Harold M. Flowers

Studies on the Koenigs-Knorr reaction : Part II. Synthesis of an α-L-linked disaccharide from tri-O-benzyl-α-L-fucopyranosyl bromide

Abstract Methyl α- L -fucopyranoside was converted into 2,3,4-tri- O -benzyl-α- L -fucopyranosyl bromide via the intermediates methyl 2,3,4-tri- O -benzyl-α- L -fucopyranoside and 2,3,4-tri- O -benzyl-1- O - p -nitrobenzoyl-β- L -fucopyranose. Koenigs-Knorr condensation of the bromide with benzyl 2-acetamido-3,4-di- O -acetyl-2-deoxy-α- D -glucopyranoside and removal of protecting groups afforded 2-acetamido-2-deoxy-6- O -α,β- L -fucopyranosyl- D -glucose 65% yield, the product containing a 7:3 ratio of α to β anomers. Optically pure 2-acetamido-2-deoxy-6- O -α- L -fucopyranosyl- D -glucose was obtained by hydrogenolysis of the intermediate crystalline benzyl 2-acetamido-2-deoxy-6- O -(tri- O -benzyl-α- L -fucopyranosyl-α- D -glucopyranoside.

Chemistry and Biochemistry of d- and l-Fucose

Publisher Summary This chapter discusses the various aspects of the chemistry of fucoses, especially the developments made in this field, and explains their metabolism and biochemistry while pointing out the possible biological functions and medical applications of fucoses and their compounds. Fucose is a 6-deoxyhexose having the “galacto” configuration—that is, it has cis-hydroxyl groups on C-3 and C-4. D-Fucose is usually found in plants and some microorganisms, in relatively simple glycosides comprising only a few sugar units. Some plant glycosides and polysaccharides crossreact with reagents for blood-group antigens, and interesting crossreactions has also been shown by methylated-fucose haptens and L-fucose-specific lectins. Great interest has been shown in the characterization of glycosyltransferases in connection with the studies on the tissue localization and mechanism of the biosynthesis of other animal glycoproteins. L-Fucose is found to inhibit the growth of some cells in culture, but only at high doses, and to inhibit the growth of implanted, mammary tumors in rats. The conversion of GDP-D-Man into GDP-Fuc serves as a preliminary stage in the biosynthesis of L-fucans, and its hydrolysis affords free L-fucose.

Studies on the Koenigs-Knorr reaction : Part I. Synthesis of 6-O-α-D-glucopyranosyl-D-galactose and 3-O-α-D-glucopyranosyl-D-galactose

Abstract Two α- D -linked disaccharides, 6- O -α- D -glucopyranosyl- D -galactose and 3- O -α- D -glucopyranosyl- D -galactose were synthesized by reaction of 2- O -benzyl-3,4,6-tri- O - p -nitrobenzoyl-α(or β- D -glucopyranosyl bromide with 1,2:3,4-di- O -isopropylidene-α- D -galactose and 4,6- O -ethylidene-1,2- O -isopropylidene-α- D -galactose, respectively, and subsequent removal of the protecting groups. No β- D -linked disaccharide was detected in the reaction mixture, irrespective of the anomeric configuration of the sugar halide used.

Selective benzylation of some D-galactopyranosides. Unusual relative reactivity of the hydroxyl group at C-4

Abstract Partial benzylation of methyl 2,3-di- O -benzyl-α- D -galactopyranoside gave methyl 2,3,6-tri- O -benzyl-α- D -galactopyranoside as the major product, whereas the isomeric 2,6-di- O -benzyl ether gave a mixture of products in which the ratio of methyl 2,4,6- to methyl 2,3,6-tri- O -benzyl-α- D -galactopyranoside was ≈4:1. The proportion of unreacted starting-material was low in both cases, whereas after a similar reaction of methyl 2,6-di- O -benzyl-β- D -galactopyranoside more than 50% of the dibenzyl ether was recovered unchanged. In this case also, considerably higher reactivity was exhibited by the hydroxyl group at C-4 than that at C-3. Acid hydrolysis of the methyl glycosides of the tribenzyl ethers afforded crystalline 2,4,6-tri- O -benzyl-α- D -galactose and syrupy 2,3,6-tri- O -benzyl- D -galactose. Structures of intermediates were established by acetylation, examination of their n.m.r. spectra, and conversion into the known 3- O and 4- O -methyl- D -galactose.

Studies on the koenigs-knorr reaction : Part IV: The effect of participating groups on the stereochemistry of disaccharide formation

Abstract Partial benzylation of methyl 2- O -benzyl-α- L -fucopyranoside afforded a mixture of methyl 2,3-, and 2,4-di- O -benzyl-α- L -fucopyranoside which were separated by means of their monoacetates. Partial benzylation of methyl α- L -fucopyranoside gave the 2,4-, and 3,4-dibenzyl ethers in the ratio of 3:2, and no 2,3-isomer could be detected in the reaction mixture. The structures of the three dibenzyl ethers were established: (a) by analysis of the n.m.r. spectra of their acetates, and (b) by methylation, removal of benzyl groups by hydrogenolysis, and characterization of the methyl ethers of the methyl glycosides. Acid hydrolysis of these compounds gave the monomethyl ethers of L -fucose, two of which were identical with known compounds, whereas the third, 4- O -methyl- L -fucose, was a new compound. Selective p -nitrobenzoylation of 2,3-, 2,4-, and 3,4-di- O -benzyl- L -fucose, followed by acetylation and treatment with hydrogen bromide in dichloromethane, gave the three possible mono- O -acetyl-di- O -benzyl-α- L -fucopyranosyl bromides, which were condensed with benzyl 2-acetamido-3,4-di- O -acetyl-2-deoxy-α- D -glucopyranoside. The disaccharide derived from the 2- O -acetyl substituted bromide was enriched in β- L -fucopyranoside, whereas the other two bromides gave mainly the α- L -linked anomer. The α-directing influence of the 3- and 4- O -acetyl substituents is not less than the β-directing influence of the 2- O -acetyl group in similar bromides; participation of acyl groups and electronic-steric influences are discussed as possible explanations for the steric course of the reaction.

