- Oxidative Deamination of N-Acetyl Neuraminic Acid: Substituent Effects and Mechanism
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A study of the mechanism of the oxidative deamination of the N-nitroso-N-acetyl sialyl glycosides leading with overall retention of configuration to the corresponding 2-keto-3-deoxy-d-glycero-d-galacto-nonulopyranosidonic acid (KDN) glycosides is described, making use of a series of differentially O-protected N-nitroso-N-acetyl sialyl glycosides and of isotopic labeling studies. No evidence is found for stereodirecting participation by ester groups at the 4- and 7-positions. Comparisons are drawn with oxidative deamination reactions of 4-amino-4-deoxy and 2-amino-2-deoxy hexopyranosides and a common mechanism is formulated involving the intermediacy of 1-oxabicyclo[3.1.0]hexyl oxonium ions following participation by the pyranoside ring oxygen. A minor reaction pathway has been uncovered by labeling studies in the β-thiosialosides that results in the exchange of the 4-O-acetyl group by the glacial acetic acid that serves as external nucleophile in the general oxidative deamination process. A mechanism is proposed for this exchange involving participation by the thioglycoside at the level of an intermediate diazoalkane.
- Buda, Szymon,Crich, David
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- FORMATION AND IDENTIFICATION OF TWO NOVEL ANHYDRO COMPOUNDS OBTAINED BY METHANOLYSIS OF N-ACETYLNEURAMINIC ACID AND CARBOXYL-REDUCED, MENINGOCOCCAL B POLYSACCHARIDE
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Methanolysis of N-acetylneuraminic acid gives, in addition to the methyl ester methyl β-ketoside, 10-15percent of a product identified by g.l.c.-m.s. as methyl 5-acetamido-2,7-anhydro-3,5-dideoxy-α-D-glycero-D-galacto-nonulopyranosonate (3).Only the major product was formed on methanolysis of the capsular, sialic acid polysaccharides from Neisseria meningitidis serogroup B and C and Escherichia coli K1 (colominic acid).Methanolysis of carboxyl-reduced, meningococcal B polysaccharide affords a major product, identified by g.l.c.-m.s. as methyl 5-acetamido-1,7-anhydro-3,5-dideoxy-β-D-glycero-D-galacto-nonulopyranoside (7).Mass-spectral data for 3 and 7 together with those of known sialic acid derivatives correlate well with previously observed fragmentation patterns.
- Lifely, Robert M.,Cottee, Frank H.
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- Synthesis of C-9 oxidised N-acetylneuraminic acid derivatives as biological probes
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Sialic acids are 9-carbon acidic sugars involved in a number of important biological processes and human diseases. As part of our ongoing interest in the development of novel sialic acids as biological probes, we have developed an efficient and simple synthesis of C-9 oxidised sialic acid derivatives. The key oxidative step involves the use of TEMPO under carefully controlled aqueous pH conditions.
- Kiefel, Milton J.,Chopra, Pradeep,Madge, Paul D.,Szyczew, Alex,Thomson, Robin J.,Grice, I. Darren,Von Itzstein, Mark
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- Design, synthesis, functional and structural characterization of an inhibitor of N-acetylneuraminate-9-phosphate phosphatase: Observation of extensive dynamics in an enzyme/inhibitor complex
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The design, synthesis and characterization of a phosphonate inhibitor of N-acetylneuraminate-9-phosphate phosphatase (HDHD4) is described. Compound 3, where the substrate C-9 oxygen was replaced with a nonlabile CH2 group, inhibits HDHD4 with a binding affinity (IC50 11 μM) in the range of the native substrate Neu5Ac-9-P (compound 1, Km 47 μM). Combined SAR, modeling and NMR studies are consistent with the phosphonate group in inhibitor 3 forming a stable complex with native Mg2+. In addition to this key interaction, the C-1 carboxylate of the sugar interacts with a cluster of basic residues, K141, R104 and R72. Comparative NMR studies of compounds 3 and 1 with Ca2+ and Mg2+ are indicative of a highly dynamic process in the active site for the HDHD4/Mg2+/3 complex. Possible explanations for this observation are discussed.
