Morphology and gelation of thermosensitive chitosan hydrogels

The morphology of physical hydrogels is often difficult to examine due to the delicate nature of the system and therefore has not been studied in detail. Chitosan/GP (glycerophosphate salt) is a significant hydrogel in the biomedical and cosmetic fields as it is thermosensitive and contains less than 5% polysaccharide. The morphology of this system was examined with laser scanning confocal microscopy (LSCM) to image the gel morphology. The images indicate that the gel is quite heterogeneous, and power spectra reveal a fractal-like morphology. A study of composition found that increasing chitosan concentration increased the amount of polymer-rich phase present in the gel, and that the smallest aggregates decreased in size.     

Rheology and dynamic light scattering of octa-ethyleneglycol-monododecylether/chitosan solutions

In this work we used rheometry and DLS to probe relaxation phenomena in solutions of chitosan and octa-ethyleneglycol-monododecylether. The dispersions had a marked pseudoplastic behavior, which became less evident, as surfactant concentration was increased. Arrhenius plots showed that systems with surfactant presented a characteristic temperature at which apparent enthalpy of activation (varying from 3 to 40 kJ mol⁻¹) changed: this change was correlated to a possible transition of colloidal aggregates to a wormlike configuration. DLS intensity correlation functions were described by KWW equation: pure chitosan solutions had relaxation rate distributions centered at a characteristic relaxation rate around 4.6 × 10⁻⁶ μS⁻¹; as surfactant was added, a new component, with a faster characteristic relaxation rate with a magnitude order of 10⁻³ μs⁻¹, appeared. It was shown that the dependence between these relaxation rates and surfactant concentration could be used to describe DLS-related relaxation phenomena as an Arrhenius-activated process, agreeing with results obtained using rheometry.     

Rheological characterisation of a novel thermosensitive chitosan/poly(vinyl alcohol) blend hydrogel

Thermosensitive hydrogels that are triggered by changes in environmental temperature thus resulting in in situ hydrogel formation have recently attracted the attention of many investigators for biomedical applications. In the current work, the thermosensitive hydrogel was prepared through the mixture of chitosan (CS), poly(vinyl alcohol) (PVA) and sodium bicarbonate. The mixture was liquid aqueous solutions at low temperature (about 4 °C), but a gel under physiological conditions. The hydrogel was characterized by FTIR, swelling and rheological analysis. The effect of hydrogel composition and temperature on both the gel process and the gel strength was investigated from which possible hydrogel formation mechanisms were inferred. In addition, the hydrogel interior morphology as well as porosity of structure was evaluated by scanning electron microscopy (SEM). The potential of the hydrogels as vehicles for delivering bovine serum albumin (BSA) were also examined. In this study, the physically crosslinked chitosan/PVA gel was prepared under mild conditions without organic solvent, high temperature or harsh pH. The viscoelastic properties, as investigated rheologically, indicate that the gel had good mechanical strength. The gel formed implants in situ in response to temperature change, from low temperature (about 4 °C) to body temperature, which was very suitable for local and sustained delivery of proteins, cell encapsulation and tissue engineering.     

Spinning of hydroalcoholic chitosan solutions

We investigated the spinning of hydroalcoholic chitosan solutions. The dope composition was optimized in order to obtain a continuous alcogel fiber by water evaporation on heating the extruded hydroalcoholic solution. This alcogel fiber was then neutralized in aqueous alkali baths and washed in water to eliminate the residual alcohol and salts before final drying. Depending on the alcohol content in the filament at the neutralization step, on specific alcohol–chitosan interactions and on the nature and concentration of the coagulation base, the process yielded semicrystalline chitosan fibers with different proportions of anhydrous and hydrated allomorphs. Contrarily to the classical annealing method, the formation of mainly anhydrous crystals was obtained without significant molecular weight decrease by neutralizing the polymer in hydrophobic conditions. The control of allomorph content was shown to be related to the hydrophobicity of the solvent (alcohol fraction) at the neutralization step.     

