Biomedical subjects

O Smidsrød

Publications and source records attributed to O Smidsrød.

At least 19 recordsLinked to original sources

PubMed2001

Preparation and characterisation of oligosaccharides produced by nitrous acid depolymerisation of chitosans.

Two chitosans with widely different chemical composition (fraction of N-acetylated units (F(A))<0.001 and F(A)=0.59), were degraded by nitrous acid, to obtain the reactive 2,5-anhydro-D-mannose- (M-) unit at the new reducing end. The fully N-acetylated and fully N-deacetylated oligomers were separated by size-exclusion chromatography. Both the chemical structure and purity were studied by one- and two-dimensional 1H and 13C NMR methods. The fully N-acetylated oligomers were found to be stable, whereas the N-deacetylated oligomers reacted intermolecularly by a Schiff base reaction between the 2-amino group on the N-deacetylated units and the M-units, facilitating the cleavage of the glycosidic bond next to the M-unit and the formation of 5-hydroxymethylfurfural (HMF).

Acetylation↗

PubMed2000

Ionic and acid gel formation of epimerised alginates; the effect of AlgE4.

AlgE4 is a mannuronan C5 epimerase converting homopolymeric sequences of mannuronate residues in alginates into mannuronate/guluronate alternating sequences. Treating alginates of different biological origin with AlgE4 resulted in different amounts of alternating sequences. Both ionically cross-linked alginate gels as well as alginic acid gels were prepared from the epimerised alginates. Gelling kinetics and gel equilibrium properties were recorded and compared to results obtained with the original non-epimerised alginates. An observed reduced elasticity of the alginic acid gels following epimerisation by AlgE4 seems to be explained by the generally increased acid solubility of the alternating sequences. Ionically (Ca(2+)) cross-linked gels made from epimerised alginates expressed a higher degree of syneresis compared to the native samples. An increase in the modulus of elasticity was observed in calcium saturated (diffusion set) gels whereas calcium limited, internally set alginate gels showed no change in elasticity. An increase in the sol-gel transitional rate of gels made from epimerised alginates was also observed. These results suggest an increased possibility of creating new junction zones in the epimerised alginate gel due to the increased mobility in the alginate chain segments caused by the less extended alternating sequences.

Alginates↗

PubMed1998

Microcapsules of alginate-chitosan--I. A quantitative study of the interaction between alginate and chitosan.

The binding of chitosan to alginate beads was studied quantitatively by using radioactive labelled fractions of chitosan. The alginate-chitosan capsules were made either by dropping a solution of sodium alginate into a solution containing chitosan or by incubating calcium alginate beads in a solution of chitosan. The first procedure yielded a binding of 0.015 microg chitosan per mm2 of capsule surface, while the latter procedure yielded over 2 microg mm(-2). The maximum obtained weight ratio of chitosan to alginate in a microcapsule after 24 h was 0.40. The binding of chitosan was markedly increased by reducing the number average molecular weight of chitosan below 20000 Da and by increasing the porosity of the alginate gel. The porosity was increased by producing homogeneous gels, and by adding calcium chloride to the chitosan solution during the membrane forming stage. The effect of calcium ions on the porosity of the gel was studied by experiments involving release of blue dextran from calcium alginate beads. The binding of chitosan was also found to increase with decreasing fraction of N-acetylations, FA, on chitosan in the range of FA = 0.3 to FA = 0, and with increasing pH in the range from pH 4 to 6. Capsules with a diameter of 500 microm had a higher weight ratio of chitosan to alginate after 24 h of binding than the capsules with the larger diameter of 1500 microm.

Acetylation↗

PubMed1997

In vitro degradation rates of partially N-acetylated chitosans in human serum.

The initial degradation rates (r) in human serum of three chitosans with FA = 0.42, 0.51, and 0.60 were determined by measuring the decrease in viscosity as a function of time. A strong increase in r with increasing FA of the chitosans was observed, with r increasing proportionally to FA4.5. With increasing concentrations of lysozyme added to the reaction mixtures of chitosan and serum, the relative increase in degradation rate of chitosans with increasing FA was almost the same as that without lysozyme added. Addition of the chitinase inhibitor allosamidin (50 microM) did not inhibit the degradation rate of chitosan (FA = 0.60) by human serum. The results suggest that chitosans are actually mainly depolymerized by lysozyme in human serum, and not by other enzymes or other depolymerization mechanisms.

