Background <p>In light of the critical role of adipogenesis and inflammation in the development of obesity, various plant-derived extracts have been reported to attenuate the differentiation of preadipocytes into mature adipocytes. This study elucidated the protective effect of ethanolic lychee seed extract (LSE) on the adipogenesis and inflammation of adipocytes.</p> Methods <p>3T3-L1 cells were treated with or without LSE under the adipogenic condition for 2 days, followed by insulin induction for an additional 6 days. Lipid droplet accumulation in the cytoplasm was evaluated using Oil Red O staining. Genes associated with adipogenesis were assessed by quantitative PCR (qPCR) in differentiated 3T3-L1 cells.</p> Results <p>The results demonstrated that LSE reduced the accumulation of lipid droplets and triglyceride in a dose-dependent manner. LSE (500&#xa0;µg/mL)-treated differentiated cells suppressed the mRNA level of adipogenesis-associated genes (<i>Pparg</i>, <i>Srebf1</i>, <i>Srebf2</i>, <i>Hif1a</i>), lipid metabolism (<i>Cd36</i>, <i>Slc2a4</i>), and insulin signaling (<i>Insr</i>, <i>Irs1</i>, <i>Akt1</i>), while significantly upregulating Adipor1 (<i>Adipor1</i>) and AMP-activated protein kinase α (<i>Ampk</i>) levels when compared to the differentiated group. Additionally, LSE significantly reduced the secretion of inflammatory cytokines, including MCP-1, consistent with the downregulation of <i>NF-κB p65</i> expression. LSE also increased the expression of antioxidant-associated genes (e.g., <i>Sod</i>) while downregulating <i>p47</i>^<i>phox</i>, which may lead to the attenuation of inflammation in mature adipocytes.</p> Conclusions <p>These findings suggest that LSE suppresses adipocyte differentiation by reducing the activation of key adipogenic transcription factors during adipogenesis, indicating that LSE or its essential constituents may be a potential therapeutic alternative for preventing obesity.</p>