Ionization constants and solubility of compounds involved in enzymatic synthesis of aminopenicillins and aminocephalosporins

The article deals with experimental determination of ionization constants and solubility for the compounds (target products, initial β-lactams, acylating agents and by-products) involved in enzymatic synthesis of some therapeutically used aminopenicillins and aminocephalosporins, namely ampicillin, amoxicillin, cephalexin, cephadroxil, cephaloglycin, cefaclor, cefprozil, cefatrizine. Methodology of investigations and the evaluation of experimental data for the determination of ionization constants and solubility of the different type electrolytes are presented. Applications of the methods based on acid–base potentiometric titration and on determination of solubility–pH dependence of assayed substances are discussed. The original data on ionization constants and solubility of amoxicillin, cefprozil, cefatrizine, cephadroxil and initial β-lactams for production of cefaclor, cefprozil and cefatrizine, as well as solubility of by-product d-(−)-p-hydroxyphenylglycine are presented. Experimentally determined parameters and constants available in the literature for all abovementioned aminopenicillins and aminocephalosporins are collected. These data might be used for choice of the conditions of both processes: the enzymatic synthesis and the isolation of the product from reaction mixture.     

Synthesis and characterization of sugar-bearing chitosan derivatives: aqueous solubility and biodegradability

The extended use of chitosan in biomedical fields has been limited by its insoluble nature in a biological solution. To endow the water solubility in a broad range of pH, chitosan derivatives were prepared by the covalent attachment of a hydrophilic sugar moiety, gluconic acid, through the formation of an amide bond. These sugar-bearing chitosans (SBCs) were further modified by the N-acetylation in an alcoholic aqueous solution. Thereafter, the effect of the gluconyl group and the degree of N-acetylation (DA) on the water solubility at different pHs and on the biodegradability of chitosan were investigated. The SBCs showed the water solubility in a broader range of pH than chitosan, whereas they were still insoluble at neutral and alkali pH. The N-acetylation of SBCs significantly affected the water solubility, for example, the SBCs with the DA, ranging from 29% to 63%, were soluble in the whole range of pH. This might result from the improved hydrophilicity by the gluconyl group, accompanied by the role of the N-acetyl group that disturbed the hydrogen bonding between amino groups of chitosan. From the biodegradation tests, determined by the decrease in the viscosity of a polymer solution exposed to lysozyme, it was evident that the gluconyl group attached to chitosan improved the biodegradability. Thus, it was possible to control the biodegradability of chitosan by adjusting the amounts of gluconyl and N-acetyl groups in the chitosan backbone. The N-acetylated SBCs, soluble in the broad range of pH, might be useful for various biomedical applications.     

Enzymatic grafting of a natural product onto chitosan to confer water solubility under basic conditions

Chitosan is a natural biopolymer whose rich amine functionality confers water solubility at low pH. At higher pH's (greater than 6. 5), the amines are deprotonated and chitosan is insoluble. To attain water solubility under basic conditions we enzymatically grafted the hydrophilic compound chlorogenic acid onto chitosan. Despite its name, chlorogenic acid is a nonchlorinated phenolic natural product that has carboxylic acid and hydroxyl functionality. The enzyme in this study was tyrosinase, which converts a wide range of phenolic substrates into electrophilic o-quinones. The o-quinones are freely diffusible and can undergo reaction with the nucleophilic amino groups of chitosan. Using slightly acidic conditions (pH = 6), it was possible to modify chitosan under homogeneous conditions. When the amount of chlorogenic acid used in the modification reaction exceeded 30% relative to chitosan's amino groups, the modified chitosan was observed to be soluble under both acidic and basic conditions, and to have a pH window of insolubility at near neutral pH. 1H NMR spectra confirmed that chitosan was chemically modified, although the degree of modification was low. Copyright 1999 John Wiley & Sons, Inc.     

