D-glucuronic Acid Residues Research Articles

Identification of hyaluronic acid in seized samples by Hypercarb chromatography-high resolution mass spectrometry.

Hyaluronic acid (HA) has been commonly used to treat osteoarthritis and joint injuries in horses and dogs since the 1970s. HA is a polysaccharide made up of alternating N-acetyl-d-glucosamine and d-glucuronic acid residues, with polymeric molecules achieving molecular weights as high as 20 MDa. High molecular weight HA forms a viscous hydrogel when dissolved in water, making HA solutions distinct from most other pharmaceutical preparations. Clear viscous solutions are often encountered during stable and kennel inspections, but in the absence of an analytical method, it is not possible to identify if these substances contain HA or other unknown compounds. This paper presents a simple method for the identification of HA in seized materials based on chemical hydrolysis followed by Hypercarb chromatography and MS/MS analysis.

Chemical Synthesis and Antitumor Evaluation of Chikusetsusaponin IVa Butyl Ester and Its Analogues

AbstractChikusetsusaponin IVa butyl ester (CS-IVa-Be) is a triterpene saponin that acts as a novel IL6R antagonist for inducing breast cancer cell apoptosis. However, the structure–activity relationship for this class of saponins remains unclear. Here, we report a gram-scale synthesis of CS-IVa-Be and the efficient preparation of eight of its analogues. CS-IVa-Be was demonstrated to have significant antitumor activities against MDA-MB-231, HepG2, and A549 cells. When one of the sugar residues at either the 3-OH or 28-COOH position of CS-IVa-Be was cleaved, or the length of the alkyl chain on the d-glucuronic acid residue of CS-IVa-Be was changed, these analogues showed varied inhibitory activities against the cancer cell lines. Notably, the carboxylic acid form of CS-IVa-Be exhibited a stronger antitumor activity against MDA-MB-231 cells. Furthermore, the carboxylic acid form of CS-IVa-Be inhibited MDA-MB-231 cell proliferation in a dose-dependent manner by arresting the cell cycle at the G2/M phase.

Biochemical and crystallographic assessments of the effect of 4,6-O-disulfated disaccharide moieties in chondroitin sulfate E on chondroitinase ABC I activity.

O-sulfated N-acetyl-d-galactosamine (GalNAc) residues in chondroitin sulfate (CS) play a crucial role in chondroitinase ABC I (cABC-I) activity. CSA containing mainly 4-O-monosulfated GalNAc was a good substrate for the enzyme, but not CSE containing mainly 4,6-O-disulfated GalNAc [GalNAc(4S,6S)]. Each CS isomer exhibits structural heterogeneity; CSE has di-sulfated disaccharide units and mono-sulfated disaccharide units. Disaccharide composition analysis of digested products revealed that mono-sulfated disaccharide units in CSE contributed to the enzyme reactivity. Although enough substrate (CSA) was present in mixtures of CSA and CSE for reaction, the reactivity was reduced depending on the amount of CSE in the mixture. These results suggested that CSE is not only resistant to enzyme digestion but also attenuates enzyme activity. To understand the mechanism of action, crystallography of cABC-I in complex with unsaturated CSE-disaccharide, ΔDi-(4,6)S, was performed. Both 4-O- and 6-O-sulfate groups in ΔDi-(4,6)S interact with Arg500, suggesting that there was a greater interaction between ΔDi-(4,6)S and Arg500 than between mono-sulfated disaccharides and Arg500. Besides, this interaction attenuated enzyme activity by interfering with a function of Arg500, which is the charge neutralization of the carboxy group of D-glucuronic acid (GlcA) residues in CS. When interacting with the CSE-disaccharide unit [GlcAβ1-3GalNAc(4S,6S)] in CS, cABC-I cannot interact with other CS-disaccharide units until it has digested the CSE-disaccharide unit. The low reactivity of cABC-I with CSE is attributable to two suggested factors: (a) resistance of E-units in CSE molecules to digestion by cABC-I, and (b) tendency of E-units in CSE molecules to attenuate cABC-I activity.

