Synthesis and biological activities of octyl 2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-beta-L-fucopyranoside

Octyl 2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-beta-L-fucopyranoside, a fucosyl pentasaccharide with a regular structure resembling the repeating unit of a natural sulfated fucan, was chemically synthesized using a convergent '2+3' strategy. Regioselective 3-O-silylation of beta-thiofucopyranoside and AgOTf-catalyzed glycosylation of the protected glycosyl trichloroacetimidate facilitated a one-pot trisaccharide synthesis. The synthesized target compound showed good antitumor activity in vivo, and promising anticoagulant activity in vitro.     

Synthesis of beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp-(1-->3)-beta-D-Glcp-(1-->6)-[beta-D-Galp-(1-->4)-beta-D-Glcp-(1-->3)]-beta-D-GlcpOLauryl, an oligosaccharide with anti-tumor activity

A concise and effective synthesis of lauryl heptasaccharide 17 was achieved from the key intermediates lauryl 2,3,4,6-tetra-O-benzoyl-beta-D-galactopyranosyl-(1-->4)-2,3,6-tri-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-2,4-di-O-benzoyl-beta-D-glucopyranoside (10) and isopropyl 2,4,6-tri-O-acetyl-3-O-allyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-2,4-di-O-acetyl-beta-D-glucopyranosyl-(1-->3)-2,4,6-tri-O-acetyl-1-thio-beta-D-glucopyranoside (15). The key trisaccharide glycosyl acceptor 10 was constructed by coupling 2,3,4,6-tetra-O-benzoyl-beta-D-galactopyranosyl-(1-->4)-2,3,6-tri-O-benzoyl-alpha-D-glucopyranosyl trichloroacetimidate (3) with lauryl 6-O-acetyl-2,4-di-O-benzoyl-beta-D-glucopyranoside (9), followed by deacetylation. The thioglycoside donor 15 was obtained by condensation of 2,4,6-tri-O-acetyl-3-O-allyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-2,4-di-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (11) with isopropyl 4,6-O-benzylidene-1-thio-beta-D-glucopyranoside (12), followed by debenzylidenation and acetylation. A bioassay of the inhibition of S180 noumenal tumors showed that lauryl heptasaccharide 17 could be employed as a potential agent for cancer treatment.     

Synthesis of mannose-containing analogues of (1-->6)-branched (1-->3)-glucohexaose

Coupling of the trisaccharide acceptor 2,4,6-tri-O-acetyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-5-O-acetyl-1,2-O-isopropylidene-alpha-D-glucofuranose (2) with the trisaccharide donor 2,3,4,6-tetra-O-benzoyl-alpha-D-annopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-2,4-di-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (1) gave an alpha-linked hexasaccharide 3, while coupling of 2 with the trisaccharide donor 2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)]-2,4-di-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (7) produced alpha- 8 and beta-linked 12 hexasaccharides in a ratio of 3:2. Deprotection of 3, 8, and 12 afforded the analogues of the immunomodulator beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-D-Glcp (A).     

Synthesis of galactose-containing analogues of (1-->6)-branched (1-->3)-glucohexaose and its lauryl glycoside

Coupling of the trisaccharide acceptor either 2,4,6-tri-O-acetyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-5-O-acetyl-1,2-O-isopropylidene-alpha-D-glucofuranose (13) or lauryl 2,4,6-tri-O-acetyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-2,5-di-O-acetyl-alpha-D-glucopyranoside (15) with the trisaccharide donor 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)]-2,4-di-O-acetyl-alpha-D-galactopyranosyl trichloroacetimidate (12) gave alpha-linked hexasaccharides 14 and 16, respectively, while coupling of either 13 or 15 with trisaccharide donor 2,3,4,6-tetra-O-benzoyl-beta-D-galactopyranosyl-(1-->3)-[2,3,4,6-tetra-O-benzoyl-beta-D-galactopyranosyl-(1-->6)]-2,4-di-O-acetyl-alpha-D-galactopyranosyl trichloroacetimidate 17 did not afford any hexasaccarides. The analogues of the immunomodulator beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-beta-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp (1) was obtained by deprotection of 14 and 16.     

