Analysis of structural components and molecular construction of soybean soluble polysaccharides by stepwise enzymatic degradation.

Soybean soluble polysaccharides (SSPS) extracted from soybean cotyledons have a pectin-like structure. The core polysaccharides after treatments with four kinds of hemicellulases and a pectinase contained approximately equal numbers of L-rhamnose and D-galacturonate residues, suggesting the presence of the rhamnogalacturonan (RG) I structure consisting of the diglycosyl repeating unit, -4)-alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1-. The lengths of RG chains were calculated as approximately 15, 28, and 100 diglycosyl repeats. The RG components linked to each other by intervention of galacturonan (GN) chains, constituting the backbone of SSPS. All arabinose residues, which constitute 21% of total SSPS sugars, were found to be in side chains from RG regions, and this was also true for galactose residues, which constitute 50% of total sugars. Of arabinose residues, 94% are present as alpha-1,3- or alpha-1,5-arabinans, and 89% of galactose residues were present as beta-1,4-galactans. Galactan chains are modified with arabinose, xylose, fucose, and glucose at the sites close to the RG regions.     

Analysis of Structural Components and Molecular Construction of Soybean Soluble Polysaccharides by Stepwise Enzymatic Degradation

Soybean soluble polysaccharides (SSPS) extracted from soybean cotyledons have a pectin-like structure. The core polysaccharides after treatments with four kinds of hemicellulases and a pectinase contained approximately equal numbers of L-rhamnouse and D-galacturonate residues, suggesting the presence of the rhamnogalacturonan (RG) I structure consisting of the diglycosyl repeating unit, -4)-α-D-GalpA-(1→2)-α-L-Rhap-(1-. The lengths of RG chains were calculated as approximately 15, 28, and 100 diglycosyl repeats. The RG components linked to each other by intervention of galacturonan (GN) chains, constituting the backbone of SSPS. All arabinose residues, which constitute 21% of total SSPS sugars, were found to be in side chains from RG regions, and this was also true for galactose residues, which constitute 50% of total sugars. Of arabinose residues, 94% are present as α-1,3- or α-1,5-arabinans, and 89% of galactose residues were present as β-1,4-galactans. Galactan chains are modified with arabinose, xylose, fucose, and glucose at the sites close to the RG regions.     

Analysis of the molecular construction of xylogalacturonan isolated from soluble soybean polysaccharides

Soluble soybean polysaccharides (SSPS) extracted from soybean cotyledons are acidic polysaccharides, and exhibited a pectin-like structure. After digesting galacturonan with polygalacturonase, two novel galacturonan (GN) fragments, which were directly linked to xylosyl oligosaccharides, were obtained. One consisted of (beta-D-Xyl)7 branched at the C-3 site of 1,4-linked (alpha-D-GalA)4, and the other consisted of (beta-D-Xyl)4 branched at the C-3 site of 1,4-linked (alpha-D-GalA)3.     

Glucuronic acid directly linked to galacturonic acid in the rhamnogalacturonan backbone of beet pectins.

Sugar-beet pulp was de-esterified and submitted to 72 h hydrolysis by 0.1 M HCl at 80 degrees C. Oligomers containing a single glucuronic acid (GlcA) moiety in addition to n(>/= 2) repeats of the dimer -->4)-alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1--> were isolated from the hydrolysate by ion-exchange and gel-permeation. Glycosyl linkage composition analysis and 1H NMR studies indicated that the GlcA was attached to O-3 of a galacturonic acid (GalA) residue, as shown for the two pentamers beta-D-GlcpA-(1-->3)-alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1-->4)-alpha-D-GalpA-(1-->2)-L-Rhap and alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1-->4)-[beta-D-GlcpA-(1-->3)]-alpha-D-GalpA-(1-->2)-L-Rhap. Substitution by GlcA was estimated as occurring on one GalA residue out of 72 in the rhamnogalacturonan fraction of the backbone of beet pectins.     

Water-soluble polysaccharides from Ginkgo biloba leaves.

