Efficient synthesis of 5-thio-D-arabinopyranose and 5-thio-D-xylopyranose from the corresponding D-pentono-1,4-lactones

5-Thio-D-arabinopyranose (5) and 5-thio-D-xylopyranose (10) were synthesized from the corresponding D-pentono-1,4-lactones. After regioselective bromination at C-5, transformation into 5-S-acetyl-5-thio derivatives, reduction into lactols and deprotection afforded the title compounds in 49 and 42% overall yield, respectively.     

Two approaches to the synthesis of 3-beta-D-glucopyranosyl-D-glucitol

Glycosidation of 1,2:5,6-di-O-isopropylidene-D-glucose with tetra-O-acetyl-glucosyl bromide in 1:1 benzene-MeNO2 afforded approximately equal amounts of the 3-O-beta-D-glycoside and the rearranged 6-O-beta-D-glycoside, while in MeCN only the latter was formed. When tetra-O-acetyl-beta-thiophenylglucoside was used as donor in CH2Cl2 in the presence of NIS/TfOH as activator, the 6-O-beta-D-glycoside and a 3-O-orthoester were formed in a 1:2 ratio at -20 degrees C, while at 20 degrees C only the former could be isolated. Glycosidation of 1-O-benzoyl-2,4-O-benzylidene-5,6-O-isopropylidene-d-glucitol with tetra-O-acetyl-glucosyl bromide in MeCN in the presence of Hg(CN)2 afforded the corresponding 3-O-alpha- and 3-O-beta-glycopyranoside in a 1:4 ratio in MeCN and 1:5 in 1:1 benzene-MeNO2, respectively. When Hg(CN)2/HgBr2 was used as promoter, the corresponding orthoester was also formed. When tetra-O-acetyl-beta-thiophenylglucoside was used as donor, the 3-O-beta-anomer and the orthoester were obtained predominantly in a 3:2 ratio together with traces of the 3-O-alpha-glycoside. Both beta-glycosides could be smoothly converted into 3-beta-D-glucopyranosyl-D-glucitol.     

Base catalysed isomerisation of aldoses of the arabino and lyxo series in the presence of aluminate

Base-catalysed isomerisation of aldoses of the arabino and lyxo series in aluminate solution has been investigated. L-Arabinose and D-galactose give L-erythro-2-pentulose (L-ribulose) and D-lyxo-2-hexulose (D-tagatose), respectively, in good yields, whereas lower reactivity is observed for 6-deoxy-D-galactose (D-fucose). From D-lyxose, D-mannose and 6-deoxy-L-mannose (L-rhamnose) are obtained mixtures of ketoses and C-2 epimeric aldoses. Small amounts of the 3-epimers of the ketoses were also formed. 6-Deoxy-L-arabino-2-hexulose (6-deoxy-L-fructose) and 6-deoxy-L-glucose (L-quinovose) were formed in low yields from 6-deoxy-L-mannose and isolated as their O-isopropylidene derivatives. Explanations of the differences in reactivity and course of the reaction have been suggested on the basis of steric effects.     

Facile synthesis of benzyl beta-D-galactofuranoside. A convenient intermediate for the synthesis of D-galactofuranose-containing molecules

Benzyl beta-D-galactofuranoside was efficiently obtained from 1,2,3,5,6-penta-O-benzoyl-alpha,beta-D-galactofuranose, via benzyl 2,3,5,6-tetra-O-benzoyl-beta-D-galactofuranoside. Conditions for the O-debenzylation were investigated in order to evaluate the synthetic application of the benzyl group as an anomeric protector of a galactofuranose moiety in synthetic strategies involving galactofuranose.     

Chemical synthesis of cholesteryl beta-D-galactofuranoside and -pyranoside

Two isomeric cholesteryl galactosides, cholesteryl beta-D-galactofuranoside and -pyranoside, have been synthesized by the Koenigs-Knorr reaction. Glycosylation of cholesterol with 2,3,5,6-tetra-O-benzoyl-D-galactofuranosyl bromide, followed by Zemplén saponification with sodium methoxide, gave cholesteryl beta-D-galactofuranoside. By using 2,3,4,6-tetra-O-acetyl-D-galactopyranosyl bromide as the glycosyl donor, followed by alkaline hydrolysis, cholesteryl beta-D-galactopyranoside was obtained. The title compounds were characterized by their IR spectra and by their (1)H and (13)C NMR spectra. Structure considerations of the two cholesteryl galactosides correlated with data in the literature, thus confirming that cholesteryl beta-D-galactopyranoside is an antigenic lipid of Lyme disease agent, Borrelia burgdorferi.     