Studies on B-antigenic sites of human erythrocytes by use of coffee bean α-galactosidase

Abstract Treatment of human type B erythrocytes, erythrocyte membranes and membrane fractions with highly purified coffee bean α-galactosidase resulted in the disappearance of B activity and enhancement of H activity with concomitant release of galactose. In the case of B erythrocytes, enzyme treatment led to the release of 3.0 × 10 6 molecules of galactose per cell, about 75% of which were found in the aqueous phase of butanol/water extracts of cell membranes. AB and O cells released 1.9 × 10 6 and 0.8 × 10 6 molecules per cell, respectively. All the galactose released from O cells came from glycolipid material lacking B activity. Application of a correction for the enzymically labile, non-B-active α-galactosides presumably also present in B cells would suggest that such B cells have approximately 2.2 × 10 6 B-antigenic sites. No release of galactose was observed when membranes and cell fractions, prepared from B erythrocytes which had been previously incubated with α-galactosidase were treated with the enzyme, showing that all the α-galactosidase-labile galactose present in B-active macromolecules in intact red cells is located at sites available to the enzyme and can be readily removed. We did not find any significant differences in the number and distribution of the B-antigenic sites present in erythrocytes from secretors and from a nonsecretor B individual.

Synthesis of O-β-d-galactopyranosyl-(1→3)-O-β-d-galactopyranosyl-(1→4)-d-glucose☆

Abstract O -β- d -Galactopyranosyl-(1→3)- O -β- d -galactopyranosyl-(1→4)- d -glucose was obtained by selective glycosidation of benzyl 4- O -(2,6-di- O -acetyl-β- d -galactopyranosyl)-2,3,6-tri- O -acetyl-β- d -glucopyranoside with tetra- O -acetyl-α- d -galactopyranosyl bromide. The disaccharide was prepared from benzyl 4- O -(3,4- O -isopropylidene-β- d -galactopyranosyl)-β- d -glucopyranoside, which was obtained in good yield by treatment of benzyl β-lactoside with acetone and p -toluenesulfonic acid.

Purification of coffee bean α-galactosidase by affinity chromatography

Abstract The preparation and properties of a specific adsorbent for the rapid and complete purification of α- d -galactosidase (α- d -galactoside galactohydrolase, EC 3.2.1.22) from green coffee beans by affinity chromatography is described. The adsorbent consists of N-e- aminocaproyl -N-e- aminocaproyl -α- d -galactopyranosylamine coupled to Sepharose. α-Galactosidase was adsorbed from a partially purified preparation obtained from a coffee bean extract, and eluted with either d -galactose or p-nitrophenyl α- d -galactopyranoside. The specific activity of the purified enzyme, tested with p-nitrophenyl α- d -galactopyranoside as substrate, was 94.5 units/mg protein, representing an 8600-fold purification of the original crude extract. Polyacrylamide gel electrophoresis at pH 8.3 revealed three protein bands, all of which possessed enzymic activity.

Synthesis of benzyl 6-O-benzoyl-3,4-O-isopropylidene-β-D-galactopyranoside and of 2-O-α-L-fucopyranosyl-D-galactose

Abstract Benzyl β- D -galactopyranoside was converted into the crystalline 3,4- O -isopropylidene derivative ( 2 ), which was selectively benzoylated to give crystalline benzyl 6- O -benzoyl-3,4- O -isopropylidene-β- D -galactopyranoside ( 8 . The structure of 8 was established by various reactions, which included the preparation of benzyl 6- O -benzoyl-β- D -galactopyranoside ( 9 ) and of benzyl 2- O -benzoyl-β- D -galactopyranoside ( 10 ). Condensation of 8 with 2,3,4-tri- O -acetyl-α- L -fucopyranosyl bromide, followed by removal of the isopropylidene benzoyl and acetate groups, afforded crystalline benzyl 2- O -α- L -fucopyranosyl-β- D -galactopyranoside ( 12 ). Catalytic hydrogenolysis of 12 gave the disaccharide 2- O -α- L -fucopyranosyl- D -galactose ( 1 ), which was identical with the natural product.

Synthesis of 2-O-α-l-fucopyranosyl-l-fucopyranose

Abstract Acetylation of l -fucose (6-deoxy- l -galactose) with acetic anhydride and sodium acetate, followed by treatment of the resulting syrup with hydrogen bromide in acetic acid, gave 2,3,4-tri- O -acetyl-α- l -fucopyranosyl bromide ( 1 ). Treatment of this compound with benzyl alcohol in the presence of silver oxide gave the crystalline benzyl 2,3,4-tri- O -acetyl-β- l -fucopyranoside ( 2 ). Catalytic de-esterification afforded benzyl β- l -fucopyranoside ( 3 ), which was converted into benzyl 3,4- O -isopropylidene-β- l -fucopyranoside ( 4 ). The Koenigs-Knorr reaction of compound 1 with 4 in the presence of mercuric cyanide afforded the crystalline disaccharide, benzyl 3,4- O -isopropylidene-2- O -(2,3,4-tri- O -acetyl-α- l -fucopyranosyl)-β- l -fucopyranoside ( 5 ). Removal of the protecting groups gave crystalline 2- O -α- l -fucopyranosyl- l -fucopyranose ( 8 ), identical with a disaccharide previously isolated from fucoidin.