Acetylation↗

PubMed1997

Alginate based new materials.

Present and future applications of alginates are mainly linked to the most striking feature of the alginate molecule; i.e. a sol/gel transition in the presence of multivalent cations, e.g. Ca2+, almost independent on temperature. These very mild conditions, combined with the fact that alginates are highly characterised and understood both in the liquid and in the gel phase, makes this biopolymer unique compared to other gelling polysaccharides. Only pectins resemble alginate in the sol/gel transition behaviour, but this system can hardly be said to be as well characterised and understood as the alginates. The properties of alginate solutions and gels suggest biomedical and pharmaceutical uses. In this paper, the question of the specifications required by a polymer for applications in some biomedical areas will be discussed.

Alginates↗

PubMed1996

Determination of enzymatic hydrolysis specificity of partially N-acetylated chitosans.

A new method for determining the specificity of hydrolysis of the linear binary heteropolysaccharide chitosan composed of (1-->4)-linked 2-acetamido-2-deoxy-beta-D-glucopyranose (GlcNAc; A-unit) and 2-amino-2-deoxy-beta-D-glucopyranose (GlcN; D-unit) residues is described. The method is based on the assignments of the 13C chemical shifts of the identity (A- or D-units) of the new reducing and non-reducing ends and the variation in their nearest neighbours, using low molecular weight chitosans with known random distribution of A- and D-units as substrate. A highly N-acetylated chitosan with fraction of acetylated units (FA) of 0.68 and a number-average degree of polymerization (DPn) of 30 was hydrolysed with hen egg-white lysozyme, showing that both the new reducing and non-reducing ends consisted exclusively of A-units, indicating a high specificity for A-units in subsites DL and EL on lysozyme. Our data suggests that the preceding unit of the reducing A-units, is invariable, and based on earlier studies, most probably an A-unit, while the unit following the non-reducing A-units can be either an A- or a D-unit. A more detailed study of the specificity of lysozyme at subsite DL was performed by hydrolyzing a more deacetylated chitosan (FA = 0.35 and DPn of 20) to a DPn of 9, showing that even for this chitosan more than 90% of the new reducing ends were acetylated units. Thus, lysozyme depolymerizes partially N-acetylated chitosans by preferentially hydrolyzing sequences of acetylated units bound to site CL, DL and EL of the active cleft, while there is no specificity between acetylated and deacetylated units to site FL. In addition, a moderately N-acetylated chitosan with fraction of acetylated units (FA) of 0.35 and a DPn of 20 was hydrolysed with Bacillus sp. No. 7-M chitosanase, showing that both the new reducing and non-reducing ends consisted exclusively of D-units. Our data suggests that the nearest neigbour to the D-unit at the reducing end is invariable, and based on earlier studies, most probably a D-unit, while the unit following the non-reducing D-units can be either an A- or a D-unit. We conclude that the Bacillus chitosanase hydrolyzes partially N-acetylated chitosan by preferentially attacking sequences of three consecutive deacetylated units, hypothetical subsites CC, DC and EC, where the cleavage occur between sugar units bound to subsites DC and EC. A hypothetical subsite FC on the chitosanase show no specificity with respect to A- and D-units. The new NMR method described herein offers a time and labour-saving alternative to the procedure of extensive hydrolysis of the binary heteropolysaccharide chitosan and subsequent isolation and characterization of the oligosaccharides.

Acetylation↗

PubMed1996

Degradation of double-stranded xanthan by hydrogen peroxide in the presence of ferrous ions: comparison to acid hydrolysis.