Melting behaviour of D-sucrose, D-glucose and D-fructose

The melting behaviour of d-sucrose, d-glucose and d-fructose was studied. The melting peaks were determined with DSC and the start of decomposition was studied with TG at different rates of heating. In addition, melting points were determined with a melting point apparatus. The samples were identified as d-sucrose, alpha-d-glucopyranose and beta-d-fructopyranose by powder diffraction measurements. There were differences in melting between the different samples of the same sugar and the rate of heating had a remarkable effect on the melting behaviour. For example, T(o), DeltaH(f) and T(i) (initial temperature of decomposition) at a 1 degrees Cmin(-1) rate of heating were 184.5 degrees C, 126.6Jg(-1) and 171.3 degrees C for d-sucrose, 146.5 degrees C, 185.4Jg(-1) and 152.0 degrees C for d-glucose and 112.7 degrees C, 154.1Jg(-1) and 113.9 degrees C for d-fructose. The same parameters at 10 degrees Cmin(-1) rate of heating were 188.9 degrees C, 134.4Jg(-1) and 189.2 degrees C for d-sucrose, 155.2 degrees C, 194.3Jg(-1) and 170.3 degrees C for d-glucose and 125.7 degrees C, 176.7Jg(-1) and 136.8 degrees C d-fructose. At slow rates of heating, there were substantial differences between the different samples of the same sugar. The melting point determination is a sensitive method for the characterization of crystal quality but it cannot be used alone for the identification of sugar samples in all cases. Therefore, the melting point method should be validated for different sugars.     

Synthesis and intramolecular reactions of Tyr-Gly and Tyr-Gly-Gly related 6-O-glucopyranose esters

6-O-(L-Tyrosylglycyl)- and 6-O-(L-tyrosylglycylglycyl)-D-glucopyranose were synthesized by condensation of the pentachlorophenyl esters of the respective di- and tripeptide with fully unprotected D-glucose. The intramolecular reactivity of the sugar conjugates was studied in pyridine-acetic acid and in dry methanol, at various temperatures and for various incubation times. The composition of the incubation mixtures was monitored by a reversed-phase HPLC method that permits simultaneous analysis of the disappearance of the starting material and the appearance of rearrangement and degradation products. To determine the influence of esterification of the peptide carboxy group on its amino group reactivity, parallel experiments were done in which free peptides were, under identical reaction conditions, incubated with D-glucose (molar ratios 1:1 and 1:5). Depending on the starting compound, different types of Amadori products (cyclic and bicyclic form), methyl ester of peptides, and Tyr-Gly-diketopiperazine were obtained.     

Enzymatic synthesis of D-glucosaminic acid from D-glucosamine

D-glucosaminic acid (2-amino-2-deoxy-D-gluconic acid), a component of bacterial lipopolysaccharides and a chiral synthon, is easily prepared on a multigram scale by air oxidation of D-glucosamine (2-amino-2-deoxy-D-glucose) catalysed by glucose oxidase.     

NMR studies of molybdate complexes of D-erythro-L-manno-octose and D-erythro-L-gluco-octose and their alditols

Different amounts and various types of bis-dinuclear tetradentate molybdate complexes of D-erythro-L-manno-octose, D-erythro-L-gluco-octose, D-erythro-L-manno-octitol and D-erythro-L-gluco-octitol were characterized by 1H and 13C NMR spectroscopy in aqueous solutions. Detailed analysis of 1H-(1)H coupling constants and NOEs, together with chemical shifts, allowed characterization of the different isomers of these complexes.     

