Nitrous acid deamination of N-deacetylated blood-group O poly(glycosyl)ceramides resulted in a complete degradation of the glycolipids to give O-beta-D-galactopyranosyl-(1 leads to 4)-2,5-anhydro-D-mannose, O-alpha-L-fucopyranosyl-(1 leads to 2)-O-beta-D-galactopyranosyl-(1 leads to 4)-2,5-anhydro-D-mannose, lactosylceramide, and small proportions of free 2,5-anhydro-D-mannose. A series of straight-chain glycolipids containing up to six glycosyl residues and comprising alternating 3-O-substituted D-galactosyl and 4-O-substituted 2-acetamido-2-deoxy-D-glucosyl residues was isolated from partial acid hydrolyzates of poly(glycosyl)ceramides. Branching points of 3,6-di-O-substituted and 6-O-substituted D-galactosyl residues were observed only in fractions containing more than six glycosyl residues. Sequential periodate oxidation of poly(glycosyl)ceramides gradually eliminated the branching points. This elimination was not complete, even after four cycles of degradation. The ability of poly(glycosyl)ceramides to precipitate with anti-I serums disappeared after two cycles of degradation. These results suggest a general structure for poly(glycosyl)ceramides. I blood-group activity of the glycolipids would depend on the periodical arrangement of branched side-chains.