To complete identification of the major components of the human macular pigment. Chemical ionization mass spectra of the macular pigment components were obtained and compared with those of zeaxanthin and lutein standards. A comparison was also made using chiral column high-performance liquid chromatography, which is capable of resolving individual stereoisomers of these carotenoids. Zeaxanthin and lutein from human blood plasma were similarly analyzed. The mass spectrometry data supported earlier work in which high-performance liquid chromatography, UV-visible spectrometry and chemical modification showed that the macular pigment comprises two carotenoids with identical properties to those of zeaxanthin and lutein. Chiral column chromatography showed that the "zeaxanthin" fraction is a mixture of two stereoisomers, zeaxanthin itself [(3R,3'R)-beta,beta-Carotene-3,3'-diol] and meso-zeaxanthin [(3R,3'S)-beta,beta-Carotene-3,3'-diol]. The other fraction is the single stereoisomer, lutein [(3R,3'R,6'R)-beta,epsilon-Carotene-3,3'-diol]. In human blood plasma, only zeaxanthin and lutein were found. The results strongly suggest that meso-zeaxanthin results from chemical processes within the retina. Noting that lutein exceeds zeaxanthin in plasma but that the combined zeaxanthin stereoisomers exceed lutein in the retina, the possibility was considered that meso-zeaxanthin is a conversion product derived from retinal lutein. Under nonphysiologic conditions, the authors demonstrate that a base-catalyzed conversion of lutein to zeaxanthin yields only the meso-(3R,3'S) stereoisomer.