Isolation and identification of products in the reaction of 2-acetamido-3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-d-arabino-hex-1-enitol with theophylline

Fusion of 2-acetamido-3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-d-arabino-hex-1-enitol with theophylline, in the presence of boron trifluoride etherate as the catalyst, caused condensation to occur. This reaction afforded a variety of products of nucleosidic character, which were successively separated by repeated chromatography on silica gel. The structures of the products were determined, on the basis of X-ray crystallographic analysis (for three compounds) and by means of n.m.r.-spectral data and mass spectrometry, as the following: 7-(2-acetamido-4,6-di-O-acetyl-2,3-dideoxy-β-d-erythro-hex-2-enopyranosyl)theophylline, the corresponding α- and β-d-threo derivatives, and 7-(2-acetamido-6-O-acetyl-2,3-dideoxy-α-d-threo-hex-2-enopyranosyl)theophylline and its β anomer.In addition to these 2′,3′-unsaturated nucleosides having the base linked at C-1′, three products of a new type, having the base attached at C-4′, were also isolated: 7-(methyl 2-acetamido-6-O-acetyl-2,3,4-trideoxy-β-d-erythro-hex-2-enopyranosid-4-yl)theophylline, and the corresponding α-d-threo and α-d-erythro isomers.The correlation of the data obtained by X-ray structure analysis and proton nuclear magnetic spectroscopy, together with their application for the determination of configuration and conformation of these compounds, are discussed. It appears that the ¹H-n.m.r. data alone do not suffice for unambiguous and correct structure determination for these classes of compounds.     

The synthesis of P1-2-acetamido-4-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-2-deoxy-α-d-glucopyranosyl P2-dolichyl pyrophosphate, (P1-di-N-acetyl-α-chitobiosyl P2-dolichyl pyrophosphate)

2-Acetamido-4-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-2-deoxy-α-d-glucopyranosyl phosphate, pure according to thin-layer and gas—liquid chromatography, optical rotation, and treatment with alkaline phosphatase and 2-acetamido-2-deoxy-β-d-glucosidase, was prepared by treatment of 2-methyl-[4-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-3,6-di-O-acetyl-1,2-dideoxy-α-d-glucopyrano]-[2,1-d]-2-oxazoline with dibenzyl phosphate, followed by the removal of the benzyl groups by catalytic hydrogenolysis, and O-deacetylation. In contrast, a sample prepared by the phosphoric acid procedure was shown to consist mainly of the β anomer. 2-Acetamido-4-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-3,6-di-O-acetyl-2-deoxy-α-d-glucopyranosyl phosphate was treated wit P¹-diphenyl P²-dolichyl pyrophosphate to give a fully acetylated pyrophosphoric diester, which was O-deacetylated to give P¹-2-acetamido-4-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-2-deoxy-α-d-glucopyranosyl P²-dolichyl pyrophosphate. This compound could be separated from the β anomer by t.l.c., and its behavior under dilute acid and alkaline conditions was investigated.     

Determination of the position of linkage of 2-acetamido-2deoxy-D-galactose and 2-acetamido-2-deoxy-D-glucose residues in oligosaccharides and glycoproteins. Synthesis of 2-acetamido-2deoxy-D-xylitol and 2-acetamido-2-deoxy-L-threitol

A method has been studied for the determination of the position of the linkage of the 2-acetamido-2-deoxy-D-galactose and 2-acetamido-2-deoxy-D-glucose residues in oligosaccharides and glycoproteins that is based on the borohydride reduction of the reducing terminal residues to the corresponding alditol derivatives periodate oxidation, borohydride reduction, hydrolysis (eventually followed by borohydride reduction), separation of the fragments as per-O-(trimethylsilyl) or per-O-(trifluoroacetyl) derivatives, and identification of the fragments as derivatives of 2-acetamido-2-deoxyglycerol, 2-acetamido-2-deoxy-L-threitol, 2-acetamido-2-deoxy-L-arabinitol, 2-acetamido-2-deoxy-D-xylitol, 2-acetamido-2-deoxy-D-galactitol, and 2-acetamido-2-deoxy-D-glucitol by gas-liquid chromatography-mass spectrometry. New syntheses for the standard compounds 2-acetamido-2-deoxy-L-threitol and 2-acetamido-2-deoxy-D-xylitol are described.     

