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Alginates consist of distinct blocks with different physical properties. This short review focuses on research carried out in Trondheim related to the early discovery of the block structure, their isolation, their different chemical and physical properties, and how they recently are utilized in diblock polysaccharides to obtain new nanostructuring properties.

Renewable resources can provide a range of different polysaccharide blocks that can be used to prepare new types of stimuli-responsive polysaccharide-based block copolymers. Alginates are natural polysaccharides widely used as biomaterials. Functional properties depend on the content and distribution of the two 4-linked monomers (β-D-mannuronate (M) and α-L-guluronate (G)). Blocks of L-guluronate (Gn) are responsible for cooperative binding of calcium ions and hydrogel formation. Incorporation of such blocks in block polysaccharide copolymers would represent a new class of engineered, Ca-sensitive biomacromolecules. Dioxyamines and dihydrazides have recently been shown to be well suited for preparation of block polysaccharide structures. Here we first show that when applied to alginate blocks (Gn and Mn) the two types are both very reactive, but the detailed distribution of acyclic (E)- and (Z)-forms and cyclic N-pyranosides, reaction kinetics, conjugate stability, and the rate of Schiff base reduction with α-picoline borane differ considerably, also compared to other polysaccharides. Hence, alginate specific protocols were developed. The linkers introduce a highly flexible joint in otherwise semiflexible Gn-based diblocks. This was demonstrated by SEC-MALS using a symmetrical Gn-b-Gn diblock, which in solution can best be described according to a broken rod model. Ca-Induced self-assembly of Gn-b-dextran diblocks was studied by dynamic light scattering, demonstrating that well defined nanoparticles could be prepared for certain combinations of chain lengths. Taken together, this approach provides a new class of engineered, stimuli-responsive block polysaccharide copolymers solely based on natural resources.

Sequential periodate-chlorite oxidation of sodium hyaluronate to 2,3-dicarboxylated hyaluronate (DCH), a novel biocompatible and highly functionalized derivative bearing additional pair of COOH groups at C2 and C3 carbons of oxidized ᴅ-glucuronic acid units, is investigated. The impact of various reaction parameters (time, oxidizer concentration, and molar amount) on DCH's composition, molecular weight, degree of oxidation, and cytotoxicity are investigated to guide the synthesis of DCH derivatives of desired properties. Subsequently, fully (99%) and partially (70%) oxidized DCH derivatives were compared to untreated sodium hyaluronate in terms of anticancer drug cisplatin loading efficacy, carrier capacity, drug release rates, and cytotoxicity towards healthy and cancerous cell lines. DCH derivatives were found to be superior in every aspect, having nearly twice the carrier capacity, significantly slower release rates, and higher efficacy. DCH is thus a highly interesting hyaluronate derivative with an adjustable degree of oxidation, molecular weight, and great potential for further modifications.

Most polysaccharides used in polysaccharide-based block copolymers are attached to the second block through the reducing end, due to the few and highly polysaccharide specific non-reducing end (NRE) functionalisation methods available. Chitin oligomers, prepared by nitrous acid degradation of chitosan (AnM) can, however, be selectively oxidised by periodate since they only possess a single vicinal diol in the NRE residue. Here, we show that both aldehydes formed after oxidation are highly reactive towards bifunctional oxyamines and hydrazide linkers. Sub-stochiometric amounts of linkers resulted in conjugation of AnM oligomers through both chain termini to yield a discrete distribution of 'polymerised' oligomers. Such chitin-based block polymers were, in contrast to chitins of the same chain lengths, water-soluble. Oxidised AnM oligomers, functionalised at both termini can also enable the preparation of more complex block polysaccharides such as ABA- or ABC-type.

International audience; Diblock oligosaccharides based on renewable resources allow for a range of new but, so far, little explored biomaterials. Coupling of blocks through their reducing ends ensures retention of many of their intrinsic properties that otherwise are perturbed in classical lateral modifications. Chitin is an abundant, biodegradable, bioactive, and self-assembling polysaccharide. However, most coupling protocols relevant for chitin blocks have shortcomings. Here we exploit the highly reactive 2,5-anhydro-D-mannose residue at the reducing end of chitin oligomers obtained by nitrous acid depolymerization. Subsequent activation by dihydrazides or dioxyamines provides precursors for chitin-based diblock oligosaccharides. These reactions are much faster than for other carbohydrates, and only acyclic imines (hydrazones or oximes) are formed (no cyclic N-glycosides). α-Picoline borane and cyanoborohydride are effective reductants of imines, but in contrast to most other carbohydrates, they are not selective for the imines in the present case. This could be circumvented by a simple two-step procedure. Attachment of a second block to hydrazide- or aminooxy-functionalized chitin oligomers turned out to be even faster than the attachment of the first block. The study provides simple protocols for the preparation of chitin-b-chitin and chitin-b-dextran diblock oligosaccharides without involving protection/deprotection strategies.

Engineered block polysaccharides is a relatively new class of biomacromolecules consisting of chemical assembly of separate block structures at the chain termini. In contrast to conventional, laterally substituted polysaccharide derivatives, the block arrangement allows for much higher preservation of inherent chain properties such as biodegradability and stimuli-responsive self-assembly, while at the same time inducing new macromolecular properties. Abundant, carbon neutral, and even recalcitrant biomass is an excellent source of blocks, opening for numerous new uses of biomass for a wide range of novel biomaterials. Among a limited range of methodologies available for block conjugation, bifunctional linkers allowing for oxyamine and hydrazide ‘click’ reactions have recently proven useful additions to the repertoire. This article focuses the chemistry and kinetics of these reactions. It also presents some new data with the aim to provide useful protocols and methods for general use towards new block polysaccharides.

