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Michael D. Buschmann - One of the best experts on this subject based on the ideXlab platform.
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Ultrastructure of hybrid chitosan-Glycerol Phosphate blood clots by environmental scanning electron microscopy.
Microscopy research and technique, 2008Co-Authors: M. Iliescu, Abdellatif Chenite, Caroline D Hoemann, Matthew S Shive, Michael D. BuschmannAbstract:Chitosan-based polymers have been extensively studied for biomedical applica- tions. Recently, liquid solutions of chitosan in a Glycerol Phosphate buffer (chitosan-GP) with phys- iological pH and osmolality were mixed with autologous blood to form hybrid chitosan-GP/blood implants that improved the repair of articular cartilage lesions in a large animal model. The mix- ture of chitosan-GP and blood forms a viscous liquid, which solidifies in minutes via normal blood coagulation as well as chitosan-mediated mechanisms. Here we have examined the ultrastructure of these chitosan-GP/blood clots as well as regular blood clots and chitosan-GP gels, the latter pro- duced by heating. Both unfixed and fixed samples of chitosan-GP/blood clots, regular blood clots, and chitosan-GP gels were investigated by environmental scanning electron microscopy (ESEM) in conjunction with energy dispersive X-ray spectrometry (EDS), the former permitting direct ob- servation of the ultrastructure in hydrated conditions simulating the natural state. By examina- tion of unfixed specimens using ESEM we found that chitosan formed a network structure in both chitosan-GP gels and chitosan-GP/blood clots; however this structure was altered by aldehyde fix- ation to produce artifactual aggregates of chitosan microparticles. We were also able to identify chi- tosan in chitosan-GP/blood clots by washing samples in low concentration NaCl solutions followed by local EDS analyses to identify excess chloride versus sodium, and thus presence of cationic chi- tosan in analyzed features. Additional results indicated that the majority of Glycerol Phosphate dif- fuses freely from chitosan-GP gels (by EDS of phosphorus) and that hyperosmotic paraformalde- hyde-based fixatives (i.e. 4% w/v) significantly disturb erythrocyte morphology in fixed whole blood clots. Microsc. Res. Tech. 71:236-247, 2008. V C 2007 Wiley-Liss, Inc.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, pKap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 degrees C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per degrees C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 degrees C to 6.35 at 37 degrees C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per degrees C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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ionization and solubility of chitosan solutions related to thermosensitive chitosan Glycerol Phosphate systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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chitosan Glycerol Phosphate blood implants improve hyaline cartilage repair in ovine microfracture defects
Journal of Bone and Joint Surgery American Volume, 2005Co-Authors: Caroline D Hoemann, Mark B Hurtig, Evgeny Rossomacha, Anik Chevrier, Matthew S Shive, Michael D. BuschmannAbstract:Background: Microfracture is a surgical procedure that is used to treat focal articular cartilage defects. Although joint function improves following microfracture, the procedure elicits incomplete repair. As blood clot formation in the microfracture defect is an essential initiating event in microfracture therapy, we hypothesized that the repair would be improved if the microfracture defect were filled with a blood clot that was stabilized by the incorporation of a thrombogenic and adhesive polymer, specifically, chitosan. The objectives of the present study were to evaluate (1) blood clot adhesion in fresh microfracture defects and (2) the quality of the repair, at six months postoperatively, of microfracture defects that had been treated with or without chitosan-Glycerol Phosphate/blood clot implants, using a sheep model. Methods: In eighteen sheep, two 1-cm2 full-thickness chondral defects were created in the distal part of the femur and treated with microfracture; one defect was made in the medial femoral condyle, and the other defect was made in the trochlea. In four sheep, microfracture defects were created bilaterally; the microfracture defects in one knee received no further treatment, and the microfracture defects in the contralateral knee were filled with chitosan-Glycerol Phosphate/autologous whole blood and the implants were allowed to solidify. Fresh defects in these four sheep were collected at one hour postoperatively to compare the retention of the chitosan-Glycerol Phosphate/blood clot with that of the normal clot and to define the histologic characteristics of these fresh defects. In the other fourteen sheep, microfracture defects were made in only one knee and either were left untreated (control group; six sheep) or were treated with chitosan-Glycerol Phosphate/blood implant (treatment group; eight sheep), and the quality of repair was assessed histologically, histomorphometrically, and biochemically at six months postoperatively. Results: In the defects that were examined one hour postoperatively, chitosan-Glycerol Phosphate/blood clots showed increased adhesion to the walls of the defects as compared with the blood clots in the untreated microfracture defects. After histological processing, all blood clots in the control microfracture defects had been lost, whereas chitosanGlycerol Phosphate/blood clot adhered to and was partly retained on the surfaces of the defect. At six months, defects that had been treated with chitosan-Glycerol Phosphate/blood were filled with significantly more hyaline repair tissue (p < 0.05) compared with control defects. Repair tissue from medial femoral condyle defects that had been treated with chitosan-Glycerol Phosphate/blood contained more cells and more collagen compared with control defects and showed complete restoration of glycosaminoglycan levels. Conclusions: Solidification of a chitosan-Glycerol Phosphate/blood implant in microfracture defects improved cartilage repair compared with microfracture alone by increasing the amount of tissue and improving its biochemical composition and cellular organization. Clinical Relevance: The use of chitosan-Glycerol Phosphate/blood implants in conjunction with microfracture can improve the structural and compositional properties of repaired cartilage. These effects may result in better integration, improved biomechanical properties, and longer durability of the repair tissue.
