Analysis by the reductive-cleavage method of linkage positions in a polysaccharide containing 4-linked D-glucopyranosyluronic residues

The fate of 4-linked D-glucopyranosyluronic residues under reductive-cleavage conditions was investigated by using the Klebsiella aerogenes type 54 strain A3 capsular polysaccharide. Treatment of the fully methylated polysaccharide with triethylsilane and trimethylsilyl trifluoromethanesulfonate in dichloromethane, followed by in situ acetylation, yielded 1,5-anhydro-2,3,4,6-tetra-O-methyl-D-glucitol, 3,4-di-O-acetyl-1,5-anhydro-2,6-di-O-methyl-D-glucitol, and 3-O-acetyl-1,5-anhydro-2,4-di-O-methyl-L-fucitol, as expected, but the expected product of reductive cleavage of the 4-linked D-glucopyranosyluronic residue, namely, methyl 3-O-acetyl-2,6-anhydro-4,5-di-O-methyl-L-gulonate, was not observed. Instead, methyl 2-O-acetyl-3,6-anhydro-4,5-di-O-methyl-L-gulonate (6) was identified as the sole product of reductive cleavage of the 4-linked D-glucopyranosyluronic residue. That compound 6 arose as a result of rearrangement during reductive cleavage rather than by reductive cleavage of a 5-linked D-glucofuranosyluronic residue, was established by reductive cleavage of the fully methylated polysaccharide following reduction of its ester groups with either lithium aluminum hydride or lithium aluminum deuteride. The products of the latter reductive cleavage were the same as before, except for the absence of 6 and the presence of 4,6-di-O-acetyl-1,5-anhydro-2,3-di-O-methyl-D-glucitol, or its 6,6-dideuterio isomer. Although the reductive-cleavage technique is suitable for the direct analysis of polysaccharides containing 4-linked D-glucopyranosyluronic residues, it does not establish whether the uronic residue is a 4-linked pyranoside or a 5-linked furanoside. The expected product is, however, derived from the 4-linked D-glucopyranosyluronic residue after sequential methylation, reduction of its ester group and reductive cleavage.     

Analysis of Macrocystis pyrifera and Pseudomonas aeruginosa alginic acids by the reductive-cleavage method.

Permethylated alginic acids comprised of 4-linked D-mannopyranosyluronic acid and 4-linked L-gulopyranosyluronic acid residues undergo reductive cleavage to yield, after acetylation, methyl 3-O-acetyl-2,6-anhydro-4,5-di-O-methyl-D-mannonate (2b) and methyl 3-O-acetyl-2,6-anhydro-4,5-di-O-methyl-D-gluconate (3b) as major products. Small amounts (ca. 13%) of ring-contracted products, namely methyl 2-O-acetyl-3,6-anhydro-4,5-di-O-methyl-D-mannonate (9) and methyl 2-O-acetyl-3,6-anhydro-4,5-di-O-methyl-D-gluconate (10), were also observed in these experiments. These results are in marked contrast to previous results on the reductive cleavage of 4-linked D-glucopyranosyluronic acid residues, wherein the ring-contracted product was formed exclusively. Formation of the ring-contracted products could be completely eliminated by reduction (LiAlH4) of ester groups in the permethylated alginic acid prior to reductive cleavage. In the latter experiments, 4,6-di-O-acetyl-1,5-anhydro-2,3-di-O-methyl-D-mannitol (5b) and 4,6-di-O-acetyl-1,5-anhydro-2,3-di-O-methyl-L-gulitol (6b) were the sole products of reductive cleavage of the 4-linked ManA and 4-linked GulA residues, respectively. However, in the previous experiments it was noted that low yields of permethylated alginic acids were obtained and that extensive depolymerization occurred under methylation conditions. Depolymerization could be avoided and higher yields of permethylated polysaccharides could be obtained, by reduction of the carboxyl groups of the alginic acids prior to methylation. Reductive cleavage of the latter polysaccharides yielded the products expected from 4-linked D-mannopyranosyl and 4-linked L-gulopyranosyl residues, namely 4-O-acetyl-1,5-anhydro-2,3,6-tri-O-methyl-D-mannitol (13b) and 4-O-acetyl-1,5-anhydro-2,3,6-tri-O-methyl-L-gulitol (14b), respectively. Using the latter analytical strategy, it was established that the Macrocystis pyrifera alginate was comprised of 60% 4-linked ManA and 40% 4-linked GulA residues, whereas the Pseudomonas aeruginosa alginate was comprised of 80% 4-linked ManA and 20% 4-linked GulA residues.     