The water-soluble polysaccharides from dried Ginkgo biloba leaves were isolated after exhaustive extraction with organic solvents. The polysaccharide mixture could be separated into a neutral (GF1) and two acidic (GF2 and GF3) polysaccharide fractions by ion exchange chromatography. According to the Mr distribution GF1 and GF3 seemed to be homogenous, whereas GF2 could be further fractionated into two subfractions (GF2a and GF2b) by gel permeation chromatography. GF1 (Mr 23,000) showed the structural features of a branched arabinan. The main chain was composed of 1,5-linked arabinose residues and three in 12 arabinose molecules were branched via C-2 or C-3. GF2a (Mr 500,000) consisted mainly of 1,2,4-branched mannose (29%), 1,4-linked glucuronic (32%) and galacturonic (8%) acid as well as terminal rhamnose (25%). After removal of ca 70% of the terminal rhamnose the remaining polysaccharide showed a decrease in 1,2,4-branched mannose and an increase in 1,2-linked mannose indicating that at least half of the rhamnose residues were linked to mannose via C-4. GF3 (Mr 40,000) consisted of 1,4-linked galacturonic (30%) and glucuronic (16) acid, 1,3,6-branched galactose (15%), 1,2-linked (5%) and 1,2,4-branched (3.5%) rhamnose as well as 1,5-linked arabinose (11%). Rhamnose (5%) and arabinose (10%) were present as terminal groups. Mild acid hydrolysis selectively cleaved arabinose and the remaining polysaccharide showed an increased amount of 1,6-linked and terminal galactose and a decreased quantity of 1,3,6-branched galactose. These results indicated that the terminal as well as the 1,5-linked arabinose were mainly connected to galactose via C-3. The GF3 polysaccharide appeared to be a rhamnogalacturonan with arabinogalactan side chains.     

Structural characterization of a tobacco rhamnogalacturonan

A rhamnogalacturonan, extracted with hot water from the aqueous ethanol insoluble residue of flue-cured bright tobacco lamina, was purified by tangential flow ultrafiltration, ion chromatography and gel filtration. It was characterized by chemical and spectroscopic methods. Fractionation revealed that the rhamnogalacturonan consisted of a series of polysaccharides with different amounts of methyl-esterified galactopyranosyluronic acid residues in the backbone and different amounts of neutral sugar residues.The main pectic polysaccharide fraction has a backbone consisting of 4-linked α-d-galactopyranosyluronic acid residues interspersed with 2-linked l-rhamnopyranosyl residues. Approximately 22% of the galactopyranosyluronic acid residues are methylated. The main chain is branched at C-4 of rhamnose with neutral sugar side chains containing terminal and 4-linked β-d-galactopyranosyl and terminal and 5-linked α-l-arabinofuranosyl residues. The average degree of polymerization of this tobacco rahamnogalacturonan was estimated to be 400.     

Isolation and characterization of an extracellular polysaccharide from Pseudomonas caryophylli CFR 1705

Extracellular polysaccharides were isolated from Pseudomonas caryophylli CFR 1705 grown on lactose containing medium. The major fraction (no.1) obtained on DEAE-cellulose chromatography was composed of rhamnose, mannose and glucose in the ratio 1:3.26:4.97, respectively, and having a molecular weight of 1.1×10⁶ Da. Methylation followed by GC-MS analysis revealed it to be a highly branched 1,4-linked hexosan with mannose and glucose as the branch-off residues at positions C-2 and C-6 of the main chain. Rhamnose was essentially found as non-reducing terminal residue.     

Molecular characterisation of soybean polysaccharides: an approach by size exclusion chromatography, dynamic and static light scattering methods

Water soluble polysaccharides from soybean (SSPS) have a pectin-like structure and are used as stabilisers in acidified beverages. Physicochemical properties such as structure, molecular weight and shape or conformation are primary factors controlling their functional properties. Two soybean polysaccharides, a native SSPS and a modified SSPS treated with beta-(1-->4)-D-galactosidase (GPase/SSPS) were studied by dynamic and static light scattering (DLS, SLS) and size exclusion chromatography (SEC). Consecutive filtrations using a range of membrane pore size removed a small fraction of macromolecular aggregates from dilute polysaccharide solutions with relatively little effect on the major component molecules as monitored by DLS and SEC measurements. Access to aggregate-free dilute solutions of SSPS and GPase/SSPS allowed the direct measurement of molecular characteristics. SLS results showed that SSPS had a weight average molecular weight of (645+/-11)x 10(3)g/mol and a radius of gyration, Rg, of (23.5+/-2.8)nm. By comparing R(g) with the hydrodynamic radius, Rh (21.1+/-0.5 nm) obtained from DLS, the structural parameter rho (Rg/Rh) was found to be 1.1, suggesting that SSPS has an overall globular shape due to a highly branched structure. The modified SSPS had a significantly lower molecular weight (287+/-18)x 10(3)g/mol but a similar radius of gyration (23.2+/-1.7 nm). The structure parameter rho of GPase/SSPS was higher (rho=1.3) because of a smaller hydrodynamic radius (17.7+/-1.8 nm). This suggests that GPase/SSPS has a much less branched structure yet still differs significantly from a linear random coil conformation (rho=1.7-2.0). The results derived from SLS and DLS are in agreement with the conclusions obtained from a chemical analysis where the reduction of molecular weight of GPase/SSPS was caused by the cleavage of galactan side chains.     