Synthesis of beta-D-GlcpNAc-(1-->3)-alpha-L-Rhap-(1-->2)-[beta-L-Xylp-(1-->4)]-alpha-L-Rhap-(1-->3)-alpha-L-Rhap,the repeating unit of the O-antigen produced by Pseudomonas solanacearum ICMP 7942

An efficient synthesis of beta-D-GlcpNAc-(1-->3)-alpha-L-Rhap-(1-->2)-[beta-L-Xylp-(1-->4)]-alpha-L-Rhap-(1-->3)-alpha-L-Rhap, the repeating unit of the O-antigen produced by Pseudomonas solanacearum ICMP 7942 and its isomer beta-D-GlcpNAc-(1-->3)-alpha-L-Rhap-(1-->4)-[beta-L-Xylp-(1-->2)]-alpha-L-Rhap-(1-->3)-alpha-L-Rhap was achieved via sequential assembly of the building blocks, allyl 2,3-O-isopropylidene-alpha-L-rhamnopyranoside (2), allyl 3,4-O-isopropylidene-alpha-L-rhamnopyranoside (3), allyl 2,4-di-O-benzoyl-alpha-L-rhamnopyranoside (6), 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-beta-D-glucopyranosyl trichloroacetimidate (7), and 2,3,4-tri-O-benzoyl-beta-L-xylopyranosyl trichloroacetimidate (12). The process was carried out in a regio- and stereoselective manner using glycosyl trichloroacetimidates as donors and unprotected or partially protected rhamnopyranosides as acceptors in the presence of a catalytic amount of trimethylsilyl trifluoromethanesulfonate (TMSOTf).     

Concise syntheses of arabinogalactans with beta-(1-->6)-linked galactopyranose backbones and alpha-(1-->3)- and alpha-(1-->2)-linked arabinofuranose side chains

4-methoxyphenyl glycosides of 2,3'-bis-alpha-L-arabinofuranosyl branched beta-D-(1-->6)-linked galactopyranosyl tetraose (16), 3',2'-bis-alpha-L-arabinofuranosyl branched beta-D-(1-->6)-linked galactopyranosyl hexaose (27), and a twentyose (42) consisting of beta-(1-->6)-linked D-galactopyranosyl pentadecaoligosaccharide backbone with alpha-L-arabinofuranosyl side chains alternately attached at C-2 and C-3 of the middle galactose residue of each consecutive beta-(1-->6)-linked galactotriose unit of the backbone, were synthesized with isopropyl 3-O-allyl-2,4-di-O-benzoyl-1-thio-beta-D-galactopyranoside (6), 2,3,4,6-tetra-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (7), 2,3,5-tri-O-benzoyl-alpha-L-arabinofuranosyl trichloroacetimidate (12), 6-O-acetyl-2,3,4-tri-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (17), 4-methoxyphenyl 2,3,4-tri-O-benzoyl-beta-D-galactopyranoside (19), and 2,6-di-O-acetyl-3,4-di-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (28) as the key synthons. Condensation of 6 with 7 gave the disaccharide donor 8, and subsequent condensation of 8 with 4-methoxyphenyl 2,3,4-tri-O-benzoyl-beta-D-galactopyranosyl-(1-->6)-2-O-acetyl-3,4-di-O-benzoyl-beta-D-galactopyranoside (9) followed by selective deacetylation afforded the tetrasaccharide acceptor 11. Coupling of 11 with 12 gave the pentasaccharide 13, its deallylation followed by coupling with 12, and debenzoylation gave the hexasaccharide 16 with beta-(1-->6)-linked galactopyranose backbone and 2- and 3'-linked alpha-L-arabinofuranose side chains. The octasaccharide 27 was similarly synthesized, while the twentyoside 42 was synthesized with tetrasaccharides 33 or 24 as the donors and 23, 36, 38, and 40 as the acceptors by consecutive couplings followed by deacylation.     

