Jatupalathika is composed of four dried fruit of Terminalia bellerica (Gaertn.) Roxb, Terminalia chebula Retz. var. chebula, Terminalia arjuna Roxb., and Phyllanthus emblica L. This research aimed to investigate the antiobesity potential of Jatupalathika aqueous extracts on the inhibition of fat accumulation and the induction of lipolysis in 3T3-L1 adipocytes, compared with individual herbs in the formula, gallic acid, and ellagic acid at a series of concentrations of 10-100 g/ml. The viability and proliferation of preadipocytes and differentiated adipocytes were analysed by the MTT technique. Lipid accumulation was measured by oil red O staining. The amount of triglycerides and glycerol released in the medium was analysed by a colorimetric method. Compared with the control, Jatupalathika extracts reduced fat accumulation (3.5 folds at 100 g/ml) and triglyceride content (3 folds at 100 g/ml), but increased glycerol release (4.5 folds at 10 g/ml). These activities could be attributed to the synergy between gallic acid and other bioactive components in the formula. Moreover, Jatupalathika extract (100 g/ml) decreased the % viability of both preadipocytes and differentiated adipocytes by 28.85 1.87% and 32.36 2.66%, respectively. It could be concluded that Jatupalathika extract effectively reduced fat accumulation by the inhibition of lipogenesis and adipogenesis and the induction of lipolysis, which could be related to the induction of cell apoptosis or a reduction in cell proliferation. This herbal formula might potentially be used in the prevention and treatment of obesity. However, further investigations on mechanism of action, in vivo effects, and toxicity of the formula need to be done.