The isolation, preliminary study of structure and physiological activity of water-soluble polysaccharides from squeezed berries of Snowball tree Viburnum opulus

Water-soluble polysaccharide fractions VO1-VO4 were isolated from the squeezed berries of snowball tree (Viburnum opulus) by successive extraction with water at various temperatures and pH and with aqueous solutions of ammonium oxalate. These fractions were purified by ion-exchange chromatography on DEAE cellulose, and the homogeneity of the purified polysaccharides was determined by gel filtration on Sephacryl S-500. Acidic polysaccharides close to pectins in their sugar composition were found in all the extracts (fractions VO1-1, VO2-1, VO3-2, and VO4-2). Residues of galacturonic acid, galactose, arabinose, and (to a lesser extent) rhamnose are their main constituents. Neutral polysaccharides composed mainly of galactose and mannose residues were additionally found in fractions extracted with acidified water (pH 4.0) and with aqueous ammonium oxalate solutions. Partial acidic hydrolysis and digestion with pectinase of acidic polysaccharides indicated that their carbohydrate backbone consists of alpha-1,4-linked residues of D-galacturonic acid. NMR spectra of acidic polysaccharides (fractions VO3-2 and VO3-3) confirmed this and demonstrated that their side oligosaccharide chains are composed of beta-1,4-linked galactopyranose residues and of terminal and 2,5- and 3,5-substituted residues of alpha-arabinofuranose at a Gal: Ara ratio of 3:1. Some polysaccharides from V. opulus were found to possess an immunostimulating activity: they enhance phagocytosis, in particular, the phagocytic index and the secretion of lysosomal enzymes with peritoneal macrophages. Calcium ions were found to be necessary for the appearance of the stimulating effect of acidic polysaccharides from V. opulus.     

Pectin Biosynthesis: GALS1 in Arabidopsis thaliana Is a β-1,4-Galactan β-1,4-Galactosyltransferase

β-1,4-Galactans are abundant polysaccharides in plant cell walls, which are generally found as side chains of rhamnogalacturonan I. Rhamnogalacturonan I is a major component of pectin with a backbone of alternating rhamnose and galacturonic acid residues and side chains that include α-1,5-arabinans, β-1,4-galactans, and arabinogalactans. Many enzymes are required to synthesize pectin, but few have been identified. Pectin is most abundant in primary walls of expanding cells, but β-1,4-galactan is relatively abundant in secondary walls, especially in tension wood that forms in response to mechanical stress. We investigated enzymes in glycosyltransferase family GT92, which has three members in Arabidopsis thaliana, which we designated GALACTAN SYNTHASE1, (GALS1), GALS2 and GALS3. Loss-of-function mutants in the corresponding genes had a decreased β-1,4-galactan content, and overexpression of GALS1 resulted in plants with 50% higher β-1,4-galactan content. The plants did not have an obvious growth phenotype. Heterologously expressed and affinity-purified GALS1 could transfer Gal residues from UDP-Gal onto β-1,4-galactopentaose. GALS1 specifically formed β-1,4-galactosyl linkages and could add successive β-1,4-galactosyl residues to the acceptor. These observations confirm the identity of the GT92 enzyme as β-1,4-galactan synthase. The identification of this enzyme could provide an important tool for engineering plants with improved bioenergy properties.     