Re-expression of glucuronyl C5-epimerase in the mutant MEF cells increases heparan sulfate epimerization but has no influence on the Golgi localization and enzymatic activity of 2-O-sulfotransferase.

Heparan sulfate (HS) is a linear and complex polysaccharide that modulates the biological activities through protein recognition and interaction. Evidence indicates that protein-binding properties of HS are largely dependent on distinctive sulfation and epimerization patterns that are modified by a series of Golgi-localized enzymes. In particular, the glucuronyl C5-epimerase (Hsepi) converts D-glucuronic acid (GlcA) residues to L-iduronic acid (IdoA) and 2-O-sulfotransferase (2OST) catalyzes sulfation at C2 position of IdoA and rarely GlcA residues. Mice lacking both Hsepi and 2OST display multiple development defects, indicating the importance of IdoA in HS. Here, to gain greater insights of HS structure-function relationships, as well as a better understanding of the regulatory mechanisms of Hsepi and 2OST, the fine structure and cellular signaling functions of HS were investigated after restoration of Hsepi in the mutant mouse embryonic fibroblast (MEF) cells. Introduction of Hsepi into the Hsepi mutant MEF cells led to robustly increased proportion of IdoA residues, which rescued the cell signaling in response to fibroblast growth factor 2. However, we found that Hsepi knockout had no influence on either cellular transport or enzymatic activity of 2OST in the MEF cells, which is not in accord with the findings suggesting that the enzymatic activity and cellular transport of 2OST and Hsepi might be differently regulated.

Extraction and characterization of xylan from sugarcane tops as a potential commercial substrate

Xylan is the major hemicellulose present in sugarcane stem secondary cell walls. Xylan is composed of xylose backbone with a high degree of substitutions, which affects its properties. In the present study, the xylan from sugarcane tops (SCT) was extracted and characterized. Compositional analysis of xylan extracted from SCT (SCTx) displayed the presence of 74% of d-xylose residues, 16% of d-glucuronic acid residues and 10% of l-arabinose. High performance size exclusion chromatographic analysis of SCTx displayed a single peak corresponding to a molecular mass of ∼57kDa. The Fourier transform infrared spectroscopic analysis of SCTx displayed the peaks corresponding to those obtained from commercial xylan. FESEM analysis of SCTx showed the granular and porous surface structure. Differential thermogravimetric analysis (DTG) of SCTx displayed two thermal degradation temperatures (Td) of 228°C, due to breakdown of the side chains of glucuronic acid and arabinose and 275°C, due to breakdown of xylan back bone. The presence of arabinose and glucuronic acid as a side chains was confirmed by the DTG and thermogravimetric analysis. The CHNS analysis of SCTx showed the presence of only carbon and hydrogen supporting its purity. The recombinant xylanase (CtXyn11A) from Clostridium thermocellum displayed a specific activity of 1394±51U/mg with SCTx, which was higher than those with commercial xylans. The thin layer chromatography and electrospray ionization mass spectroscopy analyses of CtXyn11A hydrolysed SCTx contained a series of linear xylo-oligosaccharides ranging from degree of polymerization 2-6 and no substituted xylo-oligosaccharides because of the endolytic activity of enzyme. The extracted xylan from SCT can be used as an alternative commercial substrate and for oligo-saccharide production.

Highly Specific near-Infrared Fluorescent Probe for the Real-Time Detection of β-Glucuronidase in Various Living Cells and Animals.

