Isolation and Structural Determination of Reducing Fructooligosaccharides Newly Produced in Stored Edible Burdock

Fresh edible burdock roots were stored in soil of 1 m depth underground from November to May. Three fructooligosaccharide derivatives without a terminal glucose residue, designated saccharides 1, 2 and 3, were generated in the stored burdock roots. They were purified from the sugar extract using carbon-Celite column chromatography. Saccharides 1, 2 and 3 have R-sucrose values (retention time of sucrose = 1) of 1.55, 2.15 and 2.73 by HPAEC, reducing terminal, molar ratios (reducing sugar to D-fructose) of 0.50, 0.33 and 0.25 and degrees of polymerization of 2, 3 and 4 by TOF-MS, respectively. Analyses by GLC and NMR con- firmed the three different following structures: first was inulobiose (-D-fructofuranosyl-(2↑1)--D- fructopyranose), and the two others were inulotriose (-D-fructofuranosyl-(2↑1)--D-fructofuranosyl-(2↑1)-- D-fructopyranose) and inulotetraose (-D-fructofuranosyl-(2↑1)--D-fructofuranosyl-(2↑ 1)- -D-fructo- furanosyl-(2↑1)--D-fructopyranose). The NMR spectra showed that 70 to 80% of the terminal fructose resi- due of the three saccharides is pyranosyl form, while 20 to 30% is furanosyl form. The 13 C- and 1 H-signals were also assigned by 2D-NMR including COSY, HSQC, HSQC-TOCSY and HMBC. These saccharides could be synthesized by purified burdock fructan:fructan 1-fructosyltransferase from 1-kestose to free D- fructopyranose giving inulobiose and sucrose, while elongation of fructofuranosyl units occurred at this trans- ferred fructofuranosyl residue to produce inulooligosaccharides having one, two and more additional units of fructofuranose.