Abstract
Intestinal immune system modulating arabino-3,6-galactan (ALR-5IIa-1-1) has been purified from rhizomes of Atractylodes lancea DC. In order to characterize the structure of ALR-5IIa-1-1 sequential enzymic digestion using exo-α- l-arabinofuranosidase and exo-β- d-(1→3)-galactanase was employed. After ALR-5IIa-1-1 was digested with the arabinofuranosidase to give AF-ALR-5IIa-1-1, the galactanase digestion cleaved only 10% and 23% of 3-linked and 3,6-branched Gal in AF-ALR-5Iia-1-1, respectively, and gave small amounts of intermediate size (AF-GN-2) and shorter oligosaccharide (AF-GN-3) fractions in addition to a large amount of the galactanase-resistant fraction (AF-GN-1). When AF-GN-1 was re-digested gradually with the arabinofuranosidase and galactanase, it released trace amounts of oligosaccharides in addition to a large amount of the enzyme-resistant fraction. When the final enzyme-resistant fraction from AF-GN-1 was digested simultaneously with both arabinofuranosidase and galactanase, the resistant fraction was significantly degraded into two long fragments (AF 3-GN 3-1 and 2). Mixture of digestion products from the first galactanase digestion of AF-ALR-5IIa-1-1 showed decreased intestinal immune system modulating activity which was correspond to about 30% of the activity of ALR-5IIa-1-1, but the galactanase resistant fraction (AF-GN-1) still had significant activity. Although the second gradual enzymic digestion of AF-GN-1 caused little decrease in activity, the resulting fragments (AF 3-GN 3-1 and 2) from the final enzymic digestion lost most of the activity. Component sugar, methylation and MALDI-TOF-MS analyses indicated that the oligosaccharides released from AF-ALR-5IIa-1-1 by the first galactanase digestion consisted of galactosyl di- to hexadecasaccharides mainly comprising 6-linked Gal p and Gal f some of which were partially mono- or diarabinosylated. These oligosaccharides were attached to the non-reducing terminal side of β- d-(1→3)-galactan backbone as side chains. The final fragments (AF 3-GN 3-1 and 2) were located in the inner core of ALR-5IIa-1-1, and AF 3-GN 3-2 was composed mainly of terminal, 3-linked and 6-linked Gal whereas AF 3-GN 3-1 was suggested to have a complex structure
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