Akira Yamamori

Two Novel Oligosaccharides Formed by 1F-Fructosyltransferase Purified from Roots of Asparagus (Asparagus officinalis L.)

Two novel oligosaccharides, tetra-and penta-saccharides were synthesized by fructosyl transfer from 1-kestose to 4G-β-D-galactopyranosylsucrose with a purified 1F-fructosyltransferase of asparagus roots and identified as 1F-β-D-fructofuranosyl-4G-β-D-galactopyranosylsucrose, O-β-D-fructofuranosyl-(2→1)-β-D-fructofurano-syl-O-[β-D-galactopyranosyl-(1→4)]-α-D-glucopyranoside and 1F(1-β-D-fructofuranosyl)2-4G-β-D-galactopyranosylsucrose, [O-β-D-fructofuranosyl-(2→1)]2-β-D-fructofuranosyl-O-[β-D-galactopyranosyl-(1→4)]-α-D-glucopyranoside, respectively. Both oligosaccharides were scarcely hydrolyzed by carbohydrase from rat small intestine. Human intestinal bacterial growth by 1F-β-D-fructofuranosyl-4G-β-D-galactopyranosylsucrose was compared with that by the tetrasaccharides, stachyose and nystose. Bifidobacteria utilized 1F-β-D-fructofuranosyl-4G-β-D-galactopyranosylsucrose to the same extent as stachyose or nystose. On the other hand, the unfavorable bacteria, Clostridium perfringens, Escherichia coli and Enterococcus faecalis, that produce mutagenic substances did not use the synthetic oligosaccharide.

NMR analysis of tri- and tetrasaccharides from asparagus

Detailed analysis of the 1H and 13C NMR spectra of five fructo‐oligosaccharides (1–5) isolated from Asparagus officinalis L. (Liliaceae) was carried out. Tetrasaccharides 1, 2 and 3 consist of one unit of glucose and three units of fructose, whereas trisaccharides 4 and 5 consist of one unit of glucose and two units of fructose. The 1H NMR signals of these tetrasaccharides were severely overlapped around δH 3.6–4.0 ppm and the 13C NMR signals of the corresponding signals of three fructose residues were close to each other. Therefore, it was difficult to make assignments of those signals using conventional NMR methods. Overcoming these signal‐overlapping problems required the application of some special NMR techniques. The spectral analysis was started from the anomeric proton of the glucose, since each compound had only one glucose residue and its 1H NMR spectrum showed a separate characteristic anomeric proton signal. After the 1H and 13C signals of the glucose had been assigned, the fructose residues attached to the C‐1 and/or C‐6 positions of glucose were determined from the HMBC spectrum, which also revealed further fructosyl fructose linkages. The assignments of each signal of C‐5 and C‐6 of the fructose residues overlapped in 2D spectra were achieved by the 1D INAPT method. Unresolved correlation peaks of methylene protons in HMQC could be separated by the CH2‐selected HSQC method, which gave sufficient 13C resolution to separate each CH2 signal, resulting in the determination of the chemical shift of the methylene protons. Copyright

Isolation and structural confirmation of the oligosaccharides containing α-D-fructofuranoside linkages isolated from fermented beverage of plant extracts.

Fermented beverage of plant extracts was prepared from the extracts of approximately 50 types of vegetables and fruits. Natural fermentation was carried out mainly by lactic acid bacteria (Leuconostoc spp.) and yeast (Zygosaccharomyces spp. and Pichia spp.). Two oligosaccharides containing an α-fructofuranoside linkage were detected in this beverage and isolated using carbon-Celite column chromatography and preparative HPLC. The structural confirmation of the saccharides was determined by methylation analysis, MALDI-TOF-MS, and NMR measurements. These saccharides were identified as α-D-fructofuranosyl-(2→6)-D-glucopyranose, which was isolated from a natural source for the first time, and a novel saccharide β-D-fructopyranosyl-(2→6)-α-D-fructofuranosyl-(2↔1)-α-D-glucopyranoside.

Synthesis of β-D-fructopyranosyl-(2→6)-D-glucopyranose from D-glucose and D-fructose by a thermal treatment.

The synthesis is reported of β-D-fructopyranosyl-(2→6)-D-glucopyranose that had previously been isolated from a fermented plant extract as a new saccharide. A disaccharide was predominately formed from an equal amount of D-glucose and D-fructose under melting conditions at 140 °C for 60 to 90 min. This saccharide was isolated from the reaction mixture by carbon-Celite column chromatography and preparative HPLC, and was confirmed to be β-D-fructopyranosyl-(2→6)-D-glucopyranose by TOF-MS and NMR analyses.

Structural Analysis of Two Trisaccharides Isolated from Fermented Beverage of Plant Extract

Fermented beverage of plant extract was prepared from about fifty kinds of vegetables and fruits. Natural fermentation was carried out mainly by lactic acid bacteria (Leuconostoc spp.) and yeast (Zygosaccharomyces spp.and Pichia spp.). Two novel oligosaccharides have been found from this beverage and isolated from the beverage using carbon-Celite column chromatography and preparative high performance liquid chromatography. Structure confirmation of the saccharides was provided by methylation analysis, MALDI-TOF-MS and NMR measurements. These saccharides were identified as new trisaccharides, -D-glucopyranosyl-(1 1)-D-fructofuranosyl-(2 1)-D-glucopyranoside; -Dgalactopyranosyl-(1 1)-D-fructofuranosyl-(2 1)-D-glucopyranoside.