Synthesis of methyl 3-O-alpha-D-mannopyranosyl-alpha-D-talopyranoside and methyl 3-O-alpha-D-talopyranosyl-alpha-D-talopyranoside

Methyl 2-O-benzyl-3-O-(2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl)-alpha- D-mannopyranoside (4) and methyl 2-O-benzyl-3-O-alpha-D-mannopyranosyl-alpha-D-mannopyranoside (6) were prepared from a common intermediate, namely, methyl 2-O-benzyl-4,6-O-benzylidene-3-O-(2,3,4,6-tetra-O-acetyl-alpha-D- mannopyranosyl)-alpha-D-mannopyranoside. On treatment with tert-butylchlorodiphenylsilane, in N,N-dimethylformamide in the presence of imidazole, 4 and 6 afforded methyl 2-O-benzyl-6-O-tert-butyldiphenylsilyl-3-O-(2,3,4,6-tetra-O-acetyl -alpha-D- mannopyranosyl)-alpha-D-mannopyranoside (7), and methyl 2-O-benzyl-6-O-tert-butyldiphenylsilyl-3-O-(6-O-tert- butyldiphenylsilyl-alpha-D-mannopyranosyl)-alpha-D-mannopyranoside (8), respectively. Compound 8 was converted into its 2,3-O-isopropylidene derivative (9), and oxidation of 7 and 9 with pyridinium chlorochromate, and reduction of the resulting carbonyl intermediates gave methyl 2-O-benzyl-6-O-tert-butyldiphenylsilyl-3-O-(2,3,4,6-tetra-O-acetyl -alpha-D- mannopyranosyl)-alpha-D-talopyranoside and methyl 2-O-benzyl-6-O-tert-butyldiphenylsilyl-3-O-(6-O-tert-butyldiphe nylsilyl- 2,3-O-isopropylidene-alpha-D-talopyranosyl)-alpha-D-talopyranoside , respectively. Removal of the protecting groups furnished the title disaccharides.     

Synthesis of two isomeric pentasaccharides,the possible repeating unit of the beta-glucan from the micro fungus Epicoccum nigrum Ehrenb. ex Schlecht

Two isomeric pentasaccharides, beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->6)]-beta-D-Glcp (I) and beta-D-Glcp-(1-->6)-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->3)-beta-D-Glcp-(1-->6)]-beta-D-Glcp (II), the possible repeating unit of the beta-glucan from the micro fungus Epicoccum nigrum Ehrenb. ex Schlecht, were synthesized as their 4-methoxyphenyl glycosides in a regio- and stereoselective manner. The pentasaccharide I was obtained from 3-O-selective glycosylation of 4-methoxyphenyl 4,6-O-benzylidene-beta-D-glucopyranoside (12) with 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-glucopyranosyl trichloroacetimidate (6) followed by acetylation, debenzylidenation, and 6-O-selective glucosylation with 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl trichloroacetimidate (1), and then by deprotection. The pentasaccharide II was obtained from 3-O-selective coupling of 12 with 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->6)-2,4-di-O-acetyl-3-O-allyl-alpha-D-glucopyranosyl trichloroacetimidate (10) followed by acetylation, debenzylidenation, and 6-O-selective glycosylation with 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-2,4,6-tri-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (11), and finally by deprotection.     

Synthesis of a mannose heptasaccharide existing in baker's yeast,Saccharomyces cerevisiae X2180-1A wild-type strain

A mannose heptasaccharide existing in baker's yeast, Saccharomyces cerevisiae X2180-1A wild-type strain, was effectively synthesized as its allyl glycoside via TMSOTf-promoted condensation of a disaccharide donor 13 with a pentasaccharide acceptor 12, followed by deprotection. The pentasaccharide 12 was constructed by coupling of 2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl-(1-->3)-2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->2)-3,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->2)-3,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate (9) with allyl 6-O-acetyl-3,4-di-O-benzoyl-alpha-D-mannopyranoside (10), followed by deacetylation. The tetrasaccharide 9 was obtained by coupling of 2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl-(1-->3)-2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate (5) with allyl 3,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->2)-3,4,6-tri-O-benzoyl-alpha-D-mannopyranoside (6), followed by deallylation and trichloroacetimidation. The disaccharides 6 and 13 were readily obtained by known methods.     

