Carotinoid-Glycosylester. 3. Mitteilung. Die synthese von crocetin-di-(β-D-glucosyl)-ester. Eine neue methode zur selektiven veresterung von ungeschützter β-D-glucose

Carotenoid Glycosyl Esters. The Synthesis of Crocetin-di-(β-D -glucosyl) Ester. A New Method for the Selective Esterification of Unprotected β-D -Glucose The naturally occurring crocetin-di-(β-D -glucosyl) ester is easily synthesized by the reaction of crocetin-di-imidazolide or crocetin-di-(1,2,4-triazolide) and unprotected β-D -glucose in pyridine in presence of a base (Scheme 4). Under the described experimental conditions the esterification takes place exclusively at the anomeric C-atom and furthermore produces only the β-anomer. It is the first time that an unprotected carbohydrate has been used for the selective synthesis of glucosyl esters at the anomeric C-atom. This represents the major advantage of this new method.     

Glycosylidene Carbenes. Part 19. Regioselective glycosidation of allyl 2-deoxy-2-phthalimido-D-allopyranosides

The regio- and stereoselectivity of the glycosidation of the partially protected mono-alcohols 3 and 7 , the diols 2 and 8 , and the triol 4 by the diazirine 1 have been investigated. Glycosidation of the α-D -diol 2 (Scheme 2) gave regioselectively the 1,3-linked disaccharides 11 and 12 (80%, α-D /β-D 9:1), whereas the analogous reaction with the βD -anomer 8 led to a mixture of the anomeric 1,3- and 1,4-linked disaccharides 13 (12.5%), 14 (16%), 15 (13%), and 16 (20.5%; Table 2). Protonation of the carbene by OH–C(4) of 2 is evidenced by the observation that the α-D -mono-alcohol 3 did not react with 1 under otherwise identical conditions, and that the β-D -alcohol 7 yielded predominantly the β-D -glucoside 18 (52%) besides 14% of 17 . Similarly as for the glycosidation of the diol 2 , the influence of the H-bond of HO? C(4) on the direction of approach of the carbene, the role of HO? C(4) in protonating the carbene, and the stereoelectronic control in the interception of the ensuring oxycarbenium cation are evidenced by the reaction of the triol 4 with 1 (Scheme 3), leading mostly to the α-D -configurated 1,3-linked disaccharide 19 (41%), besides its anomer 20 (16%), and some 4-substituted β-D -glucoside 21 (9%). No 1,6-linked disaccharides could be detected. In agreement with the observed reactivity, the ¹H-NMR and IR spectra reveal a strong H-bond between HO? C(3) and the phthalimido group in the α-D -, but not in the β-D -allosides. The different H-bonds in the anomeric phthalimides are in keeping with the results of molecular-mechanics calculations.     

Synthesis of New Nucleosides by coupling of chloropurines with 2- and 3-deoxy derivatives of N-methyl-D-ribofuranuronamide

The synthesis of new deoxyribose nucleosides by coupling chloropurines with modified D -ribose derivatives is reported. The methyl 2-deoxy-N-methyl-3-O-(p-toluoyl)-α-D -ribofuranosiduronamide (α-D - 8 ) and the corresponding anomer β-D - 8 were synthesized starting from the commercially available 2-deoxy-D -ribose ( 1 ) (Scheme 1). Reaction of α-D - 8 with the silylated derivative of 2,6-dichloro-9H-purine ( 9 ) afforded regioselectively the N⁹-(2′-deoxyribonucleoside) 10 as anomeric mixture (Scheme 2), whereas β-D - 8 did not react. Glycosylation of 9 or of 6-chloro-9H-purine ( 17 ) with 1,2-di-O-acetyl-3-deoxy-N-methyl-β-D -ribofuranuronamide ( 13 ) yielded only the protected β-D -anomers 14 and 18 , respectively (Scheme 3). Subsequent deacetylation and dechlorination afforded the desired nucleosides β-D - 11 , β-D - 12,15 , and 16 . The 3′-deoxy-2-chloroadenosine derivative 15 showed the highest affinity and selectivity for adenotin binding site vs. A₁ and A_2A adenosine receptor subtypes.     

