一种木葡聚糖类寡糖的化学合成及生物活性

天然木葡聚糖类寡糖是一类对植物生长具有调节作用的寡糖, 本文以3个单糖组分为原料, 经5步合成了一种木葡聚糖三糖(1)(总产率15%), 以及该三糖的糖苷缀合物1a及其异构体1b. 利用糖基化立体选择性原则, 一步偶联反应同时得到所需的α,β连接产物, 整个合成路线高效简捷. 活性测试结果表明, 3种目标寡糖在1 mg/L浓度下, 对烟草的生长均显示出一定的促进作用, 表明所合成的3种寡糖有望发展成为植物生长促进剂.     

Application of the anomeric samarium route for the convergent synthesis of the C-linked trisaccharide alpha-D-Man-(1-->3)-[alpha-D-Man-(1-->6)]-D-Man and the disaccharides alpha-D-Man-(1-->3)-D-Man and alpha-D-Man-(1-->6)-D-Man

Studies are reported on the assembly of the branched C-trisaccharide, alpha-D-Man-(1-->3)-[alpha-D-Man-(1-->6)]-D-Man, representing the core region of the asparagine-linked oligosaccharides. The key step in this synthesis uses a SmI(2)-mediated coupling of two mannosylpyridyl sulfones to a C3,C6-diformyl branched monosaccharide unit, thereby assembling all three sugar units in one reaction and with complete stereocontrol at the two anomeric carbon centers. Subsequent tin hydride-based deoxygenation followed by a deprotection step produces the target C-trimer. In contrast to many of the other C-glycosylation methods, this approach employes intact carbohydrate units as C-glycosyl donors and acceptors, which in many instances parallels the well-studied O-glycosylation reactions. The synthesis of the C-disaccharides alpha-D-Man-(1-->3)-D-Man and alpha-D-Man-(1-->6)-D-Man is also described, they being necessary for the following conformational studies of all three carbohydrate analogues both in solution and bound to several mannose-binding proteins.     

Tandem exploitation of Helix pomatia glycosyltransferases: facile syntheses of H-antigen-bearing oligosaccharides

Snails from the family Helicidae produce in their albumen glands a highly branched galactan, which consists almost exclusively of D- and L-galactose. The D-Gal residues are glycosydically beta(1-->6)- or beta(1-->3)-linked, whereas the L-Gal moieties are attached alpha(1-->2). Up until the present time, two beta(1-->6)-D-galactosyl transferases and one alpha(1-->2)-L-galactosyl transferase have been identified in a membrane preparation of these glands. These were used to synthesise various oligosaccharides by successive addition of the NDP-activated (NDP=nucleoside-5'-diphosphate) D-Gal or L-Fuc moieties, up to a heptasaccharide by starting from the disaccharide D-Gal-beta(1-->3)-D-Gal-beta(1-->OMe. Even larger oligosaccharides up to a tridecasaccharide were obtained by starting with the hexasaccharide D-Gal-[beta(1-->3)-D-Gal]4-beta(1-->4)-D-Glc as an acceptor substrate. This tandem exploitation process has high potential for the easy introduction of D-Gal and L-Fuc residues into a great variety of oligosaccharides, which can be used in ligand/acceptor studies.     

A comparison of liquid chromatography, capillary electrophoresis, and mass spectrometry methods to determine xyloglucan structures in black currants

Different separation (HPAEC, RP-HPLC, CE) and identification (MALDI-TOF-MS, ESI-MS(n)) techniques were compared to analyse oligosaccharides obtained after incubation of xyloglucan with endo-glucanase. It was possible to analyse xyloglucan oligosaccharides with each technique. Several techniques, including off line (HPAEC-MALDI-TOF-MS) or online (CE-ESI-MS(n), RP-HPLC-ESI-MS(n)) connection provided complementary information on xyloglucan structure. Online CE-MS and RP-HPLC-MS are described for the first time in xyloglucan analysis. Advantages and disadvantages of the techniques for different purposes such as structural characterisation of oligosaccharides or oligosaccharide profiling are discussed. Black currant xyloglucans had a rather simple XXXG-type structure with galactose and fucose containing side chains.     