Biological evaluation of a series of 2-acetamido-2-deoxy-D-glucose analogs towards cellular glycosaminoglycan and protein synthesis in vitro

Using primary hepatocytes in culture, various 2-acetamido-2-deoxy-D-glucose (GlcNAc) analogs were examined for their effects on the incorporation of D-[³H]glucosamine, [³⁵S]sulfate, and L-[¹⁴C]leucine into cellular glycoconjugates. A series of acetylated GlcNAc analogs, namely methyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-α-(3) and β-D-glucopyranoside (4) and 2-acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-D-glucopyranose (5), exhibited a concentration-dependent reduction of D-[³H]glucosamine, but not of [³⁵S]sulfate incorporation into isolated glycosaminoglycans (GAGs), without affecting L-[¹⁴C]leucine incorporation into total protein synthesis. These results suggest that analogs 3–5 exhibit an inhibitory effect on D-[³H]glucosamine incorporation into isolated GAGs by diluting the specific activity of cellular D-[³H]glucosamine and by competing for the same metabolic pathways. In the case of the corresponding series of 4-deoxy-GlcNAc analogs, namely methyl 2-acetamido-3,6-di-O-acetyl-2,4-dideoxy-α-(6) and β-D-xylo-hexopyranoside (7) and 2-acetamido-1,3,6-tri-O-acetyl-2,4-dideoxy-D-xylo-hexopyranose (8), compound 8 at 1.0 mM exhibited the greatest reduction of D-[³H]glucosamine and [³⁵S]sulfate incorporation into isolated GAGs, namely to ∼7% of controls, and a moderate inhibition of total protein synthesis, namely to 60% of controls. Exogenous uridine was able to restore the inhibition of total protein synthesis by compound 8 at 1.0 mM. Isolated GAGs from cultures treated with compound 8 were shown to be smaller in size (∼40 kDa) than for control cultures (∼77 kDa). These results suggest that the inhibitory effects of compound 8 on cellular GAG synthesis may be mediated by the incorporation of a 4-deoxy moiety into GAGs resulting in premature chain termination and/or by its serving as an enzymatic inhibitor of the normal sugar metabolites. The inhibition of total protein synthesis from cultures treated with compound 8 suggests a uridine trapping mechanism which would result in the depletion of UTP pools and cause the inhibition of total protein synthesis. A 1-deoxy-GlcNAc analog, namely 2-acetamido-3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-D-glucitol (9), also exhibited a reduction in both D -[³H]glucosamine and [³⁵S]sulfate incorporation into isolated GAGs by 19 and 57%, of the control cells, respectively, at 1.0 mM without affecting total protein synthesis. The inability of compound 9 to form a UDP-sugar and, hence, be incorporated into GAGs presents another metabolic route for the inhibition of cellular GAG synthesis. Potential metabolic routes for each analog's effects are presented.     

The separation of chondroitin sulfate disaccharides and hyaluronan oligosaccharides by capillary zone electrophoresis

