A novel glycosphingolipid from gram-negative aquatic bacteria.

The chloroform-methanol extractable lipids of the Gram-negative fresh-water bacteria Arcocella aquatica NO-502 and Flectobacillus major FM were found to contain an unusual ninhydrin-positive glycolipid. It was purified by two-stage silica gel-column chromatography. By the use of IR and (1)H-NMR spectroscopy, mass spectrometry and chemical-degradation experiment, the lipid was established to be 1-O-monoglycosyl ceramide, the carbohydrate moiety of which was the alpha-pyranose-ring form of 7-desoxy-7-amino-D-manno-heptulosonic acid, or 1-hydroxycarbonyl-6-deoxy-6-amino-alpha-D-mannopyranose. The ceramide portion consisted mainly (by 95% in the A. aquatica glycolipid and 80% in the F. major glycolipid) of 2-N-(2'-D-hydroxy-13'-methyltetradecanoyl)-15-methyl-4(E)-hexad ecasph ingenine. The minor molecular species differed from the major one only in fatty acid structure. The glycolipid accounted for 8 and 11% of the total lipids extracted from A. aquatica NO-502 and F. major FM cells, respectively.     

A precursor to the beta-pyranosides of 3-amino-3,6-dideoxy-D-mannose (mycosamine).

SN2-type reaction of 3-O-(1-imidazyl)sulfonyl-1,2:5,6-di-O-isopropylidene-alpha-D-gluco furanose with benzoate gave the 3-O-benzoyl-alpha-D-allo derivative 2, which was hydrolysed to give the 5,6-diol 3. Compound 3 was converted into the 6-deoxy-6-iodo derivative 4 which was reduced with tributylstannane, and then position 5 was protected by benzyloxymethylation, to give 3-O-benzoyl-5-O-benzyloxymethyl-6-deoxy-1,2-O-isopropylidene-alpha -D- allofuranose (6). Debenzoylation of 6 gave 7, (1-imidazyl)sulfonylation gave 8, and azide displacement gave 3-azido-5-O-benzyloxymethyl-3,6-dideoxy- 1,2-O-isopropylidene-alpha-D-glucofuranose (9, 85%). Acetolysis of 9 gave 1,2,4-tri-O-acetyl-3-azido-3,6-dideoxy-alpha,beta-D-glucopyranose (10 and 11). Selective hydrolysis of AcO-1 in the mixture of 10 and 11 with hydrazine acetate (----12), followed by conversion into the pyranosyl chloride 13, treatment with N,N-dimethylformamide dimethyl acetal in the presence of tetrabutylammonium bromide, and benzylation gave 3-azido-4-O-benzyl-3,6-dideoxy-1,2-O-(1-methoxyethylidene)-alpha-D -glucopyranose (15). Treatment of 15 with dry acetic acid gave 1,2-di-O-acetyl-3-azido-4-O-benzyl-3,6-dideoxy-beta-D-glucopyranose (16, 86% yield) that was an excellent glycosyl donor in the presence of trimethylsilyl triflate, allowing the synthesis of cyclohexyl 2-O-acetyl-3-azido-4-O-benzyl-3,6-dideoxy-beta-D-glucopyranoside (17, 90%).(ABSTRACT TRUNCATED AT 250 WORDS)     

Preparation and calculated conformations of the 2'-, 3'-, 4'-, and 6'-deoxy, 3'-O-methyl, 4'-epi, and 4'- and 6'-deoxy-fluoro derivatives of methyl 4-O-alpha-D-galactopyranosyl-beta-D-galactopyranoside (methyl beta-D-galabioside)

