## 3 Results  
### Effects of TJ on Cell Viability in 3T3-L1 Cells  
As a result of the cell viability test for 3T3-L1 cells of TJ (0-1000 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)), at least 80% cell viability was observed in concentrations up to 200 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\). In the main experiment, TJ was applied within the range where cytotoxicity was not observed (Figure 1). The Effects of the TJ in the cAMP Level, Intracellular FA, TG, and Glycerol Release Related to the Adipogenesis Mechanism in Differentiated 3T3-L1 Cells  
As a result of measuring the level of cAMP acting on the process of inhibiting adipogenesis in differentiated 3T3-L1 cells, we confirmed that in the DC group (701.7 \(\pm\) 26.5 pmol/mL), there were significant differences compared to that of the NC group (1077.0 \(\pm\) 48.5 pmol/mL). Moreover, PC1 (1484.7 \(\pm\) 66.9 pmol/mL), PC2 (1987.7 \(\pm\) 30.8 pmol/mL), and TJ groups (10: 892.0 \(\pm\) 35.4 pmol/mL, 50: 1095.0 \(\pm\) 81.1 pmol/mL, 100: 1367.7 \(\pm\) 50.5 pmol/mL, 200: 1581.3 \(\pm\) 65.4 pmol/mL, respectively) showed significant increases compared to that of the DC group (Figure 2A). The intracellular FA levels of PC1 (183.0 \(\pm\) 13.6 nmol/mL), PC2 (159.9 \(\pm\) 2.0 nmol/mL), and TJ groups (10: 258.9 \(\pm\) 4.1 nmol/mL, 50: 233.4 \(\pm\) 10.4 nmol/mL, 100: 212.6 \(\pm\) 8.5 nmol/mL, 200: 186.6 \(\pm\) 4.5 nmol/mL, respectively) significantly decreased compared to that of the DC group (287.7 \(\pm\) 11.1 mmol/mL) (Figure 2B). As a result of testing the TG content, we found that the level of the DC group (11.1 \(\pm\) 1.3 mM) significantly increased compared to that of the NC group (2.7 \(\pm\) 0.8 mM). On the other hand, in the PC1 (6.8 \(\pm\) 0.3 mM), PC2(4.7 \(\pm\) 1.1 mM), and TJ groups (10: 10.7 \(\pm\) 0.3 mM, 50: 9.9 \(\pm\) 0.3 mM, 100: 8.3 \(\pm\) 0.1 mM, 200: 7.1 \(\pm\) 0.2 mM, respectively), it significantly decreased relative to that of the DC group (Figure 2C). Upon testing the glycerol release, we found that in the PC1 (1.02 \(\pm\) 0.10 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)), PC2 (1.25 \(\pm\) 0.04 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)), and TJ groups (10: 0.24 \(\pm\) 0.12 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\), 50: 0.51 \(\pm\) 0.13 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\), 100: 0.71 \(\pm\) 0.05 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\), 200: 0.96 \(\pm\) 0.06 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)/mL, respectively), it significantly increased relative to that of the DC group (0.19 \(\pm\) 0.05 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)) (Figure 2D). The TJ Inhibited the Lipid Accumulation Related to the Adipogenesis Mechanism in Differentiated 3T3-L1 Cells  
As a result of measuring the lipid accumulation on the process of inhibiting adipogenesis in differentiated 3T3-L1 cells by conducting Oil Red O staining, we confirmed that the  
Figure 1: Effects of the TJ on cell viability in 3T3-L1 cells. TJ (0–1000 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro\SIUnitSymbolMicro mL}\)). The data are shown as mean \(\pm\) standard deviation (SD; n = 3). * = means significant differences from the controls were evaluated using student’s t-test: * \(p\) < 0.05. TJ, _Trapa japonica_ Flerov extract. OD value of the DC group (1.28 \(\pm\) 0.03) significantly increased relative to that of the NC group (0.07 \(\pm\) 0.02). Furthermore, the result of the PC1 (0.68 \(\pm\) 0.04), PC2 (0.40 \(\pm\) 0.09), and TJ groups (10: 1.02 \(\pm\) 0.09, 50: 0.88 \(\pm\) 0.03, 100: 0.67 \(\pm\) 0.05, 200: 0.59 \(\pm\) 0.03) significantly decreased in a dose-dependent manner relative to the DC group (Figure 3A,B). Figure 3: Effects of the TJ on lipid accumulation related to adipogenesis mechanism in 3T3-L1 adipocytes: (**A**) Oil Red O staining and (**B**) optical density of lipid accumulation. NC, normal control; DC, differentiated control; PC1, catechin 100 \(\upmu\)g/mL; PC2, catechin 200 \(\upmu\)g/mL; JT (10, 50, 100, or 200 \(\upmu\)g/mL).