Syntheses of arabinogalactans consisting of beta-(1-->6)-linked D-galactopyranosyl backbone and alpha-(1-->3)-linked L-arabinofuranosyl side chains

Two arabinogalactosyl nonasaccharides, beta-D-Galp-(1-->6)-[alpha-L-Araf-(1-->3)]-beta-D-Galp-(1-->6)-beta-D-Galp-(1-->6)-beta-D-Galp-(1-->6)-[alpha-L-Araf-(1-->5)-alpha-L-Araf-(1-->3)]-beta-D-Galp-(1-->6)-beta-D-Galp and beta-D-Galp-(1-->6)-[alpha-L-Araf-(1-->5)-alpha-L-Araf-(1-->3)]-beta-D-Galp-(1-->6)-beta-D-Galp-(1-->6)-beta-D-Galp-(1-->6)-[alpha-L-Araf-(1-->3)]-beta-D-Galp-(1-->6)-beta-D-Galp, were synthesized as their 4-methoxyphenyl glycosides with 2,3,4,6-tetra-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (1), 6-O-acetyl-2,3,4-tri-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (14), 4-methoxyphenyl 3-O-allyl-2,4-di-O-benzoyl-beta-D-galactopyranoside (2), 4-methoxyphenyl 2,3,4-tri-O-benzoyl-beta-D-galactopyranoside (5), 2,3,5-tri-O-benzoyl-alpha-L-arabinofuranosyl trichloroacetimidate (8), and 2,3,5-tri-O-benzoyl-alpha-L-arabinofuranosyl-(1-->5)-2,3-di-O-benzoyl-alpha-L-arabinofuranosyl trichloroacetimidate (11), as the key synthons. The tetra- (10) and pentasaccharide donor (13), and the tetra- (20) and pentasaccharide acceptor (22) were synthesized based on these synthons through simple transformations. Coupling of 22 with 10, and coupling of 20 with 13 and subsequent deacylation gave nonasaccharides 24 and 26, respectively, consisting of beta-(1-->6)-linked glactopyranosyl backbone and alpha-(1-->3)-linked arabinofuranosyl side chains of different size.     

Synthesis of divalent beta-(1-->6)-branched (1-->3)-glucohexaose and trivalent beta-(1-->6)-branched (1-->3)-glucotriose

Hexaose, beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp, based dimers were synthesized by twofold glycosidation of the hexaosyl trichloroacetimidate with hexylene 1,6-diol, diethylene glycol and triethylene glycol, respectively. Meanwhile, a triose, beta-1D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp, based trimer was obtained by glycosidation of the triosyl trichloroacetimidate with a glycerol-derived triol scaffold.     

Synthesis of mannose-containing analogues of (1-->6)-branched (1-->3)-glucohexaose (I)

alpha-D-Manp-(1-->3)-[alpha-D-Manp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-[alpha-D-Manp-(1-->6)]-D-Glcp and alpha-D-Manp-(1-->3)-[beta-D-Glcp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)[-alpha-D-Manp-(1-->6)]-D-Glcp were synthesized in a regio- and stereoselective way as the mannose-containing analogues of the immunomodulating beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-D-Glcp.     

A practical synthesis of alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->2)-[alpha-D-Glcp-(1-->3)]-alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->2)-alpha-D-Manp, an O-specific heterohexasaccharide fragment of Citrobacter braakii O7a, 3b, 1c

An O-specific heterohexasaccharide fragment of Citrobacter braakii O7a, 3b, 1c, alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->2)-[alpha-D-Glcp-(1-->3)]-alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->2)-alpha-D-Manp was synthesized as its methyl glycoside. Acetylation of allyl 4,6-O-benzylidene-alpha-D-mannopyranoside, followed by debenzylidenization and benzoylation gave allyl 2,3-di-O-acetyl-4,6-di-O-benzoyl-alpha-D-mannopyranoside (3), and subsequent deacetylation of 3 with CH(3)COCl-MeOH gave the monosaccharide acceptor 4. Condensation of isopropyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-glucopyranoside (6) with 4 selectively afforded the alpha-(1-->3)-linked disaccharide 7. Condensation of 7 with the (1-->3)-linked disaccharide donor 9, followed by deallylation and trichloroacetimidation, afforded the tetrasaccharide donor 12. Coupling of 12 with disaccharide acceptor 13, followed by debenzylation and deacylation, furnished the target heterohexasaccharide 16.     

1,3-Dideoxynojirimycin-3-yl glycosides of beta-(1-->3)- and beta-(1-->6)-linked gluco-oligosaccharides

Standard chemical methods involving the use of O-acetylated glycosyl trichloroacetimidates as glycosylating agents were used to prepare the five 1,3-dideoxynojirimycin-3-yl beta-(1-->3)-linked oligo-glucosides (1-5) and also the beta-(1-->6)-bonded glucobiose (gentiobiose)-based analogue 6 as potential fungicides. In the course of the work, the beta-(1-->6), beta-(1-->6)-linked analogue 8 of 6 and 6-O- and 4-O-beta-glucopyranosyl-deoxynojirimycins 7 and 9, respectively, were also produced.     