Isolation and structural study of polysaccharides from campion Silene vulgaris

Silenan SV, a pectic polysaccharide, was isolated from the aerial part of Silene vulgaris (Moench) Garke (Oberna behen (L.) Ikonn.), widespread through the European North of Russia. The polysaccharide was found to contain residues of galacturonic acid (63%), arabinose, galactose, and rhamnose as the main constituents. The results of a partial acidic hydrolysis, pectinase digestion, and NMR studies of silenan SV indicated that its molecule contains a linear alpha-1,4-D-galacturonan backbone and ramified regions. The core of the ramified regions is composed of residues of alpha-1,4-D-galacturonic acid along with 2-substituted alpha-rhamnopyranose residues. The NMR data showed that the silenan SV side chains are composed of the blocks built from the terminal alpha-1,5-linked arabinofuranose and beta-1,4-linked galactopyranose residues; these most likely are the side chains of rhamnogalacturonan, characteristic of other pectic polysaccharides. The nonreducing ends of these side chains contain alpha-arabinofuranose residues.     

Structural features of immunologically active polysaccharides from Ganoderma lucidum.

Three polysaccharides, two heteroglycans (PL-1 and PL-4) and one glucan (PL-3), were solubilized from the fruit bodies of Ganoderma lucidum and isolated by anion-exchange and gel-filtration chromatography. Their structural features were elucidated by glycosyl residue and glycosyl linkage composition analyses, partial acid hydrolysis, acetolysis, periodate oxidation, 1D and 2D NMR spectroscopy, and ESI-MS experiments. The data obtained indicated that PL-1 had a backbone consisting of 1,4-linked alpha-D-glucopyranosyl residues and 1,6-linked beta-D-galactopyranosyl residues with branches at O-6 of glucose residues and O-2 of galactose residues, composed of terminal glucose, 1,6-linked glucosyl residues and terminal rhamnose. PL-3 was a highly branched glucan composed of 1,3-linked beta-D-glucopyranosyl residues substituted at O-6 with 1,6-linked glucosyl residues. PL-4 was comprised of 1,3-, 1,4-, 1,6-linked beta-D-glucopyranosyl residues and 1,6-linked beta-D-mannopyranosyl residues. These polysaccharides enhanced the proliferation of T- and B-lymphocytes in vitro to varying contents and PL-1 exhibited an immune-stimulating activity in mice.     

Glycosylation of internal sugar residues of oligosaccharides catalyzed by alpha-galactosidase from Aspergillus fumigatus

Purified alpha-galactosidase from a thermotolerant fungus Aspergillus fumigatus IMI 385708 was found to catalyze efficiently transgalactosylation reactions using 4-nitrophenyl alpha-D-galactopyranoside as glycosyl donor. Self-transfer reactions with this substrate afforded in low yields several 4-nitrophenyl galactobiosides. Monosaccharides also served as poor glycosyl acceptors. Disaccharides and particularly higher oligosaccharides of alpha-1,4-gluco- (maltooligosaccharides), beta-1,4-gluco- (cellooligosaccharides) and beta-1,4-manno-series were efficiently galactosylated, the latter being the best acceptors that were also doubly galactosylated. With mannooligosaccharides product yields increased with polymerization degree of acceptors reaching 50% at DP of 4-6. Longer oligosaccharide acceptors were galactosylated at internal sugar residues. All galactosyl residues were transferred exclusively to the primary hydroxyl group(s) at C-6 position of oligosaccharide acceptors. This is in accordance with the inability of the enzyme to transfer galactose to beta-1,4-linked xylooligosaccharides. This is the first report of glycosyl transfer reaction to internal sugar residues of oligosaccharides catalyzed by a glycosidase. High affinity to oligosaccharide acceptors also opens a way toward enzymatic glycosylation of polysaccharides, thus modulating their physico-chemical and biological properties.     

Structure of Silenan, a Pectic Polysaccharide from Campion Silene vulgaris (Moench) Garcke

A pectic polysaccharide named silenan, [alpha]D20 +148.6 degrees (c 0.1; H2O), was isolated earlier from the aerial part of campion, Silene vulgaris (Moench) Garcke. Silenan has been shown to contain homogalacturonan segments as "smooth regions" and rhamnogalacturonan fragments as "hairy regions". The present study reveals a generalization of structural features of silenan. Silenan was subjected to enzymic digestion with pectinase, to Smith degradation, and to lithium-degradation to determine the conforming poly- and oligosaccharide fragments of "hairy regions" of silenan. The NMR-spectral data and mass-spectrometry confirmed that the core of the ramified region of silenan consisted of residues of alpha-rhamnopyranose 2-O-glycosylated with the residues of alpha-1,4-D-galactopyranosyl uronic acid. The part of the alpha-rhamnopyranose residues of the backbone are branched at O-4. On the basis of the data, the hairy regions of silenan proved to contain mainly linear chains of beta-1,3-, beta-1,4-, and beta-1,6-galactopyranan and alpha-1,5-arabinofuranan. The side chains of the ramified region were shown to have branching points represented 2,3-, 3,6-, 4,6-di-O-substituted beta-galactopyranose residues.     