Stereoselective synthesis of pyrrolo[2,3-d]pyrimidine α- and β-D-ribonucleosides from anomerically pure D-ribofuranosyl chlorides: Solid-Liquid Phase-Transfer Glycosylation and 15N-NMR Spectra

Solid-liquid phase-transfer glycosylation (KOH, tris[2-(2-methoxyethoxy)ethye]amine ( = TDA-1), MeCN) of pyrrolo[2,3-d]pyrimidines such as 3a and 3b with an equimolar amount of 5-O-[(1,1 -dimethylethyl)dimethylsilyl]-2,3-O-(1-methylethylidene)-α-D -ribofuranosyl chloride (1) [6] gave the protected β-D -nucleosides 4a and 4b , respectively, stereoselectively (Scheme). The β-D -anomer 2 [6] yielded the corresponding α-D -nucleosides 5a and 5b with traces of the β-D -compounds. The 6-substituted 7-deazapurine nucleosides 6a , 7a , and 8 were converted into tubercidin (10) or its α-D -anomer (11) . Spin-lattice relaxation measurements of anomeric ribonucleosides revealed that T₁ values of H? C(8) in the α-D -series are significantly increased compared to H? C(8) in the β-D -series while the opposite is true for T₁ of H? C(1′). ¹⁵N-NMR data of 6-substituted 7-deazapurine D -ribofuranosides were assigned and compared with those of 2′-deoxy compounds. Furthermore, it was shown that 7-deaza-2′deoxyadenosine ( = 2′-deoxytubercidin; 12 ) is protonated at N(1), whereas the protonation site of 7-deaza-2′-deoxyguanosine ( 20 ) is N(3).     

Silicon-29 NMR spectroscopy in carbohydrate chemistry: Galactose derivatives

²⁹Si NMR spectra of the O-trimethylsilyl (OTMS) derivatives of various methyl α- and β-D -galactopyranosides have been recorded. The effect of changes in the anomeric configuration provides a means of assigning the resonance of the 2-OTMS substituent. Whereas the signal of the OTMS group attached at the 6-position can be assigned readily, those of the OTMS group at the 3- or 4-position cannot be assigned unequivocally.     

Isolation,Synthesis and Derivatization of Xylodextrins

A series of xylodextrins has been produced by enzymatic or hydrothermal degradation of industrial xylans. For further synthetic use, the oligomers were converted into per-O-acetylated xylooligomers which were separated by silica gel chromatography to furnish preparative amounts of xylobiose up to xylopentaose. In a model reaction, selective anomeric deacetylation and treatment with trichloroacetonitrile furnished a xylobiosyl donor, which was converted into the β-methyl glycoside. In addition, methyl β-D -xylopyranoside was transformed into a suitable glycosyl acceptor via tosylation followed by a double displacement reaction at O-4, allowing for further chain elongation and modification at the reducing xylopyranosyl unit.     

Desoxy-nitrozucker. 13. Mitteilung. Herstellung ungeschützter und partiell geschützter 1-Desoxy-1-nitro-D-aldosen sowie Röntgenstrukturanalysen einiger ihrer Vertreter