We have developed techniques for the separation of unsulfated (2-acetamido-2-deoxy-3-O-(4-deoxy-alpha-L-threo- hex-4-enopyranosyluronicacid)-D-galactose and -D-glucose), monosulfated (2-acetamido-2-deoxy-3- O-(4-deoxy-2-O-sulfo-alpha-L-threo-hex-4-enopyranosyluronic acid)-D-galactose and 2-acetamido-2-deoxy-3-O-(4-deoxy-alpha-L-threo-hex- 4-enopyranosyluronic acid)-4-sulfo-D-galactose and -6-sulfo-D-galactose),disulfated (2-acetamido-2-deoxy-3-O-(4-deoxy-2-O-sulfo-alpha-L-threo-hex-4- enopyranosyluronic acid)-4-sulfo-D-galactose and -6-sulfo-D-galactose and 2-acet-amido-2-deoxy-3-O-(4-deoxy-alpha-L-threo-hex-4-enopy- ranosyluronic acid)-4,6-di-O-sulfo-D-galactose), and trisulfated (2-acetamido-2-deoxy-3-O-(4-deoxy-2-O- sulfo-alpha-L-threo-hex-4-enopyranosyluronic acid)-4,6-di-O-sulfo-D-galactose) isomers of chondroitin using capillary zone electrophoresis. In addition, it is possible to separate oligomers of hyaluronan by similar protocols. These techniques represent a rapid, sensitive, and reproducible technique for the assay of these molecules from digests of connective tissues.     

Synthesis of p-nitrophenyl 6-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-α-d-mannopyranoside

The reaction of p-nitrophenyl 2,3-O-isopropylidene-α-d-mannopyranoside and 2-methyl-(3,4,6-tri-O-acetyl-1,2-dideoxy-α-d-glucopyrano)-[2,1-d]-2-oxazoline gave a crystalline, 6-O-substituted disaccharide derivative which, on de-isopropylidenation followed by saponification, produced the disaccharide p-nitrophenyl 6-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-α-d-mannopyranoside. Synthesis of methyl 6-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-α-d-mannopyranoside was also accomplished by a similar reaction-sequence. The structures of these disaccharides have been established by ¹³C-n.m.r. spectroscopy.     

A convenient synthesis of p-nitrophenyl 2-deoxy-2-(thio-acetamido)-β-d-glucopyranoside, -galactopyranoside,and their 1-thio analogs as inhibitors of 2-acetamido-2-deoxy-β-d-glucosidase

The acetamido group of p-nitrophenyl 2-acetamido-2-deoxy-β-d-glucopyranoside, -β-d-galactopyranoside, and their 1-thio analogs was modified by replacement of the amide-carbonyl oxygen atom with sulfur by treatment of their fully acetylated derivatives with phosphorus pentasulfide in pyridine. The resulting p-nitrophenyl 2-deoxy-2-thioacetamido-β-d-hexopyranoside triacetates were O-deacetylated with catalytic amounts of sodium methoxide in methanol to obtain p-nitrophenyl 2-deoxy-2-thioacetamido-β-d-glucopyranoside, -β-d-galactopyranoside, and their 1-thio analogs. These derivatives inhibited 2-acetamido-2-deoxy-β-d-glucosidase from Turbatrix aceti to various extents. Also obtained in significant yields in the aforementioned reaction with phosphorus pentasulfide in pyridine were the two hitherto unreported thiazolines, namely, 2-methyl(2-acetamido-3,4,6-tri-O-acetyl-α-d-glucopyrano)[2′,1′:4,5]-2-thiazoline and 2-methyl(2-acetamido-3,4,6-tri-O-acetyl-α-d-galactopyrano)[2′,1′:4,5]-2-thiazoline.     

The inhibitory activities of 2-acetamido-2,3-dideoxy-D-hex-2-enonolactones on 2-acetamido-2-deoxy-beta-D-glucosidase.