Synthesis of (R)-2,3-epoxypropyl(1-->3)-beta-D-pentaglucoside

The title pentasaccharide was synthesized via a 2+3 strategy. The disaccharide donor, 3-O-acetyl-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranosyl trichloroacetimidate (8), was obtained by selective coupling of allyl 2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranoside with 3-O-acetyl-2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranosyl trichloroacetimidate (4), followed by deallylation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor, allyl 2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranoside (12), was prepared by coupling of allyl 2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranoside with 4, followed by deacetylation. Condensation of 8 with 12, followed by epoxidation, and deprotection, gave the target pentaoside.     

Effect of (1-->3)- and (1-->4)-linkages of fully sulfated polysaccharides on their anticoagulant activity

Chemically fully sulfated polysaccharides including xylan (-->4Xylbeta-(1-->4)Xylbeta1-->), amylose (-->4Glcalpha-(1-->4)Glcalpha1-->), cellulose (-->4Glcbeta-(1-->4)Glcbeta1-->), curdlan (-->3Glcbeta-(1-->3)Glcbeta1-->) and galactan (-->3Galbeta-(1-->3)Galbeta1-->), which have been isolated from Korean clam, were prepared, and their anticoagulant activity was investigated. The results strongly suggest that the activity might not be depending on anomeric configuration (alpha or beta) or monosaccharide species but on the glycosidic linkage, either (1-->3) or (1-->4). 1H NMR studies of these modified polysaccharides show that the neighboring sulfate groups at the C-2 and C-3 positions might have caused the conformational changes of each monosaccharide from 4C(1) to 1C(4). Furthermore, the effect of 6-sulfate residues on the anticoagulant activity was investigated using a specific desulfated reaction for the chemically fully sulfated polysaccharides. The 6-sulfate group is very important in determining anticoagulant activity of (1-->3)-linked polysaccharides, whereas the activity is not affected by presence or absence of the 6-sulfate group in (1-->4)-linked polysaccharides.     

Synthesis of alpha-Manp-(1-->2)-alpha-Manp-(1-->3)-alpha-Manp-(1-->3)-Manp, the tetrasaccharide repeating unit of Escherichia coli O9a, and alpha-Manp-(1-->2)-alpha-Manp-(1-->2)-alpha-Manp-(1-->3)-alpha-Manp-(1-->3)-Manp, the pentasaccharide repeating unit of E. coli O9 and Klebsiella O3

The tetrasaccharide repeating unit of Escherichia coli O9a, alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-D-Manp, and the pentasaccharide repeating unit of E. coli O9 and Klebsiella O3, alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-D-Manp, were synthesized as their methyl glycosides. Thus, selective 3-O-allylation of p-methoxyphenyl alpha-D-mannopyranoside via a dibutyltin intermediate gave p-methoxyphenyl 3-O-allyl-alpha-D-mannopyranoside (2) in good yield. Benzoylation (-->3), then removal of 1-O-methoxyphenyl (right arrow4), and subsequent trichloroacetimidation afforded the 3-O-allyl-2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate (5). Condensation of 5 with methyl 4,6-O-benzylidene-alpha-D-mannopyranoside (6) selectively afforded the (1-->3)-linked disaccharide 7. Benzoylation of 7, debenzylidenation, benzoylation, and deallylation gave methyl 2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->3)-2,4,6-tri-O-benzoyl-alpha-D-mannopyranoside (11) as the disaccharide acceptor. Coupling of 11 with (1-->2)-linked mannose disaccharide donor 17 or trisaccharide donor 21, followed by deacylation, furnished the target tetrasaccharide and pentasaccharide, respectively.     

