Synthesis of glycosaminoglycan oligosaccharides. Part 5: Synthesis of a putative heparan sulfate tetrasaccharide antigen involved in prion diseases

The synthesis of the putative prion-associated heparan sulfate tetrasaccharide containing two d-glucuronic acid units is reported. Key to the synthesis were the differentiation of the N-acetylated and N-unsubstituted glucosamine units, the high-yielding glucosylation at O-4 of an N-acetyl-d-glucosamine derivative and the α-selective glycosylation of the 4′-OH group of a β-d-GlcpA-(1→4)-d-GlcpNAc disaccharide building block with a disaccharide thioglycoside donor.     

Propargyl mediated intramolecular aglycon delivery (IAD): applications to the synthesis of core N-glycan oligosaccharides

The use of propargyl mediated intramolecular aglycon delivery (IAD) for the synthesis of the key Manβ(1→4)GlcNAc linkage of N-glycan oligosaccharides, including the core N-glycan pentasaccharide, is investigated. Isomerisation of a 2-O-progargyl group of manno thioglycoside donors to an allene is followed by iodonium ion mediated mixed acetal formation with the 4-OH of protected GlcNAc acceptors, and subsequent intramolecular glycosylation occurs with complete control of anomeric stereochemistry to form the Manβ(1→4)GlcNAc linkage. A variety of linear and convergent approaches (1+2, 3+1, 3+2) to the core pentasaccharide are investigated as means of probing the generality and limitations of this type of intramolecular aglycon delivery for the formation of β-mannoside linkages in complex oligosaccharides.     

Organische selenverbindungen—I : Elektronenstossinduzierte Sauerstoff—bzw. hydroxylgruppen-übertragungen auf selenfunktionen

The mass spectrometric investigation of some new sulfur and selen compounds containing ortho-nitrophenyl groups yields examples of novel unknown O- and OH-migrations to various acceptors. The transfer reactions to selen acceptors are the first described in literature. The electron-impact fragmentations of isosteric sulfur and selen compounds lead only to analogous ions in few cases.     

O-苯甲酰吡啶甲醛肟衍生物的合成及其季铵化反应

为了解除人畜常被杀虫剂有机磷酸酯所致的中毒症状,文献曾报道一系列乙酰胆碱酯酶复能剂的研究和机理探讨^[1],其中用于临床的有吡啶2-甲醛肟碘或氯甲烷盐等.ciba公司和schnekenburger等曾先后合成若干肟醚类和吡啶N-氧化物甲醛肟酯类季铵盐.作者设想合成若干吡啶甲醛肟酯季铵盐^[2,3].     

Development of a novel fluorescent tag O-2-[aminoethyl]fluorescein for the electrophoretic separation of oligosaccharides

This study describes the development of a novel fluorescent tag, O-2-[aminoethyl]fluorescein, for the separation of sugars by capillary electrophoresis with fluorescence detection using an argon ion laser. The tag was synthesised using three consecutive steps namely: esterification, alkylation and hydrolysis, specifically designed to offer a flexible way in which to make an assortment of fluorescent tags from cheap and readily available starting reagents (typically less than $1 per g of fluorescent tag). Via this flexible synthetic pathway, O-2-[aminoethyl]fluorescein was designed and synthesised with a spacer group to lower steric effects between the fluorescein backbone and the reducing end of the carbohydrate which were anticipated to improve the reactivity of the tag. The newly synthesised tag, O-2-[aminoethyl]fluorescein was evaluated against structurally similar commercial fluorescent motifs namely fluorescent 5-aminomethylfluorescein and non-fluorescent 5-aminofluorescein. Kinetic studies indicated that O-2-[aminoethyl]fluorescein showed similar labeling efficiencies as 5-aminomethylfluorescein, but were achieved in only 30 min, supporting the notion of improved reactivity of the spacer group. The sensitivity of O-2-[aminoethyl]fluorescein was evaluated using maltoheptaose with a detection limit of 1 nM obtained, which was slightly higher than that of 0.3 nM obtained with 5-aminomethylfluorescein, and was due to its lower quantum yield (0.24) when conjugated to the sugar. The separation performance of the tag was also benchmarked with the two commercial reagents using a range of corn syrup oligosaccharides, from 4 to 10 glucose units, typically found in rice starch. Separations were performed using an electrolyte containing 100 mM boric acid, tris at pH 8.65 as background electrolyte, 30 kV applied voltage, 50 μm I.D. × 40 cm (30 cm effective length) capillary. The novel tag showed better resolution of small oligosaccharides, G3 and G4, than the other two reagents, but slightly worse resolution for the longer oligosaccharides, most likely due to the monovalent charge state of the O-2-[aminoethyl]fluorescein compared to the divalent charge of the other two tags.     

