Sargassum horneri (S. horneri) is a well-known brown seaweed widely distributed worldwide. Several biological activities of S. horneri have been reported. However, its effects on lipid metabolism and the underlying mechanisms remain elusive. In the present study, we examined the inhibitory effect of the active compound "(-)-loliolide ((6S,7aR)-6-hydroxy-4,4,7a-trimethyl-5,6,7,7a-tetrahydro-1-benzofuran-2(4H)-one (HTT))" from S. horneri extract on lipid accumulation in differentiated adipocytes. MTT assays demonstrated that (-)-loliolide is not toxic to 3T3-L1 adipocytes in a range of concentrations. (-)-loliolide significantly reduced intracellular lipid accumulation in the differentiated phase of 3T3-L1 adipocytes as shown by Oil Red O staining. Western blot analysis revealed that (-)-loliolide increased the expression of lipolytic protein phospho-hormone-sensitive lipase (p-HSL) and thermogenic protein peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1). Additionally, (-)-loliolide decreased expression of adipogenic and lipogenic proteins, including sterol regulatory element-binding protein-1 (SREBP-1), peroxisome proliferator-activated receptor-γ (PPAR-γ), CCAAT/enhancer-binding protein-α (C/EBP-α), and fatty acid-binding protein 4 (FABP4) in 3T3-L1 adipocytes. These results indicate that (-)-loliolide from S. horneri could suppress lipid accumulation via regulation of antiadipogenic and prolipolytic mechanisms in 3T3-L1 cells. Considering the multifunctional effect of (-)-loliolide, it can be useful as a lipid-lowering agent in the management of patients who suffer from obesity.