Analysis of linkage positions in a polysaccharide containing nonreducing, terminal alpha-D-glucopyranosyluronic groups by the reductive-cleavage method

The fate of terminal (nonreducing) alpha-D-glucopyranosyluronic groups under reductive cleavage conditions was investigated by using the Klebsiella K2 (strain NCTC-418) capsular polysaccharide. Treatment of the fully methylated polysaccharide (1) with triethylsilane and a mixture of trimethylsilyl methanesulfonate (Me3SiOSO2CH3) and boron trifluoride etherate (BF3.Et2O) as the catalyst, resulted in complete cleavage of all glycosidic linkages to yield the expected products, namely 3-O-acetyl-1,5-anhydro-2,4,6-tri-O-methyl-D-glucitol (2), 3,4-di-O-acetyl-1,5-anhydro-2,6-di-O-methyl-D-mannitol (3), 4-O-acetyl-1,5-anhydro-2,3,6-tri-O-methyl-D-glucitol (4), and methyl 2,6-anhydro-3,4,5-tri-O-methyl-L-gulonate. Treatment of 1 with trimethylsilyl trifluoromethanesulfonate (Me3SiOSO2CF3) as the catalyst resulted in incomplete cleavage of the glycosidic linkage of the methylated D-glucopyranosyluronic group, to yield 4-O-acetyl-1,5-anhydro-2,6-di-O-methyl- 3-O-(methyl2,3,4-tri-O-methyl-alpha-D-glucopyranosyluronate )-D-mannitol (9). Reductive cleavage of 1 in the presence of BF3.Et2O resulted in incomplete cleavage of all glycosidic linkages and gave rise to all four dimers (including 9) that could be formed from a tetrasaccharide repeating unit. The proposed structures of these dimers are based upon their composition, as established by chemical ionization mass spectrometry and by the reported structure of the polysaccharide. A small proportion of 1,5-anhydro-2,4,6-tri-O-methyl-3-O-(methyl 2,3,4-tri-O-methyl-alpha-D-glucopyranosyluronate)-D-mannitol (12) was also detected in the products of the BF3.Et2O-catalyzed reductive cleavage. The presence of 12 is chemical evidence for the phase of the tetrasaccharide repeating unit in the polysaccharide. The reductive cleavage of 1 was also accomplished after reduction of its ester groups with lithium aluminum hydride. Complete cleavage of all glycosidic linkages was observed when either Me3SiOSO2CF3 or Me3SiOSO2CH3-BF3.Et2O was used to catalyze reductive cleavage, and anhydroalditols 2, 3, 4, and 6-O-acetyl-1,5-anhydro-2,3,4-tri-O-methyl-D-glucitol were produced, as expected.     

Analysis of pyruvic acid acetal containing polysaccharides by methanolysis and reductive cleavage methods.

The mass spectra of permethylated methyl 4,6-O-(1-carbomethoxyethylidene)-D-hexopyranoside and 1,5-anhydro-D-hexitol of glucose, galactose, and mannose and permethylated methyl 5,6-O-(1-carbomethoxyethylidene)-D-galactofuranoside and 1,4-anhydro-D-galactitol have been determined. The stability of each compound toward methanolysis and reductive cleavage is discussed. These techniques permit the identification of the acetalic linkages of pyruvic acid present in polysaccharides.     

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.     

