Publisher Summary Carotenoid pigments can function as effective quenchers of singlet oxygen and oxygen-centered radicals; interest has been rekindled in the utilization of these compounds as probes for mechanisms of oxygen radical generation, action, and damage. This chapter discusses the methods of extraction of carotenoid pigments, their separation by chromatographic procedures––such as thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC)––the means for identifying these pigments, and the methods for quantitating the pigments present in biological samples. It presents the general principles whereby carotenoid pigments can be extracted, separated, identified, and quantitated using both TLC and reversed phase HPLC (rp-HPLC). The latter system, in conjunction with a variety of tests carried out on the individual fractions obtained from chromatograph effluents, results in the rapid characterization and quantitation of a variety of carotenoids, particularly those from plant tissues. Similar systems are applicable to animal systems as well as in vitro systems to which carotenoids or tissue extracts have been added.