The glycosyl chlorides of the 3-O-methyl (6) and 4-deoxy-4-fluoro (8) O-benzylated derivatives of D-galactopyranose and 2,3,4,6-tetra-O-benzyl-D-glucopyranose were condensed with methyl 2,3,6-tri-O-benzoyl-beta-D-galactopyranoside to give, after deprotection, the 3'-O-methyl (23), 4'-deoxy-4'-fluoro (25), and 4'-epi (27) derivatives, respectively, of methyl beta-D-galabioside (1). The glycosyl fluorides of 2,3,4-tri-O-benzyl-D-fucopyranose and the 3-deoxy (12) and 4-deoxy (16) O-benzylated derivatives of D-galactopyranose were condensed with methyl 2,3,6-tri-O-benzyl-beta-D-galactopyranoside (21), to give, after deprotection, the 6'-deoxy (31), 3'-deoxy (34), and 4'-deoxy (37) derivatives of 1, respectively. The 2'-deoxy (41) derivative of 1 was prepared by N-iodosuccinimide-induced condensation of 3,4,6-tri-O-acetyl-D-galactal and 21 followed by deprotection. Treatment of methyl 2,3,6-tri-O-benzoyl-4-O-(2,3-di-O-benzoyl-alpha-D-galactopyranosyl)-beta -D- galactopyranoside with Et2NSF3 (DAST), followed by deprotection, provided the 6'-deoxy-6'-fluoro (46) derivative of 1. Molecular mechanics calculations yielded conformations for 23, 25, 27, 31, 34, 37, 41, and 46 with small deviations from the calculated conformation for 1 (phi H/psi H: -40 degrees/-6 degrees).     

A substrate for the fluorogenic assay of exo-beta-N-acetylmuramidase: synthesis and purification of 4-methylumbelliferyl N-acetylmuramide.

A fluorogenic substrate for exo-β-N-acetylmuramidase from Bacillus subtilis B was synthesized. 4-Methyl-2-oxo-1,2-benzopyran-7-yl 2-acetamido-4,6-O-benzylidene-2-deoxy-β-d-glucopyranoside was prepared from 4-methyl-2-oxo-1,2-benzopyran-7-yl 2-acetamido-2-deoxy-β-d-glucopyranoside, condensed with dl-2-chloropropionic acid, the benzylidene residue removed by acetolysis and the 4-methyl-2-oxo-1,2-benzopyran-7-yl 2-amino-3-O-(d-1-carboxyethyl)-2-deoxy-β-d-glucopyranoside purified by chromatography on silica gel and Sephadex G-10 and by high-voltage paper electrophoresis. The identity of the product was confirmed by pmr studies, acid hydrolysis followed by chromatography of the products, and enzymic digestion.     

The Synthesis of Novel Carbohydrates Amenable to the Synthesis of 2′,3′-Dideoxy-3′-Branched Nucleosides

Abstract

The facile synthesis of several substituted carbohydrates that are amenable for the preparation of 2′,3′-dideoxy-3′-hydroxymethyl nucleosides are reported. Elaboration of a previously reported analog, 5-O-benzoyl-3-deoxy-3-(benzyloxy)methyl-1,2-O-isopropylidene-β-D- ribofuranose (4) has provided two 2,3-dideoxy-3-branched ribose derivatives 5-O-benzoyl-2,3-dideoxy-3-(benzyloxy)methyl-1-O-methyl-β-D-ribofuranose (7) and 1.5-di-O-benzoyl-2,3-dideoxy-3-(benzyloxy)methyl-(α,β)-D-ribofuranose (10). Due to problems involved with the separation of anomeric mixtures when these carbohydrates were condensed with an heterocycle, another versatile synthon 5-O-benzoyl-3-deoxy-3-(benzyloxy)methyl-2-O-t-butyldimethylslyl-1-O- methyl-β-D-ribofuranose (12) was synthesized. The utility of this compound (12) is demonstrated in the total synthesis of 1-[3-deoxy-3-hydroxymethyl-β-D-ribofuranosyl]thymine (20).     

Synthesis of 2-deoxy-2-C-alkylglucosides of myo-inositol as possible inhibitors of a N-deacetylase enzyme in the biosynthesis of mycothiol

Two new analogues of 1-D-1-O-(2-acetamido-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol, a biosynthetic intermediate in the production of mycothiol in the Mycobacteria have been synthesized. Both the 2-deoxy-2-C-(2'-hydroxypropyl)-D-glucoside 5, and the 2-deoxy-2-C-(2'-oxopropyl)-D-glucoside 6, are derived from fully benzylated 1-D-1-O-(2-C-allyl-2-deoxy)-D-glucopyranosyl)-myo-inositol 20, readily assembled via a protected 2-C-allyl-2-deoxyglucosyl fluoride. Both 5 and 6 inhibit the incorporation of [3H]inositol by whole cells of Mycobacterium smegmatis into a number of metabolites which contain inositol.     