Synthesis of boron-containing tyrosine derivatives based on the closo-decaborate and closo-dodecaborate anions

A series of new boron-containing tyrosine derivatives [B _n H _n?1O(CH₂)₄O-4-C₆H₄CH₂CH-(NH₃)COOH]^? and [B _n H _n?1O(CH₂CH₂)₂O-4-C₆H₄CH₂CH(NH₃)COOH]^? (n = 10, 12) were prepared by ring opening of the cyclic oxonium derivatives of the decahydro-closo-decaborate and dodecahydro-closo-dodecaborate anions, respectively, with ethyl N-trifluoroacetyl-L-tyrosinate.     

Synthesis of fluorinated mucin core 2 branched oligosaccharides with the potential of novel substrates and enzyme inhibitors for glycosyltransferases and sulfotransferases

Syntheses of fluorinated mucin core 2 tri- and tetrasaccharides modified at the C-3 or C-4 position of the pertinent galactose residue are reported. These compounds were used for the study of sialyltransferases and 3-O-sulfotransferases involved in the biosynthesis of O-glycans. Our acceptor substrate specificity studies on three cloned sialyltransferases (Sia-Ts) revealed that a 3- or 4-fluoro substituent in beta1,4Gal resulted in poor acceptors for alpha2,6(N)Sia-T and alpha2,3(N)Sia-T, whereas 4-fluoro-Galbeta1,3GalNAcalpha was a good acceptor for alpha2,3(O)Sia-T. Uniquely, 4-F-Galbeta1,4GlcNAcbeta1,6(Galbeta1,3)GalNAcalpha-OBn was an inhibitor of alpha2,6(N)Sia-T activity but not alpha2,3(N)Sia-T activity. Further we found that the activities of only Gal 3-O-sulfotransferases and not sialyltransferases were adversely affected by a C-3 fluoro substituent at the other Gal terminal of mucin core 2. The strategy of building branched mucin core 2 structures by three glycosidation sequence coupling three classes of glycosyl donors with the reactivity-matching acceptors proved to be successful in syntheses of modified mucin-type core structures of O-glycan. The relative poor yields of the glycosylations using fluorinated galactosyl donors indicated that the fluorine modification dramatically decreased the donor reactivity due to electron-withdrawing effect.     

Synthesis and characterization of oligo-4-[(pyridin-3-ylimino)methyl]phenol

The oxidative polycondensation of 4-[(pyridin-3-ylimino)methyl]phenol (4-PIMP) with O₂, H₂O₂, and NaOCl was studied in an aqueous alkaline medium between 50°C and 90°C. Oligo-4-[(pyridin-3-ylimino)methyl]phenol (O-4-PIMP) prepared was characterized by ¹H-NMR, ¹³C-NMR, FT-IR, UV-VIS, size-exclusion chromatography, and elemental and thermal analyses techniques. At the optimum reaction conditions, the yield of O-4-PIMP was 18.9%, 39.4%, and 46.8% using H₂O₂, O₂, and NaOCl oxidant, respectively. According to the TG analysis, the initial degradation temperature of O-4-PIMP was 218°C, which was by 50°C higher than that of 4-PIMP. Thermal analyses of 4-PIMP and O-4-PIMP were carried out in N₂ atmosphere at 15–1000°C. The highest occupied molecular orbital, the lowest unoccupied molecular orbital, and electrochemical energy gaps of 4-PIMP and O-4-PIMP were determined from the onset potentials for n-doping and p-doping, respectively. Also, optical band gaps of 4-PIMP and O-4-PIMP were determined according to UV-VIS measurements.     

