Chemical structure of the complex pyruvylated and sulfated agaran from the red seaweed Palisada flagellifera (Ceramiales, Rhodophyta)

A homogeneous agaran fraction from Palisada flagellifera (Laurencia complex, Rhodomelaceae, Ceramiales) was obtained by aqueous room-temperature extraction, followed by ion-exchange chromatography. This galactan presents a highly complex structure with at least 18 different types of derivatives. The A units were found mostly pyruvylated, 2-sulfated (～34%), and 6-methylated (～34%), with the latter partially 2- and 2,4-sulfated. Minor amounts of β-D-galactopyranosyl units 2-, 6- and 2,6-sulfated, 6-glycosylated, and non-substituted are also present. The B-units are L-sugars composed predominantly of their cyclized derivatives, 3,6-anhydrogalactose and 3,6-anhydro-2-O-methylgalactose (～56%). The former are linked to β-D-galactosyl (6-methyl) (6-glycosylated) units, as well as to 4,6-O-(1-carboxyethylidene)-β-D-galactose 2-sulfate in the proportion of 3:1.8, respectively. A significant amount (～18%) of the α-L-galactopyranosyl units are linked to pyruvylated β-D-galactose 2-sulfate residues. An important part of the B-units (20%) is represented by α-L-galactose 6-sulfate substituted on C-3 by xylosyl, galactosyl and/or 2,3-di-O-methylgalactose units or sulfate groups that preclude their cyclization to 3,6-anhydrogalactosyl derivative. The precursor units are present in relatively low percentages. Kinetic studies suggest that in P. flagellifera agaran the cyclizable units are linked to 6-O-methyl-β-D-galactosyl and/or β-D-galactosyl units (6-glycosylated). The structural complexity of this polysaccharide is increased by the presence of 2- and 3,6-sulfated α-L-galactoses, with the latter additionally 2-O-methylated. Therefore, the major subfraction obtained from the cold extract contains structurally complex sulfated, methylated, and pyruvylated agaran.     

Pyruvated galactose and oligosaccharides from Erwinia chrysanthemi Ech6 extracellular polysaccharide

The acidic extracellular polysaccharide of Ech6 was depolymerized by fuming HCl. The pyruvated sugars were isolated and characterized by methods that included a combination of low-pressure gel-filtration and high-pH anion-exchange chromatographies, methylation linkage analyses, mass (GC-MS and MALDI-TOF MS) and 1H NMR (1D and 2D) spectroscopies. The following pyruvated sugars were obtained: 4,6-O-(1-carboxyethylidene)-D-Galp; 4,6-O-(1-carboxyethylidene)- alpha-D-Galp-(1-->4)-beta-D-GlcAp-(1-->3)-D-Galp; 4,6-O-(1-carboxyethylidene)-alpha-D-Galp-(1-->4)-alpha-D-GlcAp- (1-->3)-alpha-D-Galp-(1-->3)-L-Fucp; 4,6-O-(1-carboxyethylidene)-alpha-D-Galp-(1-->4)-beta-D-GlcAp-(1-->3) -alpha-D-Galp-(1-->3)-L-[beta-D-Glcp-(1-->4)]-Fucp. These oligosaccharides present potential haptenes for the development of specific antibodies and confirm the partial structure proposed previously for the extracellular polysaccharide from Erwinia chrysanthemi Ech6 [Yang, B. Y.; Gray, J. S. S.; Montgomery, R. Int. J. Biol. Macromol., 1994, 16, 306-312].     

Methylobacterium sp. isolated from a Finnish paper machine produces highly pyruvated galactan exopolysaccharide

The slime-forming bacterium Methylobacterium sp. was isolated from a Finnish paper machine and its exopolysaccharide (EPS) was produced on laboratory scale. Sugar compositional analysis revealed a 100% galactan (EPS). However, FT-IR showed a very strong peak at 1611 cm(-1) showing the presence of pyruvate. Analysis of the pyruvate content revealed that, based on the sugar composition, the EPS consists of a trisaccharide repeating unit consisting of D-galactopyranose and [4,6-O-(1-carboxyethylidene)]-D-galactopyranose with a molar ratio of 1:2, respectively. Both linkage analysis and 2D homo- and heteronuclear 1H and 13C NMR spectroscopy revealed the following repeating unit: -->3)-[4,6-O-(1-carboxyethylidene)]-alpha-D-Galp-(1-->3)[4,6-O-(1-carboxyethylidene)]-alpha-D-Galp-(1-->3)-alpha-D-Galp-(1-->. By enrichment cultures from various ground and compost heap samples a polysaccharide-degrading culture was obtained that produced an endo acting enzyme able to degrade the EPS described. The enzyme hydrolysed the EPS to a large extent, releasing oligomers that mainly consisted out of two repeating units.     