Schizophyllan and scleroglucan are water-soluble polysaccharides having repeating units consisting of three β-1,3-linked glucose residues in the main chain and a single β-1,6-linked glucose residue as the side chain. This polysaccharide dissolves as a triple helix in an aqueous solution and shows a cooperative order-disorder transition between the side chain and solvent molecules while retaining the triple helical conformation. Periodate and subsequent chlorite oxidations selectively modify the side chain glucose to provide the corresponding dicarboxylate units. Optical rotation measurements and differential scanning calorimetry were performed on carboxylated schizophyllan/scleroglucan (‘sclerox’) samples to investigate the effects of the degree of carboxylation on the order-disorder transition in deuterium oxide with 0.1 M NaCl. The transition curves for the sclerox samples are strongly dependent on the degree of carboxylation. The modified side chains cannot take the ordered structure, resulting in a reduction of the transition enthalpy. The transition temperature for carboxylated schizophyllan becomes lowered and the transition curve broadens with increasing the degree of carboxylation. The permanent disordered units are included in a trimer by the carboxylation to inhibit a long sequence of the ordered units. © 2017. This is the authors’ accepted and refereed manuscript to the article. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/

International audience; Reducing end activation of poly-and oligosaccharides by bifunctional dioxyamines and dihydrazides enables aniline-free and cyanoborohydride-free conjugation to aldehyde-containing molecules, particles and surfaces without compromising the chain structure. Chitosans are due to their polycationic character, biodegradability, and bioactivity important candidates for conjugation. Here, we present a kinetic and structural study of the conjugation of a dioxyamine and a dihydrazide to enzymatically produced chitooligosaccharides ranging from N,N'-diacetylchitobiose to a decamer, all having N-acetyl D-glucosamine at the reducing end. Conjugation of the dioxyamine resulted in mixtures of (E)-and (Z)-oximes and β-N-pyranoside, whereas the dihydrazide yielded cyclic N-glycosides. Reaction kinetics was essentially independent of DP. Stable secondary amines were in both cases obtained by reduction with α-picoline borane, but higher temperatures were needed to obtain acceptable reduction rate. Comparison to dextran oligomers shows that the nature of the reducing end strongly influences the kinetics of both the conjugation and reduction.

The aim of this paper was to perform a comprehensive characterization of polysaccharides isolated from the interior (IOI) and exterior (IOE) parts of the fungus Inonotus obliquus. Pre-extraction with DCM and MeOH, followed by water and alkali extraction and ethanol precipitation gave two water extracts and two alkali extracts. Neutral and acidic polysaccharide fractions were obtained after anion-exchange chromatography of the water extracts. The neutral polysaccharides (60–73 kDa) were heterogeneous and branched and consisted of a (1 → 3)-linked β-Glc backbone with (1 → 6)-linked kinks in the chain at approximately every fifth residue, with branches of (1 → 6)-linked β-Glc in addition to substantial amounts of (1 → 6)-linked α-Gal with 3-O-methylation at about every third Gal residue. The acidic polysaccharide fractions (10–31 kDa) showed similar structural motifs as the neutral fractions differing mainly by the presence of (1 → 4)-linked α-GalA and α-GlcA. β-Xyl, α-Man and α-Rha were also present in varying amounts in all fractions. No major structural differences between the IOI and IOE fractions were observed. An alkaline polysaccharide fraction (>450 kDa) was obtained from the IOI alkali extract, and consisted mainly of (1 → 3)- and (1 → 6)-linked β-Glc and (1 → 4)-linked β-Xyl. Several of the fractions showed in vitro immunomodulatory effect by increasing NO production in the murine macrophage and dendritic cell lines J774.A1 and D2SC/1. Most fractions managed to increase NO production only at the highest concentration tested (100 μg/ml), while the neutral fraction IOE-WN activated potent NO production at 10 μg/ml and was considered the most promising immunomodulating fraction in this study.

Alginate-based biomaterials and medical devices are commonly subjected to γ-irradiation as a means of sterilization, either in the dry state or the gel (hydrated) state. In this process the alginate chains degrade randomly in a dose-dependent manner, altering alginates’ material properties. The addition of free radical scavenging amino acids such as histidine and phenylalanine protects the alginate significantly against degradation, as shown by monitoring changes in the molecular weight distributions using SEC-MALLS and determining the pseudo first order rate constants of degradation. Tris buffer (0.5 M), but not acetate, citrate, or phosphate buffers had a similar effect on the degradation rate. Changes in pH itself had only marginal effects on the rate of alginate degradation and on the protective effect of amino acids. Contrary to previous reports, the chemical composition (M/G profile) of the alginates, including homopolymeric mannuronan, was unaltered following irradiation up to 10 kGy. © American Chemical Society 2014. This is the authors accepted and refereed manuscript to the article.