Convenient temporary methyl imidate protection of N-acetylglucosamine and glycosylation at O-4

This paper expands on the scope and utility of the temporary conversion of N-acetyl groups to alkyl imidates when attempting to glycosylate at O-4 of N-acetylglucosamine acceptors. The optimized synthesis of alkyl imidate protected glucosamine acceptors at position 4 and carrying various protecting groups at O-3 is described. These imidates were prepared immediately prior to glycosylation by treating the 4-OH acceptors with 0.5 M MeOTf to obtain the corresponding methyl imidates still carrying a free 4-OH group. When preparing these imidates in diethyl ether as the reaction solvent, we observed the unexpected formation of ethyl imidates in addition to the desired methyl imidates. While the 3-O-allyl acceptors were too unstable to be useful in glycosylation reactions, the 3-O-acylated methyl and ethyl imidates of glucosamine were shown to behave well during the glycosylation of the 4-OH with a variety of reaction conditions and various glycosyl donors. Glycosylation of these acceptors was successfully carried out with perbenzylated beta-thioethyl rhamnopyranoside under MeOTf promotion, while activation of this donor under NIS/TMSOTf or NIS/TfOH proved less successful. In contrast, activation of the less reactive perbenzylated alpha-thioethyl and peracetylated beta-thioethyl rhamnopyranosides with NIS/TfOH led to successful glycosylations of the 4-OH. Activation of a peracetylated rhamnosyl trichloroacetimidate by TMSOTf at low temperature also gave a high yield of glycosylation. We also report one-pot glycosylation reactions via alkyl imidate protected acceptor intermediates. In all cases the alkyl imidate products were readily converted to their corresponding N-acetyl derivatives under mild conditions.     

Synthesis of three regioisomers of the pentasaccharide part of the Skp1 glycoprotein of Dictyostelium discoideum

Three regioisomers of the linear pentasaccharide part of the Skp1 glycoprotein, found in Dictyostelium discoideum, were prepared in the form of (2-trimethylsilyl)ethyl glycosides by means of 2+3 block syntheses using the disaccharide donor at the non-reducing end, and three different trisaccharide acceptors at the reducing end. Fucosylation of (2-trimethylsilyl)ethyl 3,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-4,6-O-benzylidene-2-deoxy-2-NPhth-β-d-glucopyranoside with different fucosyl donors carrying an O-(2-naphthyl)methyl ether as a temporary-protecting group at positions C2, C3 or C4 gave rise to the protected core trisaccharides. After selective removal of the (2-naphthyl)methyl group, the resulting acceptors were glycosylated with the α(1→6) linked digalactosyl donor to yield the respective three regioisomeric pentasaccharides. Transformation of the phthalimido moiety into an N-acetyl group, followed by catalytic hydrogenation of the reducible-protecting groups furnished the free target pentasaccharides, which should be able to assist during the elucidation of the exact structure of the natural pentasaccharide.     

Synthesis of highly functionalized 2,5-disubstituted pyrrolidines via an aza-Morita-Baylis-Hillman-type reaction

An aza-Morita-Baylis-Hillman-type reaction of Michael acceptors with 5-substituted cyclic N,O-acetals derived from pyrrolidines has been investigated. It has been found that the combination of Me₂S and TMSOTf work well with unhindered and reactive enals and enones whilst the use of quinuclidine and TMSOTf is superior for more hindered Michael acceptors. The reactions lead to 2,5-trans-disubstituted pyrrolidines with good to excellent diastereoselectivity. The origin of the selectivity is discussed.     

Influence of acyl groups on glucopyranoside reactivity in Lewis acid promoted anomerisation

Lewis acid promoted anomerisation has potential in O- or S-glycoside synthesis. Herein, the anomerisation kinetics of thirty-one β-d-glucopyranosides was determined to determine how particular acyl protecting groups and their location influence reactivity towards a Lewis acid promoted reaction. The replacement of acetyl groups with benzoyl groups led to reduced reactivity when located at O-3, O-4 and O-6. However a reactivity increase was observed when the acetyl group was replaced by a benzoyl group at O-2. The 2,3,4,6-tetra-O-(4-methoxy)benzoate had an?～2-fold increase in rate when compared to the tetrabenzoate.     