Systematic synthesis of sulfated sialyl-alpha-(2 --> 3)-neolactotetraose derivatives and their acceptor specificity for an alpha-(1 --> 3)-fucosyltransferase (Fuc-TVII) involved in the biosynthesis of L-selectin ligand

Sulfated sialyl-alpha-(2 --> 3)-neolactotetraose (IV3NeuAcnLcOse4) derivatives at C-6 of GlcNAc (6-O-sulfo), terminal Gal (6'-O-sulfo), and both GlcNAc and Gal (6,6'-di-O-sulfo) residues have systematically been synthesized. (Methyl 5-acetamido-4,7,8,9- tetra-O-acetyl-3,5-dideoxy-D-glycero-alpha-D-galacto-2-nonulopyranosy lonate)-(2 --> 3)-2,4-di-O-benzoyl-6-O-levulinoyl-D-galactopyranosyl trichloroacetimidate was coupled with 2-(trimethylsilyl)ethyl (2-acetamido-2-deoxy- 3-O-benzyl-6-O-p-methoxyphenyl-beta-D-glucopyranosyl)-(1 --> 3)-(2,4,6-tri-O-benzyl-beta-D-galactopyranosyl)-(1 --> 4)-2,3,6-tri-O-benzyl-beta-D-glucopyranoside to give the suitably protected pentasaccharide which, upon selective removal of the p-methoxyphenyl and/or levulinoyl groups at C-6 of the GlcNAc and the terminal Gal residues, successive O-sulfation(s) and deprotection, afforded the desired three sulfated IV3NeuAcnLcOse4 derivatives. Acceptor specificity of the synthetic IV3NeuAcnLcOse4 probes for a human alpha-(1 --> 3)-fucosyltransferase (Fuc-TVII) was examined to study the biosynthetic pathway of L-selectin ligand. Only the 6-sulfated derivative at C-6 of GlcNAc was recognized by Fuc-TVII to give 6-O-sulfo sialyl LeX.     

Synthesis, (1-->3)-beta-D-glucanase-binding ability and phytoalexin-elicitor activity of (R)-2,3-epoxypropyl (1-->3)-beta-D-pentaglucoside

The (1-->3)-beta-D-pentaglucoside was synthesized as its (R)-2,3-epoxypropyl glycoside via 2+3 strategy. The disaccharide donor 8 was obtained by 3-selective coupling of 2 with 4, followed by deallylation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor 12 was prepared by coupling of 10 with 4, followed by deacetylation. Condensation of 8 with 12, followed by epoxidation, and deprotection, gave the target pentaoside. The results of these bioassays demonstrated that the (1-->3)-beta-D-glucanase was obviously inactivated by 15 with k(app)=3.79 x 10(-4) min(-1). At the same time, we found that the 15 was more active as compared to the laminaripentaose in eliciting phytoalexin accumulation in tobacco cotyledon tissue, and it could be kept longer time than laminaripentaose, which indicated it is much more stable than laminaripentaose.     

Identification of bound water molecules in the cyclic tetrasaccharide, cyclo-{-->6}-alpha-d-Glcp-(1-->3)-alpha-D-Glcp-(1-->6)-alpha-D-Glcp-(1-->3)-alpha-D-Glcp-(1-->)

A structural characterization of bound water molecules in the cyclic tetrasaccharide, cyclo-{-->6}-alpha-D-Glcp-(1-->3)-alpha-D-Glcp-(1-->6)-alpha-D-Glcp-(1-->3)-alpha-D-Glcp-(1-->), was carried out by NMR spectroscopy. H-1', 2'-OH, H-3', and 4'-OH of the 3-O-glycosylated residue and H-1 of the 6-O-glycosylated residue were found to cross-relax with protons of bound waters using the double-pulsed field-gradient spin-echo ROESY experiment. In the crystal structure, one water molecule is located in the center of the plate, and its temperature factor is very low, indicating that this water molecule is an intrinsic component.     

Candida albicans cell wall comprises a branched beta-D-(1-->6)-glucan with beta-D-(1-->3)-side chains