Structural analysis of underivatized neutral human milk oligosaccharides in the negative ion mode by nano-electrospray MS(n) (part 2: application to isomeric mixtures)

A complex mixture of isomeric neutral oligosaccharides from pooled human milk was analyzed by nano-electrospray ionization (ESI) in a quadrupole ion trap mass spectrometer (QIT-MS) in the negative ion mode. Since deprotonated molecules of neutral oligosaccharides follow distinct fragmentation rules, which have been elucidated by using model compounds (see [1]), spectra obtained from consecutive CID experiments allowed the differentiation of isomers out of this highly complex mixture. With this method new human milk oligosaccharides of previously unknown isomeric structures have been identified, e.g., the occurence of three isomeric fucosylated lacto-N-hexaoses could be determined precisely, which have not been described before: (1) Fuc (alpha1-->2) Gal (beta1-->3) GlcNac (beta1-->3) Gal (beta1-->4) GlcNac (beta1-->3) Gal (beta1-->4) Glc, (2) Gal (beta1-->4) GlcNAc [(alpha1-->3) Fuc] (beta1-->3) Gal (beta1-->4) GlcNac (beta1-->3) Gal (beta1-->4) Glc, (3) Gal (beta1-->4) GlcNAc (beta1-->3) Gal (beta1-->4) GlcNac [(alpha1-->3) Fuc] (beta1-->3) Gal (beta1-->4) Glc.     

Hypolipidemic effects of hydrolyzed xyloglucan

Xyloglucan is found in the primary cell walls of higher plants, where it plays important biological roles. It also exists in certain seeds, for example tamarind, as a reserve polysaccharide. Some xyloglucan oligosaccharides possess plant growth promotion activity. We prepared hydrolyzed xyloglucan from the tamarind seed xyloglucan using endo-(1 → 4)- β -glucanase and examined its effects on lipid metabolism in rats fed a high fat diet. Among the plasma lipids, total lipid, cholesterol, triglyceride and β - lipoprotein were each reduced 14 - 17% by hydrolyzed xyloglucan. Hydrolyzed xyloglucan significantly reduced both adipose tissue weight and plasma GOT. Among the hepatic lipids, total lipid, cholesterol, triglyceride and phospholipid were significantly reduced by hydrolyzed xyloglucan. These hypolipidemic effects may be important for the treatment and prevention of geriatric diseases, including diabetes and cardiac disorders.     

Construction and structural characterization of versatile lactosaminoglycan-related compound library for the synthesis of complex glycopeptides and glycosphingolipids

We have established a facile and efficient protocol for the preparative-scale synthesis of various compound libraries related to lactosaminoglycans: cell surface oligosaccharides composed of N-acetyllactosamine as a repeating disaccharide unit, based on chemical and enzymatic approaches. Substrate specificity and feasibility of a bacterial glycosyltransferase, Neisseria meningitidis beta1,3-N-acetylglucosaminyltransferase (LgtA), were investigated in order to synthesize various key intermediates suited for the construction of mammalian O-glycopeptides and glycosphingolipids containing poly-N-acetyllactosamine structures. Recombinant LgtA exhibited the highest glycosyltransferase activity with strongly basic conditions (pH = 10, glycine-NaOH buffer) and a broad range of optimal temperatures from 20 to 30 degrees C. Interestingly, it was found that LgtA discriminates L-serine and L-threonine and functions both as a core-1 beta1,3-N-acetylglucosaminyltransferase and core-2 beta1,3-N-acetylglucosaminyltransferase toward Fmoc-Ser derivatives, while LgtA showed only core-2 beta1,3-N-acetylglucosaminyltransferase activity in the presence of Fmoc-Thr derivatives. Combined use of LgtA with human beta1,4-galactosyltransferase allowed for controlled sugar extension reactions from synthetic sugar amino acids and gave synthetic lactosaminoglycans, such as a decasaccharide derivative, Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Ser-OH (6), and a dodecasaccharide derivative, Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Ser-OH (9). A partially protected pentasaccharide intermediate, GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Thr-OH (11), was applied for the microwave-assisted solid-phase synthesis of a MUC1-related glycopeptide 19 (MW = 2610.1). The findings suggest that this sugar extension strategy can be employed for the modification of lactosyl ceramide mimetic polymers to afford convenient precursors for the synthesis of various glycosphingolipids.     