Preparation of Unprotected and Partially Protected 1-Deoxy-1-nitro-D -aldoses and Some Representative X-Ray Structure Analyses The unprotected and partially protected 1-deoxy-1-nitro derivatives of α-and β-D -glucopyranose (see 15 and 14 ), β-D -mannopyranose (see 16 ), N-acetyl-β-D -glucosamine (see 17 ), β-D -galactofuranose (see 19 ), β-D -ribofuranose (see 20 ), α-D -arabinofuranose (see 21 ), 4,6-O-benzylidene-β-D -glucose (see 40 ), N-acetyl-4,6-O-benzylidene-β-D -glucosamine (see 41 ), and 4,6-O-benzylidene-β-D -galactose (see 42 ) were prepared by ozonolysis of the corresponding nitrones which were obtained from the acid-catalyzed reaction of p-nitrobenzaldehyde with the hydroxylamine 4 , the unprotected oximes 3 and 5–9 and the 4,6-O-benzylidene oximes 35–37 , respectively (Schemes 1–3). The gluco- and manno-nitrones 10 and 12 were isolated, and their ring size and their anomeric and (E/Z) configurations were determined by NMR spectroscopy and by their transformation into their corresponding nitro derivatives. The structure of the deoxynitroaldoses were determined by NMR spectroscopy, polarimetry, and, in the case of 14 , 16 , and 17 , by formation of the 4,6-O-benzylidene ( 14 → 40 ) or 4,6-O-isopropylidene ( 16 → 43 , 17 → 23 ) derivatives (Scheme 3). Acetylation of the nitroglucopyranose 14 , the 2-acetamido-nitroglucopyranose 17 , and the nitrogalactofuranose 19 gave the crystalline peracetylated nitroaldoses 22 , 24 , and 45 , respectively (Scheme 4, Figs. 1 and 3); acetylation of the nitromannopyranose 16 gave the nitro-arabino-glycal 44 (Scheme 4). The structure of the peracetylated nitroglucopyranose 22 , the nitroglucosamine 25 , the nitrogalactofuranose 45 , and the nitroribofuranose 20 were confirmed by X-ray analysis (Figs. 1 4). In all cases, including the β-D -glucopyranose derivative 22 , considerably shortening of the (endocyclic) C(1)-O bond was observed. Base-catalyzed anomerization of the β-D -configurated nitroglucopyranose 14 , the nitromannopyranose 16 , the benzylidene acetal 40 of nitroglucose, and the 2,3,4,6-tetraacetylated glucosamine derivative 24 gave the corresponding nitro-α-D -aldoses 15 , 26 , 47 , and 25 , respectively (Scheme 4).     

Glycosylidene Carbenes. Part 10. Regioselective Glycosidation of 4,6-O-Benzylidene-D-altropyranosides

Glycosidation by the diazirine 1 , the trichloroacetimidate 4 , and the bromide 5 of the altro-diol 2 , possessing an intramolecular H-bond (HO? C(3) to O? C(1)) in solution, but not in the solid state, proceeds with high and complementary regioselectivity. From 2 and 1 , one obtains mostly the 1,2-linked disaccharides 10 and 11 (β-D > α-D ), together with the 1,3-linked isomers 12 and 13 (α-D > β-D ; 1,2-/1,3-linked products ca. 9:1), the demethylated 1,3-linked disaccharides 24–27 , the trisaccharides 19–22 , the lactone azines 23 , and the hydroxyglucal 18 , while 2 reacted with 4 or 5 to yield mostly the 1,3-linked disaccharides (1,2-/1,3-linked products ca. 1:9). The disaccharides were additionally characterized as acetates (→ 14–17, 28–31 ). Yields and stereoselectivity depended upon the donor, stoichiometry, solvent, temperature, and concentration. Glycosidation of the 1,3-linked disaccharides with 1 yielded the trisaccharides 19–22 . Reaction of the β-D -altro-diol 3 with 1 gave the 1,2- and 1,3-linked disaccharides 32/33 and 34/35 in a 1:1 ratio, characterized as the acetates 36–39 , while glycosidation with 5 according to Lemieux proceeded regioselectively (1,2-/1,3-linked products 91:9). The monotosylates 6 and 7 reacted with 1 to yield the anomeric pairs 40/41 , and 42/43 of the tosylated disaccharides; the oxiranes 44 and 45 were not observed.     

Regioselective Enzyme-Mediated Glycosylation of Natural Polyhydroxy Compounds. Part 1. Galactosylation of stevioside and steviolbioside

Bovine β-1,4-galactosyltransierase is an efficient catalyst for the regioselective transfer of galactose from UPD-galactose, generated in situ with the UDP-glucose/UDP-glucose-4-epimerase system, to the kaurane glycosides stevioside ( 1 ) and Steviolbioside ( 2 ), affording the corresponding galactosyl derivatives 3 and 4 in high yields. By a combination of 2D NMR techniques (COSY, TOCSY, ROESY, HMQC, and HMBC), the structure of the products is established as 13-[(β -D -galactopyranosyl-(1 → 4)-β-D -glucopyranosyl-(1 → 2)- β -D -glucopyranosyl)oxy]kaur-16-en-19-oic acid β -D -glucopyranosyl ester ( 3 ) and 13-[(β-D -galactopyranosyl-(1 → 4)- β-D -glu-copyranosyl-(1 → 2)- β-D -glucopyranosyl)oxy]kaur-16-en-19-oic acid ( 4 ).     