Conformationally ordered, double-stranded xanthan, degraded in the presence of H2O2 and Fe2+ (at 20 degrees C) or in dilute acid (0.1 M HCl at 80 degrees C), produced xanthan variants with weight-average molecular weights (Mw) ranging from 2 x 10(6) to 5.4 x 10(4). In both cases the fraction of cleaved linkages in the glucan backbone (alpha), measured as reducing ends, increased to very high values (0.05 for Mw = 2-3 x 10(4)), demonstrating that a large number of linkages in the backbone could be cleaved without a correspondingly large reduction in Mw, in accordance with the double-stranded nature of xanthan. Extensive degradation (more than 10-fold reduction in Mw) in both cases released single-stranded, conformationally disordered oligomers; this release was accompanied by an increase in the rate of acid hydrolysis of the glucan backbone and a pronounced increase in the rate of release of glucose monomer. In contrast, there was no significant change in the rate of reducing end-group formation associated with the release of oligomers upon degradation with H2O2/Fe2+. Both types of degradation were accompanied by changes in the composition of the side chains. However, in contrast to acid hydrolysis, where the terminal beta-D-mannose is preferentially hydrolyzed, the reaction with H2O2/Fe2+ resulted in removal of both mannose and glucuronic acid at approximately equal rates. This observation can be explained by a preferential attack on the inner alpha-D-mannose, with concomitant removal of the entire side chain. Removal of side chains and the release of single-stranded oligomers by H2O2/Fe2+ strongly influenced the optical rotation and also broadened the chiroptically detected conformational transition, whereas no change in the transition temperature was observed.

Carbohydrate Conformation↗

PubMed1996

Physicochemical studies on xylinan (acetan). I. Characterization by gel permeation chromatography on sepharose Cl-2B coupled with static light scattering and viscometry.

Laboratory-made samples of the polysaccharide xylinan (acetan) were fractionated on Sepharose Cl-2B using 0.1M NaCl as eluant. The weight average molar masses and intrinsic viscosities were estimated in the fractions by multiangle laser light scattering (off-line) and capillary viscometry, respectively. The Mark-Houwink-Sakurada plot was found to be indicative of semiflexible coils (a = 0.90). The angular dependence of scattered light was interpreted by fitting with theoretically calculated "Master Curves" in terms of a wormlike chain model. The ambiguity of the interpretation of scattering curves owing to the overlapping effects of chain stiffness and polydispersity is discussed in detail. The experimental data is found to be consistent with a persistence length of Lp = 100 nm. The main proportion consists of double-stranded chains (consistent with a robust double-helix), but single- and multistranded chains also are present. Our results suggest a fractionation according to the contour length rather than the molar mass.

Acetobacter↗

PubMed1996

Dependence of the content of unsubstituted (cellulosic) regions in prehydrolysed xanthans on the rate of hydrolysis by Trichoderma reesei endoglucanase.

Removal of side chains from the bacterial polysaccharides xanthan and xylinan (acetan) results in the formation of unsubstituted cellulosic regions that are susceptible to hydrolysis by cellulases (beta-1,4-endoglucanases). In contrast to cellulose derivatives, low degrees of substitution (DS) may be obtained in xanthan without affecting the solubility in water, and longer unsubstituted regions are obtained for the same DS due to the regular distribution of side chains. By varying the fraction of cellobiosic residues carrying a side chain from 1.0 to 0.54, the viscosimetrically detected rate of hydrolysis by cellulase of conformationally disordered xanthan, increased by 4 orders of magnitude. An increase was also obtained by removing side chains from xylinan. An analysis of the kinetic data suggests that very long unsubstituted regions (more than 10 glucose residues) are required for maximum rate of hydrolysis by cellulase.

Carbohydrate Sequence↗

PubMed1996

Alginate polycation microcapsules. I. Interaction between alginate and polycation.

The interactions between alginate and polycations have been studied by using different labelling techniques. Binding of poly-L-lysine (PLL) to alginate in the gel state is mainly governed by the amount of dissociable negative charges on the bead surface. PLL was found to bind more rapidly to gel beads made from alginate with a high content of mannuronic acid. The binding was enhanced by increasing the alginate concentration on the surface by making inhomogeneous beads. When the capsules were stored in the presence of cations with high affinity for alginate (Ca2+, Sr2+), PLL was washed off. Less PLL is bound to strontium alginate than to calcium alginate beads. Two mechanisms appear to be responsible for the binding of sodium alginate to alginate PLL capsules (coating): (i) an electrostatic interaction between the soluble coating material and excess positive charges on PLL on the surface; (ii) the formation of a calcium alginate gel on the surface owing to leaching of calcium ions from the core. The stability and efficiency of the coating as a function of molecular size and sequential structure of the coating polymer have also been investigated.