Analysis of pyruvylated beta-carrageenan by 2D NMR spectroscopy and reductive partial hydrolysis

A polysaccharide rich in 4',6'-O-(1-carboxyethylidene)-substituted (i.e., pyruvylated) beta-carrageenan has been prepared by solvolytic desulfation of a polysaccharide containing predominantly pyruvylated alpha-carrageenan, which was extracted from the red seaweed, Callophycus tridentifer. The 13C and 1H NMR chemical shifts of pyruvylated beta-carrageenan have been fully assigned using 2D NMR spectroscopic techniques. The 4',6'-O-(1-methoxycarbonylethylidene) group, generated during chemical methylation of the polysaccharide, has been shown to survive under the conditions of acidic hydrolysis that cleave the 3,6-anhydro-alpha-D-galactosidic bonds in permethylated samples of both pyruvylated beta- and pyruvylated alpha-carrageenans. As a result, two novel pyruvylated carrabiitol derivatives have been prepared.     

Characterization of two novel pyruvylated glycosphingolipids containing 2'-aminoethylphosphoryl(-->6)-galactose from the nervous system of Aplysia kurodai.

Two novel acidic glycosphingolipids containing pyruvylated galactose were purified from the nervous tissue of Aplysia kurodai by successive Iatrobeads column chromatographies. By component analysis, sugar analysis, permethylation studies, fast atom bombardment-mass spectrometry, and proton magnetic resonance spectrometry, the structures of these acidic glycosphingolipids, named F-9 and FGL-I, were determined to be: [3,4-O-(S-1-carboxyethylidene)]Gal beta 1-->3 GalNAc alpha 1-->3[6'-O-(2-aminoethylphosphonyl)Gal alpha 1-->2] (2-aminoethylphosphoryl 1-->6)Gal beta 1-->4Glc beta 1-->1ceramide and [3,4-O-(S-1-carboxyethylidene)] Gal beta 1-->3GalNAc alpha 1-->3(Fuc alpha 1-->2)(2-aminoethylphosphonyl-->6 Gal beta 1-->4Glc beta 1-->1ceramide, octadeca-4-sphingenine and anteisononadeca-4-sphingenine. Thus, pyruvylated glycosphingolipids containing phosphoethanolamine in addition to or in place of 2-aminoethylphosphonate are present in the nervous system of Aplysia.     

Positional isomers of sulfated oligosaccharides obtained from agarans and carrageenans: preparation and capillary electrophoresis separation

Partial reductive hydrolysis was used to produce oligosaccharide alditols from repetitive sulfated galactans obtained from four Rhodophyta species: kappa-carrageenan (from Kappaphycus alvarezii), theta-carrageenan (Gigartina skottsbergii-alkali-treated lambda-carrageenan), agarose 6-sulfate (Gracilaria domingensis), and pyruvylated agarose 2-sulfate (Acanthophora spicifera-alkali-treated pyruvylated agaran sulfate). Each hydrolyzate was submitted to anion-exchange and gel-filtration chromatography, and the isolated oligosaccharide alditols were identified by 1D and 2D NMR spectroscopy and by ESI mass spectrometry. The positional isomers of the sulfated oligosaccharide alditols were then completely resolved by capillary electrophoresis in a borate buffer. Attempts to correlate the availability of the hydroxyl groups for borate complexation with the relative migration of the oligosaccharides are presented.     

