Smilax sieboldii, a climbing tree belonging to Smilacaceae, has been used in traditional oriental medicine for treating arthritis, tumors, leprosy, psoriasis, and lumbago. To evaluate the anti-obesity effects ofS. sieboldii(Smilacaceae), we screened methylene chloride (CH2Cl2), ethyl acetate (EtOAc), aqueous-saturatedn-butanol, and ethanol (EtOH) extracts of the whole plant at various concentrations to inhibit adipogenesis in adipocytes. The 3T3-L1 cell line with Oil red O staining with the help of fluorometry was used as an indicator of anti-obesity activity. Bioactivity-guided fractionation of the EtOH extract and subsequent phytochemical investigation of the active CH2Cl2- and EtOAc-soluble fractions resulted in the isolation of 19 secondary metabolites (1–19), including a new α-hydroxy acid derivative (16) and two new lanostane-type triterpenoids (17and18). The structures of these compounds were characterized using various spectroscopic methods. All the isolated compounds were screened for adipogenesis inhibition at a concentration of 100 μM. Of these, compounds1,2,4–9,15, and19significantly reduced fat accumulation in 3T3-L1 adipocytes, especially compounds4,7,9, and19, showing 37.05 ± 0.95, 8.60 ± 0.41 15.82 ± 1.23, and 17.73 ± 1.28% lipid content, respectively, at a concentration of 100 μM. These findings provide experimental evidence that isolates fromS. sieboldiiextracts exert beneficial effects regarding the regulation of adipocyte differentiation.