Alginates↗

PubMed1996

Alginate polycation microcapsules. II. Some functional properties.

The main cause of alginate polycation capsule breakage under physiological conditions is probably the osmotic swelling of the alginate core owing to the Donnan equilibrium set up by the negative charges of the carboxyl groups not involved in cooperative binding of counterions in the junction zones of the network. In the present paper we show how capsules can be stabilized extensively by reducing their swelling capacity in various ways. Alginate polycation capsules with good chemical and mechanical stability have been made by controlling their swelling behaviour through selection of capsule material according to chemical structure and molecular weight, as well as by controlling the kinetics of the capsule formation. Stable capsules have been made either by increasing the strength of the polyanion-polycation membrane, or by keeping a low-swelling gel network in the core. The latter capsules are made from an alginate rich in guluronic acid both in the core and in an outer coating, and with anisotropic distribution of the polymer material in the core where the concentration at the surface is higher than that in the centre of the capsule. Some functional properties of these capsules, such as porosity, have also been studied.

Alginates↗

PubMed1993

Temperature-induced conformational transition in xanthans with partially hydrolyzed side chains.

The conformational properties of xanthans with partially hydrolyzed side chains were investigated by optical rotation, CD, and differential scanning calorimetry (DSC). All variants displayed the well-known temperature-driven, cooperative order-disorder transition, and both optical rotation and DSC showed that the transition temperature was essentially independent of the content of terminal beta-mannose. It was found that up to 80% of the changes in the specific optical rotation accompanying the transition reflects conformational changes linked to the terminal beta-mannose in the side chains. Modification of the side chains also affected the CD when xanthan was in the ordered state, but in this case the data suggest that the glucuronic acid is the major component determining the magnitude of the CD signal. DSC measurements showed that the transition enthalpy (delta Hcal) increased linearly with the fraction of beta-mannose, again indicating that a significant part (up to 80%) of delta Hcal reflects conformational changes in the side chains. The conformational transition of the xanthan variants generally showed a higher degree of cooperativity (sharper transition) than unmodified, pyruvated xanthan. Calculation of the cooperativity parameter sigma by means of the Zimm-Bragg theory (OR data) or from the ratio between delta Hcal and the van't Hoff enthalpy (delta HvH) using DSC data showed a correlation between sigma and the content of beta-mannose, but the two methods gave different results when the content of beta-mannose approached 100%. The ionic strength dependence of the transition temperature, expressed as d (log I)/d(T-1m), was nearly identical for intact xanthan and a sample containing only 6% of the terminal beta-mannose. Application of the Manning polyelectrolyte theory does not readily account for the observed delta Hcal values, neither does it provide new information on the nature of the ordered and disordered conformations in xanthan.

Calorimetry, Differential Scanning↗

PubMed1993

Similar mechanisms of action of defined polysaccharides and lipopolysaccharides: characterization of binding and tumor necrosis factor alpha induction.

Little has been reported about the effects of different polysaccharides on cytokine production from human monocytes. In this study, we show that several well-defined polysaccharides, including polymers with different sizes of beta 1-4-linked D-mannuronic acid (poly-M, high-M alginate, and M-blocks) and cellulose oxidized in the C-6 position, induced human monocytes to produce tumor necrosis factor alpha (TNF-alpha). Poly-M was the most efficient polysaccharide tested and, on a weight basis, was approximately as efficient as lipopolysaccharide (LPS) from Escherichia coli. TNF-alpha production was shown to depend strongly on the molecular weights of poly-M and high-M alginate, with maximal TNF-alpha production occurring at molecular weights above 50,000 and 200,000, respectively. G-blocks, alpha 1-4-linked L-guluronic acid polymers that did not induce cytokine production from monocytes, reduced the cytokine production induced by the beta 1-4-linked polyuronic acids and LPS. Furthermore, both G-blocks and LPS were found to inhibit the binding of poly-M to monocytes, as measured by flow cytometry. In addition, we found that the binding of LPS to monocytes was inhibited by G-blocks, M-blocks, and poly-M. Our results indicate that beta 1-4-linked polyuronic acids and LPS may stimulate monocytes to produce TNF-alpha by similar mechanisms and may bind to a common receptor.