- Kim, Soong-Hoon,Constantine, Keith L.,Duke, Gerald J.,Goldfarb, Valentina,Hunt, John T.,Johnson, Stephen,Kish, Kevin,Klei, Herbert E.,McDonnell, Patricia A.,Metzler, William J.,Mueller, Luciano,Poss, Michael A.,Fairchild, Craig R.,Bhide, Rajeev S.
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- 13C-labeled N-acetyl-neuraminic acid in aqueous solution: Detection and quantification of acyclic keto, keto hydrate, and enol forms by 13C NMR spectroscopy
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Aqueous solutions of N-acetyl-neuraminic acid (Neu5Ac, 1) labeled with 13C at C1, C2, and/or C3 were analyzed by 13C NMR spectroscopy to detect and quantitfy the acyclic forms (keto, keto hydrate, enol) present at varying pHs. In addition to pyranoses, solutions contained the keto form, based on the detection of C2 signals at ~198 ppm (~0.7% at pH 2). Spectra of [2-13C] and [3-13C] isotopomers contained signals arising from labeled carbons at ~143 and ~120 ppm, respectively, which were attributed to enol forms. Solution studies of [1,2,3- 13C3]I substantiated the presence of enol (~0.5% at pH 2). Enol was not detected at pH > 6.0. A C2 signal observed at ~94 ppm was identified as C2 of the keto hydrate (~1.9% at pH 2), based partly on its abundance as a function of solution pH. Density functional theory (DFT) calculations were used to study the effect of enol and hydrate structure on JCH and JCC values involving C2 and C3 of these forms. Solvated DFT calculations showed that 2JC2,H3 in cis and trans enols have similar magnitudes but opposite signs, making this J-coupling potentially useful to distinguish enol configurations. Solvent deuterium exchange studies of 1 showed rapid incorporation of 2H from 2H2O at H3axial in the pyranoses at p 2H 8.0, followed by slower exchange at H3equatorial. The acyclic keto form, which presumably participates in this reaction, must assume a pseudo-cyclic conformation in solution in order to account for the exchange selectivity. Weak 13C signals arising from labeled species were also observed consistently and reproducibly in aqueous solutions of 13C-labeled 1, possibly arising from products of lactonization or intermolecular esterification.
- Klepach, Thomas,Carmichael, Ian,Serianni, Anthony S.
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- Semi-Synthetic Sialic Acid Probes for Challenging the Substrate Promiscuity of Enzymes in the Sialoconjugation Pathway
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A series of unusual sialic acid analogs were prepared using a semi-synthetic strategy. Truncation of natural N-acetylneuraminic acid was followed by diastereoselective carbon backbone reconstruction using Barbier-type carboligations as well as different functional group interconversions, which provided access to a variety of functional motifs in the terminal carbon backbone, including examples of saturated and unsaturated, linear and branched alkyl chains, partially deoxygenated sialic acids, sialic diacids and a first truncated legionaminic acid analog. The synthetic sialic acid probes were studied for nucleotide activation by the CMP-sialic acid synthetase from Neisseria meningitidis using a universal pH-shift assay for kinetic analysis. One-pot enzymatic nucleotide activation and sialyltransfer to lactose was performed using a selection of five probes together with an engineered α2,3-sialyltransferase from Photobacterium phosphoreum to furnish five new-to-nature analogs of the GM3 trisaccharide, which were finally utilized to test the substrate tolerance of two bacterial sialidases. The obtained set of sialic acid analogs and neo-sialocojugates provides interesting opportunities for further glycobiology studies. (Figure presented.).
- Mertsch, Alexander,Poschenrieder, Silvan,Fessner, Wolf-Dieter
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supporting information
p. 5485 - 5495
(2020/10/22)
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- Novel method for synthesizing 4, 7-dimethoxy-N-acetylneuraminic acid-fluorescein and its intermediate
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The invention discloses a novel method for synthesizing 4, 7-dimethoxy-N-acetylneuraminic acid-fluorescein. The method comprises that the 4, 7-dimethoxy-N-acetylneuraminic acid-fluorescein is synthesized from 2-cyano-6-hydroxybenzothiazole as a raw material. The method improves a yield to 60-70% and has the compound production capacity to the gram level. Compared with the existing method, the novel method greatly improves a yield, is free of expensive fluorescein raw materials and greatly reduces a production cost.
