AbstractGlycosphingolipids were isolated from total lipids of female and male human thyroids by alkaline hydrolysis, silicic acid, diethylaminoethyl‐celluose and thin layer chromatography and analyzed by gas liquid chromatography. On the basis of their mobility in two dimensions on thin layer chromatography, IR analysis, and of sugar molar ratio, four neutral glycolipids, a sulfatide, and a hematoside fraction were identified. Glucosyl, plus galactosyl ceramide, and trihexosyl ceramide were the major fractions and accounted for 33% and 28% of total neutral glycolipids, respectively. Dihexosyl ceramide was a mixture of lactosyl and digalactosyl ceramide. The acidic lower phase glycolipids comprised ceramide galactosyl sulfate as the major component of male thyroids. Hematoside was identified tentatively as a minor component of the thyroids of both sexes. Major fatty acids of all neutral glycolipid fractions were 20∶0, 22∶0, 24∶0, and 24∶1; 24∶0 and 24∶1 for sulfatides. Low proportions of α‐hydroxy fatty acids were identified. Total neutral glycosphingolipids of male thyroids were comparable in quantities with human liver but lower than kidneys, leucocytes, and platelets. Male thyroids comprised higher quantities of neutral glycosphingolipids (4.04±0.32 μmoles/g total lipid) as compared to females (2.34±0.21 μmoles/g total lipid), and much higher sulfatide than the females. These marked differences may suggest that the biosynthesis of the glycosphingolipids in the thyroid gland is under hormonal control. Similarities in glycosphingolipid composition of human thyroid and kidney are discussed in relation to a possible role played by glycolipids in ion transport, which is a common feature of the two organs.