Synthesis of alpha-galactosylated fragments related to the core-structure of the GPI anchor of Trypanosoma brucei

A series of octyl glycosides di- to tetrasaccharides related to the GPI anchor of Trypanosoma brucei was prepared. Treatment of octyl 2-O-benzoyl-4,6-O-(1,1,3,3-tetraisopropyl-1,3-disiloxane-1,3 -diyl)-alpha-D-mannopyranoside with ethyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-galactopyranoside under activation with bromine and silver trifluoromethanesulfonate afforded the alpha-linked disaccharide octyl 2-O-benzoyl-3-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-4,6-O- (1,1,3,3-tetraisopropyl-1,3-disiloxane-1,3-diyl)-alpha -D-mannospyranoside, the siloxane ring of which was regioselectively opened with a HF-pyridine complex to give the disaccharide acceptor octyl 3-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-2-O-benzoyl-4-O-(3 -fluoro-1,1,3,3-tetraisopropyl-1,3-disiloxane-3-yl)-alpha-D- mannopyranoside (4). Mannosylation of 4 with benzobromomannose (7), followed by fluoride catalyzed desilylation gave the trisaccharide octyl 2-O-benzoyl-6-O-(2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl)-3-O-(2, 3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-alpha-D-mannospyranosi de, which was deblocked via the deacylated intermediate octyl 3-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-6-O-(alpha-D-manno pyranosyl)-alpha-D-mannospyranoside to afford the octyl glycoside trisaccharide octyl 3-O-(alpha-D-galactopyranosyl)-6-O-(alpha-D-mannopyranosyl)-alpha-D-m annospyranoside. Glycosylation of 4 with 3,4,6-tri-O-acetyl-2-O-(2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl)- alpha-D-mannopyranosyl trichloroacetimidate resulted in the tetrasaccharide octyl 2-O-benzoyl-4-O-(1-fluoro-1,1,3,3-tetraisopropyl-1,3-disiloxane -3-yl)-3-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-6-O-[2-O -(2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl)-3,4,6-tri-O-acetyl-alp ha-D-mannopyranosyl]-alpha-D-mannospyranoside, sequential desilylation, deacylation and debenzylation, respectively, of which via the intermediate octyl 2-O-benzoyl-3-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-6-O-[2 -O-(2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranosyl)-3,4,6-tri-O-acetyl-a lpha-D-mannopyranosyl]-alpha-D-mannospyranoside afforded the octyl glycoside tetrasaccharide octyl 3-O-(alpha-D-galactopyranosyl)-6-O-[2-O-(alpha-D-mannopyranosyl)-alpha-D -mannopyranosyl]-alpha-D-mannospyranoside.     

Synthesis of beta-D-glucose oligosaccharides from Phytophthora parasitica

The glucohexaose, beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-beta-D-Glcp-(1-->6)]-beta-D-Glcp-(1-->3)-D-Glcp, was synthesized as its allyl glycoside via 3+3 strategy. The trisaccharide donor, 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-2,4,6-tri-O-acetyl-beta-D-glucopyranosyl-(1-->3)-2,4,6-tri-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (11), was obtained by 3-selective coupling of isopropyl 4,6-O-benzylidene-1-thio-beta-D-glucopyranoside (2) with 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-2-O-acetyl-4,6-O-benzylidene-alpha-D-glucopyranosyl trichloroacetimidate (6), followed by hydrolysis, acetylation, dethiolation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor, allyl 2,3,4,6-tetra-O-benzoyl-beta-D-glucopyranosyl-(1-->3)-2-O-acetyl-beta-D-glucopyranosyl-(1-->3)-4,6-di-O-acetyl-2-O-benzoyl-alpha-D-glucopyranoside (14), was prepared by coupling of allyl 4,6-di-O-acetyl-2-O-benzoyl-alpha-D-glucopyranoside (12) with 6, followed by debenzylidenation. Condensation of 14 with 11, followed by deacylation, gave the target hexaoside. A beta-(1-->3)-linked tetrasaccharide 29 was also synthesized with methyl 2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2,4,6-tri-O-acetyl-beta-D-glucopyranoside (25) as the acceptor and acylated beta-(1-->3)-linked disaccharide 21 as the donor.     