Crystal structures of D-tagatose 3-epimerase from Pseudomonas cichorii and its complexes with D-tagatose and D-fructose

Pseudomonas cichoriiid-tagatose 3-epimerase (P. cichoriid-TE) can efficiently catalyze the epimerization of not only d-tagatose to d-sorbose, but also d-fructose to d-psicose, and is used for the production of d-psicose from d-fructose. The crystal structures of P. cichoriid-TE alone and in complexes with d-tagatose and d-fructose were determined at resolutions of 1.79, 2.28, and 2.06 Å, respectively. A subunit of P. cichoriid-TE adopts a (β/α)₈ barrel structure, and a metal ion (Mn²⁺) found in the active site is coordinated by Glu152, Asp185, His211, and Glu246 at the end of the β-barrel. P. cichoriid-TE forms a stable dimer to give a favorable accessible surface for substrate binding on the front side of the dimer. The simulated omit map indicates that O2 and O3 of d-tagatose and/or d-fructose coordinate Mn²⁺, and that C3-O3 is located between carboxyl groups of Glu152 and Glu246, supporting the previously proposed mechanism of deprotonation/protonation at C3 by two Glu residues. Although the electron density is poor at the 4-, 5-, and 6-positions of the substrates, substrate-enzyme interactions can be deduced from the significant electron density at O6. The O6 possibly interacts with Cys66 via hydrogen bonding, whereas O4 and O5 in d-tagatose and O4 in d-fructose do not undergo hydrogen bonding to the enzyme and are in a hydrophobic environment created by Phe7, Trp15, Trp113, and Phe248. Due to the lack of specific interactions between the enzyme and its substrates at the 4- and 5-positions, P. cichoriid-TE loosely recognizes substrates in this region, allowing it to efficiently catalyze the epimerization of d-tagatose and d-fructose (C4 epimer of d-tagatose) as well. Furthermore, a C3-O3 proton-exchange mechanism for P. cichoriid-TE is suggested by X-ray structural analysis, providing a clear explanation for the regulation of the ionization state of Glu152 and Glu246.     

An improved synthesis of 5-thio-D-ribose from D-ribono-1,4-lactone

5-Thio-D-ribopyranose was synthesized from D-ribono-1,4-lactone (1) by two approaches: (i) 5-bromo-5-deoxy-D-ribono-1,4-lactone (2) was successively transformed into 5-bromo-5-deoxy, 5-S-acetyl-5-thio or 5-thiocyanato-D-ribofuranose derivatives; appropriate treatment then lead to 5-thio-D-ribopyranose (7) in 46-48% overall yield and; (ii) 2 was transformed into the 5-S-acetyl-5-thio-D-ribono-1,4-lactone derivative (11). Reduction and deprotection of 11 afforded 5-thio-D-ribopyranose (7) in 57% overall yield.     

An original chemoenzymatic route for the synthesis of beta-D-galactofuranosides using an alpha-L-arabinofuranosidase

DGalactofuranose is a widespread component of cell wall polysaccharides in bacteria, protozoa and fungi, but is totally absent in mammals. Importantly, galactofuranose is a key constituent of major cell envelope polysaccharides in pathogenic mycobacteria. In this respect, galactofuranose-based glycoconjugates are interesting target molecules for drug design. O-Glycosidases and notably beta-D-galactofuranosidases could be useful tools for the chemoenzymatic synthesis of galactofuranosides, but to date no studies of this type have been reported. Here we report the use of a GH 51 alpha-l-arabinofuranosidase for the synthesis of beta-D-galactofuranosides. We have demonstrated that this enzyme can catalyse both the autocondensation of p-nitrophenyl-beta-D-galactofuranoside and the transgalactofuranosylation of benzyl alpha-D-xylopyranoside, forming p-nitrophenyl beta-D-galactofuranosyl-(1-->2)-beta-D-galactofuranoside and benzyl beta-D-galactofuranosyl-(1-->2)-alpha-D-xylopyranoside, respectively. Both reactions were very regiospecific and the reaction involving benzyl alpha-D-xylopyranoside afforded very high yields (74.8%) of the major product. To our knowledge, this demonstration of chemoenzymatic synthesis of galactofuranosides constitutes the very first use of an O-glycosidase for the synthesis of galactofuranosides.     

A stereocontrolled synthesis of 3-acetamido-1,3,5-trideoxy- and 1,3,5,6-tetradeoxy-1,5-imino-d-glucitol

3-Acetamido-5-amino-3,5,6-trideoxy-d-glucono-1,5-lactam and 3-acetamido-5-amino-3,5-dideoxy-d-glucono-1,5-lactam were synthesized from corresponding 3-acetamido-3-deoxy-β-d-glucopyranosides in 63% and 35% overall yield, respectively. Acetylation followed by reduction led to the title 3-acetamido-3-deoxy derivatives of both deoxynojirimycin and 1,6-dideoxynojirimycin. The procedure developed is useful for a multi-gram scale.     