Alginate/chitosan hydrogels as perspective transport systems for cefotaxime

This work reports synthesis of pH-responsive alginate/chitosan hydrogel spheres with the average diameter of 2.0 ± 0.05 mm, which contain cefotaxime that is an antibiotic of the cefalosporine group. The spheres provided the cefotaxime encapsulation efficiency of 95 ± 1%. An in vitro release of cefotaxime from the spheres in the media that simulate human biological fluids in peroral delivery conditions was found to be a pH-dependent process. The analysis of cefotaxime release kinetics by the Korsmeyer–Peppas model revealed a non-Fickian mechanism of its diffusion, which may be related to intermolecular interactions occurring between the antibiotic and chitosan. Conductometry, UV spectroscopy, and IR spectroscopy were used to study complexation of chitosan with cefotaxime in aqueous media with varied pH, characterize the composition of the complexes, and calculate their stability constants. The composition of the cefotaxime–chitosan complexes was found to correspond to the 1.0:4.0 and 1.0:2.0 molar ratios of the components at pH 2.0 and 5.6, respectively. Quantum chemical modeling was used to evaluate energy characteristics of chitosan–cefotaxime complexation considering the influence of a solvent.     

The solubility of fibrinogen in dilute salt solutions

Highly purified human fibrinogen was dialyzed versus eleven different sodium salts at ionic strengths of 0.005–0.3 and pH values of 4.5–8.0. After equilibration and centrifugation of the protein solutions, fibrinogen solubilities were determined spectrophotometrically and were analyzed as functions of pH, ionic strength, and specific anion. Bell-shaped curves are obtained when fibrinogen solubility is plotted as a function of pH. The solubility exhibits a minimum at a given pH and rises at acid and alkaline values. As the ionic strength is increased, the solubility curves shift toward more acid pH values. At constant pH values between 6 and 7, fibrinogen solubility increases with an increase in ionic strength. At constant pH values below pH 6, a decrease in solubility occurs as the ionic strength is increased. The isoionic pH of a saturated aqueous fibrinogen solution has been determined to be 6.25, meaning that fibrinogen in pure water behaves as a weak acid with a mean net charge of ?0.9. At pH values acid to 6.25, the anions solubilize fibrinogen in the following order of increasing efficacy: thiocyanate, perchlorate, sulfate, citrate, bromide, nitrate, phosphate, chloride, acetate, fluoride, and formate. This order is reversed at pH values alkaline to 6.25. Anion binding parameters calculated from the solubility data indicate that those anions which most effectively solubilize fibrinogen at alkaline pH and precipitate it at acid pH have the highest apparent binding affinities for the protein. Anions with less pronounced solubility effects have lower binding affinities.     

Three solutions of the protein solubility problem.

Three simple equations are presented, which describe the variation of protein solubility (S) with changes in salt concentration, in terms of either the salt molality (M), the salt activity (ax), or the water activity (aw). Each equation yields, essentially independent, estimates of the numbers of salt ions (delta vx) and water molecules (delta vw) involved in the dissolution of a mol of the protein. The equations can be used to elucidate the physical significance of the parameters in other empirical equations for protein solubility.     

Heterogeneous molecular aggregation and fractal structure in chitosan/acetic acid systems

The influence of deacetylation degree on heterogeneous molecular aggregation has been investigated for chitosan solution in 2 wt % acetic acid aqueous solution using rheological and small-angle x-ray scattering (SAXS) methods. Three samples of chitosan, which were designated CS62, CS79, and CS96, were used, and the deacetylation degrees of these samples were 0.62, 0.79 and 0.96, respectively. Rheological properties show that the systems of CS62 and CS96 are homogeneous, and the system of CS79 has a certain heterogeneous structure with a long-time relaxation mechanism. According to the SAXS measurement, the heterogeneous system has a fractal structure and the fractal dimension is about 1.3.     

