High-Performance Liquid Chromatography with Fluorescence Detection for Simultaneous Analysis of Phytosterols (Stigmasterol, β-Sitosterol, Campesterol, Ergosterol, and Fucosterol) and Cholesterol in Plant Foods

A method based on high-performance liquid chromatography (HPLC) with fluorescence (FL) detection for the simultaneous analysis of phytosterols (stigmasterol, β-sitosterol, campesterol, ergosterol, and fucosterol) and cholesterol was developed. To fluoresceinate the sterols, they were derivatized by 1-anthroyl cyanide to the hydroxyl group at carbon 3 of each sterol skeleton. This HPLC-FL method consists of a C-30 column, an isocratic solution using acetone/acetonitril/hexane/water (71:20:4:5, v/v) as the mobile phase at 1.0 mL min−1 and fluorescence detection at an excitation of 370 nm and an emission of 470 nm. The separation of five phytosterols, cholesterol, and 1-hexacosanol as an internal standard was achieved with sufficient reproducibility and quantitative ability. Our method could evaluate the sterols of land plants such as wood ear fungus, soybean, and parsley, as well as marine algae such as Hiziki (Phaeophyta), Ogonori (Rhodophyta), and Heraiwazuta (Chlorophyta). As a result of the analysis of land plants, wood ear contained a large amount of ergosterol as a precursor of vitamin D2. Soybean contained a large amount of stigmasterol, campesterol, and β-sitosterol. Parsley contained small amounts of these sterols compared with wood ear and soybean. Among the marine algae, Hiziki, Ogonori, and Heraiwazuta contained large amounts of fucosterol, cholesterol, and β-sitosterol, respectively. The compositions of marine algae differed from those of land plants.