Reinhard Sterner - One of the best experts on this subject based on the ideXlab platform.
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Significance of the Protein Interface Configuration for Allostery in Imidazole Glycerol Phosphate Synthase.
Biochemistry, 2020Co-Authors: Andrea C. Kneuttinger, Chitra Rajendran, Nadja Anita Simeth, Astrid Bruckmann, Burkhard König, Reinhard SternerAbstract:Imidazole Glycerol Phosphate synthase (ImGPS) from Thermotoga maritima is a model enzyme for studying allostery. The ImGPS complex consists of the cyclase subunit HisF and the glutaminase subunit H...
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Kinetic mechanism of indole-3-Glycerol Phosphate synthase.
Biochemistry, 2012Co-Authors: Sandra Schlee, Susanne Dietrich, Tomasz Kurcon, Pamela Delaney, Nina M. Goodey, Reinhard SternerAbstract:The (βα)8-barrel enzyme indole-3-Glycerol Phosphate synthase (IGPS) catalyzes the multistep transformation of 1-(o-carboxyphenylamino)-1-deoxyribulose 5-Phosphate (CdRP) into indole-3-Glycerol Phosphate (IGP) in tryptophan biosynthesis. Mutagenesis data and crystal structure analysis of IGPS from Sulfolobus solfataricus (sIGPS) allowed for the formulation of a plausible chemical mechanism of the reaction, and molecular dynamics simulations suggested that flexibility of active site loops might be important for catalysis. Here we developed a method that uses extrinsic fluorophores attached to active site loops to connect the kinetic mechanism of sIGPS to structure and conformational motions. Specifically, we elucidated the kinetic mechanism of sIGPS and correlated individual steps in the mechanism to conformational motions of flexible loops. Pre-steady-state kinetic measurements of CdRP to IGP conversion monitoring changes in intrinsic tryptophan and IGP fluorescence provided a minimal three-step kinetic mo...
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Imidazole Glycerol Phosphate synthase from Thermotoga maritima. Quaternary structure, steady-state kinetics, and reaction mechanism of the bienzyme complex.
The Journal of biological chemistry, 2001Co-Authors: Silke Beismann-driemeyer, Reinhard SternerAbstract:Imidazole Glycerol Phosphate synthase, which links histidine and de novo purine biosynthesis, is a member of the glutamine amidotransferase family. In bacteria, imidazole Glycerol Phosphate synthase constitutes a bienzyme complex of the glutaminase subunit HisH and the synthase subunit HisF. Nascent ammonia produced by HisH reacts at the active site of HisF with N'-((5'-phosphoribulosyl)formimino)-5-aminoimidazole-4-carboxamide-ribonucleotide to yield the products imidazole Glycerol Phosphate and 5-aminoimidazole-4-carboxamide ribotide. In order to elucidate the interactions between HisH and HisF and the catalytic mechanism of the HisF reaction, the enzymes tHisH and tHisF from Thermotoga maritima were produced in Escherichia coli, purified, and characterized. Isolated tHisH showed no detectable glutaminase activity but was stimulated by complex formation with tHisF to which either the product imidazole Glycerol Phosphate or a substrate analogue were bound. Eight conserved amino acids at the putative active site of tHisF were exchanged by site-directed mutagenesis, and the purified variants were investigated by steady-state kinetics. Aspartate 11 appeared to be essential for the synthase activity both in vitro and in vivo, and aspartate 130 could be partially replaced only by glutamate. The carboxylate groups of these residues could provide general acid/base catalysis in the proposed catalytic mechanism of the synthase reaction.
Dominic Filion - One of the best experts on this subject based on the ideXlab platform.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, pKap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 degrees C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per degrees C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 degrees C to 6.35 at 37 degrees C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per degrees C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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ionization and solubility of chitosan solutions related to thermosensitive chitosan Glycerol Phosphate systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
N Kandani - One of the best experts on this subject based on the ideXlab platform.