Glycosides for testing glycosidases: Vinyl,l-ethoxyethyl,and l-ethoxybut-3-enyl d-glucopyranosides

The reaction of ethyl vinyl ether and 2,3,4,6-tetra-O-acetyl-β-d-glucopyranose (1) in the presence of Hg-(OAc)₂ and toluene-p-sulphonic acid as catalysts yielded the acetylated vinyl, l-ethoxyethyl, and l-ethoxybut-3-enyl glycosides in varying proportions. Crystalline l-ethoxybut-3-enyl 2,3,4,6-tetra-O-acetyl-β-d-glucopyranoside (2), vinyl 2,3,4,6-tetra-O-acetyl-α-d-glucopyranoside (3), and l-ethoxyethyl 2,3,4,6-tetra-O-acetyl-β-d-glucopyranoside (4) were isolated by chromatography. Compound 4 was also prepared by the reaction of 1 with cold acetaldehyde diethyl acetal containing a trace of acetic acid, and its α anomer (5) by the reaction of 1 with boiling acetaldehyde diethyl acetal containing a trace of acetic acid. Each deacetylated d-glucoside was cleaved by the corresponding d-glucosidase, to yield d-glucose and either acetaldehyde (from deacetylated 3-5) or but-3-enal (from deacetylated 2).     

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 lacto-N-neotetraose and lacto-N-tetraose using the dimethylmaleoyl group as amino protective group.

The disaccharide donor O-[2,3,4,6-tetra-O-acetyl-beta-D- galactopyranosyl)-(1-->4)-3,6-di-O-benzyl-2-deoxy-2-dimethylmaleimido - alpha,beta-D-glucopyranosyl] trichloroacetimidate (7) was prepared by reacting O-(2,3,4,6-tetra-O-acetyl- alpha-D-galactopyranosyl) trichloroacetimidate with tert-butyldimethylsilyl 3,6-di-O-benzyl-2-deoxy-2- dimethylmaleoylamido-glucopyranoside to give the corresponding disaccharide 5. Deprotection of the anomeric center and then reaction with trichloroacetonitrile afforded 7. Reaction of 7 with 3'-O-unprotected benzyl (2,4,6-tri-O-benzyl-beta-D-galactopyranosyl)- (1-->4)-2,3,6-tri-O-benzyl-beta-D-glucopyranoside (8) as acceptor afforded the desired tetrasaccharide benzyl (2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1-->4)-(3,6-di-O- benzyl-2-deoxy-2-dimethylmaleimido-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. Replacement of the N-dimethylmaleoyl group by the acetyl group, O-debenzylation and finally O-deacetylation gave lacto-N-neotetraose. Similarly, reaction of O-[(2,3,4,6-tetra-O-acetyl-beta- D-galactopyranosyl)-(1-->3)-4,6-O-benzylidene-2-deoxy-2-dimethylmalei mido- alpha,beta-D-glycopyranosyl] trichloroacetimidate as donor with 8 as acceptor afforded the desired tetrasaccharide benzyl (2,3,4,6-tetra-O-acetyl-beta-D- galactopyranosyl)-(1-->3)-(4,6-benzylidene-2-deoxy-2-dimethylmaleimid o- beta-D-glucopyranosyl)-(1-->3)-(2,4,6-tri-O-benzyl-beta-D-galactopyranos yl)- (1-->4)-2,3,6-tri-O-benzyl-beta-D-glucopyranoside. Removal of the benzylidene group, replacement of the N-dimethylmaleoyl group by the acetyl group and then O-acetylation afforded tetrasaccharide intermediate 15, which carries only O-benzyl and O-acetyl protective groups. O-Debenzylation and O-deacetylation gave lacto-N-tetraose (1). Additionally, known tertbutyldimethylsilyl (2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1-->3)-4,6-O-benzylide ne- 2-deoxy-2-dimethylmaleimido-beta-D-glucopyranoside was transformed into O-[2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)- (1-->3)-4,6-di-O-acetyl-2-deoxy-2-dimethylmaleimido-alpha,beta-D- glucopyranosyl] trichloroacetimidate as glycosyl donor, to afford with 8 as acceptor the corresponding tetrasaccharide 22, which is transformed into 15, thus giving an alternative approach to 1.     

