TLR2/4) and the downstream nuclear factor-κB (NF-κB) signaling pathway, which mediate the inflammatory reaction, are involved in the pathophysiological processes of inflammation. In this study, we investigated whether baicalin inhibits TLR2/4 signaling pathway in gential tract chlamydia-infected mice. The progesterone-treated animals were given intravaginally 200 mg/kg baicalin administered. Nineteen days after infection, cervical tissue were taken and expression of TLR2/4, NF-κB were determined by RT-PCR or westernblot. Nitric oxide and prostaglandin E2 production in cervical tissue were detected by enzyme-linked immunosorbent assay. It was demonstrated that baicalin significantly reduced C. trachomatis loading in BALB/c mice that were vaginally infected with the pathogen. Meanwhile, baicalin also reduced the expression of TLR2/4 and NF-κB, decreased activity of inducible nitric oxide synthase and cyclooxgenase-2 in cervical tissue. Our results suggest that baicalin inhibits the TLR2/4 signaling pathway in cervical tissue of gential tract chlamydia-infected mice. On the basis of these data and our previous observations, we conclude that further evaluation of baicalin for prevention and treatment of sexually transmitted chlamydial infection is warranted.