Chemoenzymatic synthesis of the 3-sulfated Lewisa pentasaccharide

The sulfated pentasaccharide benzyl O-(3-O-sulfo-beta-D-galactopyranosyl)-(1-->3)-O-[(alpha-L-fucopyranosyl)-(1-->4)]-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1-->3)-O-(beta-D-galactopyranosyl)-(1-->4)-O-beta-D-glucopyranoside sodium salt was synthesized using a chemo-enzymatic approach. Lacto-N-tetraose, obtained from two disaccharides [4-methoxybenzyl O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1-->3)-4,6-O-benzylidene-2-deoxy-2-phtalimido-beta-D-glucopyranoside and benzyl 2,6-di-O-acetyl-beta-D-galactopyranosyl-(1-->4)-2,3,6-tri-O-acetyl-beta-D-glucopyranoside], was regioselectively sulfated at the 3 OH position of the terminal galactose using the stannylene procedure. The fucosylation of the sulfated tetrasaccharide was performed using soluble or immobilized fucosyltransferase FucT-III to give the title compound.     

Synthesis and conformational analysis of the repeating units of bacterial spore peptidoglycan

Deprotection of the fully blocked disacharide allyl O-(2-amino-4,6-O-benzylidene-3-O-[(R)-1-carboxyethyl]-2-deoxy-beta-D-glucopyranosyl-1',2-lactam)-(1-->4)-2-acetamido-3,6-di-O-benzyl-2-deoxy-beta-D-glucopyranoside by selective de-O-allylation and parallel removal of the benzylidene and O-benzyl groups is described. The resulting beta-muramyl lactam-(1-->4)-GlcNAc disaccharide is characterised as the per-O-acetylated derivative by 1H and 13C NMR spectroscopy and X-ray structure analysis. Conformational analysis about glycosidic bond of repeating units of bacterial spore cortex is based on experimental data and molecular modelling.     

Novel phosphonoglycosphingolipids containing pyruvylated galactose from the nervous system of Aplysia kurodai

We have reported the existence of a phosphonoglycosphingolipid containing a pyruvylated galactose, FGL-IIb, in nerve fibers of Aplysia kurodai (Araki, S., Abe, S., Ando, S., Kon, K., Fujiwara, N. & Satake, M. (1989) J. Biol. Chem. 264, 19922-19927). We have now isolated two other pyruvylated galactose-containing phosphonoglycosphingolipids, named FGL-V and FGL-IIa, from the nervous tissue of Aplysia, and characterized them as [3,4-O-(S-1-carboxyethylidene)]Gal beta 1----3GalNAc alpha 1----3[6'-O-(2- aminoethylphosphonyl)Gal alpha 1----2] (2-aminoethylphosphonyl----6) Gal beta 1----4Glc beta 1----1 ceramide and [3,4,O-(S-1-carboxyethylidene)] Gal beta 1----3GalNAc alpha 1----3[6'-O-(2-aminoethylphosphonyl)Gal alpha 1----2]Gal beta 1----4Glc beta 11----ceramide, respectively. Their major aliphatic components are palmitic acid, octadeca-4-sphingenine and anteisononadeca-4-sphingenine. Thus, the nervous system of Aplysia contains several pyruvylated phosphonoglycolipids.     

The structure of the agaran sulfate from Acanthophora spicifera (Rhodomelaceae, Ceramiales) and its antiviral activity. Relation between structure and antiviral activity in agarans

The sulfated agaran isolated by water extraction from the red seaweed, Acanthophora spicifera (Rhodomelaceae, Ceramiales), is made up of A-units highly substituted with sulfate groups on C-2 (28-30%), sulfates on C-2 and 4,6-O-(1'-carboxyethylidene) groups (9-15%), and only the C-2 sulfate groups (5-8%) with small amounts of C-6 sulfate, 6-O-methyl, and nonsubstituted residues. B-units are formed mainly by 3,6-anhydro-alpha-L-galactose (15-16%) and its precursor, alpha-L-galactose 6-sulfate (10-17%), together with lesser amounts of 3,6-anhydro-alpha-L-galactose 2-sulfate, alpha-L-galactose 2,6-disulfate, alpha-L-galactose 2,3,6-tri-sulfate, alpha-L-galactose 2,6-disulfate 3-xylose, 2-O-methyl-alpha-L-galactose, and unsubstituted alpha-L-galactose. Small, but significant quantities of beta-D-xylose were found in all the fractions, together with small amounts to traces of D-glucose. Some of the fractions have high antiviral activity. Attempts to correlate structure and antiviral activity in agarans are presented.     