Tomofuji Y., Yoshiba K., Christensen B., et al. Single-chain conformation of carboxylated schizophyllan, a triple helical polysaccharide, in dilute alkaline aqueous solution. Polymer, 185, 121944. © 2019 Elsevier Ltd https://doi.org/10.1016/j.polymer.2019.121944., Synchrotron-radiation small-angle X-ray scattering measurements were carried out for a schizophyllan (SPG) sample with the weight-average molar mass Mw of 340 kg mol−1 and five carboxylated SPG (sclerox) samples with different degrees of substitution (DS) ranging from 0.18 to 0.45 in 200 mM aqueous NaOH including 10 mM NaCl to determine Mw, the second virial coefficient A2, the particle scattering function P(q), and the radius of gyration ⟨S2⟩. Positive A2 values indicated that this alkaline solvent was a good solvent for all polysaccharide samples investigated. The resultant Mw values (~100 kg mol−1) were much smaller than that for the trimer in aqueous NaCl at neutral pH, indicating all SPG and sclerox samples dissolved as single chains in the alkaline solvent. Both P(q) and ⟨S2⟩ were consistently explained by the wormlike chain model. The obtained parameters were almost independent of DS. While the chain stiffness (12–18 nm) in terms of the Kuhn segment length or twice the persistence length was similar to those for the other single chain polysaccharides, the helix pitch per residue (0.1 nm) was quite smaller than the trimer state (0.3 nm). This shrunken main-chain helical structure is most likely due to hydrophobic interactions between helical main chain and side groups.

Degradation of alginate remains a critical issue to allow predictable biological performance upon implantation of alginate-based materials. Therefore, the objective of the current study is to compare the effects of gamma-irradiation (dry state, 20-80 kGy), partial (1 and 4%) periodate oxidation (aqueous solution), and autoclaving (dry state) on the molecular weight of alginate, as well as the degradation behavior of alginate-based composites. The results show that gamma-irradiation is by far the most destructive technique characterized by strongly reduced molecular weights and rapid loss of composite integrity upon soaking in simulated body fluid. Partial periodate oxidation is less destructive as characterized by more moderate decreases in molecular weight, but the production of hydrolytically labile bonds compromises the integrity of the resulting composites. Autoclaving is shown to be a powerful tool to reduce the molecular weight of alginate in a controllable and mild manner without compromising the integrity of the resulting alginate-hydroxyapatite composites, simply by increasing the number of repetitive autoclaving cycles.

The molecular weight distributions of artificially aged transformer papers were studied using SEC–MALLS with 0.5 % DMAc/LiCl as solvent, providing chain length distributions and averages. The slow dissolution and presence of non-cellulosic contaminants tend in some cases to introduce errors in the DPw estimates, and a modified data processing procedure was developed to correct for extraneous scattering. Data were compared to the intrinsic viscosities obtained in 0.5 M Cuen and the ‘viscosimetric DP’ calculated thereof according to ISO 5351. DPvisc was 2–3 times lower than the weight average DP (DPw), but closer to or slightly above the number average DP (DPn) obtained by SEC–MALLS in the case of pure cellulose. Ageing of transformer papers were in some cases associated with changes in the polydispersity (DPw/DPn). The apparent degradation rate (defined as δ(1/DP)/δt) gradually decreased with time, resulting in a tendency for a ‘level-off’ DP in the range 200–300 for DPvisc and DPn, and roughly 1,000 for DPw.

Biomaterials based on peptide-coupled alginates must provide both optimal biological environments and tuneable stability/degradation profiles. The present work investigates the degradation pattern and kinetics of peptide-coupled alginates prepared via the periodate oxidation reductive amination route. Alginates degrade slowly (non-enzymatically) under physiological conditions by acid-catalysed hydrolysis and alkali-catalysed β-elimination, both operating simultaneously but dominated by the latter. While periodate oxidation alone largely increases the rate of β-elimination, substitution restores the susceptibility towards β-elimination to that of the parent alginate. For acid hydrolysis the rate of depolymerization is proportional to the degree of substitution, being approximately one order of magnitude larger than the parent alginate, but still lower than for the corresponding materials with fully reduced dialdehydes. Results also suggest a composition-dependent preference for substitution at C2 or C3. These results demonstrate how the various chemistries introduced by the coupling provide useful means to tune the biodegradability profiles. © 2016. This is the authors’ accepted and refereed manuscript to the article. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/

Water-soluble (1→3)-β- d -glucans with 1,6-linked branches (SBG), originally isolated from the cell walls of Saccharomyces cerevisiae and partially depolymerised to a weight average degree of polymerisation (DP w ) in the range 120–160 for optimal performance in wound healing applications, were studied by dynamic light scattering (DLS), SEC MALLS and AFM. Results indicate that dilute aqueous SBG solutions (1 μg/ml to 3 mg/ml) contain higher order structures with a very wide size distribution in water (10–500 nm), corresponding to a mixture of single chains, multi-chain aggregates including triple-stranded motifs, and particulate materials. The latter were enriched in longer chains compared to non-particulate fractions. The size distribution of SBG aggregates shifted to slightly lower values upon heating, but showed hysteresis upon cooling. AFM images prepared from very dilute aqueous solution (1–5 μg/ml) analysis showed by comparison to other (1→3)-β- d -glucans that some of the structures were the triple helical species coexisting with larger aggregates and single chains, in contrast to carboxymethylated SBG, which contained predominantly single chains. The ability to control the aggregation behaviour of SBG enables tailoring of the physical, and possibly bioactive, properties of SBG preparations.

Alginates in the format of scaffolds provide important functions as materials for cell encapsulation, drug delivery, tissue engineering and wound healing among others. The method for preparation of alginate-based foams presented here is based on homogeneous, ionotropic gelation of aerated alginate solutions, followed by air drying. The method allows higher flexibility and better control of the pore structure, hydration properties and mechanical integrity compared to foams prepared by other techniques. The main variables for tailoring hydrogel properties include operational parameters such as degree of aeration and mixing times and concentration of alginate, as well as macromolecular properties such as the type of alginate (chemical composition and molecular weight distribution). Exposure of foams to γ-irradiation resulted in a dose-dependent (0-30 kGy) reduction in molecular weight of the alginate and a corresponding reduction in tensile strength of the foams.