Synthesis of new quinuclidine derivatives via Pd-mediated cross-coupling and cross-benzannulation reactions

New functionalized quinuclidines were prepared via palladium-catalyzed addition reactions of terminal alkynes (donors) to internal alkynes (acceptors). The enantiopure terminal alkynes were derivatives of quincoridine and quincorine, two semi-natural Cinchona alkaloids. The processes exhibited high chemoselectivity and excellent diastereoselectivity, the E-enynes being obtained as single products in almost all cases. The synthesis of new tetra and pentasubstituted benzene derivatives in good yields by [2+2+2] benzannulation of the diynes, obtained by the palladium-catalyzed homodimerization of 10,11-didehydro quincoridine and 10,11-didehydro quincorine, with terminal alkynes and in fair yield by [4+2] benzannulation of an enyne derivative of 10,11-didehydro quincoridine with 2,4-hexane-diyne are reported.     

Preparation of linear oligosaccharides by a simple monoprotective chemo-enzymatic approach

A monoprotective approach, involving acetyl ester as unique protective group in oligosaccharides synthesis, has been developed. Starting from peracetylated monosaccharides and glycals, by using an efficient and selective chemo-enzymatic ‘one-pot’ strategy (a regioselective hydrolysis catalyzed by immobilized lipases followed by a chemical acyl migration), different carbohydrate acceptors, only protected with acetyl ester, can be achieved. If combined with the use of an acetylated glycosyl donor, the glycosylation reaction with these glycosyl acceptors leads to peracetylated oligosaccharides. These compounds can be directly used as intermediates for the synthesis of glycopeptides used as antitumoral vaccines and, at the end of the process, can be easily fully deprotected in only one step. Thus, these key building blocks have been successfully used in glycosylation reactions for an efficient construction of peracetylated disaccharides, such as the biological relevant lactosamine, in multigram scale. Subsequently, glycosylation with the 3OH-tetraacetyl-α-d-galactose, used as carbohydrate acceptor, allowed the synthesis of a peracetylated N-trisaccharidic precursor of the lacto-N-neo-tetraose antigen. Extending this strategy to a 3OH-di-acetyl galactal, one peracetylated precursor of the T tumor-associated carbohydrate antigen has been synthesized.This efficient approach, characterized by the use of the acetyl ester as only protecting group during all the synthetical steps expected, represents an easy and efficient alternative to the classical synthetic methods in carbohydrate chemistry that involve several protecting group manipulation.     

Synthesis of amine-functionalized heparin oligosaccharides for the investigation of carbohydrate-protein interactions in microtiter plates

The synthesis of well-defined oligosaccharides is crucial for the establishment of structure-activity relationships for specific sequences of heparin, contributing to the understanding of the biological role of this polysaccharide. It is highly convenient that the synthetic oligosaccharides contain an orthogonal functional group that allows selective conjugation of the probes and expands their use as chemical tools in glycobiology. We present here the synthesis of a series of amine-functionalized heparin oligosaccharides using an n+2 modular approach. The conditions of the glycosylation reactions were carefully optimized to produce efficiently the desired synthetic intermediates with an N-benzyloxycarbonyl-protected aminoethyl spacer at the reducing end. The use of microwave heating greatly facilitates O- and N-sulfation steps, avoiding experimental problems associated with these reactions. The synthesized oligosaccharides were immobilized in 384-well microtiter plates and successfully probed with a heparin-binding protein, the basic fibroblast growth factor FGF-2. The use of hexadecyltrimethylammonium bromide minimized the amount of sugar required for attachment to the solid support. Using this approach we quantified heparin-protein interactions, and surface dissociation constants for the synthetic heparin derivatives were determined.     