SYNTHESIS AND IN VITRO ANTI-HCV ACTIVITY OF β-D- and L-2′-DEOXY-2′-FLUORORIBONUCLEOSIDES

Based on the discovery of β-D-2′-deoxy-2′-fluorocytidine as a potent anti-hepatitis C virus (HCV) agent, a series of β-D- and l-2′-deoxy-2′-fluororibonucleosides with modifications at 5 and/or 4 positions were synthesized and evaluated for their in vitro activity against HCV and bovine viral diarrhea virus (BVDV). The introduction of the 2′-fluoro group was achieved by either fluorination of 2,2′-anhydronucleosides with hydrogen fluoride-pyridine or potassium fluoride, or a fluorination of arabinonucleosides with DAST. Among the 27 analogues synthesized, only the 5-fluoro compounds, namely β-D-2′-deoxy-2′,5-difluorocytidine (5), had anti-HCV activity in the subgenomic HCV replicon cell line, and inhibitory activity against ribosomal RNA. As β-D-N4-hydroxycytidine (NHC) had previously shown potent anti-HCV activity, the two functionalities of the N4-hydroxyl and the 2′-fluoro were combined into one molecule, yielding β-D-2′-deoxy-2′-fluoro-N4-hydroxycytidine (12). However, this nucleoside showed neither anti-HCV activity nor toxicity. All the l-forms of the analogues were devoid of anti-HCV activity. None of the compounds showed anti-BVDV activity, suggesting that the BVDV system cannot reliably predict anti-HCV activity in vitro.     

Mouse anti-ceramide antiserum: a specific tool for the detection of endogenous ceramide.

Ceramide is a pivotal molecule in signal transduction and an essential structural component of the epidermal permeability barrier. The epidermis is marked by a high concentration of ceramide and by a unique spectrum of ceramide species: Besides the two ceramide structures commonly found in mammalian tissue, N-acylsphingosine and N-2-hydroxyacyl-sphingosine, six additional ceramides differing in the grade of hydroxylation of either the sphingosine base or the fatty acid have been identified in the epidermis. Here we report on the characterization of an IgM-enriched polyclonal mouse serum against ceramide. In dot blot assays with purified epidermal lipids the antiserum bound to a similar extent to N-acyl-sphingosine (ceramide 2), N-acyl-4-hydroxysphinganine (ceramide 3), and N-(2-hydroxyacyl)-sphingosine (ceramide 5), whereas no specific reaction was detected with glycosylceramides, sphingomyelin, free sphingosine, phospholipids, or cholesterol. In contrast, a monoclonal IgM antibody, also claimed to be specific for ceramide, was shown to bind specifically to sphingomyelin and therefore was not further investigated. In thin-layer chromatography immunostaining with purified lipids a strong and highly reproducible reaction of the antiserum with ceramide 2 and ceramide 5 was observed, whereas the reaction with ceramide 1 and ceramide 3 was weaker and more variable. Ceramide 2 and ceramide 5 were detected in the nanomolar range at serum dilutions of up to 1:100 by dot blot and thin-layer immunostaining. In thin-layer chromatography immunostaining of crude lipid extracts from human epidermis, the antiserum also reacted with N-(2-hydroxyacyl)-4-hydroxysphinganine (ceramide 6) and N-(2-hydroxyacyl)-6-hydroxysphingosine (ceramide 7). Furthermore, the suitability of the antiserum for the detection of endogenous ceramide by immunolight microscopy was demonstrated on cryoprocessed human skin tissue. Double immunofluorescence labeling experiments with the anti-ceramide antiserum and the recently described anti-glucosylceramide antiserum (Brade et al., 2000, Glycobiology 10, 629) showed that both lipids are concentrated in separate epidermal sites. Whereas anti-ceramide stained the dermal and basal epidermal cells as well as the corneocytes, anti-glucosylceramide staining was concentrated in the stratum granulosum. In conclusion, the specificity and sensitivity of the reagent will enable studies on the subcellular distribution and biological functions of endogenous ceramide.     

