This article is available online at http://www.jlr.org Glycosphingolipid (GSL) is composed of sugar chain and ceramide, the latter of which consists of a fatty acid and a sphingoid. GSLs, which are expressed on the outer side of the lipid bilayer in animal cells, form patches with sphingomyelin (microdomains) and play a foundational role in intercellular adhesion, cellular recognition, differentiation/growth, and immune response ( 1-3 ). However, a comprehensive understanding of GSL function has not yet been attained because of the structural complexity in the sugar chain of GSLs even in vertebrates and invertebrates ( 4, 5 ). We have analyzed the GSL structures in lower animals on the assumption that their complexity plays an important role in maintaining the biological activity of multicellular organisms.In invertebrates, structural analyses of GSLs have been performed in several phyla and are well known in Arthropoda, Mollusca, and Nematoda. The immune response by ␣ -galactosylceramide from a marine sponge ( 3 ) and the induction of cytokine secretion by zwitterionic GSLs from a parasitic nematode ( 6 ) are interesting in terms of GSL function, and investigation of invertebrate GSL structures could therefore be a productive research area. In Arthropoda, structural analyses of GSLs have begun for fl ies (Diptera Insecta) ( 7,8 ), and a characteristic arthro-series sugar chain (GlcNAc  3Man  4Glc  Cer; At 3 Cer) has been identifi ed. In Drosophila , mactosyl ceramide (MacCer) and arthrotriaosylceramide (At 3 Cer) are biosynthesized by Abstract Neutral glycosphingolipids containing one to six sugars in their oligosaccharide chains have been isolated from cysts of the brine shrimp Artemia franciscana . The structures of these glycolipids were identifi ed by methylation analysis, partial acid hydrolysis, gas-liquid chromatography, combined gas-liquid chromatography-mass spectrometry, matrix-assisted laser desorption/ionization time-of-fl ight mass spectrometry, and proton nuclear magnetic resonance spectroscopy to be Glc  1-Cer, Man  1-4Glcand GalNAc  1-4(GlcNAc ␣ 1-2Fuc ␣ 1-3)GlcNAc  1-3Man  1-4Glc  1-Cer (CHS). Two glycosphingolipids, CPS and CHS, were characterized as novel structures. Because Artemia contains a certain series of glycosphingolipids (-Fuc ␣ 3Man  -4Glc  Cer), which differ from the core sugar sequences reported thus far, we tentatively designated the glycosphingolipids characterized as nonarthro-series ones. Furthermore, CHS exhibited a hybrid structure of arthro-series and nonarthro-series sugar chain. Two novel glycosphingolipids were characterized from the brine shrimp Artemia franciscana ; one was composed of arthrotetraose and a branching fucose attached to N -acetylglucosamine residue, and the other was composed of CPS with an additional N -acetylglucosamine residue attached to the branching fucose.