The 2-Amino Group of 8-Aza-7-deaza-7-bromopurine-2,6-diamine and Purine-2,6-diamine as Stabilizer for the Adenine–Thymine Base Pair in Heterochiral DNA with Strands in Anomeric Configuration

Stabilization of DNA is beneficial for many applications in the fields of DNA therapeutics, diagnostics, and materials science. Now, this phenomenon is studied on heterochiral DNA, an autonomous DNA recognition system with complementary strands in α-D and β-D configuration showing parallel strand orientation. The 12-mer heterochiral duplexes were constructed from anomeric (α/β-D) oligonucleotide single-strands. Purine-2,6-diamine and 8-aza-7-deaza-7-bromopurine-2,6-diamine 2′-deoxyribonucleosides having the capability to form tridentate base pairs with dT were used to strengthen the stability of the dA–dT base pair. T_m data and thermodynamic values obtained from UV melting profiles indicated that the 8-aza-7-deaza 2′-deoxyribonucleoside decorated with a bromo substituent is so far the most efficient stabilizer for heterochiral DNA. Compared with that, the stabilizing effect of the purine-2,6-diamine 2′-deoxyribonucleoside is low. Global changes of helix structures were identified by circular dichroism (CD) spectra during melting.     

Glyconothio-O-lactones. Part III. Thermolysis of a 4,5-Dihydro-1,2,3- and a 2,5-Dihydro-1,3,4-thiadiazole

Addition of CH₂N₂ to 2,3:5,6-di-O-isopropylidene-1-thio-mannono-1,4-lactone ( 1 ) gave the 2,5-dihydro-1,3,4-thiadiazole 2 and the 4,5-dihydro-1,2,3-thiadiazole 3 . First-order kinetics were observed for the thermolysis of 3 (Scheme 3) at 80–110° in C₆D₅Cl solution and of 2 (Scheme 3) at 20–35° in CDC1₃, respectively. The 1,2,3-thiadiazole 3 led to mixtures of the thiirane 9 , the starting thionolactone 1 , the thiono-1,5-lactone 8 , and the enol ether 7 , while the isomeric 1,3,4-thiadiazole 2 led to mixtures of the anomeric thiiranes 9 and 12 , the O-hydrogen S,O,O-ortholactone α-D - 14 , the S-methyl thioester 15 , the S,S,O-ortholactone 13 , and the 2,3:5,6-di-Oisopropylidene-mannono-1,4-iactone ( 16 ). Pure products of the thermolysis were isolated by semipreparative supercritical fluid chromatography (SFC), whereas preparative HPLC led to partial or complete decomposition. Thus, the β-D -mannofuranosyl β-D -mannofuranoside 10 , contaminated by an unknown S species, was isolated by preparative HPLC of the crude product of thermolysis of 3 at 115–120° and partially transformed in CD₃OD solution into the symmetric di(α-D -mannofuranosyl) tetrasulfide 11 . Its structure was evidenced by X-ray analysis. Similarly, HPLC of the thermolysis product of 2 gave the enethiol 17 , the sulfide 19 , and the mercapto alcohol 18 as secondary products. Thermolysis of the thiirane 9 at 110–120° (Scheme 4) led to the anomeric thiirane 12 which was transformed into mixtures of the enethiol 17 and the enol ether 7. Addition of H₂O to 17 and 7 gave the corresponding hemiacetals 18 and 20. The mechanism of the thermolysis of the dihydrothiadiazoles 2 and 3 , and the thiiranes 9 and 12 is discussed.     

Anomeric DNA Strand Displacement with α-D Oligonucleotides as Invaders and Ethidium Bromide as Fluorescence Sensor for Duplexes with α/β-, β/β- and α/α-D Configuration

DNA strand displacement is a technique to exchange one strand of a double stranded DNA by another strand (invader). It is an isothermal, enzyme free method driven by single stranded overhangs (toeholds) and is employed in DNA amplification, mismatch detection and nanotechnology. We discovered that anomeric (α/β) DNA can be used for heterochiral strand displacement. Homochiral DNA in β-D configuration was transformed to heterochiral DNA in α-D/β-D configuration and further to homochiral DNA with both strands in α-D configuration. Single stranded α-D DNA acts as invader. Herein, new anomeric displacement systems with and without toeholds were designed. Due to their resistance against enzymatic degradation, the systems are applicable to living cells. The light-up intercalator ethidium bromide is used as fluorescence sensor to follow the progress of displacement. Anomeric DNA displacement shows benefits over canonical DNA in view of toehold free displacement and simple detection by ethidium bromide.     