Synthesis of 7-O-galloyl-D-sedoheptulose

A facile synthetic approach to 7-O-galloyl-D-sedoheptulose (1), a natural product with notable immunosuppressant activity, was developed. The starting material, 2,7-anhydro-d-sedoheptulose (2), was converted in three steps into 1,3,4,5-tetra-O-benzyl-d-sedoheptulose (5), a key intermediate that allows specific functionalization at C-7 of the sedoheptulpyranose. After regioselective esterification of 5 with 3,4,5-tri-O-benzyl galloyl acid, followed by catalytic debenzylation (Pd-C), 1 was obtained in an overall yield of 60%. The spectroscopic data and TLC behavior of 1 were found to be identical to that of the natural product.     

Anomeric selectivity in the synthesis of galloyl esters of d-glucose

The anomeric selectivity of the ester formation between d-glucopyranose and gallic acid was investigated under a variety of conditions. A new protocol was established that allows performing the reaction under conditions where mutarotation is very slow. Highly α- or β-selective transformations are possible when starting with α- or β-d-glucopyranose, respectively. Due to the kinetic anomeric effect, a high α-selectivity is more difficult to achieve than a high β-selectivity. The new methodology presented in this article was compared with established procedures for the synthesis of gallotannins. In addition to the advantages of a high yield and an easy purification protocol, the new procedure uniquely allowed for a highly selective synthesis of α-products.     

The inhibitory effects of flavonoids and antiestrogens on the Glut1 glucose transporter in human erythrocytes

Flavonoids and isoflavonoids are potent inhibitors of glucose efflux in human erythrocytes. Net changes of sugars inside the cells were measured by right angle light scattering. The inhibitory potency of hydroxylated flavonoids depends on the pH of the medium. The apparent affinity is maximal at low pH where the molecule is in the undissociated form. The following K(i)-values at pH 6.5 in microM have been obtained: phloretin 0.37+/-0.03, myricetin 0.76+/-0.42, quercetin 0.93+/-0.28, kaempferol 1.33+/-0.17, isoliquiritigenin 1.96, genistein 3.92+/-0.62, naringenin 8.88+/-1.88, 7-hydroxyflavone 17.58+/-3.15 and daidzein 18.62+/-2.85. Flavonoids carrying hydroxyl groups are weak acids and are deprotonated at high pH-values. From spectral changes pK-values between 6.80 (naringenin) and 7.73 (myricetin) have been calculated. No such pK-value could be obtained from quercetin which was rather unstable at alkaline pH. Flavone itself without a hydroxyl group does not demonstrate any absorbance changes at different pH-values and no significant change in inhibition of glucose transport with pH (K(i)-value around 35 microM). In this respect it is similar to the antiestrogens diethylstilbestrol, tamoxifen and cyclofenil with K(i)-values for glucose efflux inhibition of 2.61+/-0.30, 6.75+/-2.03 and 3.97+/-0.54 microM. Except for phloretin, the flavonoids investigated have planar structures. The inhibitory activity in glucose efflux of planar flavonoids increases exponentially with the number of hydroxyl groups in the molecule.     

The structure of the antigenic polysaccharide produced by Eubactrium saburreum T15

The antigenic polysaccharide was obtained from the cell wall of Eubacterium saburreum strain T15 by trypsin digestion followed by gel permeation and ion-exchange chromatography. Its structure was determined using acid hydrolysis, methylation analysis, and 1D and 2D NMR spectroscopy. It contained L-threo-pent-2-ulose (Xul), D-fucose (Fuc), and D-glycero-D-galacto-heptose (Hep) in 2:3:3 ratio. Methylation analysis indicated an octasaccharide repeating-unit containing five branches. The 1H and 13C signals in NMR spectra of the sugar residues were assigned by COSY, HOHAHA, and HMQC 2D experiments, and the sequence of sugar residues in the repeating unit was determined by NOESY and HMBC experiments. The polysaccharide also contains two O-acetyl groups in the repeating unit, located on the Hep residue. The repeating structure can be written as: [see text for equation]. This is a novel structure in bacterial cell-wall polysaccharides from Gram-positive bacteria.     

