The identity of the major phospholipid component of human lens membrane extracts, previously referred to as the 'unknown phospholipid', was recently proposed to be 4,5-dihydrosphingomyelin (DHS). Using high-performance liquid-chromatographic separation with atmospheric pressure chemical ionization and electrospray ionization mass-spectrometric detection, we report here the first identification of the molecular species of DHSs and sphingomyelins (SPMs) of the human eye lens. The most abundant molecular species were palmitic and tetracosenoic (24:1) DHSs, representing 57.8 and 23.3% of human lens DHSs, respectively. Lesser amounts of hexacosanoic, hexacosenoic, tetracosanoic, docosanoic, docosenoic, stearic, palmitoleic, myristic and other DHSs were found. The most abundant normal SPM molecular species in the human lens were similar to those of DHS. Palmitic SPM represented 53.9% of the uncorrected SPM peak areas, while tetracosenoic SPM represented 17.6% of the SPM in the human lens. The sphingolipids of the human lens were determined to be composed of 76.9% DHS species and 23.1% SPM species. Commercially available SPM was also found to contain significant amounts of DHS species.