4-Methylumbelliferyl 5-Acetamido-3,4,5-trideoxy-α-D-manno-2-nonulopyranosidonic Acid: Synthesis and Resistance to Bacterial Sialidases

The synthesis of 4-methylumbelliferyl α-D -glycoside 13 of N-acetyl-4-deoxyneuraminic acid and its behaviour towards bacterial sialidases is described. N-Acetyl-4-deoxyneuraminic acid ( 1 ) was transformed into its methyl ester 2 and then acetylated to give the anomeric pentaacetates 3 and 4 of methyl 4-deoxyneuraminate and the enolacetate 5 (Scheme). A mixture 3/4 was treated with HCl/AcCl to give the glycosyl chloride, which was directly converted into the 4-methylumbelliferyl α-D -glycoside 9 of methyl 7-O,8-O,9-O,N-tetraacetylneuraminate and into the 2,3-dehydrosialic acid 11 . The ketoside 9 was de-O-acetylated to 12 with NaOMe in MeOH. Saponification (NaOH) of the methyl ester 12 followed by acidification gave the free 13 , which was also converted into the sodium salt 14 by passage through Dowex 50 (Na⁺). The 4-deoxy α-D -glycoside 13 is not hydrolyzed at significant rates by Vibrio cholerae and Arthrobacter ureafaciens sialidase. Neither the free N-acetyl-4-deoxyneuraminic acid ( 1 ), nor the α-D -glycoside 13 inhibit the activity of these sialidases.     

Anomeric spiroannelated 1,4-diazepine 2,5-diones from furano exo-glycals: towards a new class of spironucleosides

The first synthesis of 1,4-diazepine 2,5-dione peptides containing a β-amino acid in which the β carbon is also the anomeric carbon of a furanoid sugar is described. These new anomeric spirosugars obtained with a stereoselective control in the d-gulo, d-manno, d-allo and d-ribo series can be regarded as the first members of a new class of spironucleosides. In the course of our study, two symmetrical tetrameric cyclopeptides comprising two identical sugar β-amino acid and α-amino acid residues were also isolated, these structures could be of interest as new potential host molecules.     

Synthesis and Enzymatic Evaluation of Substrates and Inhibitors of β-Glucuronidases

The phosphono and the tetrazolyl analogues 4 and 5 of 4-methylumbelliferyl β-D -glucuronide (=(4-methyl-2-oxo-2H-1-benzopyran-7-yl β-D -glucopyranosid)uronic acid; 6 ) were synthesized and evaluated as substrates of β-glucuronidases. Similarly, the phenylcarbamate 7 and its phosphono analogue 8 were prepared and evaluated as inhibitors. To examine the diastereoselectivity of the phosphorylation, we also synthesized the protected L -ido-D -gluco-, and D -galacto-configurated phospha-glycopyranuronates 12, 13, 21, 22, 34 and 35 . Two strategies were followed. In the first one, the glucuronic acid 19 was decarboxylated to 11 and further transformed, via 20 , into the trichloroacetimidate 10 (Scheme 2). Phosphorylation of 10 with (MeO)₃P yielded the diastereoisomers 12 and 13 , the diastereoselectivity depending on the solvent. In MeCN, 12 and 13 were obtained in a ratio of 1:1, while in non-participating solvents the L -ido 12 was by far the major diastereoisomer. The acetate 11 was inert to (MeO)₃P, but reacted with (PhO)₃P to the anomeric mixture 21/22 , in keeping with a stabilizing 1,3-interaction in the intermediate phosphonium salt. Similarly, the phospha-galacturonates 34 and 35 were prepared from the galactoside 23 via the enol ether 26 , the lactone 27 , and the acetates 28/29 that were also transformed into the trichloroacetimidate 33 (Scheme 3). In the second, higher-yielding strategy, phosphorylation of the pentodialdehyde 39 to 40/41 was followed by hydrolysis and acetylation to the phospha-glucuronates 43/44 (Scheme 4). Transesterification to 45/46 , selective deacetylation to 48/49 , and formation of the trichloroacetimidates 50/51 were followed by glycosidation and deprotection to 4 . The tetrazole 5 was prepared from the lactones 54/55 via the N-benzylamides 57/58 that were treated with TfN₃ to give the N-benzyltetrazoles 59/60 (Scheme 4). These were transformed into the trichloroacetimidates 63/64 , glycosylated to 65 , and deprotected. The O-carbamoylhydroximo-lactone 7 derived from the glucuronate 67/68 , and the phosphonate analogue 8 were prepared by established methods. The phosphonate 4 is slowly hydrolyzed by the E. coli β-glucuronidase, but neither 4 nor the tetrazole 5 are affected by the bovine liver β-glucuronidase (Table 4). The phenylcarbamate 7 of D -glucarhydroximo-1,5-lactone, but not its phosphonate analogue 8 , is an inhibitor (K_I = 8 m?M ) of the E. coli β-glucuronidase. The bovine liver β-glucuronidase is inhibited strongly by 7 (IC₅₀ = 0.2 m?M ) and weakly by 8 (IC₅₀ = 2mM ).     