Water-soluble (1→3)-β-D-glucans with 1,6-linked branches (SBG), originally isolated from the cell walls of Saccharomyces cerevisiae and partially depolymerised for optimal performance in wound healing applications, were studied by size exclusion chromatography (SEC) with multi-angle laser light scattering (MALLS) detector and a viscosity detector at both high and ambient column temperatures. The strongly aggregating materials could be dispersed as single chains in water following partial carboxymethylation (degree of substitution (DS) 0.51 or higher). Lower DS (0.23) also dispersed as single chains provided a column temperature of 80 °C was applied. Reduction of reducing ends prior to carboxymethylation was required to avoid alkaline peeling and hence to obtain correct molecular weight distributions of the native material. DS was determined using (13)C NMR and potentiometric titration (range 0.23-0.91). Further analysis of CM-SBG in the single chain state suggested a randomly coiled behaviour with marginal influence of the branches in terms of macromolecular dimensions, which were close to those of CM-curdlan. The result of the investigation is a simple and reliable protocol for preparing undegraded and un-aggregated SBG derivatives, which are well suited as a standard analysis of the molecular weight distribution of SBG-like molecules.

The localisation of pectin in Sphagnum moss leaves and its role in preservation has been investigated. Light microscopy using ruthenium red to detect pectin in whole and sectioned Sphagnum papillosum leaves revealed it is abundant in hyaline cell walls, fibrils, papillae, chlorophyllous cell walls and thickenings around hyaline cell pores. Transmission electron microscopy of ultrathin cell walls labelled with poly- l -lysine colloidal gold revealed pectin was distributed throughout the cell wall. The preservative/microbiocidal properties of these pectins are explained by the acid-dissociation properties of galacturonic acid carboxyls and their incorporation in the unique cell arrangement of the Sphagnum leaf. Liquid from a salmon fillet absorbed into S. papillosum leaves and incubated at room temperature for 22 h had a pH around 4.85, was dominated by Lactobacillus sp. and smelled fresh compared to experimental controls. Chlorite-treated Sphagnum leaves could have a potential as a food tray pad that absorbs liquid and prevents the growth of spoilage bacteria inside it.

Limited periodate oxidation (0–8%) of alginates results in enhanced degradability, extending the range of applications of alginates as biomaterials, e.g. in tissue engineering. Oxidation produces dialdehydes that are highly sensitive to alkaline β-elimination, even under physiological conditions (pH 7.4, 37 °C). Although all periodate is consumed during limited oxidation, not all of the resulting dialdehydes are equally degradable, leading to a characteristic level off in molecular weight upon prolonged degradation. A significant fraction (20–50%) of the oxidized residues were resistant even at pH 10.4. We tentatively assign this finding to the presence of particularly stable intramolecular hemiacetals known to form in periodate oxidized alginates (POA). The enhanced degradation of POA persists into the gel state, allowing tailoring of more biodegradable alginate gels. The activation energy for β-elimination of dialdehydes was 88 kJ/mol compared to 113 kJ/mol for unoxidized residues, presumably reflecting the absence of strain in the non-cyclic transition state of the former.

Fully de-N-acetylated chito-oligomers were prepared by de-amination of chitosan using nitrous acid (HNO2). The dimer to hexamer fractions of chitosan oligomers were fractionated with preparative size-exclusion chromatography (SEC) and characterized using 1H NMR spectroscopy. The purified oligomers were polymerized by reductive N-alkylation by the action of NaCNBH3, in a reaction between the terminal 2,5-anhydro- d -mannofuranose (M-unit) and the primary amines of the repeating units (1 → 4) bound β- d -glucosamine (D-unit) resulting in a covalent bond and formation of secondary amines. The polymerized products were studied by SEC-RI and SEC-MALLS-RI and were confirmed to be branched reaching molecular weights of up to 10 000 g/mol. Analysis by 1H NMR spectroscopy confirmed that the chitosan oligomers had been branched by formation of secondary amines. Fully de-N-acetylated chitosan (FA

We studied the effect of the mole fraction ( F M ) of β- d -mannuronate residues in alginate chains and the weight-average molecular weight ( M w ) of alginates on intestinal immunological activity through Peyer's patch cells of C3H/HeJ mice. Over most of the range of M w (30,000–690,000), alginates with high mannuronate content (high-M alginates, F M 0.69–0.86) showed immunological activity, but alginates with F M lower than 0.31 did not. For high-M alginates with F M = 0.78 extracted from Laminaria japonica , those with M w lower than 200,000 showed the highest activity.

The paper presents a new investigation of a native and a hydrophobically modified hyaluronan (HMHA) C8NH-HA100 interactions with surfactants SDS, DTAB, and a dye polarity probe Coomassie Brilliant Blue (CBB), and their macromolecular properties in aqueous environment using isothermal titration calorimetry (ITC) and SEC-MALLS. A novel alkylated HA derivative was prepared by the introduction of octyl chains onto –OH groups of the HA d-glucuronic acid unit via carbamate bond at a high degree of substitution preserving the HA carboxylic groups. Large and abrupt endothermic enthalpy changes (ΔH) during the titration of HA by DTAB evidence a strong electrostatic interaction and a formation of complex aggregates between HA and DTAB micelles. Heat effects of HA–SDS titration are moderate and reveal a lowering of the surfactant CMC in the presence of HA. The C8NH-HA100 displays binding with CBB in terms of exothermic enthalpy changes, which is ascribed to hydrophobic interactions, while such binding is not observed for HA. The assessment of the polysaccharides backbone stiffness and the coefficient of the Mark–Houwink–Sakurada plot (MHS), a, by SEC-MALLS shows a little alternation of the HA backbone stiffness after its modification.

