Ján Hirsch

Hydration properties of xylan‐type structures: an FTIR study of xylooligosaccharides

An FTIR study of the effect of hydration as a function of relative humidity on the structurally different xylooligosaccharide models is presented. The IR spectral pattern changes are strongly affected by the type and position of the glycosidic linkages and reflects the hydration properties of the models.

13C-n.m.r. spectra of xylo-oligosaccharides and their application to the elucidation of xylan structures

Abstract 13 C-N.m.r. spectra of thirteen xylo-oligosaccharides [a complete series of α- and β- d -xylopyranosyl derivatives of methyl α- d -xylopyranoside, β- d -xylopyranosyl derivatives of methyl 4- O -β- d -xylopyranosyl- d -xylopyranoside, methyl O -α- d -xylopyranosyl-(1→3)- O -β- d -xylopyranosyl-(1→4)- d -xylopyranoside, and a branched methyl β-xylotetraoside] have been interpreted. The data obtained have been used for the carbon signal assignment in the spectra of a number of red-algal xylans. 13 C-N.m.r. spectroscopy is shown to be a rapid and convenient method for the structural analysis of xylose-rich polysaccharides.

Sequential synthesis and 13C-N.m.r. spectra of methyl β-glycosides of (1→4)-β-d-xylo-oligosaccharides

Abstract The reaction of 2,3-di- O -acetyl-4- O -benzyl-α,β- d -xylopyranosyl bromide ( 2 ) with methyl 2,3-di- O -acetyl-β- d -xylopyranoside gave methyl O -(2,3-di- O -acetyl-4- O -benzyl-β- d -xylopyranosyl)-(1→4)-2,3-di- O -acetyl-β- d -xylopyranoside ( 22 ). Catalytic hydrogenolysis of 22 exposed HO-4′ which was then condensed with 2 . This sequence of reactions was repeated three more times to afford, after complete removal of protecting groups, a homologous series of methyl β-glycosides of (1→4)-β- d -xylo-oligosaccharides. 13 C-N.m.r. spectra of the synthetic methyl β-glycosides (di- to hexa-saccharide) are presented together with data for six other, variously substituted, homologous series of (1→4)- d -xylo-oligosaccharides.

Production, Refining, Structural Characterization and Fermentability of Rice Husk Xylooligosaccharides

Oligosaccharides produced by hydrothermal processing of rice husks (xylooligosaccharides and glucooligosaccharides) were refined by membrane processing (operating in diafiltration and concentration modes), subjected to xylanase treatment to reduce the average molar mass, and subjected to further purification by ultrafiltration (operating in concentration mode) and ion exchange. The purified products were assayed for composition, molar mass distribution and structural characterization by HPLC, HPAEC-PAD, HPSEC, MALDI-TOF-MS and NMR ((1)H and (13)C). The fermentability of the purified product by fecal inocula was assessed on the basis of the time courses of pH and oligosaccharide concentrations. Succinate, lactate, formiate, acetate, propionate and butyrate were the major products resulting from fermentation experiments.

An approach to the systematic synthesis of (1→4)-β-d-xylo-oligosaccharides☆

Abstract The first approach to a general synthesis of (1→4)-β- d -xylo-oligosaccharides is described. Thus, 1,2,3-tri- O -acetyl-4- O -benzyl-β- d -xylopyranose was treated with hydrogen bromide in dichloromethane to give the corresponding glycosyl bromide 3 , which was condensed with 1,2,3-tri- O -acetyl-β- d -xylopyranose. The β-linked disaccharide formed, bearing at O-4 of its non-reducing end the selectively removable benzyl group, was submitted to hydrogenolysis to afford a disaccharide nucleophile 21 , having HO-4′ free, which was again treated with 3 . This sequence of reactions was repeated two more times and the isolated products 17–20 were deprotected to give a homologous series of lower xylo-oligosaccharides (from the disaccharide to the pentasaccharide), demonstrating thus the feasibility of stepwise construction of xylo-oligosaccharide molecules. The corresponding α-linked oligosaccharides 5–8 , present in the mixtures from the condensation reactions, were also isolated and characterized. 13 C-N.m.r. spectral characteristics for six homologous series of variously substituted (1→4)- d -xylo-oligosaccharides are presented.