Favorable chitosan/cellulose film combinations for copper removal from aqueous solutions

Three well-formed film combinations of chitosan, in the beta form and cellulose acetate biopolymer, having different proportions, have been synthesized and characterized by X-ray diffraction and infrared spectroscopy. The film having a 1.0/0.50 proportion presented 6.87 mmol of nitrogen atoms per gram of synthesized hybrid, with the highest affinity for adsorbing copper from aqueous solutions at 298+/-1K. The isotherm obtained in this adsorption showed a saturation plateau that corresponds to 1.92 mmol of copper per gram of hybrid. The energetic effects caused by copper ion interaction were determined through calorimetric titration at the solid-liquid interface in aqueous solution and gave a net thermal effect that enabled the calculation of the exothermic enthalpic values and the equilibrium constant. The complete thermodynamic data showed that the system is favored by exothermic enthalpies, negative Gibbs free energies and positive entropies.     

Polyelectrolyte microstructure in chitosan aqueous and alcohol solutions

This work deals with chain ordering in aqueous and water-alcohol solutions of chitosan. The so-called polyelectrolyte peak is investigated by small-angle synchrotron X-ray scattering. The polyelectrolyte microstructure was characterized by the position of the maximum of the polyelectrolyte scattering peak qmax, which scales with the polymer concentration cp as qmax approximately cp alpha. An evolution of the power law exponent alpha is observed as a function of the degree of acetylation (DA) of chitosan, which is responsible for changes of both the charge density (f) and the hydrophobicity of the polymer chains. The results highlighted the two organization regimes of the theory of Dobrynin and Rubinstein, investigated here for the first time for a natural polymer. At low DAs, alpha approximately 1/2, in agreement with a pearl necklace organization where the structure is controlled by the string between pearls. For higher DA, alpha approximately 1/3, and the correlation revealed by the polyelectrolyte peak is controlled by the pearls. This analysis offers a way to study quantitatively the balance between solvophobic-solvophilic interactions that play an important role in the solution properties of natural polymers. In addition, the role of several parameters acting on the interaction balance were evidenced, such as the nature of the counterion, the composition of the solvent (amount of alcohol in the aqueous solution), and the screening of Coulombic forces by salt addition. Finally, the nanostructure transition from a polyelectrolyte solution to a physical gel is discussed. The gel state is reached when the solvophobic interactions are favored, but depending on the gelation route the polyelectrolyte ordering could be preserved or not.     

Insertion-duplication mutagenesis of Salmonella enterica and related species using a novel thermosensitive vector

We constructed a novel temperature-sensitive vector as a tool for gene disruption by insertion-duplication mutagenesis (IDM) in Salmonella enterica and related species. A phoN insertion mutant was proven highly stable during growth in LB medium and during infection of macrophage cells in the absence of selection pressure. By progressive shortening of a phoN fragment, the minimal length for effective insertional mutagenesis driven by homologous recombination was determined to be 50 bp, allowing to disrupt even short genes that could not yet be subjected to site-specific IDM. We also showed that plasmid excision from the chromosome restores the wild-type genotype with a reliability of 98%. Intracellular recovery of the excised vector provides the option to switch between two genotypes and thus to rapidly attribute the observed mutant phenotype to the targeted gene. In addition, a fragment library was used to measure the integration rate at various chromosomal sites that varies greatly by at least 2.5 magnitudes, independently from the length of the cloned fragment.     

Modification of heavy metal uptake efficiency by modified chitosan/anionic surfactant systems

The presence of toxic heavy metals in natural environments entails a potential health hazard for humans. Metal contaminants in these environments are usually tightly bound to colloidal particles and organic matter. On the other hand, the potential of these metals towards chelation by different chelating agents presents a good characteristic for their removal from the environment. On this basis, two chitosan/anionic surfactant complexes were prepared and evaluated for their ability to remove heavy metals from aqueous solutions. The experimental results of the uptake of metal ions including Cu²⁺, Sn²⁺, Co²⁺ and Ni²⁺ are reported in this study. The results show that modified chitosan with short‐spacer group cross‐linkers has a higher potential for heavy metal uptake than long‐chain cross‐linker‐modified chitosan. Also, increasing the electronegativity of the heavy metals increases their uptake from the medium. Increasing the time of exposure of the heavy metals to the modified polymer increases the efficiency of the metal uptake process.     

