Isolation and Purification of Six Bioactive Compounds from the Seeds of Trigonella foenum-graecum L. using High-Speed Counter-Current Chromatography

It is difficult to simultaneously isolate and purify flavonoids and polyhydroxystilbenes due to their similar polarities. In this study, we isolated and purified these compounds from Trigonella foenum-graecum L. seed extracts using high-speed counter-current chromatography (HSCCC) in two steps with two different kinds of solvent systems. In the first step, n-hexane:ethyl acetate:methanol:water (1:5:1:5, v/v) was used as the two-phase solvent system, desoxy-rhaponticin and rhapontigenin were purified, and orientin, vitexin, isovitexin, and rhaponticin were eluted together. In the second step, ethyl acetate:water (1:1, v/v) was used as the two-phase solvent system, and orientin, vitexin, rhaponticin, and isovitexin were isolated and purified. Purities of the isolated compounds were determined using HPLC, while UV spectra, ¹ H-NMR, and ¹³ C-NMR aided in structure identification.