Preparation and Characterization of the Enzymatic Degradation Products of the Exopolysaccharide From Klebsiella K13

Oligosaccharides were prepared from the exopolysaccharide of Klebsiella K13 by enzymatic degradation and characterized. A phage-borne depolymerase enzyme was used to degrade the exopolysaccharide of Klebsiella K13. Bio-Gel P4 and P6 were used to purify the oligosaccharide products. The purified oligosaccharides were characterized by HPLC, mass spectroscopy, infrared spectroscopy, and NMR spectroscopy. The monosaccharide constituents of these enzymatic degradation products include D-glucose, D-galactose, D-mannose, and D-glucuronic acid. It was concluded that a pentasaccharide repeating unit with the following structure, as well as its dimer and trimer, was released from the exopolysaccharide: 3,4-O-(1-carboxyethylidene)-β-D-Galp-(1→4)-α-D-GlcpA-(1→3)-β-D-Manp-(1→4)-α-D-Glcp-(1→3)-D-Glcp     

Determination of the hyperfine coupling constant and zero-field splitting in the ESR spectrum of Mn(2+) in calcite

The hyperfine coupling constant (A in units of energy or A' in units of magnetic field) and zero-field splitting (D in units of energy or D' in units of magnetic field) in the ESR spectrum of Mn(2+) in calcite were determined. The spreading of the non-central allowed transitions |3/2, m --> |5/2, m, |1/2, m --> |3/2, m, |-3/2, m --> |-1/2, m and |-5/2, m --> |-3/2, m was analysed and the experimental transitions were attributed. Particular relevance was given to the difference between the two types of resonances, shoulder or divergence, and to their origin (M, m). The analysis explains the presence of a weak shoulder between each of the five central doublets |-1/2, m --> |1/2, m. Six independent methods for calculating the hyperfine coupling constant and five methods for calculating the zero-field splitting, based on the analysis of the allowed and forbidden transitions, were provided. The values of the hyperfine coupling constant range from -93.9 to -94.6 G and those of the zero-field splitting range from -79.5 to -80.5 G. A critical evaluation of the advantages and drawbacks of the 11 methods is included: the best value for A' is -94.30 G and that for D' is -79.95 G.     

Synthesis of a 2,3,4-triglycosylated rhamnoside fragment of rhamnogalacturonan-II side chain A using a late stage oxidation approach

Pectic polysaccharide RG-II, a key component of plant primary cell walls, is known to exist as a dimer formed by means of borate diester cross-links between apiosyl residues of one of its constituent side-chain oligosaccharides. Described herein is the strategy for the synthesis of the branched tetrasaccharide alpha-d-GalA-(1-->2)-[beta-D-GalA-(1-->3)]-[alpha-L-Fuc-(1-->4)]-alpha-L-Rha-OMe, an RG-II fragment that is linked to the apiosyl residue that is thought to be responsible for the borate complexation in RG-II dimer. Iterative glycosylation of the rhamnoside acceptors derived from the key 2,3-orthoacetate of methyl 4-O-methoxybenzyl-alpha-d-rhamnopyranoside afforded the protected tetrasaccharide. The target dicarboxylic acid saccharide was subsequently prepared by removal of protecting groups followed by TEMPO-mediated oxidation of galactopyranosyl residues to galactopyranosyluronic acids.     

Glycosynthase-based synthesis of xylo-oligosaccharides using an engineered retaining xylanase from Cellulomonas fimi

Glycosynthases are synthetic enzymes derived from retaining glycosidases in which the catalytic nucleophile has been replaced. The mutation allows irreversible glycosylation of sugar acceptors using glycosyl fluoride donors to afford oligosaccharides without any enzymatic hydrolysis. Glycosynthase technology has proven fruitful for the facile synthesis of useful oligosaccharides, therefore the expansion of the glycosynthase repertoire is of the utmost importance. Herein, we describe for the first time a glycosynthase, derived from a retaining xylanase, that synthesizes a range of xylo-oligosaccharides. The catalytic domain of the retaining endo-1,4-beta-xylanase from Cellulomonas fimi (CFXcd) was successfully converted to the corresponding glycosynthase by mutation of the catalytic nucleophile to a glycine residue. The mutant enzyme (CFXcd-E235G) was found to catalyze the transfer of a xylobiosyl moiety from alpha-xylobiosyl fluoride to either p-nitrophenyl beta-xylobioside or benzylthio beta-xylobioside to afford oligosaccharides ranging in length from tetra- to dodecasaccharides. These products were purified by high performance liquid chromatography in greater than 60% combined yield. 1H and 13C NMR spectroscopic analyses of the isolated p-nitrophenyl xylotetraoside and p-nitrophenyl xylohexaoside revealed that CFXcd-E235G catalyzes both the regio- and stereo-selective synthesis of xylo-oligosaccharides containing, exclusively, beta-(1 --> 4) linkages.     