Preferential hydration and solubility of proteins in aqueous solutions of polyethylene glycol

This paper is focused on the local composition around a protein molecule in aqueous mixtures containing polyethylene glycol (PEG) and the solubility of proteins in water + PEG mixed solvents. Experimental data from literature regarding the preferential binding parameter were used to calculate the excesses (or deficits) of water and PEG in the vicinity of β-lactoglobulin, bovine serum albumin, lysozyme, chymotrypsinogen and ribonuclease A. It was concluded that the protein molecule is preferentially hydrated in all cases (for all proteins and PEGs investigated). The excesses of water and deficits of PEG in the vicinity of a protein molecule could be explained by a steric exclusion mechanism, i.e. the large difference in the sizes of water and PEG molecules.

The solubility of different proteins in water + PEG mixed solvent was expressed in terms of the preferential binding parameter. The slope of the logarithm of protein (lysozyme, β-lactoglobulin and bovine serum albumin) solubility versus the PEG concentration could be predicted on the basis of experimental data regarding the preferential binding parameter. For all the cases considered (various proteins, various PEGs molecular weights and various pHs), our theory predicted that PEG acts as a salting-out agent, conclusion in full agreement with experimental observations. The predicted slopes were compared with experimental values and while in some cases good agreement was found, in other cases the agreement was less satisfactory. Because the established equation is a rigorous thermodynamic one, the disagreement might occur because the experimental results used for the solubility and/or the preferential binding parameter do not correspond to thermodynamic equilibrium.     

Neurocognitive systems related to real-world prospective memory

Background

Prospective memory (PM) denotes the ability to remember to perform actions in the future. It has been argued that standard laboratory paradigms fail to capture core aspects of PM.

Methodology/Principal Findings

We combined functional MRI, virtual reality, eye-tracking and verbal reports to explore the dynamic allocation of neurocognitive processes during a naturalistic PM task where individuals performed errands in a realistic model of their residential town. Based on eye movement data and verbal reports, we modeled PM as an iterative loop of five sustained and transient phases: intention maintenance before target detection (TD), TD, intention maintenance after TD, action, and switching, the latter representing the activation of a new intention in mind. The fMRI analyses revealed continuous engagement of a top-down fronto-parietal network throughout the entire task, likely subserving goal maintenance in mind. In addition, a shift was observed from a perceptual (occipital) system while searching for places to go, to a mnemonic (temporo-parietal, fronto-hippocampal) system for remembering what actions to perform after TD. Updating of the top-down fronto-parietal network occurred at both TD and switching, the latter likely also being characterized by frontopolar activity.

Conclusion/Significance

Taken together, these findings show how brain systems complementary interact during real-world PM, and support a more complete model of PM that can be applied to naturalistic PM tasks and that we named PROspective MEmory DYnamic (PROMEDY) model because of its dynamics on both multi-phase iteration and the interactions of distinct neurocognitive networks.     

Contractile injection systems of bacteriophages and related systems

Contractile tail bacteriophages, or myobacteriophages, use a sophisticated biomolecular structure to inject their genome into the bacterial host cell. This structure consists of a contractile sheath enveloping a rigid tube that is sharpened by a spike‐shaped protein complex at its tip. The spike complex forms the centerpiece of a baseplate complex that terminates the sheath and the tube. The baseplate anchors the tail to the target cell membrane with the help of fibrous proteins emanating from it and triggers contraction of the sheath. The contracting sheath drives the tube with its spiky tip through the target cell membrane. Subsequently, the bacteriophage genome is injected through the tube. The structural transformation of the bacteriophage T4 baseplate upon binding to the host cell has been recently described in near‐atomic detail. In this review we discuss structural elements and features of this mechanism that are likely to be conserved in all contractile injection systems (systems evolutionary and structurally related to contractile bacteriophage tails). These include the type VI secretion system (T6SS), which is used by bacteria to transfer effectors into other bacteria and into eukaryotic cells, and tailocins, a large family of contractile bacteriophage tail‐like compounds that includes the P. aeruginosa R‐type pyocins.