Treatment of 2-acetamido-2-deoxy-D-mannono-1,4-lactone with dicyclohexylamine in ethanolic solution afforded an unsaturated 1,4-lactone, 2-acetamido-2,3-dideoxy-D-erythro-hex-2-enono-1,4-lactone (1), in good yield. 2-Acetamido-2,3-dideoxy-D-threo-hex-2-enono-1,4-lactone (2) was similarly prepared from 2-acetamido-2-deoxy-D-galactono-1,4-lactone. An unsaturated 1,5-lactone, 2-acetamido-2,3-dideoxy-D-threo-hex-2-enono-1,5-lactone (4), was obtained through the oxidation of 2-acetamido-2-doexy-4,6-0-isopropylidene-D-galactopyranose with silver carbonate on Celite, followed by mild hydrolysis. The inhibitory activity of four isomeric 2-acetamido-2,3-dideoxy-D-hex-2-enonolactones [1, 2, 4, and 2-acetamido-2,3-dideoxy-D-erythro-hex-2-enono-1,5-lactone (3)] was assayed against 2-acetamido-2-deoxy-beta-D-glucosidase from bull epididymis. Only the erythro lactones 1 and 3 are weak competitive inhibitors, whereas the threo lactones 2 and 4 are practically inactive. The 1,4-lactone 1 inhibited 2-acetamido-2-deoxy-beta-D-glucosidase more strongly than the 1,5-lactone 3. The lactones 1-4 were found to be quite stable in aqueous solution or under inhibitory-assay conditions. In addition, two 2-acetamido-2-deoxy-D-glycals, 2-acetamido-1,5-anhydrohex-1-enitol (7) were tested; both are 10 times as active as 1.     

Studies on the epimerization of 2-acetamido-2-deoxyhexoses: preparation of 2-acetamido-2-deoxy-d-[2-3H]-glucose and -mannose

Epimerization of either 2-acetamido-2-deoxy-d-glucose (1) or 2-acetamido-2-deoxy-d-mannose (2) in basic tritium oxide gave 2-acetamido-2-deoxy-d-[2-³H]-glucose (3) and 2-acetamido-2-deoxy-d[2-³H]mannose (4). In both cases, compound 3 was isolated in higher proportion and higher specific activity than 4. The mechanism of the epimerization of 1 and 2 is discussed.     

Synthesis of glycopeptide derivatives containing the 2-acetamido-N-(L-aspart-4-oyl)-2-deoxy-β-D-glucopyranosylamine linkage and having the amino acid sequences 32–34, 33–35,33–37, and 33–38 of bovine ribonuclease

The synthesis is described of the glycotripeptide derivatives 2-acetamido-3,4,6-tri-O-acetyl-N-[N-(benzyloxycarbonyl)-L--seryl-L-nitroarginyl-L-aspart-4-oyl]-2-deoxy-β-D-glucopyranosylamine, 2-acetamido-3,4,6-tri-O-acetyl-N-[N-(benzyloxycarbonyl)-L-seryl-L-nitroarginyl-L-aspart-1-oyl-(1-p-nitrobenzyl ester)-4-oyl]-2-deoxy-β-D-glucopyranosylamine, and 2-acetamido-3,4,6-tri-O-acetyl-N-[N-(benzyloxycarbonyl)-L-nitroarginyl-L-aspart-1-oyl-(L-leucine methyl ester)-4-oyl]-2-deoxy-β-D-glucopyranosylamine, and of the glycopentapeptide and glycohexapeptide derivatives 2-acetamido-3,4,6-tri-O-acetyl-N-[N-(benzyloxycarbonyl)-L-nitroarginyl-L-aspart-1-oyl-(L-leucyl-L-threonyl-threonyl-N^ε-tosyl-L-lysine-(p-nitrobenzyl ester)-4-oyl]-2-deoxy-β-D-glycopyranosylamine and 2-acetamido-3,4,6-tri-O-acetyl-N-[N-(benzyloxycarbonyl)-L-nitroarginyl-L-aspart-1-oyl-(L-leucyl-L-threonyl-N^ε-tosyl-L-lysyl-L-aspartic 1,4-di-p-nitrobenzyl ester)-4-oyl]-2-deoxy-β-D-glucopyranosylamine.     