Conservation of key members in the course of the evolution of the insulin signaling pathway

Our understanding of the evolution of the insulin signaling pathway (ISP) is still incomplete. One intriguing unanswered question is the explanation of the emergence of the glucostatic role of insulin in mammals. To find out whether this is due to the development of new sets of signaling transduction elements in these organisms, or to the establishment of new interactions between pre-existing proteins, we rebuilt putative orthologous ISPs in 17 eukaryotic organisms. Then, we computed the conservation of orthologous ISPs at different levels, from sequence similarity of orthologous proteins to co-evolution of interacting domains. We found that the emergence of glucostatic role in mammals can neither be explained by the development of new sets of signaling elements, nor by the establishment of new interactions between pre-existing proteins. The comparison of orthologous IRS molecules indicates that only in mammals have they acquired their complete functionality as efficient recruiters of effector sub-pathways.     

Synthesis of D- and L-myo-inositol 2,4,5-trisphosphate and trisphosphorothioate: structural analogues of D-myo-inositol 1,4,5-trisphosphate

The preparation of D- and L-myo-inositol 2,4,5-trisphosphate is described, together with the phosphorothioate counterparts. The known chiral diols D- and L-1,4-di-O-benzyl-5,6-bis-O-p-methoxybenzyl-myo-inositol were regioselectively protected at the 3-position using a benzyl group via a 2,3-O-stannylene acetal. Removal of the p-methoxybenzyl groups of each enantiomer gave D- and L-1,3,6-tri-O-benzyl-myo-inositol. Phosphitylation with bis(benzyloxy)diisoproplyaminophosphine and 1H-tetrazole gave the trisphosphite intermediate for each enantiomer. Oxidation with 3-chloroperoxybenzoic acid gave the fully protected D- and L-myo-inositol 2,4,5-trisphosphates. Sulphoxidation of the D- and L-2,4,5-trisphosphite intermediates gave the fully protected D- and L-myo-inositol 2,4,5-trisphosphorothioate compounds. The fully protected trisphosphates were deblocked using hydrogenolysis and the phosphorothioates were deprotected using sodium in liquid ammonia. The individual compounds were then purified using ion exchange chromatography to afford pure D- and L-myo-inositol 2,4,5-trisphosphates together with the corresponding phosphorothioates.     

Towards alpha- or beta-D-C-glycosyl compounds by tin-catalyzed addition of glycosyl radicals to acrylonitrile and vinylphosphonate,and flexible reduction of tetra-O-acetyl-alpha-D-glucopyranosyl bromide with cyanoborohydride

Photo-induced radical addition of acetylated alpha-D-glucopyranosyl bromide (1). to acrylonitrile or diethyl vinylphosphonate, in the presence of catalytic amounts of tri-n-butyltin chloride and sodium (or tetra-n-butylammonium) cyanoborohydride in excess, allowed efficient preparations of alpha-configurated nonononitrile and 2-(alpha-D-glucopyranosyl)-ethylphosphonate (79, 70% yields, respectively). These conditions led to 2-(alpha-D-manno-, and galactopyranosyl)-ethylphosphonates in 68 and 76% yields. Similarly, radical addition of acetylated 1-bromo-beta-D-glucopyranosyl chloride (2). to acrylonitrile or diethyl vinylphosphonate afforded mainly intermediate chlorides which, upon radical reduction with excess tri-n-butyltin hydride, afforded the corresponding beta anomers (40 and 38%, respectively) by sequential C-C and C-H bond formation. Stereocontrol relies on the alpha-stereoselective quenching of D-glycopyranos-1-yl radicals. We found also that UV light irradiation of 1 with excess NaBH(3)CN in tert-butanol afforded either 1,3,4,6-tetra-O-acetyl-2-deoxy-alpha-D-arabino-hexopyranose (65% after crystallization) or, when 10% mol thiophenol was added, 2,3,4,6-tetra-O-acetyl-1,5-anhydro-D-glucitol (79%). These are simple, tin-free, and easily controlled conditions, which compare well with known preparations of these reduced sugars.