Reversed-Phase Separation and the Identification of Unknown Polar Compounds

Reversed-Phase Chromatography is a standard technique for the separation of many natural compounds and is an extremely useful tool for a variety of industrial, food, law enforcement and medical applications. This manuscript describes an experiment for the separation of unknown polar compounds using inexpensive reversed-phase SPE C18 columns. Students perform a chromatographic separation on a sample of an unknown polar compound contaminated with benzophenone. Students then determine the identity of their compound with melting point, GC-MS, IR, 1H and 13C NMR spectroscopy, and comment on the purity of the final sample. The use of unknowns increases students’ individual responsibility and prevents using a partner’s data analysis. This experiment was developed for use in a first year foundation laboratory but it would also be suitable for implementation in a sophomore organic chemistry laboratory.