Synthesis of the tetrasaccharide core region of antigenic lipo-oligosaccharides characteristic of Mycobacterium kansasii

The oligosaccharide core region, beta-D-Glcp-(1----3)-beta-D-Glcp-(1----4)-alpha-D-Glcp- 1----1)-alpha-D-Glcp (1), of the lipo-oligosaccharide-type antigens isolated from M. kansasii has been synthesised from 2,3,2',3',4',6'-hexa-O-benzyl-6-O-(1-phenylethyl)-alpha, alpha-trehalose (4). Compound 4 was obtained by LiAlH4-AlCl3-type hydrogenolysis of 2,3,2',3',4',6'-hexa-O-benzyl-4,6-O-(S)-(1-phenylethylidene)-alpha , alpha-trehalose. The beta-laminaribiosyl part of the molecule was built-up by sequential glycosylation steps using 2,4,6-tri-O-acetyl-3-O-allyl-alpha-D-glucopyranosyl bromide in the presence of HgBr2 and methyl 2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranoside promoted by methyl triflate. The complete a priori 13C-n.m.r. spectrum assignment of 1 was achieved by applying 2D methods.     

Regioselective synthesis of 1I,1II,5I,5II,6I,6I,6II,6II-2H8-cellobiose

Partially deuteriated 1,5,6,6-(2)H(4)-d-glucose and 1(I),1(II),5(I),5(II),6(I),6(I),6(II),6(II)-(2)H(8)-d-cellobiose were synthesized in high yields and on a large scale from d-glucose. (2)H enrichment at C-5 and C-6 of each glucopyranosyl unit in excess of 85% and 90%, respectively, was realized by (1)H-(2)H exchange in (2)H(2)O containing deuteriated Raney Ni. Nucleophilic addition of LiAlD(4) to 5,6,6-(2)H(3)-2,3,4,6-tetra-O-benzyl-d-gluconolactone led to a 98% (2)H enrichment at C-1. Deuteriated cellobiose is of interest as building block for the synthesis of a model compound of cellulose I.     

A facile and effective synthesis of alpha-(1-->6)-linked mannose di-, tri-, tetra-, hexa-, octa-, and dodecasaccharides, and beta-(1-->6)-linked glucose di-, tri-, tetra-, hexa-, and octasaccharides using sugar trichloroacetimidates as the donors and unprotected or partially protected glycosides as the acceptors

Reaction of 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl trichloroimidate with allyl alpha-D-mannopyranoside in the presence of TMSOTf selectively gave allyl 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl-(1-->6)-alpha-D-mannopyranoside through an orthoester intermediate. Benzoylation of 3, followed by deallylation, and then trichloroimidation afforded the disaccharide donor 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroimidate, while benzoylation of 3 followed by selective removal of acetyl groups yielded the disaccharide acceptor allyl alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranoside. Coupling of 5 with 6 gave the tetrasaccharide allyl 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranoside, which were converted into the tetrasaccharide donor 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroimdate and the tetrasaccharide acceptor allyl alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->6)-2,3,4-tri-O-benzoyl-alpha-D-mannopyranoside, respectively, by the same strategies as used for conversion of 3 into 5 and 6. Condensation of 5 with 13 gave the hexasaccharide 14, while condensation of 12 with 13 gave the octasaccharide 17. Dodecasaccharide 21 was obtained by the coupling of 12 with the octasaccharide acceptor 20. Similar strategies were used for the syntheses of beta-(1-->6)-linked glucose di-, tri-, tetra-, hexa-, and octamers. Deprotection of the oligosaccharides in ammonia-saturated methanol yielded the free alpha-(1-->6)-linked mannosyl and beta-(1-->6)-linked glucosyl oligomers.     

Synthesis of trisaccharide methyl glycosides related to fragments of the capsular polysaccharide of Streptococcus pneumoniae type 18C.

