The anti-obesity activity of the water extract of soy milk fermented with Lactobacillus paracasei subsp. paracasei NTU 101 (W101) was investigated. A high-fat diet (HFD) was used to induce obesity in rats, and the effects of daily W101 feeding (8 weeks) were observed. The rats fed the HFD and supplemented with low-dose W101 (LW101, 15 mg per kg body weight per day) or high-dose W101 (HW101, 150 mg per kg body weight per day) had significantly reduced final body weight in comparison with that of the HFD group. W101 decreased the formation of lipid plaques in the aorta, reduced the adipocyte cross-sectional area and diameter, and reduced the levels of CCAAT/enhancer-binding protein β (C/EBPβ), peroxisome proliferator associated receptor γ (PPARγ), and C/EBPα. Regarding lipogenesis regulation in adipocytes, W101 suppressed heparin-releasable lipoprotein lipase (HR-LPL) in adipose tissues and inhibited lipid absorption, thereby reducing lipogenesis. Lactobacillus paracasei subsp. paracasei NTU 101-fermented soy milk may be used to develop health foods that prevent obesity.