Supercritical fluid CO2 extraction followed by polystyrene/divinylbenzene stationary phase complementary supercritical fluid chromatography and reversed-phase liquid chromatography separation of unsaturated fatty acids.

Supercritical fluid chromatography (SFC) has emerged as a powerful separation technique with extensive applications in pharmaceutical analysis, environmental monitoring, and food safety testing. In this study, weakly polar unsaturated fatty acids were successfully extracted and separated via a polystyrene/divinylbenzene (PS-DVB) stationary phase complementary supercritical fluid chromatography and reversed-phase liquid chromatography (RPLC) approach. The weakly polar sample was obtained using supercritical fluid extraction (SFE) of Floccularia luteovirens powder. The extracted weakly polar sample was first separated using a PS-DVB stationary phase SFC, yielding 304.7 mg of the target fraction Fr2. Then, the established complementary SFC and RPLC method was applied to the separation and purification of compounds from the target fraction Fr2. Finally, three unsaturated fatty acids with purities exceeding 95 % were successfully prepared. This approach has significant potential for analyzing and purifying weakly polar natural products on the same stationary phase using SFC and RPLC.