Novel alginates prepared by independent control of chain stiffness and distribution of G-residues : Structure and gelling properties

The carbonyl content of a pectic polysaccharide from Sphagnum papillosum (sphagnan) and periodate oxidised alginates was investigated using three different carbonyl labelling strategies combined with size-exclusion chromatography (SEC) with multi-angle laser light scattering (MALLS) and on-line fluorescence or off-line tritium detection. The labelling strategies were tritium incorporation via NaB3H4 reduction, and fluorescent labelling with carbazole carbonyl oxyamine (CCOA), or 2-aminobenzamide (2-AB), respectively. Carbonyl quantification was based on labelled pullulan, dextran and alginate standards possessing only the reducing end carbonyl group. As a result the carbonyl distribution in the polysaccharides could be determined. In sphagnan it was found that the carbonyl content increased with increasing molecular weight, whereas in periodate oxidised alginate the carbonyl content was as expected independent of the molecular weight. The methods proved useful for carbonyl detection in water soluble polysaccharides in general. The tritium incorporation method was preferred for alkali stable polysaccharides, while the CCOA method was most suitable for acid stable polysaccharides with low carbonyl content. The 2-AB method is applicable for all polysaccharides tested with varying carbonyl content; however, it lacks the ability to detect ketone functionalities.

Chitosans with different chemical compositions ( F A = 0.05–0.65) and different molecular weights ( M w = 36.000–460.000) were studied by size-exclusion chromatography combined with multi-angle laser light scattering (MALS) and viscosity detectors to provide R G – M and [ η ]– M data as basis for further analysis using the wormlike chain model. In both cases intrinsic persistence lengths ( q 0 ) in the range 5.1–7.6 nm were obtained, with little or no detectable dependence on F A . These values are significantly lower than values obtained for alginates (12 nm), including homopolymeric mannuronan, using the same approach. This finding is also corroborated by differences in the Smidsrod B -parameter, confirming that chitosans are less extended than alginates, despite the similarity in basic chain geometry (cellulose type) and linear charge density. Partial periodate oxidation of chitosans led to a pronounced increase in the chain flexibility as shown by a gradual decrease in persistence length, approaching 2 nm for the most oxidised samples.

The purpose of this research was to evaluate if pectin-like polysaccharides, collectively known as sphagnan, extracted by acid hydrolysis from the leaves of Sphagnum moss have a unique ability to react with phenylhydrazine, o-phenylenediamine or collagen. A previous assay for determination of carbonyl groups in sphagnan by reaction with phenylhydrazine was disproved due to spectrophotometric interference from furans. The actual carbonyl content of sphagnan is estimated to be much less than previously thought. NMR spectroscopy showed that small amounts of o-phenylenediamine and/or its oxidation products bind to sphagnan probably via imine formation, but evidence of quinoxaline formation was inconclusive. Sphagnan-gelatin mixtures formed complex coacervates at pH 2.0-4.8 at low ionic strength, which is typical of electrostatic polyelectrolyte interactions, rather than covalent carbonyl-amine reactions. Measurements of hydrothermal stability and collagenase-degradation of sphagnan-treated hide powder collagen suggest that sphagnan is a poor tanning agent. The results indicate the suggested preservative properties of sphagnan are not related to tanning.

Three water-soluble, non-ionic extracellular polysaccharides (EPS) obtained from lactic acid bacteria (S. thermophilus THS, L. helveticus K16 and S. thermophilus ST1) were subjected to a comparative study by means of multidetector size-exclusion chromatography, providing distributions and averages of molar masses, radii of gyration and intrinsic viscosities. All polysaccharides displayed random coil character. Further analysis of the data reveals differences in chain stiffness and extension that could be well correlated to structural features. The calculated persistence lengths ranged from 5 to 10 nm and fall within the range typical for many single-stranded bacterial or plant polysaccharides. The ST1 polysaccharide had the highest molar mass but the lowest persistence length, which is attributed to the presence of the flexible (1→6)-linkage in the main chain.

Mild acid hydrolysis was used to release polysaccharides from the chlorite-treated leaves of Sphagnum papillosum . These polysaccharides, collectively known as sphagnan, were physically and chemically characterised. No evidence was found of the presence of a previously described 5-keto- d -mannuronic acid (5-KMA) monomer in sphagnan, or in the chlorite-treated leaves from which sphagnan was extracted. After fractionation by anion-exchange chromatography the majority of polysaccharides were similar to rhamnogalacturonan I-type pectins. They were highly branched and had a weight average molecular weight of 3.9 × 10 g/mol.

A series of alginates isolated from the stem and leaf of a brown algae (Laminaria hyperborea), bacterial mannuronan, in vitro epimerized mannuronans, and periodate oxidized alginates were analyzed by size-exclusion chromatography (SEC) combined with online multiangle laser light scattering (MALS) and viscometry (collectively abbreviated SMV). Selected samples were also analyzed off-line using low-angle laser light scattering and capillary viscometry. Excellent agreement between the two methods was obtained for properly purified samples. In contrast, abnormal results were obtained for some industrial samples due to the presence of particulate material. Naturally occurring alginates and in vitro epimerized mannuronans were found to obey essentially the same RG-M and [eta]-M relations, and hence, the same Mark-Houwink-Sakurada (MHS) equations (valid for I = 0.10 M): 20 000 g/molM100 000 g/mol, [eta] = 0.0054 .M(1.00); 100 000 g/molM1 000 000 g/mol, [eta] = 0.071 .M(0.89). Application of the wormlike chain model to the [eta]-M data obtained by SMV yielded persistence lengths (q) of 15 nm for all alginates at an ionic strength of 0.17 M. Intrinsic viscosities corresponding to infinite ionic strength were estimated on the basis of Smidsrød's B-parameter, and the wormlike chain model then yielded q = 12 nm. Periodate oxidized alginates showed, in contrast, a pronounced decrease in persistence length with increasing degree of oxidation, reaching values below 4 nm at 44% oxidation. Periodate oxidation also resulted in some depolymerization, even in the presence of a free-radical scavenger.