Purification,Characterization and Mechanistic Study of β-Glucosidase from Flavobacterium meningosepticum (ATCC 13253)

A β-glucosidase (EC 3.2.1.21) from Flavobacterium meningosepticum has been purified and characterized. Purity was enhanced at least 530-fold from crude cell extract with 16.6% yield. The estimated molecular mass of the purified enzyme is 150 kDa by gel filtration and 78 kDa by SDS-PAGE. This dimeric enzyme has a pI = 9.0 and an optimal activity at pH 5.0 and temperature of 50 °C. Divalent metal ions (Hg²⁺, Cu²⁺, Ca²⁺, Mg²⁺) and EDTA have negligible effect on the enzyme activity. The enzyme exhibited a high specificity on the glycon portion of aryl-β-D -glycosides. NMR spectroscopy revealed the enzyme catalyzed hydrolysis of p-nitrophenyl-β-D -glucopyranoside with the retention of anomeric configuration, indicating that a double displacement mechanism was involved. A preliminary study of the Bronsted relationship showed a concave-downward plot, which is consistent with the two-step mechanism.     

Molluscicidal Saponins from Talinum tenuissimum DINTER

Two new saponins, β-D -glucopyranosyl 3-O[O-βD -xylopyranosyl-(1→3)-O-(β-D -glucopyranosyluronic acid)]oleanolate ( 1 ) and 3-O-[O-β-D -xylopyranosyl-(1→3)-O-(β-D-glucopyranosyluronic acid)]oleanolic acid ( 2 ), have been isolated from the tubers of Talinum tenuissimum. The structures have been established mainly by ¹³C-NMR and FAB-MS. The monodesmosidic saponin 2 exhibits very strong molluscicidal activity against the schistosomiasis-transmitting snail Biomphalaria glabrata.     

Stereoselective Synthesis of Indazole 2′-Deoxy-β-D-ribonucleosides: Glycosylation of the Nucleobase Anion

The glycosylation of indazolyl anions derived from 4a , b with 2-deoxy-3,5-bis-O-(4-methylbenzoyl)-α-D -erythro-pentofuranosyl chloride ( 5 ) is described. The reaction was Stereoselective – exclusive β-D -anomer formation – but regioisomeric N¹- and N²-(2′-deoxy-β-D -ribofuranosides) (i.e. 6a and 7a , resp., and 6b and 7b , resp.) were formed in about equal amounts. They were deprotected to yield 8a , b and 9a , b . Compound 1 , related to 2′-deoxyadenosine ( 3 ), and its regioisomer 2 were obtained from 8b and 9b , respectively, by catalytic hydrogenation. The anomeric configuration as well as the position of glycosylation were determined by 1D NOE-difference spectroscopy. The first protonation site of 1 and 2 was found to be the NH₂ group. The N-glycosylic bond of 1H-indazole N¹-(2′-deoxyribofuranosides) is more stable than that of the parent purine nucleosides. Compound 1 is no substrate for adenosine deaminase.