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Paragraph 0036-0037
(2018/05/07)
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- A facile microwave-assisted protocol for rapid synthesis of N-acetylneuraminic acid congeners
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We developed a simple, rapid and efficient microwave irradiation-assisted protocol that is 1- to 2-orders of magnitude faster than conventional techniques, providing an expedient access to the sialic acid congeners Neu5Ac1Me (1), Neu5Acβ1,2Me2 (2), Neu5Ac1Me O-peracetate (3) and 4,5-oxazoline of Neu5Ac2en1Me O-peracetate (4).
- Saludes, Jonel P.,Sahoo, Dhananjaya,Monreal, I. Abrrey
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supporting information
p. 507 - 510
(2014/02/14)
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- Tuning mechanism-based inactivators of neuraminidases: Mechanistic and structural insights
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3-Fluorosialosyl fluorides are inhibitors of sialidases that function by the formation of a long-lived covalent active-site adduct and have potential as therapeutics if made specific for the pathogen sialidase. Surprisingly, human Neu2 and the Trypanosoma cruzi trans-sialidase are inactivated more rapidly by the reagent with an equatorial fluorine at C3 than by its axial epimer, with reactivation following the same pattern. To explore a possible stereoelectronic basis for this, rate constants for spontaneous hydrolysis of the full series of four 3-fluorosialosyl fluorides were measured, and ground-state energies for each computed. The alpha (equatorial) anomeric fluorides hydrolyze more rapidly than their beta anomers, consistent with their higher ground-state energies. However ground-state energies do not explain the relative spontaneous reactivities of the 3-fluoro-epimers. The three-dimensional structures of the two 3-fluoro-sialosyl enzyme intermediates of human Neu2 were solved, revealing key stabilizing interactions between Arg21 and the equatorial, but not the axial, fluorine. Because of changes in geometry these interactions will increase at the transition state, likely explaining the difference in reaction rates. Understanding reactivity and selectivity: The mechanistic basis for the surprisingly different rates of inactivation and reactivation of human and Trypanosoma cruzi sialidases by the two 3-fluoro epimers of 2,3-difluorosialic acid was probed through spontaneous hydrolysis kinetics, computational analysis, and X-ray crystallography.
- Buchini, Sabrina,Gallat, Francois-Xavier,Greig, Ian R.,Kim, Jin-Hyo,Wakatsuki, Soichi,Chavas, Leonard M. G.,Withers, Stephen G.
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supporting information
p. 3382 - 3386
(2014/04/03)
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- Chemical insight into the emergence of influenza virus strains that are resistant to Relenza
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A reagent panel containing ten 4-substituted 4-nitrophenyl α-d-sialosides and a second panel of the corresponding sialic acid glycals were synthesized and used to probe the inhibition mechanism for two neuraminidases, the N2 enzyme from influenza type A virus and the enzyme from Micromonospora viridifaciens. For the viral enzyme the logarithm of the inhibition constant (Ki) correlated with neither the logarithm of the catalytic efficiency (kcat/Km) nor catalytic proficiency (kcat/Kmkun). These linear free energy relationship data support the notion that these inhibitors, which include the therapeutic agent Relenza, are not transition state mimics for the enzyme-catalyzed hydrolysis reaction. Moreover, for the influenza enzyme, a correlation (slope, 0.80 ± 0.08) is observed between the logarithms of the inhibition (Ki) and Michaelis (Km) constants. We conclude that the free energy for Relenza binding to the influenza enzyme mimics the enzyme-substrate interactions at the Michaelis complex. Thus, an influenza mutational response to a 4-substituted sialic acid glycal inhibitor can weaken the interactions between the inhibitor and the viral neuraminidase without a concomitant decrease in free energy of binding for the substrate at the enzyme-catalyzed hydrolysis transition state. The current findings make it clear that new structural motifs and/or substitution patterns need to be developed in the search for a bona fide influenza viral neuraminidase transition state analogue inhibitor.
- Shidmoossavee, Fahimeh S.,Watson, Jacqueline N.,Bennet, Andrew J.
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supporting information
p. 13254 - 13257
(2013/09/24)
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- Rapid and clean microwave-assisted synthesis of N-acetylneuraminic acid methyl ester and its β-methyl glycoside
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The effect of microwave irradiation on the synthesis of N-acetylneuraminic acid methyl ester and its β-methyl glycoside is investigated. On a 1 g scale, a high yield (94%) of the methyl ester was obtained after 15 min at 80 °C. Acceleration of the glycosidation reaction was achieved, with the β-methyl glycoside isolated in good yield following optimisation of reaction conditions to 15 min at 12°C.