Selective chemical depolymerization of rhamnogalacturonans

A method was developed to selectively methyl esterify and then cleave GalA residues in pectic polysaccharides. The method was optimized using a rhamnogalacturonan (RG) from Arabidopsis mucilage as a model compound. The carboxyl group of the GalA residues in the RG was selectively methyl esterified using tetrabutylammonium fluoride and iodomethane in Me(2)SO containing 8% water. A 1D HMQC NMR method to determine the degree of methyl esterification was developed using (13)C-iodomethane as the methylating agent. The methyl-esterified pectins were fragmented by beta-elimination in 0.2M sodium borate, pH7.3, at 125 degrees C. The resulting oligoglycosyl fragments, which contain a nonreducing 4-deoxy-beta-l-threo-hex-4-enepyranosyluronic acid residue, were characterized using MALDI-TOF mass spectrometry, monosaccharide composition analysis, and 1D and 2D (1)H and (13)C NMR spectroscopy. Application of this method to branched RG from potato generated low-molecular-weight fragments containing two residues from the RG backbone and a single side chain. In contrast, the fragments obtained when RG is treated with RG lyase contain a minimum of four backbone residues. The chemical method thus facilitates the release and structural characterization of the side-chain structures of RG obtained from various plant sources. The method also provides a convenient method for generating fully or partially methyl-esterified homogalacturonans.     

The isolation, preliminary structural studies, and physiological activity of water-soluble polysaccharides from the squeezed berries of the snowball treeViburnum opulus

Water-soluble polysaccharide fractions VO1–VO4 were isolated from the squeezed berries of the snowball tree (Viburnum opulus) by successive extraction with water at various temperatures and pH and with aqueous solutions of ammonium oxalate. These fractions were purified by ion-exchange chromatography on DEAE cellulose, and the homogeneity of the purified polysaccharides was determined by gel filtration on Sephacryl S-500. Acidic polysaccharides close to pectins in their sugar composition were found in all the extracts (fractions VO1-1, VO2-1, VO3-2, and VO4-2). Residues of galacturonic acid, galactose, arabinose, and (to a lesser extent) rhamnose are their main constituents. Neutral polysaccharides composed mainly of galactose and mannose residues were additionally found in fractions extracted with acidified water (pH 4.0) and with aqueous ammonium oxalate solutions. Partial acidic hydrolysis and digestion with pectinase of acidic polysaccharides indicated that their carbohydrate backbone consists of α-1,4-linked residues ofD-galacturonic acid. NMR spectra of acidic polysaccharides (fractions VO3-2 and VO3-3) confirmed this and demonstrated that their side oligosaccharide chains are composed of β-1,4-linked galactopyranose residues and of terminal and 2,5- and 3,5-substituted residues of α-arabinofuranose at a Gal : Ara ratio of 3 : 1. Some polysaccharides fromV. opulus were found to possess an immunostimulating activity: they enhance phagocytosis, in particular, the phagocytic index and the secretion of lysosomal enzymes with peritoneal macrophages. Calcium ions were found to be necessary for the appearance of the stimulating effect of acidic polysaccharides fromV. opulus.     

Structural study of bergenan, a pectin from Bergenia crassifolia

A pectin polysaccharide named bergenan was isolated from the freshly collected leaves of the leather bergenia Bergenia crassifolia by extraction with an aqueous solution of ammonium oxalate. The main component of its carbohydrate chain was shown to be the residues of D-galacturonic acid (about 80%). In addition, the polysaccharide contains residues of galactose, arabinose, and rhamnose; their total content is less than 15%. It was shown that the bergenan samples from bergenia leaves collected at different vegetation periods (from July to September) do not substantially differ either in monosaccharide composition or in the viscosity of aqueous solutions they form. The results of enzymatic hydrolysis by alpha-1,4-galacturonase (pectinase), partial acidic hydrolysis, NMR spectroscopy, and methylation with subsequent analysis of the results by GC-MS indicate that the bergenan macromolecule contains the regions of a linear alpha--1,4-D-galactopyranosyluronan and rhamnogalacturonan-I (RG-1). Galacturonan responds for a greater part of the macromolecule. A considerable amount of its constituent galacturonic acid residues are present as methyl esters. The side chains in RG-I are attached to the rhamnopyranose residues of the main carbohydrate chain by 1,4-link and are composed of the residues of terminal arabinofuranose and galactopyranose, 1,5-linked (-arabinofuranose, and 1,4-and 1,6-linked beta-galactopyranose. The branching points of the side chains of the RG-I molecule are 3,4- and 3,6-di-O-substituted galactose residues.     