β-Glucuronidase (GLU) is an important biomarker for primary cancers and intestinal metabolism of drugs or endogenous substances; however, an effective optical probe for near-infrared (NIR) monitoring in vivo is still lacking. Herein, we design an enzyme-activated off-on NIR fluorescent probe, HC-glu, based on a hemicyanine keleton, which is conjugated with a d-glucuronic acid residue via a glycosidic bond, for the fluorescent quantification and trapping of endogenous GLU activity in vitro and in vivo. The newly developed NIR probe exhibited prominent features including prominent selectivity, high sensitivity, and ultrahigh imaging resolution. It has been successfully used to detect and image endogenous GLU in various hepatoma carcinoma cells, tumor tissues, and tumor-bearing mouse models, for cancer diagnosis and therapy. Moreover, it could detect the in vivo activity of GLU in the intestinal tracts of animals including mice and zebrafish, where GLU performs a vital biological function and is mainly distributed. It could also evaluate real intestinal distribution and real-time variations of GLU in development and growth, all of which are very helpful to guide rational drug use in the clinic. Our results fully demonstrated that HC-glu may serve as a promising tool for evaluating the biological function and process of GLU in living systems.

Characterisation of three fungal glucuronoyl esterases on glucuronic acid ester model compounds

The glucuronoyl esterases (GEs) that have been identified so far belong to family 15 of the carbohydrate esterases in the CAZy classification system and are presumed to target ester bonds between lignin alcohols and (4-O-methyl-)d-glucuronic acid residues of xylan. Few GEs have been cloned, expressed and characterised to date. Characterisation has been done on a variety of synthetic substrates; however, the number of commercially available substrates is very limited. We identified novel putative GEs from a wide taxonomic range of fungi and expressed the enzymes originating from Acremonium alcalophilum and Wolfiporia cocos as well as the previously described PcGE1 from Phanerochaete chrysosporium. All three fungal GEs were active on the commercially available compounds benzyl glucuronic acid (BnGlcA), allyl glucuronic acid (allylGlcA) and to a lower degree on methyl glucuronic acid (MeGlcA). The enzymes showed pH stability over a wide pH range and tolerated 6-h incubations of up to 50 °C. Kinetic parameters were determined for BnGlcA. This study shows the suitability of the commercially available model compounds BnGlcA, MeGlcA and allylGlcA in GE activity screening and characterisation experiments. We enriched the spectrum of characterised GEs with two new members of a relatively young enzyme family. Due to its biotechnological significance, this family deserves to be more extensively studied. The presented enzymes are promising candidates as auxiliary enzymes to improve saccharification of plant biomass.

When structure matters – how heparan sulfate epimerization affects animal development

Animal development requires concerted actions of different signaling molecules in order to ensure correctly timed and localized growth and differentiation of tissues. The extracellular matrix often provides guiding cues for signaling proteins such as the glycosaminoglycan heparan sulfate (HS), which is an important regulator of development. HS biosynthesis is a complex series of events, including chain initiation, chain elongation and chain modification. Chain modification comprises N -deacetylation of the N -acetyl-glucosamin (GlcNAc) residues, epimerization of D-glucuronic acid (GlcA) residues to L-iduronic acid (IdoA) and O -sulfation of all sugar units at different positions. None of these reactions is performed to completion along the HS chain, generating cell-type specific patterns of sulfation. It is well-known that loss of HS leads to severe developmental defects and during recent years it has become evident that even alterations in the HS sulfation pattern strongly affect animal development. Interestingly, these patterns do not only depend on the expression of the respective sulfotransferases, but are also strongly affected by N -deacetylation and epimerization of the HS chains. This review focuses on recent work on the HS C5-glucuronyl epimerase (protein GLCE, gene Glce , earlier named Hsepi ) in mouse cartilage development and relates these findings to earlier studies on GLCE function. The sulfation pattern of HS was altered in all Glce deficient tissues, but extent and nature of the changes were different between cell types. In addition, loss of Glce affected multiple signaling pathways depending on the cell type, stressing the importance of HS as a general regulator during animal development. Promoting ligand/receptor interactions as well as shaping morphogen gradients, two important roles of HS, are clearly affected by the loss of Glce in these studies and are further discussed here.