Modifications at C-3 and C-4 of 2-amino-2-deoxy-d-glucose

Modifications at C-3 and C-4 of 2-amino-2-deoxy-d-glucose have been developed. A 3,4-double bond was introduced into benzyl 2-acetamido-2-deoxy-3,4-di-O-Methylsulfonyl-α-d-glucopyranoside by treatment with NaI and Zn. Epoxidation of the double bond with m-chloroperoxybenzoic acid gave an exo-epoxide exclusively. The stereochemistry of the epoxidation product has been confirmed by an alternative synthesis. An analysis of the ¹H-n.m.r. spectra indicates that both the 3,4-unsaturated derivatives and the epoxide exist in the °H₁ (d) conformation. Nucleophilic reagents (F^?, I^?) opened the 3,4-epoxide to give 4-substituted derivatives having the d-gulo configuration. Thus, 2-acetamido-1,3,6-tri-O-acetyl-2,4-dideoxy-4-iodo-α-d-gulopyranose and 2-acetamido-1,3,6-tri-O-acetyl-3,4-dideoxy-4-fluoro-α-d-gulopyranose have been synthesized. Reduction of the double bond in the key intermediate and deprotection gave 2-acetamido-2,3,4-trideoxy-d-glucopyranose. Some of the derivatives were active as inhibitors of growth of mouse, mammary adenocarcinoma cells in culture.     

Biosynthesis of a P1-2-acetamido-2-deoxy-D-glucosyl P2-polyisoprenyl pyrophosphate by calf pancreas microsomes

Incubation of UDP-[¹⁴C]-$\text{N}$-acetylglucosamine with calf pancreas microsomes in the presence of Mn⁺⁺ and potassium thiocyanate gave a labeled glycolipid, tentatively identified as $\text{P}$¹-2-acetamido-2-deoxy-$\text{D}$-glucosyl $\text{P}$²-dolichyl pyrophosphate on the basis of cochromatography with synthetic $\text{P}$¹-2-acetamido-2-deoxy-α-$\text{D}$-glucopyranosyl $\text{P}$²-dolichyl pyrophosphate, similar chemical and enzymic hydrolyses of the biosynthetic and synthetic compounds, and stimulation of the biosynthesis by addition to the incubation mixture o dolichyl phosphate or a crude lipid fraction extracted from microsomes.     

Assignment of the stereochemistry and anomeric configuration of structurally informative product ions derived from disaccharides: infrared photodissociation of glycosyl-glycolaldehydes in the negative ion mode

Using mass spectrometry in the negative ion mode, m/z 221 ions are frequently observed as product ion substructures derived from reducing disaccharides having 2, 4, or 6 linkages. The ions have been shown to be glycosyl-glycolaldehydes. All 16 stereochemical variants of their pyranosides were prepared and evaluated by infrared photodissociation, in addition to HexNAc-glycolaldehyde variants (m/z 262) of 2-acetamido-2-deoxy-d-glucose and 2-acetamido-2-deoxy-d-galactose. The stereochemistry and anomeric configuration of these ions were differentiated in the gas phase using a Fourier transform ion cyclotron resonance spectrometer with infrared multiphoton dissociation at 10.6 μm. Results were compared to those obtained by collision-induced dissociation. In some cases, differentiation was far preferable using infrared photodissociation; in others, collision-induced dissociation was preferred. Using an instrument that interfaced a linear trap with a Fourier transform ion cyclotron resonance spectrometer, either dissociation technique could be used to optimally discriminate between isomers. With infrared photodissociation, spectral differences were highly statistically significant, even between pairs of isomers having spectra that appeared to be visually somewhat similar (p <1 × 10⁻⁹, student’s t-test for key discriminatory ions). Comparisons among different instruments suggest that physical standards of the stereochemical variants of these ions will be required for their detailed structural assignments in unknowns, as some variation was observed among instruments, both using infrared photodissociation and collision-induced dissociation.     