In vitro lipofection with novel asymmetric series of 1,2-dialkoylamidopropane-based cytofectins containing single symmetric bis-(2-dimethylaminoethane) polar headgroups

Novel N,N'-diacyl-1,2-diaminopropyl-3-carbamoyl[bis-(2-dimethylaminoethane)] bivalent cationic lipids were synthesized and evaluated for in vitro transfection activity against a murine melanoma cell line. In the absence of the helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine), only the dioleoyl derivative 22 (1,2lb5) elicited transfection activity. The transfection activity of this lipid was reduced when formulated with DOPE. Contrary to that, the dimyristoyl derivative 19 (1,2lb2) mediated no activity when used alone but induced the highest levels of marker gene expression in the presence of DOPE. In an effort to correlate the transfection activity with cationic lipid structures, the physicochemical properties of cationic lipids in isolation and of lipoplexes were studied with surface tensiometry, photon correlation spectroscopy, gel electrophoresis mobility shift assay, and fluorescence techniques. In regard to the lipoplex properties, gel electrophoresis mobility shift assay and EtBr exclusion fluorescence assay revealed that the 1,2lb5 was the only lipid to associate and condense plasmid DNA, respectively. Photon correlation spectroscopy analysis found that 1,2lb5/DNA complexes were of relatively small size compared to all other lipoplexes. With respect to the properties of isolated lipids, Langmuir monolayer studies and fluorescence anisotropy on cationic lipid dispersions verified high two-plane elasticity and increased fluidity of the transfection competent dioleoyl derivative 1,2lb5, respectively. The results indicate that high transfection activity is mediated by cationic lipids characterized by an expanded mean molecular area, high molecular elasticity, and increased fluidity.     

Synthesis and evaluation of N-substituted 1,4-oxazepanyl Sordaricins as selective fungal EF-2 inhibitors

Sordaricin analogues possessing 6-methoxy-7-methyl-1,4-oxazepane moiety instead of the sugar part were synthesized and evaluated. It was found that N-substituents on the oxazepane ring had influence on biological activity. In particular, N-(2-methylpropenyl) derivative 12p exhibited potent in vitro antifungal activity. Furthermore, 12p maintained significant activity (MIC 0.25 microg/mL) against Candida albicans SANK51486 even in the presence of 20% horse serum.     

Synthesis of benzyl 2-acetamido-2-deoxy-3-O-β-d-fucopyranosyl-α-d-galactopyranoside and benzyl 2-acetamido-6-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-2-deoxy-3-O-β-d-fucopyranosyl-α-d-galactopyranoside

Condensation of benzyl 2-acetamido-4,6-O-benzylidene-2-deoxy-α-d-galactopyranoside with 2,3,4-tri-O-acetyl-α-d-fucopyranosyl bromide in 1:1 nitromethane-benzene, in the presence of powdered mercuric cyanide, afforded benzyl 2-acetamido-4,6-O-benzylidene-2-deoxy-3-O-(2,3,4-tri-O-acetyl-β-d-fucopyranosyl)-α-d-galactopyranoside (3). Cleavage of the benzylidene group of 3 with hot, 60% aqueous acetic acid afforded diol 4, which, on deacetylation, furnished the disaccharide 5. Condensation of diol 4 with 2-methyl-(3,4,6-tri-O-acetyl-1,2-di-deoxy-α-d-glucopyrano)-[2,1-d]-2-oxazoline in 1,2-dichloroethane afforded the trisaccharide derivative (7). Deacetylation of 7 with Amberlyst A-26 (OH^?) anion-exchange resin in methanol gave the title trisaccharide (8). The structures of 5 and 8 were confirmed by ¹³C-n.m.r. spectroscopy.     

