Leptine I, a glycoalkaloid only known to occur in the foliage of the wild potato species Solanum chacoense (Bitt.), is a potent feeding deterrent to the economically serious insect pest, the Colorado potato beetle (Leptinotarsa decemlineata Say). In order to demonstrate, systematically, the effectiveness of leptine I, incorporation into synthetic beetle diet trials is necessary. We describe a preparative procedure for the partial purification of leptine I by a series of steps, starting with a solid-phase C18 extraction, followed by sequential silica gel chromatography, and finally reversed-phase preparative HPLC. This preparation yields a white powder, containing leptine I as the sole glycoalkaloid, with an overall purity of greater than 65%, and is entirely suitable for incorporation into synthetic diets.