Even though size exclusion chromatography (SEC) with post column addition of calcofluor (SEC-calcofluor) has been used for the determination of cereal β-glucan molar mass in foods for many years, there is a lack of systematic evaluation of the method. To address this issue a set of suitable β-glucan standards were generated by preparative SEC and their molar mass characteristics were determined by analytical multi-detection SEC (refractive index (RI), light scattering). Each standard was then analysed by SEC-calcofluor at three different labs. As a direct comparison, the analyses were repeated with a RI detector. For SEC-calcofluor accurate measurements of weight average molar mass (Mw) can be made for β-glucan populations within 10-500×10(3)g/mol. Above this molar mass threshold there is an increasing tendency for underestimation of Mw. Precipitation of some β-glucan-calcofluor complexes may have delayed their transport into the detector.Copyright © 2015 Elsevier Ltd. All rights reserved. Inter-laboratory evaluation of SEC-post-column calcofluor for determination of the weight-average molar mass of cereal β-glucan | Request PDF. Available from: https://www.researchgate.net/publication/273507433_Inter-laboratory_evaluation_of_SEC-post-column_calcofluor_for_determination_of_the_weight-average_molar_mass_of_cereal_b-glucan [accessed Sep 18 2018].

Carboxylated schizophyllan (sclerox) samples of different degrees of oxidation were molecularly characterized by size exclusion chromatography equipped with a multi-angle light scattering detector (SEC-MALS) in 0.10 M aqueous NaCl solution. The molar mass distribution obtained by SEC-MALS shows that sclerox of low degree of oxidation is dissolved mainly as the trimer, whereas the trimer and single chain coexist in solution of sclerox of high degree of oxidation. The trimer of sclerox is much more flexible than the fully ordered triple helix of the parent schizophyllan and easily dissociates into single chains upon heating. © 2015. This is the authors’ accepted and refereed manuscript to the article. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/

Oat β-glucan was partially degraded with (1→4)-β- d -glucan 4-glucanohydrolase for different periods of time. Weight average molecular weight and weight average intrinsic viscosity were obtained from SEC-RI-RALLS-Visc and ranged from 2200 to 213,900 g/mol and 7 to 316 ml/g, respectively. The viscosity equation determined [ η ] w =1.06×10 −2 M w 0.86 indicated an extended random coil conformation. When the coil was modelled as a worm-like chain a persistent length of 3.65 nm was obtained. The hydrolysis products identified after extensive degradation were glucose, cellobiose, laminaribiose, 4-O-β-laminaribiosyl d -glucose, 4-O-β-laminaribiosyl d -cellobiose and 3-O-β-cellobiosyl d -cellobiose showing that other enzyme activities were present. The depolymerization kinetics suggested that longer sequences of consecutive (1→4)-linkages represent fast hydrolysable sites and the presence of higher relative proportions of (1→3)-linkages restricts the affinity of the enzyme.

Polysaccharides such as scleroglucan and schizophyllan contain periodate-resistant β-1,3-linked d -glucan backbones with regularly distributed side chains consisting primarily of single β-1,6-linked d -glucose residues that are susceptible to periodate oxidation. Side chain oxidation results in the formation of dialdehyde groups and the production of formic acid. During the initial periodate oxidation an increase in the storage modulus (G′) associated with the formation of gels, was observed when the reduced concentration, Cp [η] was above 3. These gels appeared to be physically stable, whereas aldehyde reduction (e.g. with NaBH4) or oxidation to carboxyl (e.g. with NaClO2 in acetic acid) dissolved the gels. The mechanism of cross-linking can be explained by the formation of intermolecular hemiacetal linkages. The gels were further characterised rheologically in terms of the initial increase in the storage modulus and the stability when stored in seawater at 55 and 90°C, respectively.

Six (1→3)(1→4)-β- d -glucan standards (A–F) isolated from barley were analysed by size-exclusion chromatography (SEC) in five different laboratories with varying columns, solvent conditions and detector systems (low- and multi-angle light scattering and viscometry). Static (batch) measurements by capillary viscometry and laser light scattering were included. Fairly consistent results were obtained for the weight average molecular weights (Mw), radii of gyration (RG) and intrinsic viscosities [η], demonstrating that the β-glucans may serve as useful standards or reference materials in the study of cereal β-glucans. Average values for Mw were: A,E: 114,000 (±11%); B,C: 374,000 (±9%), D,F: 228,000 (±13%). Some inconsistencies regarding the polydispersity (Mw/Mn) could be ascribed to the influence of peak broadening in certain column/solvent systems. The study further demonstrated that individual researchers tended to use different processing parameters, especially refractive index increments (dn/dc), due to ambiguities in the literature or to differing experimental values. The need for consistent parameters and processing methods is clearly demonstrated.