Heparosan-glucuronate 5-epimerase: Molecular cloning and characterization of a novel enzyme

Iduronic acid (IdoA) is a critical component of heparan sulfate in its interaction with functional proteins. Heparosan-N-sulfate-glucuronate 5-epimerase (HNSG-5epi) converts d-glucuronic acid (GlcA) residues in N-sulfated heparosan (NS-heparosan), as an intermediate in heparan sulfate biosynthesis, to IdoA. In the present study, the authors discovered a different 5-epimerase, designated HG-5epi (heparosan-glucuronate 5-epimerase), that is involved in acharan sulfate biosynthesis and possesses novel substrate specificity. A candidate cDNA of HG-5epi was cloned from the cDNA library of Achatina fulica. The cloned cDNA contained a whole coding region that predicts a type II transmembrane protein composed of 601 amino acid residues. The amino acid sequence of HG-5epi is homologous to that of HNSG-5epi. Recombinant HG-5epi was expressed in insect cells and its enzymatic properties characterized. As expected, HG-5epi epimerizes GlcA residues in heparosan, but not in NS-heparosan. Conversion of IdoA to GlcA was also catalyzed by HG-5epi when completely desulfated N-acetylated heparin was used as the substrate, indicating a reversible reaction mechanism. At equilibrium of the epimerization, the proportion of IdoA in the reaction product reached up to 30% of total hexuronic acid. To our knowledge, this is the first report to describe an enzyme that catalyzes the epimerization of non-sulfated heparosan. This new enzyme may be applied to the study of synthetic heparan sulfate-related polysaccharides having certain biological and pharmacological activities. In addition, a new method using anion-exchange HPLC connected to a post-column fluorescent labeling system was developed for analyzing hexuronic acid isomers.

Structure of Ulvan Isolated from the Edible Green Seaweed, Ulva pertusa

Ulvan, rhamnan sulfate, was extracted from the edible green seaweed, Ana-aosa (Ulva pertusa), which is grown on the coast of the Okinawa Islands. The yield of ulvan was 8.5% (W/W), and the total carbohydrates, uronic acid and sulfuric acid and ash contents were 67.3%, 23.8%, 19.7% and 22.6%, respectively. L-Rhamnose, D-xylose and D-glucose residues were identified by liquid chromatography, and their molar ratio was 4.0:0.1:0.3. D-Glucuronic and L-idulonic acid residues were also identified in molar ratio of 1.0:0.2. The NMR (13C and 1H) and methylation analysis revealed terminal β-D-glucruonic acid, terminal α-L-idulonic acid, 1,3-linked α-L-rhamnose, 1,4-linked α-L-rhamnose, 1,2,4-linked α-L-rhamnose, 1,3,4-linked α-L-rhamnose, 1,2,3,4-linked α-L-rhamnose and 1,3,4-linked β-D-xylose. The sulfate groups were attached at the C-2 and C-3 positions of the 1,4-linked α-L-rhamnose as well as C-3 of the 1,4-linked β-D-xylose residues. The chemical structure of the ulvan from Ulva pertusa was determined.

Chondroitin Sulfate Synthase-2 Is Necessary for Chain Extension of Chondroitin Sulfate but Not Critical for Skeletal Development

Chondroitin sulfate (CS) is a linear polysaccharide consisting of repeating disaccharide units of N-acetyl-D-galactosamine and D-glucuronic acid residues, modified with sulfated residues at various positions. Based on its structural diversity in chain length and sulfation patterns, CS provides specific biological functions in cell adhesion, morphogenesis, neural network formation, and cell division. To date, six glycosyltransferases are known to be involved in the biosynthesis of chondroitin saccharide chains, and a hetero-oligomer complex of chondroitin sulfate synthase-1 (CSS1)/chondroitin synthase-1 and chondroitin sulfate synthase-2 (CSS2)/chondroitin polymerizing factor is known to have the strongest polymerizing activity. Here, we generated and analyzed CSS2−/− mice. Although they were viable and fertile, exhibiting no overt morphological abnormalities or osteoarthritis, their cartilage contained CS chains with a shorter length and at a similar number to wild type. Further analysis using CSS2−/− chondrocyte culture systems, together with siRNA of CSS1, revealed the presence of two CS chain species in length, suggesting two steps of CS chain polymerization; i.e., elongation from the linkage region up to Mr ∼10,000, and further extension. There, CSS2 mainly participated in the extension, whereas CSS1 participated in both the extension and the initiation. Our study demonstrates the distinct function of CSS1 and CSS2, providing a clue in the elucidation of the mechanism of CS biosynthesis.

