Chromatographic separation of carotenoids.

The carotenoids are extremely reactive and consequently unstable due to their long system of conjugated double bonds. Several precautions, such as protection against light and oxygen, use of low temperature and antioxidants, analysis in the shortest possible time, should be taken during isolation and chromatography. The food samples, preferably fresh, are homogenized and immediately extracted with a suitable organic solvent. Saponification has been employed in order to hydrolyze the carotenoid esters, remove fatty material and destroy chlorophyll. This optional step facilitates subsequent carotenoid separation, identification and quantification. The separation of carotenoids is usually carried out by column chromatography, thin layer chromatography and high performance liquid chromatography, in analytical or preparative scale, on many stationary phases such as silica-gel, alumina, MgO, Ca(OH)2 and reversed-phase material (C18 and C30). The choice of the most suitable chromatographic method depends on the amount of sample, carotenoid composition, resolution, speed and purity required. Examples of carotenoid separation in different stationary phases will be shown and discussed.