The synthesis is reported of methyl 3-O-(4-O-beta-D-galactopyranosyl-alpha-D- glucopyranosyl)-alpha-L-rhamnopyranoside (1), methyl 2-O-alpha-D-glucopyranosyl-4-O-beta-D-glucopyranosyl-beta-D- galactopyranoside (3), methyl 3-O-(4-O-beta-D-galactopyranosyl-alpha-D-glucopyranosyl)-alpha-L- rhamnopyranoside 3"-(sn-glycer-3-yl sodium phosphate) (2), and methyl 2-O-alpha-D-glucopyranosyl-4-O-beta-D- glucopyranosyl-beta-D-galactopyranoside 3-(sn-glycer-3-yl sodium phosphate) (4), which are trisaccharide methyl glycosides related to fragments of the capsular polysaccharide of Streptococcus pneumoniae type 18C ([----4)-beta-D- Glcp-(1----4)-[alpha-D-Glcp-(1----2)]-[Glycerol-(1-P----3)]-beta-D-Galp - (1----4)-alpha-D-Glcp-(1----3)-alpha-L-Rhap-(1----]n). Ethyl 4-O-acetyl-2,3,6-tri-O-benzyl-1-thio-beta-D-glucopyranoside (10) was coupled with benzyl 2,4-di-O-benzyl-alpha-L-rhamnopyranoside (6). Deacetylation of the product, followed by condensation with 2,4,6-tri-O-acetyl-3-O-allyl-alpha-D-galactopyranosyl trichloroacetimidate (18), gave benzyl 2,4-di-O-benzyl-3-O-[2,3,6-tri-O- benzyl-4-O-(2,4,6-tri-O-acetyl-3-O-allyl-beta-D-galactopyranosyl)-alpha- D- glucopyranosyl]-alpha-L-rhamnopyranoside (19). Acetolysis of 19, followed by methylation, deallylation (----22), and further deprotection afforded 1. Condensation of methyl 2,4-di-O-benzyl-3-O-[2,3,6-tri-O-benzyl-4-O-(2,4,6-tri- O-acetyl-beta-D-galactopyranosyl)-alpha-D-glucopyranosyl]-alpha-L- rhamnopyranoside (22) with 1,2-di-O-benzyl-sn-glycerol 3-(triethyl-ammonium phosphonate) (24), followed by oxidation and deprotection, yielded 2. Condensation of ethyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-glucopyranoside (27) with methyl 3-O-allyl-4,6-O-benzylidene-beta-D-galactopyranoside (28), selective benzylidene ring-opening of the product, coupling with 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (31), and deallylation afforded methyl 6-O-benzyl-4-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-2-O- (2,3,4,6-tetra-O-benzyl-alpha-D-glucopyranosyl)-beta-D-galactopyranoside (33). Deprotection of 33 gave 3, and condensation of 33 with 24, followed by oxidation and deprotection, gave 4.     

Glycosidation of 2,5-anhydro-3,4-di-O-benzyl-D-mannitol with different glucopyranosyl donors: a comparative study

2,5-Anhydro-3,4-di-O-benzyl-D-mannitol was glycosylated using different donors such as tetra-O-acetyl-alpha-D-glucopyranosyl bromide in the presence of Hg(CN)(2), the corresponding beta-thiophenylglycoside in the presence of NIS and TfOH as well as the alpha- and beta-trichloroimidate with TMSOTf as promoter. The resulting mixtures were analyzed by HPLC and the following main components were isolated and characterized: 2,5-anhydro-3,4-di-O-benzyl-1-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-d-mannitol; 6-O-acetyl-2,5-anhydro-3,4-di-O-benzyl-1-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-D-mannitol; 2,5-anhydro-3,4-di-O-benzyl-1,6-bis-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-D-mannitol; 2,5-anhydro-3,4-di-O-benzyl-1-O-[-2-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-3,4,6-tri-O-acetyl-beta-D-glucopyranosyl]-6-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-D-mannitol and 2,5-anhydro-3,4-di-O-benzyl-1,6-bis-O-(3,4,6-tri-O-acetyl-1,2-O-ethylidene-2'-yl-alpha-D-glucopyranosyl)-D-mannitol. The latter compound representing a bis-orthoester might be a common intermediate in all the investigated reactions, as its rearrangement and/or decomposition can yield all of the isolated compounds.     

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).     