A galabiose-based two-dimensional scaffold for the synthesis of inhibitors targeting P- and P-antigen binding proteins

A disaccharide scaffold based on galabiose (Galα1-4Gal) was synthesized. Four different acceptors were evaluated in the α-galactosylation and a relationship between the nucleophilicity, yield, and α/β-selectivity was found. The scaffold contains two orthogonal derivatisation sites, i.e. at O-2′ and the anomeric position, and as proof of concept, one derivatised galabioside was synthesized. Compounds based on this galabiose-scaffold are potential inhibitors of P- and P^k-antigen binding proteins.     

alpha-(2,6)-Sialyltransferase-catalyzed sialylations of conformationally constrained oligosaccharides

It is demonstrated that conformationally restricted oligosaccharides can act as acceptors for glycosyltransferases. Correlation of the conformational properties of N-acetyl lactosamine (Galbeta(1-4)GlcNAc, LacNAc) and several preorganized derivatives with the corresponding apparent kinetic parameters of rat liver alpha-(2,6)-sialyltransferase-catalyzed sialylations revealed that this enzyme recognizes LacNAc in a low energy conformation. Furthermore, small variations in the conformational properties of the acceptors resulted in large differences in catalytic efficiency. Collectively, our data suggest that preorganization of acceptors in conformations that are favorable for recognition by a transferase may improve catalytic efficiencies.     

Chemoenzymatic synthesis of sialylated oligosaccharides for their evaluation in a polysialyltransferase assay

A series of sialylated β-d-Gal-(1→3)-α-d-GalNAc-octyl containing oligosaccharides representative of those found on mucin type complex O-glycans were synthesized by a chemoenzymatic approach for use in the kinetic characterization of recently cloned polysialyltransferases. Enzymatic incorporation of N-acetylneuraminic acid (sialic acid) into the synthetic acceptors was accomplished by 2,3-(N) and (O)-sialyltransferases to give the target compounds 6-10 in a practical yield.     

6-O-Benzyl- and 6-O-silyl-N-acetyl-2-amino-2-N,3-O-carbonyl-2-deoxyglucosides: effective glycosyl acceptors in the glucosamine 4-OH Series. effect of anomeric stereochemistry on the removal of the oxazolidinone group

[reaction: see text] The 4-OH groups of both alpha- and beta-methyl glycosides of N-acetylglucosamine, protected with an oxazolidinone spanning the nitrogen and O-3, and bearing benzyl or silyl protection on O-6, show excellent reactivity as acceptors in couplings to a range of glycosyl donors. The enhanced reactivity of these acceptors is attributed in part to the tied back nature of the oxazolidinone, which reduces hindrance around the nucleophilic oxygen. The N-acetyloxazolidinone function also reduces the tendency seen in simple N-acetylglucosamines toward amide glycosylation, and removes the possibility of problematic hydrogen bonding networks. In the beta-, but not the alpha-, series selective hydrolysis of the N-acetyloxazolidinone directly to the N-acetylglucosamine was possible with barium hydroxide, a feature attributed to chelate formation between the acetamide carbonyl group and the glycosidic oxygen in the beta-series.     

Synthesis and structural characterization of novel tetranuclear organotitanoxane derivatives

Synthesis of the novel titanoxane compounds, [(TiCl)(TiOH){(Ti)[μ-(η(5)-C(5)Me(4)SiMe(2)O-κO)](2)(μ-O)}(2)(μ-O)] (4) and [{Ti[μ-(η(5)-C(5)Me(4)SiMe(2)O-κO)](μ-O)}(4)] (5) by controlled reaction of the dinuclear titanium oxo complex [{Ti{μ-(η(5)-C(5)Me(4)SiMe(2)O-κO)}Cl](2)(μ-O)] (1) with 2 equiv of LiOH is reported. Complex 4 is innovative and remarkable. It is one of the rare known examples of tetranuclear stable terminal hydroxo titanium complexes, with an open-chained structure, which coincides with the transient metal monohydroxo proposed in the stepwise pathway employed to justify the formation of the hexanuclear complex [{Ti[μ-(η(5)-C(5)Me(4)SiMe(2)O-κO)](μ-O)}(6)] (3) from 1. (1)H DOSY experiments were used to characterize complex 4. In addition, the structures of compound 5 and of precursor 1 were determined by single-crystal X-ray diffraction studies.