- Chopra, Pradeep,Thomson, Robin J.,Grice, I. Darren,Von Itzstein, Mark
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p. 6254 - 6256,3
(2012/12/11)
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- Rapid and clean microwave-assisted synthesis of N-acetylneuraminic acid methyl ester and its β-methyl glycoside
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The effect of microwave irradiation on the synthesis of N-acetylneuraminic acid methyl ester and its β-methyl glycoside is investigated. On a 1 g scale, a high yield (94%) of the methyl ester was obtained after 15 min at 80 °C. Acceleration of the glycosidation reaction was achieved, with the β-methyl glycoside isolated in good yield following optimisation of reaction conditions to 15 min at 12°C.
- Chopra, Pradeep,Thomson, Robin J.,Grice, I. Darren,Von Itzstein, Mark
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p. 6254 - 6256
(2013/01/15)
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- Solution phase synthesis of (1→5)-amide linked sugar amino acid dimers derived from sialic acids
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Carboxy-protected amino derivatives and amino-protected carboxy derivatives of three different C-2 analogs as well as a 2,3-dehydro NeuAc were prepared. These monomers were coupled in solution using BOP activation of the carboxy terminus to form (1→5)-amide linked dimers of sialyl amino acid derivatives.
- Gervay, Jacquelyn,Flaherty, Terrence M.,Nguyen, Can
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p. 1493 - 1496
(2007/10/03)
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- Synthesis of 13C enriched sialyllactones and their characterization using isotope edited inverse detected NMR spectroscopy
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13C enriched monosaccharide based sialyllactones were prepared in order to perform isotope edited NMR experiments for their characterization. In the inverse detected experiments only those protons long-range coupled to the enriched nuclei are detected, providing a potentially powerful tool for studying in vitro sialyllactone formation.
- Gervay, Jacquelyn,Mamuya, Nellie N.,Barber, Andrew R.
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p. 1865 - 1868
(2007/10/03)
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- Ring opening of sialyllactones with glycine esters: Synthesis of selectively protected glycinyl-NeuAc saccharopeptides
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Two different classes of sialyllactones were prepared as potential substrates for ring opening reactions with naturally occurring amino acids. The sialyllactones underwent reaction with Glycine ethyl ester hydrochloride salt to afford selectively protected glycine-NeuAc adducts. The reactions could be performed on NeuAc derivatives capable of serving as glycosylation donors. These compounds represent a new class of saccharopeptides, composed of sugar amino acids and naturally occurring amino acids.
- Gervay, Jacquelyn,Ramamoorthy,Mamuya, Nellie N.
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p. 11039 - 11048
(2007/10/03)
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- De novo Synthesis of Carbohydrates and Related Natural Products, 34 Synthesis of N-Acetyl-β-D-neuraminic Acid Derivatives via Inverse-Type Hetero-Diels-Alder Reaction
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Inverse-type hetero-Diels-Alder reaction-based diastereoselective synthesis of 3-deoxy-2-glyculosonates is performed with the help of chiral carbon substituents in the 2-position of the 1-oxa-1,3-diene required as heterodiene.This is demonstrated for the
- Haag-Zeino, Brigitte,Schmidt, Richard R.
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p. 1197 - 1203
(2007/10/02)
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- General methods for modification of sialic acid at C-9. Synthesis of N-acetyl-9-deoxy-9-fluoroneuraminic acid.