Synthesis of the tetrasaccharide related to the repeating unit of the antigen from Shigella dysenteriae type 5

Starting from L-rhamnose, D-mannose and 2-amino-2-deoxy-D-glucose hydrochloride, two disaccharide blocks, namely, ethyl 2,4-di-O-benzyl-3-O-[(R)-1-(methoxycarbonyl)ethyl]-alpha-L-rhamnopyranos yl-(1-->3)-2-O-acetyl-4,6-di-O-benzyl-1-thio-alpha-D-mannopyranoside and 2-(trimethylsilyl)ethyl 2-O-acetyl-3,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-->3)-4,6-di-O-benzy l-2-deoxy-2-phthalimido-beta-D-glucopyranoside, were synthesised and then allowed to react in the presence of N-iodosuccinimide and trifluoromethane sulfonic acid to give a tetrasaccharide derivative. This compound was converted into 2-(trimethylsilyl)ethyl 2,4-di-O-benzyl-3-O-[(R)-1-(methoxycarbonyl)ethyl]-alpha-L-rhamno- pyranosyl-(1-->3)-2-O-acetyl-4,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-- >4)-2-O-acetyl-3,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-->3)-2-acetamid o-4,6-di-O-benzyl-2-deoxy-beta-D-glucopyranoside, which on hydrogenolysis, afforded the methyl ester 2-(trimethylsilyl)ethyl glycoside of the tetrasaccharide related to the repeating unit of the O-antigen from Shigella dysenteriae type 5.     

Synthesis of a novel asparagine-linked heptasaccharide structure via p-methoxybenzyl-assisted beta-mannosylation

Synthesis of a core heptasaccharide asparagine N4-[alpha-D-mannopyranosyl-(1 --> 6)-[(alpha-D-mannopyranosyl)-(1 --> 3)]-[(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1 --> 2)]-(beta-D-mannopyranosyl)-(1 --> 4)-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1 --> 4)-[(alpha-L-fucopyranosyl)-(1 --> 6)]-2-acetamido-2-deoxy-beta-D-glucopyranosyl]-L-asparagine (1a) found from CHO glycosylation mutant cell LEC 14 is described. The structure of 1a is highly novel in terms of the presence of an extra GlcNAc residue linked to the 2-position of beta-linked mannose. The synthesis was performed using p-methoxybenzyl-assisted intramolecular aglycon delivery as the key transformation. 4,6-O-TIDPS-protected thiomannoside methyl 2-O-p-methoxybenzyl-4,6-O-(1,1,3,3-tetraisopropyl)disiloxanylid ene-3-O-trimethylsilyl-1-thio-alpha-D-mannopyranoside was adopted for this particular purpose, which afforded beta-mannoside p-methoxyphenyl 2,3-O-(p-methoxybenzylidene)-4,6-O-(1,1,3,3-tetraisopropyl)+ ++disiloxanylidene-beta-D-mannopyranosyl-(1 --> 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-beta-D-glucopyranoside stereoselectively in 75% yield.     

Structural studies on the specific capsular polysaccharide from Rhizobium trifolii, TA-1

The repeating unit of the specific capsular polysaccharide from the bacterium Rhizobium trifolii (TA)-1 has been shown to contain (a) terminal 4,6-O-(1-carboxyethylidene)-D-galactose (1 residue), (b) (1 → 3)-linked 4,6-O-(1-carboxyethylidene)-D-glucose (1 residue), (c) (1 → 4)-(1 → 6)-linked D-glucose (1 residue), (d) (1 → 4)-linked D-glucuronic acid (1 residue), and (e) (1 → 4)-linked D-glucose (4 residues). The pyruvylated sugars were shown to be positioned sequentially, and at least one other unit was interposed between them and the branch point.     

Identification of pyruvated monosaccharides in polysaccharides by gas-liquid chromatography mass spectrometry