Biological Assay↗

PubMed1992

Chitosan cross-linked with Mo(VI) polyoxyanions: a new gelling system.

A procedure for preparing homogeneous chitosan gels by in situ molybdate cross-linking is described. The gels are obtained by dispersing solid MoO3 in a buffered chitosan solution and the polymer is cross-linked by formation of heavily negatively charged molybdate polyoxyanions. The resulting ionic gels are very transparent, thermoirreversible and can be made at low polymer concentrations. Depending on the ionic strength, these gels are able to swell several times their original size in aqueous solutions. Estimates of the degree of cross-linking reveal a very open pore structure which is confirmed by electron micrographs of the gel.

Anions↗

PubMed1991

13C-n.m.r. studies of the acetylation sequences in partially N-deacetylated chitins (chitosans).

Chitosans obtained under homogeneous conditions of N-deacetylation, with degrees of N-deacetylation between 46% and 94%, were depolymerised and their 125-MHz 13C-n.m.r. spectra have been interpreted. The sequence of 2-acetamido-2-deoxy-beta-D-glucopyranose (GlcNAc) and 2-amino-2-deoxy-beta-D-glucopyranose (GlcN) residues influenced the chemical shifts, and the diad and triad frequencies have been calculated. Chitosans that were N-deacetylated under homogeneous and heterogeneous conditions gave values for the diad and triad frequencies that were consistent with a random arrangement of GlcN and GlcNAc residues.

Acetylation↗

PubMed1991

Determination of the degree of N-acetylation and the distribution of N-acetyl groups in partially N-deacetylated chitins (chitosans) by high-field n.m.r. spectroscopy.

The composition and sequence of 2-acetamido-2-deoxy-beta-D-glucose (GlcNAc) and 2-amino-2-deoxy-beta-D-glucose (GlcN) residues in partially N-deacetylated chitosans, prepared under homogeneous and heterogeneous conditions, have been determined by 1H-n.m.r. spectroscopy. It was necessary to depolymerise the chitosan slightly by treatment with nitrous acid before spectroscopy. A sequence-dependent deshielding of H-1 of the GlcNAc residues made it possible to determine the proportions of the four possible diads. Chitosan prepared by N-deacetylation under homogeneous conditions gave values for the diad frequencies that were roughly consistent with a random distribution of the N-acetyl groups. Samples prepared under heterogeneous conditions have a frequency of the GlcNAc-GlcNAc diad slightly higher than for a random (Bernoullian) distribution. The chitosans, prepared under both homogeneous and heterogeneous conditions, with a degree of acetylation of 50% were soluble at neutral pH.

Acetylation↗

PubMed1991

Induction of cytokine production from human monocytes stimulated with alginate.

Alginates are polysaccharides with gel-forming properties composed of 1,4-linked beta-D-mannuronic acid (M), alpha-L-guluronic acid (G), and alternating (MG) blocks. Alginate can be used as a matrix for implanted cells in vivo. In this study, we have examined the ability of alginates and their components to stimulate human monocytes to produce tumor necrosis factor-alpha, interleukin-6, and interleukin-1. Alginates stimulated the monocytes to produce high levels of all three cytokines. Low G alginates were approximately 10 times more potent in inducing cytokine production compared with high G alginates. The M-blocks and the MG-blocks, but not the G-blocks, stimulated the cytokine production. The results demonstrate that the mannuronic acid residues are the active cytokine inducers in alginates.

Alginates↗

PubMed1990

Alginate as immobilization matrix for cells.

In recent years, entrapment of cells within spheres of Ca2+ alginate has become the most widely used technique for immobilizing living cells. This versatile method includes applications ranging from immobilization of living or dead cells in bioreactors, immobilization of plant protoplasts for micropropagation and immobilization of hybridoma cells for production of monoclonal antibodies, to entrapment of animal cells for implantation of artificial organs. This review evaluates the potential of this method on the basis of the current knowledge of structural and functional relationships in alginate gels.

Alginates↗