Separation and identification of enzymatic sucrose hydrolysis products by high-performance anion-exchange chromatography with pulsed amperometric detection

An accurate carbohydrate analysis method, namely high-performance anion-exchange chromatography with pulsed amperometric detection was successfully applied to the study of sucrose hydrolysis under enzymatic (baker's yeast invertase) conditions. The hydrolysis was monitored by determining sucrose degradation and the corresponding formation of D-glucose, D-fructose and five intermediate fructans using a CarboPac PA-100 (Dionex) analytical anion-exchange column. Highly reproducible results were obtained. The unknown fructans were collected from a semi-preparative CarboPac PA-100 (Dionex) column, neutralized and then desalted on a column containing mixed bed resin AG 501-X8 (D) before identification of the chemical structure. This procedure permitted us to obtain about 20 microg of pure product which is not enough for NMR analysis. Detailed GC-MS analytical data of the methylated compounds indicated that these oligosaccharides were beta-D-Fru-(2 --> 1)-beta-D-Fru-(2 --> 1)-alpha-D-glucopyranoside (1-kestose), beta-D-Fru-(2 --> 6)-alpha-D-glucopyranoside (6-beta fructofuranosylglucose), beta-D-Fru-(2 --> 1)-beta-D-fructofuranoside (inulobiose), beta-D-Fru-(2 --> 6)-beta-D-Fru-(2 --> 1)-alpha-D-glucopyranoside (6-kestose) and beta-D-Fru-(2 --> 6)-alpha-D-Glc-(1 --> 2)-beta-D-fructofuranoside (neokestose) coeluating with a disaccharide.     

The 2-naphthylmethyl (NAP) group in carbohydrate synthesis: first total synthesis of the GlyCAM-1 oligosaccharide structures

Total syntheses of the GlyCAM-1 (glycosylation-dependent cell adhesion molecule-1) oligosaccharide structures: [alpha-NeuAc-(2 --> 3)-beta-Gal-(1 --> 4)-[alpha-Fuc-(1 --> 3)]-beta-(6-O-SO3Na)-GlcNAc-(1 --> 6)]-[alpha-NeuAc-(2 --> 3)-beta-Gal-(1 --> 3)]-alpha-GalNAc-OMe (1) and [alpha-NeuAc-(2 --> 3)-beta-Gal-(1 --> 4)-[alpha-Fuc-(1 --> 3)]-beta-GlcNAc-(1 --> 6)]-[alpha-NeuAc-(2 3)-beta-Gal-(1 --> 3)]-alpha-GalNAc-OMe (2) through a novel sialyl LewisX tetrasaccharide donor are described. Employing sequential glycosylation strategy, the starting trisaccharide was regio- and stereoselectively constructed through coupling of a disaccharide imidate with the monosaccharide acceptor phenyl-6-O-naphthylmethyl-2-deoxy-2-phthalimido-1-thio-beta-D-glucopyranoside with TMSOTf as a catalyst without affecting the SPh group. The novel sialyl Lewisx tetrasaccharide donor 3 was then obtained by alpha-L-fucosylation of trisaccharide acceptor with the 2,3,4-tri-O-benzyl-1-thio-beta-L-fucoside donor. The structure of the novel sialyl Lewisx tetrasaccharide was established by a combination of 2D DQF-COSY and 2D ROESY experiments. Target oligosaccharides 1 and 2 were eventually constructed through heptasaccharide which was obtained by regioselective assembly of advanced sialyl Lewisx tetrasaccharide donor 3 and a sialylated trisaccharide acceptor in a predictable and controlled manner. Finally, target heptasaccharides 1 and 2 were fully characterized by 2D DQF-COSY, 2D ROESY, HSQC, HMBC experiments and FAB mass spectroscopy.     