Oat beta-glucan was partially degraded by lichenase for different periods of time. Weight-average molecular weights were obtained from SEC-RI-RALLS-Visc and ranged from 5300 to 257,200. Weight-average intrinsic viscosities ranged from 7 to 160 mL/g. The determined viscosity equation, [eta] = 5.47 x 10(-3) M 0.82, indicated an extended coil conformation. When the coil was modeled as a wormlike chain a persistent length of 2.30 nm was obtained. 1H and 13C NMR spectroscopy were used to identify the structural units in the partially degraded beta-glucan. The identification of the disaccharide laminaribiose in the hydrolysates may suggest the presence of a new structural feature in cereal beta-glucan.

The triple-stranded polysaccharide scleroglucan was subjected to free-radical depolymerisation with Fenton's reagent (H2O2/Fe2+), and partially degraded samples in the molecular weight (Mw) range from 8 200 000 to 5200 were studied. Analyses of the chemical composition of partially degraded samples by 13C nuclear magnetic resonance spectroscopy and by determination of reducing end-groups suggested that the degradation was random. The decrease in Mw followed the kinetics expected for a multiple-stranded polymer. Following an apparently initial stable regime, degradation occurred with an exponential decrease (exponent (ν) ∼3.6) in Mw. Partially degraded samples displayed bimodal molecular weight distributions. This was attributed to the release of single-stranded oligomers having a degree of polymerisation (DP) too low to remain incorporated in the triplex structure (DP

κ- and ι-carrageenan that were partially hydrolyzed in dilute acid while being in the ordered conformation (induced by LiI or LiCl, respectively) gave rise to bimodal molecular weight distributions (MWDs) as shown by size-exclusion chromatography (SEC) and gel filtration. Optical rotation measurements showed that the high molecular weight (MW) fraction remained conformationally ordered, whereas the low-MW fraction was in the disordered state. The relative amount of the low-MW fraction increased with degradation time. By inducing the disordered conformation (in 0.01 M LiCl) the weight average molecular weight (Mw) of the partially hydrolyzed samples decreased 1.5−3 times. These results are interpreted in favor of a double-stranded ordered conformation. Partial hydrolysis of this structure yields a metastable, partially double-stranded structure containing chain breaks that are “unexposed” due to partial overlap of the individual chain fragments. When the degree of polymerization (DP) of such fragments decr...

Defatted leaves from the medicinal tree Cola cordifolia were extracted with 50% EtOH, 50 °C and 100 °C water. The polysaccharide rich extracts were fractionated and the structure of the polysaccharides elucidated. Linkage analysis of the polysaccharides indicates a rhamnogalacturonan type I backbone where both Rha and parts of GalA are substituted in position 3, indicating a highly branched polymer with short side chains. The purified fractions were tested for complement fixation, macrophage stimulating activity and anti-adhesion activity towards Helicobacter pylori. Here we report on complex and polydisperse types of pectins (Mw: 3-1300 kDa) as well as the presence of low Mw (3 kDa) acidic oligosaccharides. The fractions showed a moderate complement fixing activity and no macrophage activating effects after LPS removal. Anti-adhesion activity towards H. pylori was not found.

A new and flexible method for preparation of dry macroporous alginate foams with the capability of absorbing physiological solutions has been developed, which may find use within areas such as wound healing, cell culture, drug delivery and tissue engineering. The present study demonstrates how the gelation rate of the alginate and degree of ionic crosslinking can be utilized to control the physical foam properties. The rate of released Ca(2+)/Sr(2+) gelling ions available for interaction with the alginate was influenced by the concentration and physical characteristics of CaCO₃/SrCO₃ particles. The method of preparation of such foams allows, as described herein, tailoring of the pore structure, hydration properties and mechanical integrity in a manner not possible by other techniques.

Two chitosan samples (fraction of acetylated units (FA) 0.15 and 0.52) were fractionated by preparative size exclusion chromatography (SEC). The molecular weights and molecular weight distributions of the fractions were analyzed by analytical size exclusion chromatography coupled to an on-line low angle laser light scattering detector and a differential refractive index detector (SEC-LALLS-DRI), and their intrinsic viscosities were determined. The exponent (a) of the Mark-Houwink-Kuhn-Sakurada (MHKS) equation was found to be 0.92 ± 0.07 and 1.1 ± 0.1, respectively, at I = 0.1 and pH 4.5. No variation in FA related to molecular weight was found. Reversible interaction between chitosans and different column packings strongly influenced the log M-V relationships. This interaction was generally most pronounced for the low-FA chitosan, suggesting that the protonated amino groups are involved. Ammonium acetate buffer reduced this effect and the use of a new type of SEC-packing seemed to eliminate it. The more highly acetylated chitosan also had a more pronounced tendency towards concentration dependent self-association, which most probably involve intermolecular hydrophobic interactions between the acetyl groups.

Three different types of 15μm macroporous polymer particles (MP) based on poly(styrene- co -divinylbenzene), all with an extreme degree of monodispersity, were prepared by the method of ‘activated swelling’. The type and relative content of low molecular weight compounds used as pore-forming agents (porogens) ensured that the total pore volume was confined to large pores with radii larger than 300 A, and with an increasingly larger fraction of the pore volume shifted towards very large pores (> 2000A). A hydrophilic layer was covalently linked to the particles to eliminate their Hydrophobic character, and they were subsequently tested in aqueous size exclusion chromatography (SEC). Favourable Chromatographic properties generally associated with monosized particles were obtained, and column packing as well as the separations were performed with very low back pressures. The particles were found to be suitable for SEC of a series of high molecular weight polysaccharides, irrespective of their charge (anionic, cationic and neutral). The shift in pore size distribution towards very large pores resulted in a corresponding shift in the range of molecular weights (strictly, hydrodynamic volumes) where separation took place. For particles with 70% of the pore volume confined to pores with radii larger than 2000A, it was estimated that the molecular weight range of polysaccharides of the pullulan type accessible to SEC was extended up to ca. 10 8 . The corresponding value for an extended, rod-like, triple-helical polysaccharide (scleroglucan) was 10 7 .