Structural analysis of colanic acid from Escherichia coli by using methylation and base-catalysed fragmentation. Comparison with polysaccharides from other bacterial sources

Essentially the same methanolysis products were obtained after methylation of the slime and capsular polysaccharides from Escherichia coli K12 (S53 and S53C sub-strains) and the slime polysaccharides from E. coli K12 (S61), Aerobacter cloacae N.C.T.C. 5290 and Salmonella typhimurium SL1543. These were the methyl glycosides of 2-O-methyl-l-fucose, 2,3-di-O-methyl-l-fucose, 2,3-di-O-methyl-d-glucuronic acid methyl ester, 2,4,6-tri-O-methyl-d-glucose, 2,4,6-tri-O-methyl-d-galactose and the pyruvic acid ketal, 4,6-O-(1'-methoxycarbonylethylidene)-2,3-O-methyl-d-galactose. All were identified as crystalline derivatives from an E. coli polysaccharide. The structure of the ketal was proved by proton-magnetic-resonance and mass spectrometry, and by cleavage to pyruvic acid and 2,3-di-O-methyl-d-galactose. All these polysaccharides are therefore regarded as variants on the same fundamental structure for which the name colanic acid is adopted. Although containing the same sugar residues, quite different methanolysis products were obtained after methylation of the extracellular polysaccharide from Klebsiella aerogenes (1.2 strain). The hydroxypropyl ester of E. coli polysaccharide, when treated with base under anhydrous conditions, underwent beta-elimination at the uronate residues with release of a 4,6-O-(1'-alkoxycarbonylethylidene)-d-galactose. Together with the identification of 3-O-(d-glucopyranosyluronic acid)-d-galactose as a partial hydrolysis product, this establishes the nature of most, if not all, of the side chains as O-[4,6-O-(1'-carboxyethylidene)-d-galactopyranosyl]-(1-->4)-O-(d-glucopyranosyluronic acid)-(1-->3)-d-galactopyranosyl...     

Synthesis and mass spectra of 4-O-acetyl-1,5-anhydro-2,3,6-tri-O -(methoxycarbonylmethyl)-D-glucitol and the positional isomers of 4- O-acetyl-1,5-anhydro-di-O-(methoxycarbonylmethyl)-O-methyl-D-glucitol and 4-O-acetyl-1,5-anhydro-O-(methoxycarbonylmethyl)-di-O-methyl-D-glucitol .

Reductive cleavage of fully methylated, partially O-carboxymethylated cellulose had previously been shown to produce 4-O-acetyl-1,5-anhydro-2,3,6-tri-O-methyl-, -2-O-(methoxycarbonylmethyl)-3,6-di-O-methyl-, -3-O-(methoxycarbonylmethyl)-2,6-di-O-methyl-, -6-O-(methoxycarbonylmethyl)-2,3-di-O-methyl-, -2,3-di-O-(methoxycarbonylmethyl)-6-O-methyl-, -2,6-di-O-(methoxycarbonylmethyl)-3-O-methyl-, -3,6-di-O-(methoxycarbonylmethyl)-2-O-methyl-, and -2,3,6-tri-O-(methoxycarbonylmethyl)-D-glucitol. Described herein is the independent synthesis of these derivatives, except for the first, which had been reported. In addition, their 1H-n.m.r. spectra, chemical-ionization (NH3) mass spectra, and electronionization mass spectra are tabulated.     

Isolation and analysis by the reductive-cleavage method of linkage positions and ring forms in the Mycobacterium smegmatis cell-wall arabinogalactan