Twelve bacterial polysaccharides of known structure containing a representative range of pyruvated monosaccharides, were methanolysed, trimethylsilylated, and analysed by g.l.c. and g.l.c.-m.s. Except for 3,4-O-(1-carboxyethylidene)-L-rhamnose, which was unusually labile, the pyruvic acid substituents were largely retained during methanolysis and the Me3Si derivatives of the resulting pyruvated methyl glycosides gave distinctive g.l.c. peaks with characteristic mass spectra. The pyranose rings of 4,6-O-(1-carboxyethylidene)-D-glucose, 4,6-O-(1-carboxyethylidene)-D-mannose, 4,6-O-(1-carboxyethylidene)-D-galactose, and 3,4-O-(1-carboxyethylidene)-D-galactose survived the methanolysis, but that of 2,3-O-(1-carboxyethylidene)-D-glucuronic acid was cleaved to give the methyl ester of 2,3-O-(1-carboxyethylidene)-aldehydo-D-glucuronic acid dimethyl acetal. In the case of 2,3-O-(1-carboxyethylidene)-D-galactose, cleavage of the pyranose ring was less complete; under the conditions used in these experiments two-thirds of the pyranose rings were intact while one-third were cleaved to give the methyl ester of 2,3-O-(1-carboxyethylidene)-aldehydo-D-galactose dimethyl acetal. A very small amount of 3,4-O-(1-carboxyethylidene)-L-rhamnose from one polysaccharide retained its pyruvic acid substituent after gentle methanolysis to give the methyl ester of 3,4-O-(1-carboxyethylidene)-aldehydo-L-rhamnose dimethyl acetal. Susceptibility to cleavage of the pyranose ring during methanolysis appears to be a property of pyruvated monosaccharides with trans-fused 1,3-dioxolane rings.     

Extracellular polysaccharides of Erwinia futululu,a bacterium associated with a fungal canker disease of Eucalyptus spp

Extracellular polysaccharides (EPSs) produced by an Erwinia spp. associated with a fungal canker disease of Eucalyptus were fractionated into two polysaccharides, one that was identified with that produced by Erwinia stewartii. The other has a similar structure, but with one terminal Glc residue replaced by pyruvic acid to give 4,6-O-[(R)-1-carboxyethylidene)-Galp. Their structures were determined using a combination of chemical and physical techniques including methylation analysis, periodate oxidation, low-pressure gel filtration and anion-exchange chromatographies, high-pH anion-exchange chromatography, mass spectrometry and 1D and 2D 1H NMR spectroscopy. The new polysaccharides, identified as EPS Futululu FF-1 and FF-2, have the following structures:The molecular weights of the polysaccharides range from 1.3-2.1x10(6) and their hydrodynamic properties are those of polydisperse, polyanionic biopolymers with pseudoplastic, non-thixotropic flow characteristics in aqueous solutions.     

Sulfated polysaccharides from the red seaweed Georgiella confluens.

The water-soluble polysaccharides from Georgiella confluens, collected in Antarctica, were fractionated with cetrimide. The complexed material was subjected to fractional solubilization in solutions of increasing sodium chloride concentration. The initially extracted polysaccharide and the major fraction isolated, soluble in 0.5 M NaCl, were studied. These are sulfated xylogalactans naturally rich in methylated sugar residues, comprising of 3,6-anhydro-2-O-methyl-L-galactose, 2-O-methyl-L-galactose and 6-O-methyl-D-galactose. Structural analysis was carried out by methylation, ethylation, desulfation-ethylation, desulfation-methylation, Smith degradation, 13C NMR spectroscopy and determination of the absolute configuration of monosaccharides by gas chromatography of diastereomeric derivatives produced by reductive amination. The results indicated the presence of an agaran backbone with an unusual substitution pattern: sulfation mainly at the 3-position of the alpha-L-galactose units and the presence of xylose side chains at the 4-position of the beta-D-galactose residues.     

The structure of a galactan sulfate from the red seaweed Bostrychia montagnei

The sulfated, methylated galactan isolated from the red seaweed Bostrychia montagnei, showed an unusually narrow structural dispersion. This agaran has the defining linear backbone of alternating 3-linked beta-D-galactopyranosyl units and 4-linked alpha-L-galactopyranosyl and 3,6-anhydrogalactopyranosyl residues. The D-units have C-6 methylation, C-6 single stubs of xylopyranosyl and minor to trace amounts of (possible) C-6 linked single stubs of galactopyranosyl. These units are mainly sulfated on C-4 with lesser sulfation at C-6 and minor at C-2. The L-residues are mainly methylated on C-2 of the 3,6-anhydrogalactopyranosyl and sulfated on C-3 of the L-galactopyranosyl; minor amounts of 2,3- and 3,6-disulfated and 2-O-methyl or 2-O-glycosyl 3-sulfated L-galactopyranosyl were also found.     