Phospholipids in Mediterranean cephalopods

Polar lipids of the cephalopods Eledone moschata, Sepia officinalis and Todarodes sagittatus mantle, represent 50.5%, 66.1% and 74.2% of wet tissue respectively. On the other hand the polar lipids of these three species of cephalopods constitute of 80.8%, 94.8% and 93.7% of phospholipids, respectively. The main phospholipids identified were phosphatidylcholine (52.2, 51.3 and 58.4% of total phospholipids respectively in the above mentioned species), phosphatidylethanolamine (18.1, 19.7 and 23.9%), sphingomyelin (10.7, 15.2 and 6.7%), lyso-phosphatidylcholine (3.1, 3.8 and 1.8%) and the unusual lipid ceramide aminoethylphosphonic acid (15.9, 10 and 9.2%). The 56.8% of phosphatidylcholine in Eledone moschata, the 46% in Sepia officinalis and the 74.1% in Todarodes sagittatus refer to the structure of 1,2-diacyl-glycerocholine and the remaining percentage refer to the structure of 1-o-alkyl-2-acyl-glycerocholine or 1-o-alkyl-1-enyl-2-acyl-glycerocholine. The 87.2% of phosphatidylethanolamine in Eledone moschata, the 81% in Sepia officinalis and the 90.7% in Todarodes sagittatus refer to the structure of 1,2-diacyl-glyceroethanolamine and the remaining percentage refer to the structure of 1-o-alkyl-2-acyl-glyceroethanolamine or 1-o-alkyl-1-enyl-2-acyl-glyceroethanolamine. The major saturated fatty acids in phosphatidylcholine and phosphatidylethanolamine were C16:0 (30.3-67.5% and 23.2-54.5%) and C18:0 (3.6-17% and 15.4-28%), respectively, while the major unsaturated fatty acids in these lipids were C18:1n-9, n-7 (1.0-7.3% and 5.3-10.5%), C20:5n-3 (1.5-9.8% and 4,5-15.8%) and C22:6n-3 (12.5-42.0% and 7.0-11.3%), respectively.     

Synthesis of 1,2,4-tri-O-acetyl-5-deoxy-5-C-[(R and S)-methoxy-phosphinyl]-3-O-methyl-α- and -β- d-xylopyranose,and their structural analysis by 400-MHz,proton nuclear magnetic resonance spectroscopy

5-Deoxy-5-iodo-1,2-O-isopropylidene-3-O-methyl-α- d-xylofuranose, prepared quantitatively from its 5-Op-tolylsulfonyl precursor, readily gave the 5-C-(diethoxy-phosphinyl) derivative. Treatment of this compound with sodium dihydrobis(2-methoxyethoxy)aluminate, followed by hydrogen peroxide, mineral acid, and hydrogen peroxide, yielded 5-deoxy-5-C-(hydroxyphosphinyl)-3-O-methyl-α,β- d-xylopyranoses in 65% overall yield. The structures of these sugar analogs were effectively established on the basis of the mass and 400-MHz, ¹H-n.m.r. spectra of the four title compounds, derived by treatment with diazomethane and then acetic anhydride in pyridine. 5-C-[(S)-(1-Acetoxyethenyl)phosphino]-1,2,4-tri-O-acetyl-5-deoxy-3-O-methyl-β- d-xylopyranose was also isolated and characterized.     

Liposome/DNA systems: correlation between association, hydrophobicity and cell viability

Small unilamellar vesicles associated with plasmid DNA showed maximum association efficiency for a cationic mixture of egg phosphatidylcholine (EPC):1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE):di-1,2-dioleoyl-3-trimethyl ammonium propane (DOTAP) (16:8:1 molar ratio) [65%], followed by neutral lipids EPC:1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE):cholesterol (Chol) (2:2:1 molar ratio) [30%], and a polymerized formulation 1,2-bis(10,12-tricosadiynoyl)sn-glycero-3-phosphocholine (DC8,9PC):DMPE:Chol (2:2:1 molar ratio) [11%]. The hydrophobicity factor (HF) for these formulations followed the trend DC8,9PC:DMPE:CHOL < EPC:DMPE:Chol < EPC:DOPE DOTAP, and DNA association did not alter this trend. Results suggest that the higher the HF value, the more fluid the membrane and the higher the efficiency of DNA association. On the other hand, no differences were observed in cell toxicity with lipids up to 1 mg/ml in VERO cells.     