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Methyl 5-acetamido-3,5-dideoxy-2-O-methyl-D-glycero-D-galacto-2-nonulopyrano sate was converted into the 9-O-trityl derivative and the remaining hydroxyl groups were protected as benzyl ethers. Removal of the trityl group, followed by treatment with dieth
- Sharma,Petrie 3rd.,Korytnyk
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- Stereoselective total syntheses of the naturally occurring enantiomers of N-acetylneuraminic acid and 3-deoxy-D-manno-2-octulosonic acid. A new and stereospecific approach to sialo and 3-deoxy-D-manno-2-octulosonic acid conjugates
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The total syntheses of the title compounds have been achieved. A critical element of these syntheses was the concept of using a furan ring as a surrogate for a carboxylic acid. The furyl diene 22 reacted with the R and S enantiomers of 2-(phenylseleno)pro
- Danishefsky,DeNinno,Chen
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p. 3929 - 3940
(2007/10/02)
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- SYNTHESES OF 2-O-GLYCOSYL DERIVATIVES OF N-ACETYL-D-NEURAMINIC ACID
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Syntheses of N-acetyl-D-neuraminic acid derivatives are reported.Methyl 4,7,8,9-tetra-O-acetyl-N-acetyl-2-chloro-2-deoxy-β-D-neuraminate (3) was prepared directly from methyl N-acetyl-β-D-neuraminate (2) in good yield.Koenigs-Knorr reaction of 3 with an excess of methanol gave the methyl α-glycoside of methyl N-acetyl-D-neuraminate (4). 2,3-O-Isopropylidene-D-ribono-1,4-lactone, 2,3-O-isopropylideneuridine, and 5-fluor-2,3-O-isopropylideneuridine reacted with 3 to give anomeric mixtures of methyl N-acetyl-D-neuraminate derivatives.The stereochemistry of these compounds was confirmed from n.m.r. and c.d. spectra, and measurements of the rate of hydrolysis of the glycosidic bond.
- Ogura, Haruo,Furuhata, Kimio,Itoh, Masayoshi,Shitori, Yoshiyasu
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- Transformations with N-Acetylneuraminic Acid, 3. Synthesis of 7-epi-, 8-epi- and 7,8-bis-epi-N-Acetylneuraminic Acid and their Behaviour Towards the Cytidin Monophosphate Sialic Acid Synthetase
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From methyl-5-acetylamino-7,8-anhydro-4,9-O-bis-(t-butyldimethylsilyl)-3,5-dideoxy-β-D-glycero-D-galacto-2-nonulopyranosidonic acid methylester (1) the derivatives 1a and 1b were obtained by removing the 9-O-(t-butyldimethylsilyl)group with Bu4NF, followed by acetylation.Treatment of 1b with 80percent acetic acid and acetanhydride/pyridine yields the 8-epi-N-acetylneuraminic acid derivative 2a and the 7-epi-N-acetylneuraminic acid derivative 3a in a ratio 3:1 (Scheme 1).The structure elucidation of 2b was achieved by converting 2b via the 4,9-bis-O-(tBDMSi)-8-O-tosyl-derivative 2d into the epoxide 1 (Scheme 2).Using the same sequence the epoxides 4 and 5 were transformed into the N-acetylneuraminic acid derivative 6a and the 7,8-bis-epi-N-acetylneuraminic acid derivative 7a (Scheme 3).After treatment with sodium hydroxide and 0.025 m HCl and Dowex 50H(+) the 8-epi-, 7-epi- and 7,8-bis-epi-N-acetylneuraminic acid 2, 3, and 7 were obtained.These three compounds were tested with CMP-N-acetylneuraminic acid synthetase. - Keywords: Acetolysis of methyl-9-O-acetyl-5-acetylamino-7,8-anhydro-4-O-(tBDMSi)-3,5-dideoxy-β-D-(and L)-glycero-D-galacto-2-nonulosidonic acid methylester, methyl-9-O-acetyl-5-acetylamino-7,8-anhydro-4-O-(tBDMSi)-3,5-dideoxy-β-D-glycero-L-altro-2-nonulosidonic acid methylester; Configurational changes of sialic acid
- Zbiral, Erich,Brandstetter, Hannelore H.
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- STUDIES ON SIALIC ACIDS I. DETERMINATION OF ANOMERIC CONFIGURATION OF NEURAMINIC ACID DERIVATIVES BY CIRCULAR DICHROISM
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CD spectra were recorded for methyl α- and β-glycosides of D-neuraminic acid, and the band at the wave-length lower than 200 nm was attributed to the acetamido group.The Cotton effect at higher wave-length around 220 nm arose from the n-?* transition of the carboxyl group.Thus α-linked glycosides showed a negative band, while β-glycosides gave arise to a positive band.
- Ogura, Haruo,Furuhata, Kimio
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p. 4265 - 4268
(2007/10/02)
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