The Mycobacterium smegmatis arabinogalactan polysaccharide has been isolated from the cell wall by saponification and extraction to remove lipids and subsequent solubilization by treatment with lysozyme. Analysis for neutral sugars demonstrated the presence of D-arabinose and D-galactose in a ratio of 3:1, respectively. Reductive cleavage of the fully methylated polysaccharide in the presence of triethylsilane and trimethylsilyl trifluoromethanesulfonate and subsequent acetylation in situ gave six partially methylated 1,4-anhydroalditol acetates as the major products and three partially methylated 1,5-anhydroalditol acetates as minor products. Partially methylated 1,5-anhydroalditol acetates were not formed when reductive cleavage was accomplished with triethylsilane and a mixture of trimethylsilyl methanesulfonate and boron trifluoride etherate as the catalyst, demonstrating that the polysaccharide is exclusively comprised of furanosyl residues. The partially methylated anhydroalditols so produced were identified by comparison to authentic standards. Their identifies are consistent with the presence in the M. smegmatis arabinogalactan of an octasaccharide repeating unit comprised of a nonreducing terminal D-arabinofuranosyl group, a 2-O-linked D-arabinofuranosyl residue, three 5-O-linked D-arabinofuranosyl residues, a 3,5-di-O-linked D-arabinofuranosyl residue, a 5-O-linked D-galactofuranosyl residue, and a 6-O-linked D-galactofuranosyl residue.     

Model studies on the analysis of pyruvic acid acetal-containing polysaccharides by the reductive-cleavage method

The 4,6-O-(1-methoxycarbonylethylidene), -(hydroxyisopropylidene), and -(methoxyisopropylidene) acetals of methyl 2,3-di-O-methyl-alpha-D-glucopyranoside were subjected to reductive cleavage in the presence of triethylsilane and trimethylsilyl methanesulfonate-boron trifluoride etherate (Me3SiOMs-BF3.Et2O), BF3.Et2O, or trimethylsilyl trifluoromethanesulfonate (Me3SiOSO2CF3) and the mole fractions of products were determined as a function of reaction time. The 4,6-(1-methoxycarbonylethylidene) acetal was quite stable to reductive-cleavage conditions but isomerization of the initial R,S mixture of diastereomers to the more-stable S diastereoisomer was noted. In addition, a slow, regiospecific, reductive ring-opening of the acetal was observed to give 6-O-[1-(methoxycarbonyl)ethyl] derivatives. The 4,6-(hydroxyisopropylidene) acetal was very unstable under reductive-cleavage conditions. Both Me3SiOMs-BF3.Et2O and Me3SiOSO2CF3 catalyzed complete removal of the group, via the intermediate 6-[1-(hydroxymethyl)ethyl] ether, but BF3.Et2O gave a mixture of products. The 4,6-(methoxyisopropylidene) acetal was also very labile under reductive-cleavage conditions; Me3SiOMs-BF3.Et2O catalyzed complete removal of the acetal, via the intermediate 6-[1-(methoxymethyl)ethyl]ether, but the intermediate ether was quite stable in the presence of either BF3.Et2O or Me3SiOSO2CF3. It is concluded from these studies that polysaccharides bearing 4,6-O-(1-carboxyethylidene) substituents can be analyzed directly by sequential permethylation and reductive cleavage. It is proposed that the identity of the substituted monomer and the positions of substitution of the acetal can be determined by sequential permethylation, ester reduction, and reductive cleavage.     

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.     

Synthèses et réductions de sucres fonctionnels mono et diinsaturés

Attempts to prepare 1,2:5,6 and 2,3:5,6 di-unsaturated sugars starting from 3,4,6-tri-O-acetyl-1,5-anhydro-1,2-dideo xy-d-arabino-hex-1-enitol or from ethyl 4,6-di-O-acetyl-1,5-anhydro-2,3-dideoxy-α-d-erythro-hex-2-enopyranoside led to 1,5-anhydro-1,2,6-trideoxy-l-threo-hex-5-enitol and its 3,4-diacetate. Hydrogenation and hydrogenolysis of the unsaturated chloro and fluoro derivatives afforded 1,5-anhydro-1,2,6-trideoxy-d-arabino-hexitol and ethyl 4-O-acetyl-2,3,6-trideoxy-α-d-erythro-hexopyranoside.     

