Toll-like receptors (TLRs) play an important role in initiating innate immune responses. The stimulation of TLRs by microbial components triggers the activation of two downstream signaling pathways: myeloid differential factor 88 (MyD88)- and toll-interleukin-1 receptor domain-containing adapter inducing interferon-$eta$ (TRIF)-dependent signaling pathways. Wood vinegar (WV), a byproduct from charcoal production, has been evaluated for its various biological activities including anti-oxidant, anti-bacterial, and anti-inflammatory activities. To evaluate the therapeutic potential of WV, we examined its effect on signal transduction via the TRIF-dependent pathway of TLRs induced by TLR3 and TLR4 agonists. Oak wood vinegar (OWV) inhibited nuclear factor-$kappa$B and interferon regulatory factor 3 activation induced by lipopolysaccharide(LPS) or polyinosinic-polycytidylic acid (poly [I:C]). OWV also inhibited LPS- or poly[I:C]-induced interferon inducible protein-10(IP-10). These results suggest that OWV can modulate TRIF-dependent signaling pathway of TLRs for anti-viral, anti-bacterial, and anti-inflammatory activities.