The structure of immunogenic and immunomodulatory cell wall glucans of Candida albicans is commonly interpreted in terms of a basic polysaccharide consisting of a beta-D-(1-->3)-linked glucopyranosyl backbone possessing beta-D-(1-->6)-linked side chains of varying distribution and length. This proposed molecular architecture has been re-evaluated by the present study on the products of selective enzymolysis of insoluble C. albicans glucan particles (GG). High resolution 1H (400 and 700 MHz) and 13C (100 and 175 MHz) NMR analyses were performed on a soluble beta-glucan preparation (GG-Zym) obtained by GG digestion with endo-beta-D-(1-->3)-glucanase and on its high- (Pool 1) and low-molecular weight (Pool 2) sub-fractions. The resonances typical of uniformly beta-D-(1-->6)- and beta-D-(1-->3)-linked linear glucans, together with additional multiplets assigned to short-chain oligoglucosides, were detected in GG-Zym. Pool 1 (46.3+/-6.4% of GG-Zym content) consisted of beta-D-(1-->6)-linked glucopyranosyl polymers, with short beta-D-(1-->3)-branched side chains of 2.20+/-0.02 units (branching degree (DB)=0.14+/-0.03). Pool 2 was a mixture of glucose and linear short-chain beta-D-(1-->3)-oligoglucosides. Further digestion of Pool 1 by beta-D-(1-->6)-glucanase yielded a mixture of glucose and short beta-D-(1-->6)-linked, either linear or beta-D-(1-->3,6) branched, oligomers. These endoglucanase digestion patterns were consistent with the presence in C. albicans cell wall glucans of beta-D-(1-->6)-linked glucopyranosyl backbones possessing beta-D-(1-->3)-linked side chains, a structure very close to that of beta-D-(1-->6)-glucan from Saccharomyces cerevisiae yeast. This finding may provide the grounds for further elucidation of the cell wall structure and a better understanding of the biological properties of C. albicans beta-glucans.     

Synthesis of beta-(1-->6)-branched beta-(1-->3) glucohexaose and its analogues containing an alpha-(1-->3) linked bond with antitumor activity

A beta-(1-->6)-branched beta-(1-->3)-glucohexaose, present in many biologically active polysaccharides from traditionally herbal medicines such as Ganoderma lucidum, Schizophyllum commune and Lentinus edodes, was synthesized as its lauryl glycoside 32, and its analogues 18, 20 and 33 containing an alpha-(1-->3) linked bond were synthesized. It is interesting to find that coupling of a 3,6-branched acylated trisaccharide trichloroacetimidate donor 9 with 3,6-branched acceptors 13 and 16 with 3'-OH gave the alpha-(1--> 3)-linked hexasaccharides 17 and 19, respectively, in spite of the presence of C-2 ester capable of neighboring group participation. However, coupling of 9 with 4-methoxyphenyl 4,6-O-benzylidene-beta-D-glucopyranoside (27) selectively gave beta-(1-->3)-linked tetrasaccharide 28. Simple chemical transformation of the tetrasaccharide 28 gave acylated tetrasaccharide trichloroacetimidate 29. Coupling of 29 with lauryl (1-->6)-linked disaccharide 26 with 3-OH gave beta-(1-->3)-linked hexasaccharide 30 as the major product. Bioassay showed that in combination with the chemotherapeutic agent cyclophospamide (CPA), the hexaose 18 at a dose of 0.5-1mg/kg substantially increased the inhibition of S(180) for CPA, but decreased the toxicity caused by CPA. Some of these oligosaccharides also inhibited U(14) noumenal tumor in mice effectively.     

A practical synthesis of beta-D-GlcA-(1-->3)-beta-D-Gal-(1-->3)-beta-D-Gal-(1-->4)-D-Xyl,a part of the common linkage region of a glycosaminoglycan

A practical synthesis of beta-D-GlcA-(1-->3)-beta-D-Gal-(1-->3)-beta-D-Gal-(1-->4)-beta-D-Xyl-(1-->OMe) was achieved by coupling of methyl 2,3,4-tri-O-acetyl-alpha-D-glucopyranosyluronate trichloroacetimidate with a trisaccharide acceptor. The trisaccharide acceptor was obtained by condensation of 3-O-allyl-2,4,6-tri-O-benzoyl-beta-D-galactopyranosyl-(1-->3)-2,4,6-tri-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate with methyl 2,3-di-O-benzoyl-beta-D-xylopyranoside, followed by deallylation. The beta-(1-->3)-linked disaccharide was prepared readily with p-methoxyphenyl 3-O-allyl-2,4,6-tri-O-benzoyl-beta-D-galactopyranoside as the key synthon. The alpha-(1-->3)-linkage was formed in considerable amount with galactose mono- and disaccharide trichloroacetimidate donors with C-2 neighboring group participation.