An approach to stereoselective preparation of 3-C-glycosylated d- and l-glucals

An approach to stereoselective synthesis of α- or β-3-C-glycosylated l- or d-1,2-glucals starting from the corresponding α- or β-glycopyranosylethanals is described. The key step of the approach is the stereoselective cycloaddition of chiral vinyl ethers derived from both enantiomers of mandelic acid. The preparation of 1,5-anhydro-4,6-di-O-benzyl-2,3-dideoxy-3-C-[(2,3,4,6-tetra-O-benzyl-β-d-glucopyranosyl)methyl]-l-arabino-hex-1-enitol, 1,5-anhydro-4,6-di-O-benzyl-2,3-dideoxy-3-C-[(2,3,4,6-tetra-O-benzyl-β-d-glucopyranosyl)methyl]-d-arabino-hex-1-enitol, and 1,5-anhydro-4,6-di-O-benzyl-2,3-dideoxy-3-C-[(2,3,4-tri-O-benzyl-α-l-fucopyranosyl)methyl]-d-arabino-hex-1-enitol serves as an example of this approach.     

Pyruvic-acid-containing polysaccharide in the cell wall of Bacillus polymyxa AHU 1385

Three acidic polymer fractions with molecular masses of about 16 kDa, 35 kDa and 70 kDa were isolated from lysozyme digests of N-acetylated cell walls of Bacillus polymyxa AHU 1385 by ion-exchange chromatography and gel chromatography. These fractions, containing mannosamine, glucosamine and pyruvic acid in a molar ratio of about 1:1:1 together with glycopeptide components, were characterized as polysaccharide-linked glycopeptides with one, two and more polysaccharide chains. On the other hand, treatment of the cell walls with glycine/HC1 buffer, pH 2.5, at 100 degrees C for 10 min followed by separation of water-soluble products on ion-exchange chromatography gave three polysaccharide fractions, PS-I-III, which contained different amounts of pyruvic acid (0,0.6 and 0.9 residue/mannosamine residue) along with equimolar amounts of mannosamine and glucosamine. Pyruvate-free polysaccharides similar to PS-I were also obtained from PS-II, PS-III and polysaccharide-linked glycopeptides by treatment with 10 mM HC1 at 100 degrees C for 1 h. Results of analyses of these polysaccharide preparations by 1H-NMR and 13C-NMR measurement and methylation, together with data from characterization of fragments obtained by hydrogen fluoride hydrolysis, lead to the most likely structure, ----3)[4,6-O-(1-carboxyethylidene)]ManNAc(beta 1----4)GlcNac(beta 1----, for the acidic polysaccharide of this strain.     

Boat conformations synthesis, NMR spectroscopy, and molecular dynamics of methyl 4,6-O-benzylidene-3-deoxy-3-phthalimido-alpha-D-altropyranoside derivatives

Addition of the elements of phthalimide to methyl 2,3-anhydro-4,6-O-benzylidene-alpha-D-mannopyranoside (1) under fusion conditions has yielded methyl 4,6-O-benzylidene-3-deoxy-3-phthalimido-alpha-D-altropyranoside (2). The conformation of the pyranose ring of 2 has been shown to be non-chair by 1H NMR spectroscopy, in contrast to the conformations of related derivatives having smaller substituents at C-3. Molecular dynamics simulations of 2 in explicit chloroform-d solvent have indicated four principal conformational possibilities. Of these, the 7C5/1S5 chair/skew boat form 2d has the lowest potential energy, and is largely consistent with the observed vicinal 1H-1H NMR coupling constants.     

Synthesis of 1,5-Anhydro-D-Mannitol Nucleosides with a Purine Base Moeety

Abstract

D-Mannitol nucleosides with a purine base moiety have been conveniently synthesized strating from 1,5-anhydro-4,6-O-benzylidene-D-glucitol. The 3-OH function of 1,5-anhydro-4,6-O-benzylidene-D-glucitol was selectively protected with t-butyldimethylsilyl group and subsequently converted to the corresponding 0-triflate derivative for the introduction of the nucleobase moietes. These nucleoside derivatives were transformed to 1,5-Anhydro-6-O-MMTr-2-(N⁶-benzoyladenin-9-yl)-2-deoxy-3-O-TBDMS-D-mannitol and 1,5-Anhydro-6-O-MMTr-2-(N²-isobutyryl-guanin-9-yl)-2-deoxy-3-O-TBDMS-D-mannitol, useful as the building blocks for oligonucleotide synthesis. Also, the synthesis of the corresponding fully deprotected anhydrohexitol nucleosides were achieved for evaluation of antiviral activity test.     

