Yukmijihwang-tang (YMT) is a traditional herbal medicine known to enhance memory in brain injury models. The aims of this study were to evaluate the inhibitory effect of YMT on osteoclast differentiation and to determine its molecular mechanism of action. YMT dose-dependently inhibited receptor activator for nuclear factor-κB (NF-κB) ligand (RANKL)-induced tartrate-resistant acid phosphatase (TRAP) activity and the formation of multinucleated osteoclasts in RAW264.7 cells. In addition, quantitative reverse transcription-polymerase chain reaction showed that YMT significantly decreased RANKL-induced expression of osteoclast differentiation-specific genes (TRAP, matrix metalloproteinase-9, cathepsin K, and the d2 isoform of vacuolar ATPase V(0) domain). Furthermore, YMT inhibited RANKL-induced phosphorylation of mitogen-activated protein kinases (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38), phosphorylation of I-κBα, phosphorylation of NF-κB p65, and the expression of transcription factors Fra-2 and nuclear factor of activated T-cells, cytoplasmic 1. Furthermore, YMT inhibited the bone-resorptive activity of differentiated osteoclasts, suggesting that YMT inhibits osteoclast differentiation by suppressing RANKL-induced signaling molecules and transcription factors that affect the regulation of genes for osteoclast differentiation. As such, YMT may have therapeutic potential in bone diseases.