Abstract
Eight different glycosphingolipids with four to six sugars per molecule have been isolated from human plasma of blood-group O Leb secretores. Purfication was achieved by preparative highperformance thin-layer chromatography using four different solvent system, by gas-liquid chromatograhical analysis of the sugars as their alditol revealing integral numbers of sugars and by the hemagglutination inhibition test. Two blood-group Lea-active ceramide pentasaccharides (glycolipids 4,5) and three blood-Leb-active ceramide hexasaccharides (glycolipids 6, 7 and 8) were obtained, each revealing glucose, N-acetylglucosamine and fucose in molar rations of 1:2:1:1 (glycolipids 4,5) or 1:2:1:2 (glycolipids 6, 7 and 8) respectively. 0.027 μg of glycolipids 6, 7 and 8 completly inhibited the agglutination of human O Le(a-b+) erythrocytes by 50 μl of 4 hemagglutinating units of caprine anti-Leb sera. Honegeneity of glylipids 6, 7 and 8 was confirmed by the exclusion of contamination of the hypothetical Led antigen being a structural isomer of the Leb antigen: A crude neutral glycolipid fraction enriched in fucolipds from the plasma of O Le(a-b-) secretors showed a high-performance thin-layer chromatographical migration behaviour completely different from that of glycolipids 6, 7 and 8. Glycolipids 4 and 5 exhibited distinct Lea blood-group activities (0.4 μg of glycolipid 4 or 0.6 μ of glycopid 5 per 50 μl of 4 hemagglutinating units of caprine anti-Lea serum), but were slightly contaminated by blood-group H activity or blood-group Leb (7 μg of glycolipid 4/100 μl human anti-H serum and 1.7 μg of glycolipid 5/50 μl caprine anti-Leb serum). 0.01 μg, 0.0006 μg or even only 0.0001 μg of glycolipid 6, 7 or 8 respectively are sufficient for inclubation to convert 9x107 O Le(a-b-) erythrocytes into Lebpositive cells. This means in the case of glycolipid 8. that maximally 400 molecules per erythrocyle on the cell surface are sufficient to acheive an agglutinability by anti-Leb sera. In addition, two different globosides (glycolipids 1, 2) and a new ceramide tetrasaccharide (glycolipid 3) were isolated. The latter substance contains glucose, galactose and N-acetylglucosamine in molar ratios of 1:2:1, but differs from the lacto-N-neotetraosylceramide by showing a distictly slower high-performance thin-layer chromatographical migration behaviour.
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