Structural and thermal characterization of hemicelluloses isolated by organic solvents and alkaline solutions from Tamarix austromongolica

Three organosolv and three alkaline hemicellulosic fractions were prepared from lignocellulosic biomass of the fast-growing shrub Tamarix austromongolica ( Tamarix Linn.). Sugar analysis revealed that the organosolv-soluble fractions contained a higher content of glucose (33.7–6.5%) and arabinose (14.8–5.6%), and a lower content of xylose (62.2–54.8%) than the hemicellulosic fractions isolated with aqueous alkali solutions. A relatively high concentration of alkali resulted in a decreasing trend of the xylose/4- O-methyl- d-glucuronic acid ratio in the alkali-soluble fractions. The results of NMR analysis supported a major substituted structure based on a linear polymer of β-(1 → 4)-linked d-xylopyranosyl residues, having ramifications of α- l-arabinofuranose and 4- O-methyl- d-glucuronic acid residues monosubstituted at O-3 and O-2, respectively. Thermogravimetric analysis revealed that one step of major mass loss occurred between 200–400 °C, as hemicelluloses devolatilized with total volatile yield of about 55%. It was found that organosolv-soluble fractions are more highly ramified, and showed a higher thermal stability than the alkali-soluble fractions.

Spectral Studies on the Binding Behavior of Cationic Dyes and Surfactants with Bacterial Polysaccharide of Klebsiella K43

The binding behavior of acidic capsular polysaccharide (SPS), isolated from Klebsiella serotype K43, with oppositely charged dyes and surfactants have been studied by way of absorbance and emission spectroscopic measurements. Each repeating unit of the SPS consists of three D-mannose, one D-galactose, and one D-glucuronic acid residue. The anionic polysaccharide exhibited chromotropic character and induced strong metachromasy in the cationic dye, pinacyanol chloride (PCYN) through the formation of a 1:1 stoichiometric complex. Evaluation of thermodynamic parameters, viz., changes in free energy (ΔG), enthalpy (ΔH), and entropy (ΔS), for the formation of dye-polymer complex and studies on the effect of different cosolvents were also evaluated to shed light on the binding nature as well as the extent of stability of the dye-polymer complex. Fluorescence of the cationic dye acridine orange (AO) was quenched with the progressive addition of SPS, which was found to be of Stern-Volmer type. Cationic surfactants in their pure form as well as in the mixed state with nonionic surfactant (Tween-20), replaced the dye bound to the polymer matrices; thus the original band intensities of the dyes could be reverted. Such studies revealed the involvement of both electrostatic as well as hydrophobic interactions between the dye-polymer as well as surfactant-polymer aggregates.

Chondroitin Sulfate Synthase-2/Chondroitin Polymerizing Factor Has Two Variants with Distinct Function*

Chondroitin sulfate (CS) is a polysaccharide consisting of repeating disaccharide units of N-acetyl-D-galactosamine and d-glucuronic acid residues, modified with sulfated residues at various positions. To date six glycosyltransferases for chondroitin synthesis have been identified, and the complex of chondroitin sulfate synthase-1 (CSS1)/chondroitin synthase-1 (ChSy-1) and chondroitin sulfate synthase-2 (CSS2)/chondroitin polymerizing factor is assumed to play a major role in CS biosynthesis. We found an alternative splice variant of mouse CSS2 in a data base that lacks the N-terminal transmembrane domain, contrasting to the original CSS2. Here, we investigated the roles of CSS2 variants. Both the original enzyme and the splice variant, designated CSS2A and CSS2B, respectively, were expressed at different levels and ratios in tissues. Western blot analysis of cultured mouse embryonic fibroblasts confirmed that both enzymes were actually synthesized as proteins and were localized in both the endoplasmic reticulum and the Golgi apparatus. Pulldown assays revealed that either of CSS2A, CSS2B, and CSS1/ChSy-1 heterogeneously and homogeneously interacts with each other, suggesting that they form a complex of multimers. In vitro glycosyltransferase assays demonstrated a reduced glucuronyltransferase activity in CSS2B and no polymerizing activity in CSS2B co-expressed with CSS1, in contrast to CSS2A co-expressed with CSS1. Radiolabeling analysis of cultured COS-7 cells overexpressing each variant revealed that, whereas CSS2A facilitated CS biosynthesis, CSS2B inhibited it. Molecular modeling of CSS2A and CSS2B provided support for their properties. These findings, implicating regulation of CS chain polymerization by CSS2 variants, provide insight in elucidating the mechanisms of CS biosynthesis.