Carbohydrate esterases of family 2 are 6-O-deacetylases

Three acetyl esterases (AcEs) from the saprophytic bacteria Cellvibrio japonicus and Clostridium thermocellum, members of the carbohydrate esterase (CE) family 2, were tested for their activity against a series of model substrates including partially acetylated gluco‐, manno‐ and xylopyranosides. All three enzymes showed a strong preference for deacetylation of the 6‐position in aldohexoses. This regioselectivity is different from that of typical acetylxylan esterases (AcXEs). In aqueous medium saturated with vinyl acetate, the CE‐2 enzymes catalyzed transacetylation to the same position, i.e., to the primary hydroxyl group of mono‐ and disaccharides. Xylose and xylooligosaccharides did not serve as acetyl group acceptors, therefore the CE‐2 enzymes appear to be 6‐O‐deacetylases.

Stereoselective synthesis and 13C-N.m.r. spectra of two isomeric methyl β-glycosides of trisaccharides related to arabinoxylan

Abstract Methyl 2-O-acetyl-4-O-(2,3,4-tri-O-acetyl-β- d -xylopyranosyl)-β- d -xylopyranoside (4) and methyl 2,3-di-O-acetyl-4-O-(2,4-di-O-acetyl-β- d -xylopyranosyl)-β- d -xylopyranoside were separately condensed with 2,3,5-tri-O-benzoyl-α- l -arabinofuranosyl bromide under modified Koenigs-Knorr conditions, to give high yields ( > 90%) of trisaccharide derivatives containing a (1→3)-linked α- l -arabinofuranosyl residue. Removal of the protecting groups gave methyl 3-O- and 3′-O-α- l -arabinofuranosyl-β- d -xylobioside. 13C-N.m.r. data for these compounds are presented and the homo- and hetero-nuclear 2D-n.m.r. spectra of 4 are discussed.

Improved synthesis of an aldobiouronic acid related to hardwood xylans, and preparation of a derivative thereof suitable for linking to proteins

Treatment of 1,3,4-tri-O-acetyl-alpha-D-xylopyranose with methyl 2,3-di-O-benzyl-l-chloro-l-deoxy-4-O-methyl-alpha, beta-D-glucopyranuronate in the presence of silver trifluoromethanesulfonate was highly stereoselective to give the alpha-linked aldobiouronic acid derivative (4) in 86% yield, after hydrogenolysis of the crude product of the coupling and chromatography. Compound 4 was acetylated and the fully protected substance was converted to the corresponding glycosyl chloride. Reaction of the latter with p-nitrophenol under phase-transfer catalysis afforded, after deacetylation, p-nitrophenyl 2-O-(methyl 4-O-methyl-alpha-D-glucopyranosyluronate)-beta-D-xylopyranoside.

Functional Cloning and Expression of the Schizophyllum commune Glucuronoyl Esterase Gene and Characterization of the Recombinant Enzyme

The gene encoding Schizophyllum commune glucuronoyl esterase was identified in the scaffold 17 of the genome, containing two introns of 50u2009bp and 48u2009bp, with a transcript sequence of 1179u2009bp. The gene was synthesized and cloned into Pichia pastoris expression vector pGAPZα to achieve constitutive expression and secretion of the recombinant enzyme in soluble active form. The purified protein was 53u2009kD with glycosylation and had an acidic pI of 3.7. Activity analysis on several uronic acids and their derivatives suggests that the enzyme recognized only esters of 4-O-methyl-D-glucuronic acid derivatives, even with a 4-nitrophenyl aglycon but did not hydrolyze the ester of D-galacturonic acid. The kinetic values were K m 0.25u2009mM, V max 16.3u2009μM·min−1, and k cat 9.27u2009s−1 with 4-nitrophenyl 2-O-(methyl 4-O-methyl-α-D-glucopyranosyluronate)-β-D-xylopyranoside as the substrate.

Hydrogenolysis of 3,5-0-benzylidene acetals with the LiAlH4-AlCl3 reagent in methyl d-xylofuranosides

Abstract The hydrogenolysis of methyl 3,5-0-benzylidene-α- and -β-D-xylofuranoside derivatives with the LiAlH4-AlCl3 reagent gave 5-benzyl ethers as main products. In some cases the attack of the reagent occured at the ring oxygen of the furenoside skeleton to yield 5-0-benzyl-1-0-methylxylitol derivatives. The structure of the synthesized compounds was proved by 13C-NMR spectroscopy. Unambiguous assignment of lines in the 13C-NMR spectra of numerous partially methylated methyl α-and β- d -xylofuranoside derivatives has been made.