Evidence that the acidic polysaccharide secreted by Agrobacterium radiobacter (ATCC 53271) has a seventeen glycosyl-residue repeating unit.

The extracellular anionic polysaccharide produced by the bacterium Agrobacterium radiobacter (ATCC 53271) contains D-galactose, D-glucose, and pyruvic acid in the molar ratio 2:15:2. Analysis of the methylated polysaccharide indicated the presence of terminal, non-reducing glucosyl, 3-, 4-, 6-, 2,4-, and 4,6-linked glucosyl residues, 3-linked 4,6-O-[(S)-1-carboxyethylidene]glucosyl residues, and 3-linked galactosyl residues. Partial acid hydrolysis of the methylated polysaccharide, followed by reduction with NaB2H4 and then O-ethylation, gave a mixture of alkylated oligoglycosyl alditols that were separated by reversed-phase h.p.l.c. and analyzed by 1H-n.m.r. spectroscopy, g.l.c.-m.s., and glycosyl-linkage composition analysis. Smith degradation of the polysaccharide gave three diglycosyl alditols that were separated by semi-preparative, high-pH anion-exchange chromatography, and were analyzed by 1H-n.m.r. spectroscopy, g.l.c.-m.s., and glycosyl-linkage composition analysis. The polymer obtained by NaBH4 reduction of the periodate-oxidized polysaccharide was methylated, and the noncyclic acetals were hydrolyzed with aq. 90% formic acid to generate a mixture of partially O-methylated mono- and di-glycosyl alditols. The partially O-methylated oligoglycosyl alditols were O-ethylated. The resulting alkylated oligoglycosyl alditols were separated by reverse-phase h.p.l.c. and then characterized by 1H-n.m.r. spectroscopy, g.l.c.-m.s., and glycosyl-linkage composition analysis. The results from the studies described here provide strong evidence that the acidic polysaccharide secreted by A. radiobacter (ATCC 53271) has a heptadecasaccharide repeating unit.     

Semisynthetic epsilon-isorhodomycins: their synthesis using glycals and their structure-activity relationship

Syntheses and structure-activity relationships of 7-O-(3-amino-2,3,6-trideoxy-a-L-lyxo- (18), -L-arabino- (20) and -L-ribo- hexopyranosyl)-epsilon-isorhodomycins (25) and their 3'-dimethylamino derivatives 22, 23 and 26 are described. Condensation (trimethylsilyl triflate, molecular sieves 4 A, 10:1 dichloromethane-acetone, -15 degrees) of epsilon-isorhodomycinone (epsilon-isoRMN, 6) with 1,5-anhydro-4-O-p-nitrobenzoyl-3-trifluoroacetamido-L-lyxo- (5) -L-arabino- (9) or -L-ribo-hex-l-enitols (10) afforded mainly the 7-O-a-glycosyl-epsilon-isoRMNs 7, 11, and 12. Similar glycosylation of 6 with 1,5-anhydro-3-azido-4-O-p-nitrobenzoyl-2,3,6-trideoxy-L-arabino-hex-1-++ +enitol (15) yielded a-glycoside 16. Removal (M NaOH) of the p-nitrobenzoyl and trifluoroacetyl groups from 7, 11, and 12 gave the 7-O-(3-amino-2,3,6-trideoxy-a-L-hexopyranosyl)-epsilon-isoRMNs 18, 20, and 25. Reductive alkylation (CH2O, NaCNBH3) of these products afforded the 3'-N,N-dimethyl analogues 22, 23, and 26. The cytotoxic effect (IC50) of the semisynthetic epsilon-isorhodomycins was tested in vitro in leukemia cell line L1210.     