Synthesis of some second-generation substrate analogues of early intermediates in the biosynthetic pathway of glycosylphosphatidylinositol membrane anchors

1-D-6-O-(2-Amino-2-deoxy-alpha-D-glucopyranosyl)-2-O-octyl-myo-inositol 1-(1,2-di-O-hexadecanoyl-sn-glycerol 3-phosphate) (23) and the corresponding 2-O-hexadecyl-D-myo-inositol compound 24 have been prepared as substrate analogues of an early intermediate in the biosynthetic pathway of glycosylphosphatidylinositol (GPI) membrane anchors. 1-D-6-O-(2-Amino-2-deoxy-alpha-D-glucopyranosyl)-myo-inositol 1-(1,2-di-O-octyl-sn-glycerol 3-phosphate) has also been prepared as a substrate analogue. Biological evaluation of the analogues 23 and 24 revealed that they are neither substrates nor inhibitors of GPI biosynthetic enzymes in the human (HeLa) cell-free system but are potent inhibitors at different stages of GPI biosynthesis in the Trypanosoma brucei cell-free system.     

An efficient synthesis of 3-fluoro-5-thio-xylofuranosyl nucleosides of thymine, uracil, and 5-fluorouracil as potential antitumor or/and antiviral agents

1,2:5,6-Di-O-isopropylidene-alpha-D-glucofuranose by the sequence of mild oxidation, reduction, fluorination, periodate oxidation, borohydride reduction, and sulfonylation gave 3-deoxy-3-fluoro-1,2-O-isopropylidene-5-O-p-toluenesulfonyl-alpha-D-xylofuranose (5). Tosylate 5 was converted to thioacetate derivative 6, which after acetolysis gave 1,2-di-O-acetyl-5-S-acetyl-3-deoxy-3-fluoro-5-thio-D-xylofuranose (7). Condensation of 7 with silylated thymine, uracil, and 5-fluorouracil afforded nucleosides 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) thymine (8), 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) uracil (9), and 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) 5-fluorouracil (10). Compounds 8, 9, and 10 are biologically active against rotavirus infection and the growth of tumor cells.     

Characterization of cerebrosides from the thermally dimorphic mycopathogen Histoplasma capsulatum: expression of 2-hydroxy fatty N-acyl (E)-Delta(3)-unsaturation correlates with the yeast-mycelium phase transition

Cerebroside (monohexosylceramide) components were identified in neutral lipids extracted from both the yeast and mycelial forms of the thermally dimorphic mycopathogen Histoplasma capsulatum. The components were purified from both forms and their structures elucidated by 1- and 2-dimensional nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and low energy tandem collision-induced dissociation mass spectrometry (ESI-MS/CID-MS). Both components were characterized as beta-glucopyranosylceramides (GlcCers) containing (4E,8E)-9-methyl-4,8-sphingadienine as the long-chain base, attached to 18-carbon 2-hydroxy fatty N-acyl components. However, while the fatty acid of the yeast form GlcCer was virtually all N-2'-hydroxyoctadecanoate, the mycelium form GlcCer was characterized by almost exclusive expression of N-2'-hydroxy-(E)-delta(3)-octadecenoate. These results suggest that the yeast-mycelium transition is accompanied by up-regulation of an as yet uncharacterized ceramide or cerebroside 2-hydroxy fatty N-acyl (E)-delta(3)-desaturase activity. They also constitute further evidence for the existence of two distinct pathways for ceramide biosynthesis in fungi, since glycosylinositol phosphorylceramides (GIPCs), the other major class of fungal glycosphingolipids, are found with ceramides consisting of 4-hydroxysphinganine (phytosphingosine) and longer chain 2-hydroxy fatty acids. In addition to identification of the major glucocerebroside components, minor components (< 5%) detectable by molecular weight differences in the ESI-MS profiles were also characterized by tandem ESI-MS/CID-MS analysis. These minor components were identified as variants differing in fatty acyl chain length, or the absence of the sphingoid 9-methyl group or (E)-delta(8)-unsaturation, and are hypothesized to be either biosynthetic intermediates or the result of imperfect chemical transformation by the enzymes responsible for these features. Possible implications of these findings with respect to chemotaxonomy, compartmentalization of fungal glycosphingolipid biosynthetic pathways, and regulation of morphological transitions in H.capsulatum and other dimorphic fungi are discussed.     