Inverting character of family GH115 α-glucuronidases

α-Glucuronidases of glycoside hydrolase family 115 of the xylose-fermenting yeast Pichia stipitis and wood-destroying fungus Schizophyllum commune liberate 4- O-methyl- d-glucuronic acid residues from aldouronic acids and glucuronoxylan. The specific activities of both enzymes depended on polymerization degree of the acidic xylooligosaccharides and were inhibited by linear β-1,4-xylooligosaccharides. These results suggest interaction of the enzyme with several xylopyranosyl residues of the xylan main chain. Using 1H NMR spectroscopy and reduced aldopentaouronic acid (MeGlcA 3Xyl 4-ol) as a substrate, it was found that both enzymes are inverting glycoside hydrolases releasing 4- O-methyl- d-glucuronic acid (MeGlcA) as its β-anomer.

Rhamnogalacturonan I domains from ginseng pectin

Five rhamnogalacturonan I (RG-I) domains RG-I-1, RG-I-2, RG-I-3A, RG-I-3B and RG-I-4 were isolated from ginseng pectin by endo-polygalacturonase hydrolysis and a combination of ion-exchange and gel-permeation chromatography. The five domains all contain galacturonic acid, rhamnose, galactose and arabinose as main components and their rhamnose/galacturonic acid is from 0.26 to 0.64, among the range of RG-I. The molecular weights of RG-I-1 (5 kDa), RG-I-2 (4 kDa) and RG-I-3B (6 kDa) are smaller than those of RG-I-3A (45 kDa) and RG-I-4 (60 kDa). 13C NMR spectra of all domains showed RG-I features. RG-I-2 and RG-I-3B contained RG-I domains linked with highly methyl-esterified and acetylated homogalacturonan domains and the side chains probably belonging to type I and type II arabinogalactans, while RG-I-3A and RG-I-4 may have the side chains of type I arabinogalactans. 4-O-methyl-β- d-glucuronic acid residues were present at non-reducing terminals of RG-I-2.

Structure of acidic o-specific polysaccharide from the marine bacterium Cellulophaga baltica

The structure of an acidic O-specific polysaccharide from the marine bacterium Cellulophaga baltica was established by chemical methods of analysis and NMR spectroscopy. The polysaccharide was shown to consist of repeating tetrasaccharide units containing two mannose residues, one N-acetyl-D-glucosamine residue, and one D-glucuronic acid residue. An O-acetyl group was also found in the polysaccharide in nonstoichiometric amount. Thus, this polysaccharide had the following structure: [carbohydrate structure: in text].