A polysaccharide fraction from the red seaweed Champia novae-zealandiae Rhodymeniales,Rhodophyta

The polysaccharide recovered after extraction of Champia novae-zealandiae is a galactan with alternating 3-linked d-galactopyranosyl units sulfated at the 2-position, and 4-linked galactopyranosyl units sulfated at both the 2- and 3-positions that are predominantly of the l- and partly of the d-configuration. Other minor substitution includes 6-O-methyl ether or 4,6-pyruvate acetal on the 3-linked residues. Techniques used in determining the structure include infrared and ¹³C-NMR spectroscopy, and GC-MS analysis of alditol acetate derivatives produced by reductive hydrolysis/acetylation of native, methylated, and/or desulfated samples. These results are of particular interest because 4-linked 2,3-desulfated galactosyl residues have not been encountered as major constituents of red algal polysaccharides.     

Synthesis of modified tetrasaccharide sequences of N-glycoproteins

The tetrasaccharides O-alpha-D-mannopyranosyl-(1----3)-O-[alpha-D- mannopyranosyl-(1----6)]-O-(4-deoxy-beta-D-lyxo-hexopyranosyl)-(1- ---4)-2- acetamido-2-deoxy-alpha, beta-D-glycopyranose (22) and O-alpha-D-mannopyranosyl-(1----3)-O-[alpha-D-mannopyranosyl-(1----6)]-O- beta-D-talopyranosyl-(1----4)-2-acetamido-2-deoxy-alpha, beta-D- glucopyranose (37), closely related to the tetrasaccharide core structure of N-glycoproteins, were synthesized. Starting with 1,6-anhydro-2,3-di-O-isopropylidene-beta-D-mannopyranose, the glycosyl donors 3,6-di-O-acetyl-2-O-benzyl-2,4-dideoxy-alpha-D-lyxo- hexopyranosyl bromide (10) and 3,6-di-O-acetyl-2,4-di-O-benzyl-alpha-D-talopyranosyl bromide (30), were obtained in good yield. Coupling of 10 or 30 with 1,6-anhydro-2-azido-3-O-benzyl-beta-D-glucopyranose to give, respectively, the disaccharides 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-4-O-(3,6-di-O-acetyl-2-O-benzyl-4 -deoxy- beta-D-lyxo-hexopyranosyl)-beta-D-glucopyranose and 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-4-O-(3,6-di-O-acetyl-2,4-di-O-ben zyl- beta-D-talopyranosyl)-beta-D-glucopyranose was achieved with good selectivity by catalysis with silver silicate. Simultaneous glycosylation of OH-3' and OH-6' of the respective disaccharides with 2-O-acetyl-3,4,6-tri-O-benzyl-alpha-D-mannopyranosyl chloride yielded tetrasaccharide derivatives, which were deblocked into the desired tetrasaccharides 22 and 37.     

Synthesis of 2,3- or 1,2-unsaturated derivatives of 2-deoxy-2-trifluoromethylhexopyranoses

The attempted conversion, by treatment with CsF/TBFA in MeCN, of acetylated derivatives of 2-chlorodifluoromethyl-2-deoxyhexopyranoses into their corresponding 2-trifluoromethyl derivatives was always accompanied by an elimination reaction. Thus, representative educts with the D-gluco- and D-manno-configuration gave derivatives of 2,3-dideoxy-2-trifluoromethyl-D-erythro-hex-2-enopyranose and 1,5-anhydro-2-deoxy-2-trifluoromethyl-d-arabino-hex-1-enitol, respectively. X-ray analyses are given for 1,3,4,6-tetra-O-acetyl-2-chlorodifluoromethyl-2-deoxy-alpha-D-mannopyranose and 4,6-di-O-acetyl-2,3-dideoxy-2-trifluoromethyl-alpha-D-erythro-hex-2-enopyranose.