Swelling behaviour of alginic acid gel beads with different chemical composition, molecular weight and size was studied in acetate buffer at pH4. A correlation was observed between the swelling behaviour in this buffer and the equilibrium properties of alginic acid gels. High contents of long l -guluronic acid blocks (G-blocks), known to give a high acid gel strength, reduced the rate of swelling and also the amount of solubilized alginate molecules leaching out of the gel beads. Compared to the original alginate, the leaching molecules had a lower average molecular weight, higher content of mannuronic acid residues and a reduced average length of G-blocks. Swelling capacity, rate of swelling and solubility of alginic acid seemed to depend on a balance between the tendency of homopolymeric blocks to form intermolecular junction zones, and the tendency of alginate to reduce the chemical potential of water. As expected, swelling rate increased with increasing temperature and decreasing bead size.

Samples of carboxylated scleroglucan, sclerox, with a degree of carboxylation in the side-chains varying from 10 to 50% were made in order to investigate the potential of using carboxylation as a means to control subsequent gelation by heavy metal ions. The storage and loss moduli of aqueous solutions of sclerox samples as a function of time were determined at T = 20 °C and 30 °C after addition of trivalent chromium ions. The rheological characterization showed a transition from a viscoelastic liquid to a gel with a characteristic time depending on the chromium concentration and the degree of carboxylation. For 20% carboxylation, the rate of change in the storage modulus could be controlled within nearly three orders of magnitude from 10 −3 to 1 Pa min −1 , using total chromium concentrations ranging from 0.3 to 6 m m . This rate was decreased by a factor of 2–5 by reducing the degree of carboxylation to 10%. The present study indicates that carboxylation of scleroglucan can be used to control the rate of gelation mediated by heavy metal ion complexation. This finding suggests that extension of the application range of this polysaccharide from an efficient viscosifer for polymer flooding to profile modification of high temperature oil reservoirs is attainable.

The effect of the length and chemical composition of the side-chains in comb-like branched polysaccharides on gelation with trivalent metal ions has been studied using xanthan and xylinan (acetan) with intact and truncated side-chains. Partial or complete removal of the terminal β-d-mannose, or removal of up to 22% of the trisaccharide side-chains of xanthan using partial acid hydrolysis, has only small effects on the Cr3+-induced gelation. In contrast, replacement of β-d-mannose by the trisaccharide α-l-Rhap-(1 → 6)-α-d-Glcp-(1 → 6)-α-d-Glcp-(1 → 4)- to yield the polysaccharide xylinan totally inhibits the gelation with Cr3+ ions. Removal of the trisaccharide by partial acid hydrolysis, which leads to a series of polymers with structures converging towards the partially hydrolysed xanthans, restores the gelling ability with Cr3+ ions. These observations seem to support the gelation model where Cr3+-glucuronic acid interactions are involved in the cross-linking of chains. It is further suggested that this interaction can be suppressed due to steric hindrance caused by the bulky side-chains in xylinan.

Degradation of single- and multiple-stranded polysaccharides by acid hydrolysis or free radical depolymerisation with H 2 O 2 Fe 2+ was monitored by viscosity measurements. Single-stranded polysaccharides (alginate, hydroxyethyl cellulose, carboxymethyl cellulose and κ-carrageenan in its disordered conformation) gave the expected linear relationship between 1 η sp ( 1 a ) and the degradation time, where a is the Mark-Houwink-Sakurada exponent. Double-stranded xanthan and triple-stranded scleroglucan showed a different pattern. Following an initial period with apparently slow degradation, a second regime was entered where the apparent degradation rate was much higher, and where ηsp followed the power law, ηsp ∼ t−va. However, the estimated values of the parameter v differed from those calculated by a Monte Carlo method for double- (xanthan) and triple-(scleroglucan) stranded polymers. κ-Carrageenan in its iodide-induced ordered conformation was very stable in acid as compared to the disordered conformation. In the ordered state the apparent degradation rate was initially constant, but increased in later stages. However, the double-logarithmic plot of ηsp versus time showed that there was neither a pronounced stable regime nor a regime following the power law. The degradation of gellan resulted in a rapid and linear increase in 1 η sp with degradation time, both in the ordered and disordered conformation. The same type of degradation kinetics was obtained for welan.

The bark of Cola cordifolia used in Malian traditional medicine contains unusual types of polysaccharides with immunomodulating activities. We report for the first time on the structure of a polymer designated CC1P1 having the repeating structure [2→)[α-D-Gal(1→3)]α-L-Rha(1→4)α-d-GalA(1→] as determined by NMR and GC/MS. α-Linked Gal is unusual in pectins. The Mw of 135 kDa was determined by SEC-MALLS. CC1P2 (1400 kDa), another polymer, having the same backbone, but this was substituted with α-4-OMe-GlcA, α-2-OMe-Gal and α-Gal as terminal units. CC1P1 shows a high complement-fixing activity, IC₅₀ being 2.2 times lower than the positive pectin control PMII (IC₅₀ appr. 71 μg/mL) while IC₅₀ of CC1P2 is 1.8 times lower. The simple structure of CC1P1 did not activate macrophages, while CC1P2 (100 μg/mL) showed the same potency as the positive controls PMII (100 μg/mL) and LPS (500 ng/mL). No cytotoxicity was detected.