Application of pentafluorophenyldimethylsilyl derivatization for gas chromatography-electron-capture detection of supercritically extracted sterols

We report a convenient method of extracting and detecting sterols in environmental water samples. Particle-associated sterols were extracted onto glass-fibre filters then the filters were supercritical fluid extracted (at 33 MPa, 80°C for 30 min) followed by pentafluorophenyldimethylsilyl (flophemesyl) derivatization for gas chromatographic-electron-capture detection (GC-ECD). Optimal derivatization of epicoprostanol, coprostanol, cholesterol, dihydrocholesterol, stigmasterol, β-sitosterol and stigmastanol with neat flophemesyl chloride was achieved under mild conditions (i.e., room temperature, 15 min) without the need for added solvent. These sterols gave linear GC-ECD calibrations (r² ≤ 0.99) while detection limits in the final solutions were 0.1-0.6 μg/ml. Overall recoveries for the supercritical extraction and derivatization steps were 80-110% (mean 92%). The procedure was applied to raw sewage and to river water samples and 0-55%-higher results were obtained over those achieved by modified Bligh and Dyer extraction. Flophemesyl derivatization of sterols allows selective ECD, improved separation from other compounds such as polychlorinatedbiphenyls that may be found in environmental samples, and reduced clean-up requirements (e.g., removing lipids by saponification which may be needed when flame ionization detection is used). Characteristic mass spectra of the flophemesyl derivatives are reported; these were obtained in order to distinguish between sterol classes and individual sterols.