Glycosidation at C-4 of 2-acetamido-2-deoxy-D-glucopyranose derivatives by koenigs-knorr type condensations

The condensation of 2,3,4,6-tetra-O-benzyl-D-glucopyranosyl bromide and 2,3,4,6-tetra-O-benzyl-D-mannopyranosyl chloride with benzyl 2-acetamido-3,6-di-O-benzyl-2-deoxy-α-D-glucopyranoside (1), under Koenigs-Knorr conditions, gave the fully benzylated derivatives of benzyl 2-acetamido-2-deoxy-4-O-α-D-glucopyranosyl-α-D-glucopyranoside, benzyl 2-acetamido-2-deoxy-4-O-β-D-glucopyranosyl-α-D-glucopyranoside, and benzyl 2-acetamido-2-deoxy-4-O-α-D-mannopyranosyl-α-D-glucopyranoside. Three further compounds, namely, benzyl 2-acetamido-3-O-benzyl-2-deoxy-6-O-(2,3,4,6-tetra-O-benzyl-D-glucopyranosyl)-α-D-glucopyranoside, benzyl 2-acetamido-3-O-benzyl-2-deoxy-6-O-(2,3,4,6-tetra-O-benzyl-D)-mannopyranosyl)-α-D-glucopyranoside, and benzyl 2-acetamido-3-O-benzyl-2-deoxy-4,6-di-O-(2,3,4,6-tetra-O-benzyl-D-mannopyranosyl)-α-D-glucopyranoside, were formed by reaction of the respective glycosyl halide with benzyl 2-acetamido-3-O-benzyl-2-deoxy-α-D-glucopyranoside present as contaminant in 1.     

Synthesis and reactions of O-acetylated benzyl alpha-glycosides of 6-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-N-acetylmuramoyl-L- alanyl-D-isoglutamine esters: the base-catalysed isoglutamine in equilibrium glutamine rearrangement in peptidoglycan-related structures

Condensation of benzyl 2-acetamido-6-O-(2-acetamido-3,4,6-tri-O-acetyl-2- deoxy-3-O-[(R)-1-carboxyethyl]-alpha-D-glucopyranoside (2) and its 4-acetate (4) with L-alanyl-D-isoglutamine benzyl ester via the mixed anhydride method yielded N-(2-O-[benzyl 2-acetamido-6-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D- glucopyranosyl)-2,3-dideoxy-alpha-D-glucopyranosid-3-yl]-(R)-lacto yl)-L- alanyl-D-isoglutamine benzyl ester (5) and its 4-acetate (6), respectively. Condensation by the dicyclohexylcarbodi-imide-N-hydroxysuccinimide method converted 2 into benzyl 2-acetamido-6-O-(2-acetamido-3,4,6-tri-O-acetyl- 2-deoxy-beta-D-glucopyranosyl)-3-O-[(R)-1-carboxyethyl]-2-deoxy-alpha-D- glucopyranoside 1',4-lactone (7). In the presence of activating agents, 7 underwent aminolysis with the dipeptide ester to give 5. Zemplén O-deacetylation of 5 and 6 led to transesterification and alpha----gamma transamidation of the isoglutaminyl residue to give N-(2-O-[benzyl 2-acetamido-6-O-(2- acetamido-2-deoxy-beta-D-glucopyranosyl)-2,3-dideoxy-alpha-D-glucopyr anosid-3- yl]-(R)-lactoyl)-L-alanyl-D-isoglutamine methyl ester (8) and -glutamine methyl ester (9). Treatment of 6 with MgO-methanol caused deacetylation at the GlcNAc residue to give a mixture of N-(2-O-[benzyl 2-acetamido-6-O-(2-acetamido-2- deoxy-beta-D-glucopyranosyl)-4-O-acetyl-2,3-dideoxy-alpha-D-glucopyra nosid-3- yl]-(R)-lactoyl)-L-alanyl-D-isoglutamine methyl ester (11) and -glutamine methyl ester (12). Benzyl or methyl ester-protection of peptidoglycan-related structures is not compatible with any of the reactions requiring alkaline media. Condensation of 2 with L-alanyl-D-isoglutamine tert-butyl ester gave N-(2-O-[benzyl 2-acetamido- 6-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D-glucopyranosyl)-2,3-d ideoxy- alpha-D-glucopyranosid-3-yl]-(R)-lactoyl-L-alanyl-D-isoglutamine tert-butyl ester (16), deacetylation of which, under Zemplén conditions, proceeded without side-reactions to afford N-(2-O-[benzyl 2-acetamido-6-O-(2-acetamido-2-deoxy-beta-D- glucopyranosyl)-2,3-dideoxy-alpha-D-glucopyranosid-3-yl]-(R)-la cotyl)-L- alanyl-D-isoglutamine tert-butyl ester (17).     