Differentiation of type 1 and type 2 chain linkages of native glycosphingolipids by positive-ion fast-atom bombardment mass spectrometry with collision-induced dissociation and linked scanning.

Two underivatized glycosphingolipids, Le(b) and Le(y), isomeric in carbohydrate structure (Fuc alpha 1-->2Gal beta 1--> 3[Fuc alpha 1-->4]GlcNAc beta 1-->3Gal beta 1-->4Glc beta 1-->1Cer and Fuc alpha 1-->2Gal beta 1-->4[Fuc alpha 1-->3]GlcNAc beta 1-->3Gal beta 1--> 4Glc beta 1-->1Cer, respectively), were analyzed by positive-ion fast-atom bombardment (FAB) mass spectrometry with high energy collision-induced dissociation (CID) and linked scanning. The two isomers were distinguishable by the abundance of product ions derived from the non-reducing terminal tetrasaccharide fragment via sequential beta-eliminations of vicinally linked saccharide residues. Following earlier studies from other laboratories, which have dealt primarily with positive-ion FAB-CID mass spectrometry of simple model oligosaccharides, these results exemplify the practical application of two-sector methodology to underivatized complex glycoconjugates commonly encountered in the biomedical field.     

Rapid analysis of unsaturated acidic xylooligosaccharides from kraft pulps using capillary zone electrophoresis

Several acidic xylooligosaccharides containing unsaturated “hexenuronic acid” units, i.e. 4-deoxy-L -threo-hex-4-enopyranosy-lurinic acid (4-ΔU) units, were separated as their alditol derivatives by capillary zone electrophoriesis in 438 mM borate buffer (pH 10.3) and were detected selectively at the μM level on-column UV detection at 232 nm. These acidic oligosaccharides were obtained from birch and pine kraft pulps on enzymatic hydrolysis with endoxylanases and subsequent treatment with other Trichoderma reesei enzymes. Under the conditions empolyed, acidic 4-ΔU-containing xylooligosaccharides with a molecular size renging from trisaccharides up to nonasaccharides could be separated. Oligosaccharides with higher molecular mass were detected first. Two 4-ΔU-xylotetraose isomers, with the 4-ΔU-group linked to different xylose units in the iligosaccharide backbone, could be resolved from each other with a resolution of about 1. By using a disaccharide (4-ΔU α-(1 → 4) linked to N-acetyl glucosamine) as a model compound the minimum detectable concentration was determined as 10 μM.     

Structural analysis of underivatized neutral human milk oligosaccharides in the negative ion mode by nano-electrospray MS(n) (part 1: methodology)

Underivatized neutral oligosaccharides from human milk were analyzed by nano-electrospray ionization (ESI) using a quadrupole ion trap mass spectrometer (QIT-MS) in the negative-ion mode. Under these conditions neutral oligosaccharides are observed as deprotonated molecules [M-H]- with high intensity. CID-experiments of these species with the charge localized at the reducing end lead to C-type fragment ions forming a "new" reducing end. Fragmentations are accompanied by cross-ring cleavages that yield information about linkages of internal monosaccharides. Several isomeric compounds with distinct structural features, such as different glycosidic linkages, fucosylation and branching sites were investigated. The rules governing the fragmentation behavior of this class of oligosaccharides were elucidated and tested for a representative number of certain isomeric glycoforms using the MS/MS and MS(n) capabilities of the QIT. On the basis of the specific fragmentation behavior of deprotonated molecules, the position of fucoses and the linkage type (Gal beta-->3 GlcNAc or Gal beta1-->4 GlcNAc) could be determined and linear and branched could be differentiated. Rules could be established which can be applied in further investigations of these types of oligosaccharides even from heterogenous mixtures.     

