| Summary: | <i>Carthamus tinctorius</i> L. (Compositae; safflower or Hong Hua) has been used in Korean traditional medicine for maintaining the homeostasis of body circulation. Phytochemical investigation was performed on the florets of <i>C. tinctorius</i> by liquid chromatography–mass spectrometry (LC/MS), which afforded two dihydrophaseic acid glucosides (<b>1</b> and <b>2</b>). Isolated compounds were structurally confirmed using a combination of spectroscopic methods including 1D and 2D nuclear magnetic resonance and high-resolution electrospray ionization mass spectroscopy. Their absolute configurations were established by quantum chemical electronic circular dichroism calculations and enzymatic hydrolysis. The anti-adipogenesis activity of the isolated compounds was evaluated using 3T3-L1 preadipocytes. Treatment with the dihydrophaseic acid glucoside (<b>1</b>) during adipocyte differentiation prevented the accumulation of lipid droplets and reduced the expression of adipogenic genes, <i>Fabp4</i> and <i>Adipsin</i>. However, compound <b>2</b> did not affect adipogenesis. Our study yielded a dihydrophaseic acid glucoside derived from <i>C. tinctorius,</i> which has potential advantages for treating obesity.
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