Synthesis of phosphodiesters of 2-acetamido-2-deoxy-D-glucose--fragments of polymers of capsules from Escherichia coli K51 and Neisseria meningitidis X

Hydrogenphosphonate method was used for synthesis of 4-nitrophenyl 2-acetamido-3- and 4-nitrophenyl 2-acetamido-4-(2-acetamido-2-deoxy-alpha-D-glucopyranosyl phosphate)-2-deoxy-beta-D-glucopyranosides. The glycosides, phosphate diester fragments of the title bacteria capsular antigens, were obtained by H-phosphorylation of the suitably protected 2-acetamido-2-deoxy-beta-D-glucopyranosides with 2-acetamido-3,4,6-tri-O-benzoyl-2-deoxy-alpha-D-glucopyranosyl H-phosphonate in the presence of trimethylacetyl chloride followed by oxidation and deprotection.     

The synthesis and properties of benzylated oxazolines derived from 2-acetamido-2-deoxy-D-glucose

2-Methyl-(2-acetamido-3,4,6-tri-O-benzyl-1,2-dideoxy-α-D-glucopyrano)-[2,1-d]-2-oxazoline,2-methyl-(2-acetamido-6-O-acetyl-3,4-di-O-benzyl-1,2-dideoxy-α-D-glucopyrano)-[2,1-d]-2-oxazoline,and 2-methyl-(2-acetamido-4-O-acetyl-3,6-di-O-benzyl-1,2-dideoxy-α-D-glucopyrano)-[2,1-d]-2-oxazoline were synthesized from the allyl 2-acetamido-3,4,6-tri-O-benzyl-2-deoxy-D-glucopyranosides, and from the 3,4-di-O-benzyl or 3,6-di-O-benzyl analogs, respectively, both the α and β anomer being used in each case. The preparation of allyl 2-acetamido-3,4,6-tri-O-benzyl- and 3,6-di-O-benzyl-2-deoxy-β-D-glucopyranoside is also described. Treatment of the tri-O-benzyl oxazoline with dibenzyl phosphate gave a pentabenzylglycosyl phosphate, from which all the benzyl groups were removed by catalytic hydrogenation, giving 2-acetamido-2-deoxy-α-D-glucopyranosyl phosphate. The corresponding β anomer was not detectable. Treatment of the 3,4-, or 3,6-, di-O-benzyl oxazoline with allyl 2-acetamido-3,4-di-O-benzyl-α-D-glucopyranoside readily gave disaccharide products from which the protecting groups were removed, to give the (1→6)-linked isomer of di-N-acetylchitobiose. Under both acidic and basic conditions, this isomer was less stable than the (1→4)-linked compound.Attempts to employ the 3,6-di-O-benzyl oxazoline for the formation of (1→4)-linked disaccharides, by treatment with either anomer of allyl 2-acetamido-3,6-di-O-benzyl-2-deoxy-D-glucopyranoside, were not very successful, presumably owing to hindrance by the bulky benzyl groups.     