Triterpenoid saponins from Ardisia mamillata

Two saponins were isolated from the roots of Ardisia mamillata HANCE. Their structures were established on the basis of MALDI-TOFMS, 1H, 13C NMR and 2D NMR (COSY, HOHAHA, HETCOR, HMBC and ROESY) spectra, and on chemical evidence, to be ardisimamilloside A, 3-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl-(1 --> 4)-[beta-D-glucopyranosyl-(1 --> 2)]-alpha-L-arabinopyranosyl]-3beta, 16alpha,28alpha-trihydroxy-13beta,28-epoxy-oleanan+ ++-30-al; and ardisimamilloside B, 3-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl-(1 --> 4)-[beta-D-glucopyranosyl-( 1 --> 2)]-alpha-L-arabinopyranosyl]-beta3-hydroxy-13beta,28- epoxy-oleanan-16-oxo-30-al.     

An efficient access to protected disialylated glycohexaosyl threonine present on the leukosialin of activated T-lymphocytes

The total synthesis of the threonine-linked core 2 class disialylated hexasaccharide in a completely protected form was accomplished for the first time. The L-threonine conjugate, N-(9-fluorenylmethoxycarbonyl)-O-[(5-acetamido-4,7,8,9-tetra-O-ben zyl-3,5-dideoxy-D-glycero-alpha-D-galacto-2-nonulopyranosylonic acid)-(2-->3)-(2,6-di-O-benzyl-beta-D-galactopyranosyl)-(1-->4)-2-acetam ido-2-deoxy-3,6-di-O-benzyl-beta-D-glucopyranosyl-(1-->6)-[(5-acetamido- 4,7,8,9-tetra-O-benzyl-3,5-dideoxy-D-glycero-alpha-D-galacto-2-nonulo pyranosylonic acid)-(2-->3)-2,6-di-O-benzyl-beta-D-galactopyranosyl-(1-->3)]-2-acetami do-2-deoxy-alpha-D-galactopyranosyl-(1d-->4c:1f-->4e)-dilactone ]-L-threonine allyl ester was synthesized via stereocontrolled glycosylations employing readily accessible monosaccharidic blocks; t-butyl-diphenylsilyl-2-azido-2-deoxy-3,6-di-O-benzyl-beta-D-gluco pyranose, N-(9-fluorenylmethoxycarbonyl)-O-(2-azido-6-O-t-butyldimethylsilyl -2-deoxy-alpha-D-galactopyranosyl)-L-threonine allyl ester, 8, 9 and N-(9-fluorenylmethoxycarbonyl)-O-(2-azido-4,6-O-benzylidene-3-O-ch loroacetyl-2-deoxy-alpha-D-galactopyranosyl)-L-threonine allyl ester. For the introduction of the amino acid, the azide group was used to temporarily mask the amino group of GalNAc so as to obtain an alpha-glycosidic linkage without participation from the C-2 substituent. The threonine was attached to the sugar unit at the monosaccharide stage to avoid loss of oligosaccharide at a later stage. The Fmoc and allyl ester protected amino acid at the reducing end facilitates efficient glycopeptide synthesis on solid-phase support.     

Structure of phosphonoglycosphingolipid containing pyruvylated galactose in nerve fibers of Aplysia kurodai

A phosphonoglycosphingolipid, designated as FGL-IIb, was identified in nerve fibers of Aplysia kurodai by two-dimensional thin layer chromatography (Abe, S., Araki, S., and Satake, M. (1986) Biomed. Res. (Tokyo) 7, 47-51). FGL-IIb was isolated from the nervous system of A. kurodai by Iatrobeads column chromatography using three solvent systems. Pyruvic acid was identified by thin layer chromatography as its 2,4-dinitrophenylhydrazone and established by permethylation studies to be attached as a ketal to O-3 and O-4 of the terminal galactose of the oligosaccharide chain in FGL-IIb. By sugar analysis, permethylation studies, fast atom bombardment-mass spectrometry, and proton magnetic resonance spectrometry, the structure of FGL-IIb was concluded to be [3,4-O-(1-carboxyethylidene)]Gal beta 1----3GalNAc alpha 1----3(Fuc alpha 1----2) (2-aminoethylphosphonyl----6)Gal beta 1----4Glc beta 1----1ceramide. Its major aliphatic components were palmitic acid, octadeca-4-sphingenine and anteisononadeca-4-sphingenine. This is the first report of the occurrence of pyruvylated galactose as a constituent of animal sphingolipid.