Spectral studies with lactoperoxidase and thyroid peroxidase: interconversions between native enzyme, compound II, and compound III

Spectral scans in both the visible (650-450 nm) and the Soret (450-380 nm) regions were recorded for the native enzyme, Compound II, and Compound III of lactoperoxidase and thyroid peroxidase. Compound II for each enzyme (1.7 microM) was prepared by adding a slight excess of H2O2 (6 microM), whereas Compound III was prepared by adding a large excess of H2O2 (200 microM). After these compounds had been formed it was observed that they were slowly reconverted to the native enzyme in the absence of exogenous donors. The pathway of Compound III back to the native enzyme involved Compound II as an intermediate. Reconversion of Compound III to native enzyme was accompanied by the disappearance of H2O2 and generation of O2, with approximately 1 mol of O2 formed for each 2 mol of H2O2 that disappeared. A scheme is proposed to explain these observations, involving intermediate formation of the ferrous enzyme. According to the scheme, Compound III participates in a reaction cycle that effectively converts H2O2 to O2. Iodide markedly affected the interconversions between native enzyme, Compound II, and Compound III for lactoperoxidase and thyroid peroxidase. A low concentration of iodide (4 microM) completely blocked the formation of Compound II when lactoperoxidase or thyroid peroxidase was treated with 6 microM H2O2. When the enzymes were treated with 200 microM H2O2, the same low concentration of iodide completely blocked the formation of Compound III and largely prevented the enzyme degradation that otherwise occurred in the absence of iodide. These effects of iodide are readily explained by (i) the two-electron oxidation of iodide to hypoiodite by Compound I, which bypasses Compound II as an intermediate, and (ii) the rapid oxidation of H2O2 to O2 by the hypoiodite formed in the reaction between Compound I and iodide.     

Synthesis of cerebroside, lactosyl ceramide, and ganglioside GM3 analogs containing beta-thioglycosidically linked ceramide

Coupling of the sodium salt of 2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranose, -beta-D-galactopyranose, O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1----4)-2,3,6-tri-O- acetyl- 1-thio-beta-D-glucopyranose, or O-(methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-alpha-D-galacto -2- nonulopyranosylonate)-(2----3)-O-(2,3-di-O-acetyl-6-O-bezoyl -beta-D- galactopyranosyl)-(1----4)-3-O-acetyl-2,6-di-O-benzoyl-1-thio-beta-D- glucopyranose, which were prepared from the corresponding 1-S-acetates, 1, 3, 6, and 9, with (2S,3R,4E)-2-azido-3-O-benzoyl-1-O-(p-tolylsulfonyl)-4-oc tadecene-1,3-diol (12) derived by tosylation of 11, gave the corresponding beta-thioglycosides 13, 17, 21, and 25, respectively in good yield. The beta-thioglycosides obtained were converted, via selective reduction of the azide group, condensation with octadecanoic acid, and removal of the protecting groups, into the title compounds.     

Reactions of the class II peroxidases, lignin peroxidase and Arthromyces ramosus peroxidase, with hydrogen peroxide. Catalase-like activity, compound III formation, and enzyme inactivation

The reactions of the fungal enzymes Arthromyces ramosus peroxidase (ARP) and Phanerochaete chrysosporium lignin peroxidase (LiP) with hydrogen peroxide (H(2)O(2)) have been studied. Both enzymes exhibited catalase activity with hyperbolic H(2)O(2) concentration dependence (K(m) approximately 8-10 mm, k(cat) approximately 1-3 s(-1)). The catalase and peroxidase activities of LiP were inhibited within 10 min and those of ARP in 1 h. The inactivation constants were calculated using two independent methods; LiP, k(i) approximately 19 x 10(-3) s(-1); ARP, k(i) approximately 1.6 x 10(-3) s(-1). Compound III (oxyperoxidase) was detected as the majority species after the addition of H(2)O(2) to LiP or ARP, and its formation was accompanied by loss of enzyme activity. A reaction scheme is presented which rationalizes the turnover and inactivation of LiP and ARP with H(2)O(2). A similar model is applicable to horseradish peroxidase. The scheme links catalase and compound III forming catalytic pathways and inactivation at the level of the [compound I.H(2)O(2)] complex. Inactivation does not occur from compound III. All peroxidases studied to date are sensitive to inactivation by H(2)O(2), and it is suggested that the model will be generally applicable to peroxidases of the plant, fungal, and prokaryotic superfamily.     