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rheological characterisation of thermogelling chitosan Glycerol Phosphate solutions
Carbohydrate Polymers, 2001Co-Authors: Abdellatif Chenite, Michael D. Buschmann, D Wang, C Chaput, N KandaniAbstract:Abstract In this study we demonstrate that chitosan solutions can be neutralised up to physiological pH (∼7.2) using β-Glycerol Phosphate without creating immediate gel-like precipitation and furthermore that subsequent heating of these solutions induces hydrogel formation. The addition of the particular basic salt, Glycerol Phosphate, provides the correct buffering and other physicochemical conditions including control of hydrophobic interactions and hydrogen bonding which are necessary to retain chitosan in solution at neutral pH near 4°C and furthermore to allow gel formation upon heating to 37°C. Rheological investigation evidenced the endothermic gelation of chitosan/β-Glycerol Phosphate solutions and allowed the establishment of a sol/gel diagram. The gelation process appears to be governed by delicate interplay between the pH and the temperature. The role of β-Glycerol Phosphate is discussed in the light of relevant literature particularly those indicating the role of Glycerol and polyols in the stabilisation of proteins and polysaccharides.
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Rheological characterisation of thermogelling chitosan/Glycerol-Phosphate solutions
Carbohydrate Polymers, 2001Co-Authors: Abdellatif Chenite, Michael D. Buschmann, D Wang, C Chaput, N KandaniAbstract:Abstract In this study we demonstrate that chitosan solutions can be neutralised up to physiological pH (∼7.2) using β-Glycerol Phosphate without creating immediate gel-like precipitation and furthermore that subsequent heating of these solutions induces hydrogel formation. The addition of the particular basic salt, Glycerol Phosphate, provides the correct buffering and other physicochemical conditions including control of hydrophobic interactions and hydrogen bonding which are necessary to retain chitosan in solution at neutral pH near 4°C and furthermore to allow gel formation upon heating to 37°C. Rheological investigation evidenced the endothermic gelation of chitosan/β-Glycerol Phosphate solutions and allowed the establishment of a sol/gel diagram. The gelation process appears to be governed by delicate interplay between the pH and the temperature. The role of β-Glycerol Phosphate is discussed in the light of relevant literature particularly those indicating the role of Glycerol and polyols in the stabilisation of proteins and polysaccharides.
Marc Lavertu - One of the best experts on this subject based on the ideXlab platform.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, pKap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 degrees C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per degrees C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 degrees C to 6.35 at 37 degrees C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per degrees C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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Ionization and Solubility of Chitosan Solutions Related to Thermosensitive Chitosan/Glycerol-Phosphate Systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.
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ionization and solubility of chitosan solutions related to thermosensitive chitosan Glycerol Phosphate systems
Biomacromolecules, 2007Co-Authors: Dominic Filion, Marc Lavertu, Michael D. BuschmannAbstract:Chitosan is a linear cationic biopolymer composed of glucosamine and N-acetyl-glucosamine that is only soluble in acidic aqueous solutions and precipitates when neutralized. However, it was recently discovered that chitosan dissolved in solutions containing Glycerol Phosphate was soluble at near neutral pH and produced a sol-gel transition when heated. Understanding this unique thermogelling system requires improved characterization of the ionization and solubility behaviors of chitosan, in particular dependencies on temperature, salt, chitosan concentration, and fD, where fD is the fraction of glucosamine monomers (deacetylated monomers) in chitosan. In the current study we performed temperature-controlled titration and dilution experiments on chitosan solutions with fD of 0.72, 0.85, and 0.98 at concentrations ranging from 1.875 to 30 mM of its glucosamine monomer and with 0 to 150 mM added salt. Light transmittance measurements were performed during titration to indicate precipitation. We found the apparent proton dissociation constant of chitosan, p Kap, to (1) decrease strongly with increased temperature, (2) increase strongly with increased salt, (3) increase strongly with increased chitosan concentration in low-salt conditions, and (4) decrease weakly with increasing fD. All of the above influences on chitosan pKap were accurately predicted using a mean-field Poisson-Boltzmann (PB) cylindrical cell model where the only adjustable parameter was the temperature-dependent chitosan intrinsic monomeric dissociation constant pK0(T). The resulting chitosan pK0 values at 25 °C were in the range from 6.63 to 6.78 for all chitosans and salt contents tested. The temperature dependence of chitosan ionization was found to strongly reduce pK0(T) by 0.023 units per °C, for example, resulting in a reduction of chitosan pK0(T) from 7.1 at 5 °C to 6.35 at 37 °C for fD of 0.72 in 150 mM salt. A similar temperature-dependent reduction of the pKa of the glucosamine monomer was found (-0.027 units per °C) while the pKa of Glycerol Phosphate did not change significantly with temperature. The latter result suggested that chitosan solutions heated in the presence of Glycerol Phosphate will become partly neutralized by transferring protons to Glycerol Phosphate and thereby allow attractive interchain forces to form a physically cross-linked gel under the appropriate conditions. Additionally, the degree of ionization of chitosan when it precipitates upon addition of a strong base was measured to be in the range from 0.25 to 0.55 and was found to (1) be insensitive to temperature, (2) increase strongly with increased salt, and (3) increase strongly with fD. The salt effect was accounted for by the PB model, while the influence of fD appeared to be due to acetyl groups impeding attractive chain-to-chain association to increase solubility and require reduced ionization levels to precipitate.