Structure of a fucoidan from the brown seaweed Fucus serratus L

A fucoidan consisting of L-fucose, sulfate and acetate in a molar proportion of 1:1:0.1 and small amounts of xylose and galactose were isolated from the brown seaweed Fucus serratus L. The fucoidan structure was investigated by 1D and 2D 1H and 13C NMR spectroscopy of its desulfated and de-O-acetylated derivatives as well as by methylation analysis of the native and desulfated polysaccharides. A branched structure was suggested for the fucoidan with a backbone of alternating 3- and 4-linked alpha-L-fucopyranose residues, -->3)-alpha-L-Fucp-(1-->4)-alpha-L-Fucp-(1-->, about half of the 3-linked residues being substituted at C-4 by trifucoside units alpha-L-Fucp-(1-->4)-alpha-L-Fucp-(1-->3)-alpha-L-Fucp-(1-->. Minor chains built up of 4-linked alpha-fucopyranose and beta-xylose residues were also detected, but their location, as well as the position of galactose residues, remained unknown. Sulfate groups were shown to occupy mainly C-2 and sometimes C-4, although 3,4-diglycosylated and some terminal fucose residues may be nonsulfated. Acetate was found to occupy C-4 of 3-linked Fuc and C-3 of 4-linked Fuc in a ratio of about 7:3.     

Molecular organization of the alkali-insoluble fraction of Aspergillus fumigatus cell wall

Physical and biological properties of the fungal cell wall are determined by the composition and arrangement of the structural polysaccharides. Cell wall polymers of fungi are classically divided into two groups depending on their solubility in hot alkali. We have analyzed the alkali-insoluble fraction of the Aspergillus fumigatus cell wall, which is the fraction believed to be responsible for fungal cell wall rigidity. Using enzymatic digestions with recombinant endo-beta-1,3-glucanase and chitinase, fractionation by gel filtration, affinity chromatography with immobilized lectins, and high performance liquid chromatography, several fractions that contained specific interpolysaccharide covalent linkages were isolated. Unique features of the A. fumigatus cell wall are (i) the absence of beta-1,6-glucan and (ii) the presence of a linear beta-1, 3/1,4-glucan, never previously described in fungi. Galactomannan, chitin, and beta-1,3-glucan were also found in the alkali-insoluble fraction. The beta-1,3-glucan is a branched polymer with 4% of beta-1,6 branch points. Chitin, galactomannan, and the linear beta-1, 3/1,4-glucan were covalently linked to the nonreducing end of beta-1, 3-glucan side chains. As in Saccharomyces cerevisiae, chitin was linked via a beta-1,4 linkage to beta-1,3-glucan. The data obtained suggested that the branching of beta-1,3-glucan is an early event in the construction of the cell wall, resulting in an increase of potential acceptor sites for chitin, galactomannan, and the linear beta-1,3/1,4-glucan.     

Structure and potential immunological activity of a pectin from Centella asiatica (L.) Urban

S3A was a RG-I pectin isolated from Centella asiatica that contained Rha, Ara, Gal, Glc and GalA in molar ratio of 1.0:0.6:1.5:0.2:1.1 and had been found to have a backbone composed mainly of the disaccharide repeat unit, -->4)-alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1-->. Based on methylation analysis, NaIO4 oxidation, partial acid hydrolysis and lithium-treatment, the structural features were elucidated. Side chains of S3A were predominantly linked to O-4 of 1,2,4-linked alpha-L-Rhap. The side chains are comprised of arabinosyl chains, galactosyl chains, arabinogalactosyl chains and short glucosyl chains. A total of 45% Rhap in the backbone was substituted by side chains. The arabinosyl residues were mostly distributed in the arabinosyl side chains. According to the immunological results of S3A and its degraded derivatives, S3A had no immunological activity, but its derivatives had immuno-stimulating activities to some extent.