Syntheses of an alpha-D-Gal-(1-->6)-beta-D-Gal diglyceride, as lipase substrate

Two different routes were explored to afford 3-O-(6-O-alpha-D-galactopyranosyl-beta-D-galactopyranosyl)-1,2-di-O-dodecanoyl-sn-glycerol. In the first one, the key step was the glycosylation of the 3-O-(2,3,4-tri-O-benzyl-beta-D-galactopyranosyl)-1,2-O-isopropylidene-sn-glycerol acceptor with 2-pyridyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-galactopyranoside as the donor. In the second one, the key step was the coupling of 2,3,4-tri-O-acetyl-6-O-(2,3,4,6-tetra-O-benzyl-alpha-D-galactopyranosyl)-D-galactopyranosyl trichloroacetimidate donor with 1,2-O-isopropylidene-sn-glycerol. Even though the number of steps was the same in both pathways, the first one afforded a better overall yield (12.4%) than the second one (6.5%). This eight-step synthesis allowed the preparation of the expected glycolipid, which was used as substrate for recombinant GPLRP2 galactolipase using the monomolecular film technique.     

Synthesis of a spacer-containing repeating unit of the capsular polysaccharide of Streptococcus pneumoniae type 23F.

The synthesis is reported of 3-aminopropyl 4-O-(4-O-beta-D-glucopyranosyl-2-O-alpha-L-rhamnopyranosyl-beta-D- galactopyranosyl)-beta-L-rhamnopyranoside 3'-(glycer-2-yl sodium phosphate) (25 beta), which represents the repeating unit of the capsular polysaccharide of Streptococcus pneumoniae type 23F (American type 23) [(----4)-beta-D-Glcp-(1----4)-[Glycerol-(2-P----3)] [alpha-L- Rhap-(1----2)]-beta-D-Galp-(1----4)-beta-L-Rhap-(1----)n). 2,4,6-Tri-O-acetyl-3-O-allyl-alpha-D-galactopyranosyl trichloroacetimidate (5) was coupled with ethyl 2,3-di-O-benzyl-1-thio-alpha-L-rhamnopyranoside (6). Deacetylation of the resulting disaccharide derivative, followed by benzylidenation, and condensation with 2,3,4-trio-O-acetyl-alpha-L-rhamnopyranosyl trichloroacetimidate (10) afforded ethyl 4-O-[3-O-allyl-4,6-O-benzylidene-2-O-(2,3,4-trio-O-acetyl- alpha-L-rhamnopyranosyl)-beta-D-galactopyranosyl]-2,3-di-O-benzyl-1-thio - alpha-L-rhamnopyranoside (11). Deacetylation of 11, followed by benzylation, selective benzylidene ring-opening, and coupling with 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl trichloroacetimidate (15) gave ethyl 4-O-[3-O-allyl-6-O-benzyl-4-O-(2,3,4,6- tetra-O-acetyl-beta-D-glucopyranosyl)-2-O-(2,3,4-tri-O-benzyl-alpha-L- rhamnopyranosyl)-beta-D-galactopyranosyl]-2,3-di-O-benzyl-1-thio-alpha-L - rhamnopyranoside (16). Deacetylation of 16 followed by benzylation, deallylation, and acetylation yielded ethyl 4-O-[3-O-acetyl-6-O-benzyl-4-O-(2,3,4,6-tetra-O-benzyl-beta-D-glucopy ran osyl)- 2-O-(2,3,4-tri-O-benzyl-alpha-L-rhamnopyranosyl)-beta-D-galactopyranosyl ]-2,3- di-O-benzyl-1-thio-alpha-L-rhamnopyranoside (20). The glycosyl bromide derived from 20, when coupled with 3-benzyloxycarbonylamino-1-propanol, gave the beta-glycoside (21 beta) as the major product. Deacetylation of 21 beta followed by condensation with 1,3-di-O-benzylglycerol 2-(triethylammonium phosphonate) (27), oxidation, and deprotection, afforded 25 beta.     

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.