Modulation of the Heparanase-inhibiting Activity of Heparin through Selective Desulfation, Graded N-Acetylation, and Glycol Splitting

Heparanase is an endo-beta-glucuronidase that cleaves heparan sulfate (HS) chains of heparan sulfate proteoglycans on cell surfaces and in the extracellular matrix (ECM). Heparanase, overexpressed by most cancer cells, facilitates extravasation of blood-borne tumor cells and causes release of growth factors sequestered by HS chains, thus accelerating tumor growth and metastasis. Inhibition of heparanase with HS mimics is a promising target for a novel strategy in cancer therapy. In this study, in vitro inhibition of recombinant heparanase was determined for heparin derivatives differing in degrees of 2-O- and 6-O-sulfation, N-acetylation, and glycol splitting of nonsulfated uronic acid residues. The contemporaneous presence of sulfate groups at O-2 of IdoA and at O-6 of GlcN was found to be non-essential for effective inhibition of heparanase activity provided that one of the two positions retains a high degree of sulfation. N-Desulfation/ N-acetylation involved a marked decrease in the inhibitory activity for degrees of N-acetylation higher than 50%, suggesting that at least one NSO3 group per disaccharide unit is involved in interaction with the enzyme. On the other hand, glycol splitting of preexisting or of both preexisting and chemically generated nonsulfated uronic acids dramatically increased the heparanase-inhibiting activity irrespective of the degree of N-acetylation. Indeed N-acetylated heparins in their glycol-split forms inhibited heparanase as effectively as the corresponding N-sulfated derivatives. Whereas heparin and N-acetylheparins containing unmodified D-glucuronic acid residues inhibited heparanase by acting, at least in part, as substrates, their glycol-split derivatives were no more susceptible to cleavage by heparanase. Glycol-split N-acetylheparins did not release basic fibroblast growth factor from ECM and failed to stimulate its mitogenic activity. The combination of high inhibition of heparanase and low release/potentiation of ECM-bound growth factor indicates that N-acetylated, glycol-split heparins are potential antiangiogenic and antimetastatic agents that are more effective than their counterparts with unmodified backbones.

Polysaccharides from Sesamum indicum meal: Isolation and structural features

Defatted Sesamum indicum seed cake was extracted, following two separate sequences, and the effects of extraction medium on yield and composition of the extracts were compared. Polysaccharides extracted sequentially with dilute acid and alkali represented 250 mg/g of defatted meal. The isolated polymers contained arabinan, rhamnogalacturonan I (RG I) and arabinogalactan proteins. Polysaccharides extracted during chlorite treatment and with dilute alkali had a higher proportion of rhamnose, suggesting a more branched variety of polymer. Three extracts, which were further characterized by size exclusion chromatography, gave two overlapping peaks. Structural characterisation of hemicellulosic polysaccharides, isolated with KOH, using specific enzyme hydrolysis, ion exchange chromatography (HPAEC) and matrix assisted laser desorption ionisation-time-of-flight (MALDI-TOF) mass spectroscopy, showed that sesame meal xyloglucan (XG) contained XXXG, XXFG and XXLG, and XLLG (named according to Fry et al., 1993) as the major building sub-units in the ratio of 1:0.9:0.3. Hydrolysis with endo-β-(1 → 4)- d-xylanase and analysis of the xylan derived oligosaccharides showed the presence of monomeric xylose (40%), xylobiose (46%) and acidic xylan oligosaccharides containing 4- O-methyl- d-glucuronic acid residues (14%).

Structural elucidation of hemicelluloses from Vetiver grass

Structure of the hemicelluloses extracted from Vetiver grass ( Vetiveria zizanioides Nash) leaves was studied. The monosaccharide compositions and the position of the linkages between monosaccharides in the hemicelluloses were defined by TFA hydrolysis and methylation analysis, respectively. 13C NMR and FT-IR spectroscopic methods gave details of the anomeric linkage configuration and confirmed the structure of the hemicelluloses. The proposed structure of the hemicelluloses from the Vetiver grass is an arabinoxylan mainly consisting of (1→4)-β- d-xylan backbone substituted in O-2 and/or O-3 by single α- l-arabinose residue, single α- d-glucuronic acid residue and/or short chains of sugar residues containing arabinose, xylose and galactose. In addition, the (1→4)-linked β- d-xylopyranosyl residues in the backbone may contain substituted phenolic acids (e.g. ferulic acid and p-coumaric acid). The substituted phenolic acids are esterified via their carboxyl groups to the C-5 hydroxyl of the α- l arabinofuranosyl residues in side chain.