Synthesis of omega-(methoxycarbonyl)alkyl and 9-(methoxycarbonyl)-3,6-dioxanonyl glycopyranosides for the preparation of carbohydrate-protein conjugates.

omega-(Methoxycarbonyl)alkyl glycopyranosides of D-mannose having C4, C7, C9, C12, and C15 carbon chains, L-fucose and 2-acetamido-2-deoxy-D-mannose having C7 and C9 carbon chains, D-xylose and 2-acetamido-2-deoxy-L-fucose having a C9 carbon chain, and 9-(methoxycarbonyl)-3,6-dioxanonyl glycopyranosides of D-mannose, 2-acetamido-2-deoxy-D-mannose, and L-fucose were synthesized as intermediates for coupling to human serum albumin in order to examine the effect of chain length and hydrophobicity of the spacer arm on the binding specificity of lectins. 8-(Methoxycarbonyl)octyl glycosides of beta-D-Man-(1----2)-alpha-D-Man, alpha-D-Man-(1----2)-alpha-D-Man, alpha-D-ManNAc-(1----2)-alpha-D-Man, beta-D-GlcNAc-(1----2)-alpha-D-Man, and their 6-O-positional isomers, beta-D-Man-(1----6)-alpha-D-Man, alpha-D-Man-(1----6)-alpha-D-Man, alpha-D-ManNAc-(1----6)-alpha-D-Man, and beta-D-GlcNAc-(1----6)-alpha-D-Man, were also synthesized.     

Use of 3-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-2,4,6-tri-O-acetyl-α-d-galactopyranosyl bromide as a glycosyl donor: Synthesis of p-nitrophenyl 3-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-β-d-galactopyranoside

Acetolysis of methyl 3-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-2,4,6-tri-O-acetyl-α-d-galactopyranoside afforded 3-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-1,2,4,6-tetra-O-acetyl-d-galactopyranose (2). Treatment of 2 in dichloromethane with hydrogen bromide in glacial acetic acid gave 3-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)- 2,4,6-tri-O-acetyl-α-d-galactopyranosyl bromide (3). The α configuration of 3 was indicated by its high, positive, specific rotation, and supported by its ¹H-n.m.r. spectrum. Reaction of 3 with Amberlyst A-26-p-nitrophenoxide resin in 1:4 dichloromethane-2-propanol furnished p-nitrophenyl 3-O-(2-acetamido-3,4,6- tri-O-acetyl-2-deoxy-β-d-glucopyranosyl)-2,4,6-tri-O-acetyl-β-d-galactopyranoside (7). Compound 7 was also obtained by the condensation (catalyzed by silver trifluoromethanesulfonate-2,4,6-trimethylpyridine) of 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-β-d-glucopyranosyl bromide with p-nitrophenyl 2,4,6-tri-O-acetyl-β-d-galactopyranoside, followed by the usual deacylation-peracetylation procedure. O-Deacetylation of 7 in methanolic sodium methoxide furnished the title disaccharide (8). The structure of 8 was established by ¹³C-n.m.r. spectroscopy.     

Acetonation of 2-(acylamino)-2-deoxy-d-glucoses

2-Acetamido-2-deoxy-d-glucose and 2-(benzyloxycarbonylamino)-2-deoxy-d-glucose were each treated with 2,2-dimethoxypropane in N,N-dimethylformamide containing a trace of p-toluenesulfonic acid. The new 5,6-O-isopropylidene derivatives 2-acetamido-2-deoxy-5,6-O-isopropylidene-d-glucofuranose, 2-acetamido-1,4-anhydro-2-deoxy-5,6-O-isopropylidene-d-arabino-hex-1-enitol, 2-acetamido-2-deoxy-3,4:-5,6-di-O-isopropylidene-aldehydo-d-glucose-dimethyl acetal, and 2-(benzyloxycarbonylamino)-2-deoxy-5,6-O-isopropylidene-d-glucofuranose were isolated. The formation of these furanoid acetals may be important in ascertaining the mechanism of this unique acetonation accompanied by glycosidation.