Enamine and pyrrole formation from 2-amino-2-deoxy-D-glucose and dimethyl acetylenedicarboxylate

Addition of 2-amino-2-deoxy-β-D-glucopyranose to dimethyl acetylenedicarboxylate afforded an almost quantitative yield of amorphous 2-deoxy-2-(1,2-dimethoxycarbonylvinyl)amino-D-glucose (5). Acetylation of this adduct gave crystalline 1,3,4,6-tetra-O-acetyl-2-deoxy-2-[(Z)-1,2-dimethoxycarbonylvinyl]amino-α-D-glucopyranose (6a); the corresponding β-D anomer (6b) was obtained by addition of 1,3,4,6-tetra-O-acetyl-2-amino-2-deoxy-β-Dglucopyranose to dimethyl acetylenedicarboxylate. O-Deacetylation of tetra-acetate 6a with barium methoxide in methanol occurred selectively at C-1, yielding enamine 6c derived from 3,4,6-tri-O-acetyl-2-amino-2-deoxy-α-D-glucopyranose. Conversion of the crude adduct 5 into 3-methoxycarbonyl-5-(D-arabino-tetrahydroxybutyl)-2-pyrrolecarboxylic acid (7) took place by heating in water or in slightly basic media in yields up to 83%. Acetylation of 7 gave the tricyclic derivative 8, and its periodate oxidation afforded 5-formyl-3-methoxycarbonyl-2-pyrrolecarboxylic acid (9). Oxidation of 9 with alkaline silver oxide yielded 3-methoxy-carbonyl-2,5-pyrroledicarboxylic acid (10).     

Synthesis and preliminary in vivo evaluation of new 2-Aryl-6-methyl-1,2-dihydro-1H-pyridin-4-ones and 2-Aryl-6-methylpiperidin-4-ols, as potential anti-amnesiant agents

A revisited synthesis of 2-aryl-6-methyl-1,2-dihydro-1H-pyridin-4-ones and their saturated analogues 2-aryl-6-methylpiperidin-4-ols is described. A five steps procedure, using the sulfinimine chemistry, to prepare a key intermediate beta-(6-chloronicotinic)-beta-amino ester is also reported. In vivo spontaneous working memory activity of these compounds has been investigated in the mouse. Results obtained with 2-(3-chlorophenyl)-6-methyl-1,2-dihydro-1H-pyridin-4-one 9b, the most effective derivative in this model, have been reported.     

Synthesis of gem-di-C-substituted derivatives of carbohydrates by way of nucleophilic-addition reactions of aldohexofuranoid 3-C-methylene derivatives

Nucleophilic Michael-type additions to aldohexofuranoid 3-C-methylene derivatives, namely, 3-deoxy-1,2:5,6-di-O-isopropylidene-3-C-nitromethylene-α-d-ribo-hexofuranose and 3-C-[cyano(ethoxycarbonyl)methylene]-3-deoxy-1,2:5,6-di-O-isopropylidene-α-d-ribo-hexofuranose employing phase-transfer catalysis, afforded novel gem-di-C-substituted sugars. The conversion of 3-deoxy-1,2:5,6-di-O-isopropylidene-3-C-methyl-3-C-nitromethyl-α-d-allo-hexofuranose into a 3-C-hydroxymethyl-3-C-methyl derivative with titanium trichloride, and that of the nitromethyl groups of 3-deoxy-1,2:5,6-di-O-isopropylidene-3,3-di-C-nitromethyl-α-d-ribo-hexofuranose, and 3-deoxy-1,2:5,6-di-O-isopropylidene-3-C-methyl-3-C-nitromethyl- and -3-C-nitromethyl-α-d-allo-hexofuranose into cyano groups with phosphorus trichloride in pyridine is also described.