Synthesis of the C-linked disaccharide alpha-D-Man-(1-->4)-D-Man employing a SmI(2)-mediated C-glycosylation step: en route to cyclic C-oligosaccharides

Investigations are reported on the assembly of the C-linked disaccharide alpha-D-Man-(1-->4)-D-Man, representing the first steps in our projected synthesis of a cyclic C-oligomer containing repeating units of this C-dimer. The key step in this synthesis uses a SmI(2)-mediated coupling of 2,3,4,6-tetra-O-benzyl-alpha-D-mannopyranosyl 2'-pyridyl sulfone with a C4-formyl branched mannopyranoside unit, affording the C-disaccharide derivative with complete stereocontrol at the two new stereogenic centers. Subsequently, a modified tin hydride based deoxygenation produced the target carbohydrate analogue. The synthesis of the C4-formyl monosaccharide makes use of a stereoselective radical-based allylation followed by double bond migration and ozonolysis.     

Synthesis of heparin oligosaccharides and their interaction with eosinophil-derived neurotoxin

A convenient route for the synthesis of heparin oligosaccharides involving regioselective protection of D-glucosamine and a concise preparation of rare L-ido sugars from diacetone α-D-glucose is described. Stereoselective coupling of a D-glucosamine-derived trichloroacetimidate with a 1,6-anhydro-β-L-idopyranosyl 4-alcohol gave the desired α-linked disaccharide, which was used as repeating unit for dual chain elongation and termination. Stepwise assembly from the reducing to the non-reducing end with a D-glucosamine-derived monosaccharide as starting unit furnished the oligosaccharide skeletons having different chain lengths. A series of functional group transformations afforded the expected heparin oligosaccharides with 3, 5 and 7 sugar units. Interaction of these oligosaccharides with eosinophil-derived neurotoxin (EDN), a cationic ribonuclease and a mediator produced by human eosinophils, was further investigated. The results revealed that at 5 μg mL(-1), the heptasaccharide has sufficiently strong interference to block EDN binding to Beas-2B cells. The tri- and pentasaccharides have moderate inhibitory properties at 50 μg mL(-1) concentration, but no inhibition has been observed at 10 μg mL(-1). The IC(50) values of the tri-, penta- and heptasaccharides are 69.4, 47.2 and 0.225 μg mL(-1), respectively.     

Synthetic studies on glycosphingolipids from protostomia phyla: synthesis of amphoteric glycolipid analogues from the porcine nematode,Ascaris suum

A novel amphoteric glycosphingolipid, cholinephosphoryl-(-->6)-beta-D-GlcpNAc-(1-->3)-beta-D-Manp-(1-->4)-beta-D-Glcp-(1-->)-Cer, isolated from the porcine parasitic nematode, Ascaris suum, may be expected to be involved in host-parasite interactions. This glycosphingolipid analogue containing octyl residue in place of ceramide was synthesized as follows: The key reaction of this synthetic procedure is the formation of a intramolecular aglycon delivery (IAD) approach for beta-selective mannosylation. Then, a coupling of phosphocholine group at the position C-6' of 16 was attempted using 2-chloro-2-oxo-1,3,2-dioxaphospholane, followed by reaction of the resulting cyclic phosphate intermediate with anhydrous trimethylamine to give 17. Subsequent debenzylation and debenzylidenation afforded target compound (2).     

Structural characterization of neutral oligosaccharide mixtures through a combination of capillary electrochromatography and ion trap tandem mass spectrometry

A CEC/ESI-MS/MS combined system has been developed for the separation and on-line structural analysis of neutral oligosaccharides. Various types of isomeric oligosaccharides were first successfully separated by CEC using polar monolithic columns, while the on-line tandem mass spectrometry has been explored to differentiate and elucidate the structures of isomeric oligosaccharides. The experimentally obtained tandem spectra usually provide sequence, branching, and linkage information. Oligosaccharide isomers with a different monomeric composition and branching showed different patterns of glycosidic linkage cleavage (B- and Y-ion series), allowing us to deduce their sequence and branching points. Isomers with different linkages were distinguished by identifying cross-ring fragment ions (A-ion series). While (1-->4) linkages yielded dominant (0,2)A ions, (1-->6) linkages showed an extensive and complete cross-ring cleavage series: (0,2)A, (0,3)A, and (0,4)A ions. Although the anomeric configurations and monosaccharide identification are rarely obtained from tandem MS, the relevant mixture components can be completely resolved with high-efficiency CEC columns featuring a polar functionality.