Effect of solvent-dependent viscoelastic properties of chitosan membranes on the permeation of 2-phenylethanol

The viscoelastic behaviour of chitosan was followed by dynamic mechanical analysis (DMA) while the sample was immersed in gradient compositions of water/ethanol mixtures. The swelling equilibrium of chitosan membranes, both crosslinked with genipin or not, increased linearly with the water content. Increasing the water content, it was simultaneously observed a peak in the loss factor (around 25 vol.%) and a reduction of the storage modulus, which was attributed to the α-relaxation of chitosan. This was the first time that the glass transition dynamics in a polymer was monitored in immersion conditions where the composition of the plasticizer in the bath is changed in a controlled way. The water content at which tan δ presented a maximum increased with both increasing frequency and increasing crosslinking density. The permeability decreased steadily with the ethanol content, reaching very low values around the glass transition. Therefore we hypothesize that conformational mobility of the polymeric chains may play an important role in the diffusion properties of molecules trough polymeric matrices.     

Effects of lyophilization on the stability of bacteriophages against different serogroups of Shiga toxin-producing Escherichia coli

Lyophilization is commonly used to effectively preserve the stability of bacteriophages (phages) in long-term storage. However, information regarding the lyophilization of phages specific to Shiga toxin-producing Escherichia coli (STEC) strains is scarce. The objective of this study was to determine the effects of lyophilization with different cryoprotectants (sucrose and trehalose) and concentrations (0.1 M and 0.5 M) on the stability of seven lytic phages specific to STEC O157 and top 6 non-O157 strains during 6-month storage at −80 °C. The titers of lyophilized phages specific to STEC O26 (S1 O26) and STEC O121 (Pr121lvw) did not exhibit significant reduction after 6-month storage regardless of the use of cryoprotectants. Phages lytic against STEC O103 (Ro103C3lw) and STEC O145 (Ro145clw) with 0.1 M sucrose retained similar titers after lyophilization and frozen storage for 6 months (P > 0.05). Despite subtle differences, these results indicated that most of the selected phages had similar titer retention with the same cryoprotectants. Additionally, lytic activities of the phages against their primary hosts were not affected after lyophilization and 6-month frozen storage. Moreover, no detectable damage was observed on the lyophilized phage structures. These findings provide valuable insight into the use of lyophilization to preserve phages lytic against STEC strains.     

Phase Transitions of Frozen Camu-camu (Myrciaria dubia (H.B.K.) McVaugh) Pulp: Effect of Cryostabilizer Addition

Camu-camu is a tropical fruit with very high vitamin C content and commercialized as frozen pulp. Enthalpies of freezing, temperatures of the onset of ice melting, and glass transition temperatures of the maximally freeze-concentrated phase () of camu-camu pulp and of samples containing maltodextrin (DE20) and sucrose were measured by differential scanning calorimetry. Maltodextrin exhibited the largest freeze stabilization potential, increasing from −58.2 °C (natural pulp) to −39.6 °C when 30% (w/w) maltodextrin DE 20 was added. Sucrose showed negligible effect on but enhanced considerably the freezing point depression and less amount of ice was formed.     

Moisture sorption characteristics and thermophysical properties of trehalose-PBS mixtures

The goal of the study was to quantify the thermophysical properties and the moisture sorption characteristics of the trehalose-PBS (phosphate-buffered saline) from the desiccation preservation perspective. A moisture sorption study was undertaken to determine the desorption isotherms of the trehalose-PBS mixtures. The Brunauer, Emmett, and Teller (BET)-equation and the Guggenheim, Anderson, and de Boer equation were used to quantify the desorption data. The glass transition temperature of the mixtures of trehalose-PBS, equilibrated at different relative humidities was studied using a differential scanning calorimeter. Fourier transform infrared spectroscopy was used to study the molecular interaction between the trehalose and PBS mixtures. The results showed that the addition of PBS to the trehalose mixture causes a shift from the type II isotherm to a type III isotherm (characterized by BET equation) which may have detrimental effect on cell desiccation. The results showed that an increase in PBS mass fraction in the trehalose-PBS mixture causes a decrease in the glass transition temperature (Tg) of the mixture and also a decrease in the hydrogen bonding capacity of the trehalose glasses. The addition of PBS to trehalose posed some challenges and should be subject to further optimization to use it in desiccation preservation of biologics.     

Dissolution of sucrose crystals in the anhydrous sorbitol melt

The dissolution of a sugar (sucrose as a model) with higher melting point was studied in a molten food polyol (sorbitol as a model) with lower melting point, both in anhydrous state. A DSC and optical examination revealed the dissolution of anhydrous sucrose crystals (mp 192 degrees C) in anhydrous sorbitol (mp 99 degrees C) liquid melt. The sucrose-sorbitol crystal mixtures at the proportions of 10, 30, 60, 100 and 150 g of sucrose per 100 g of sorbitol were heat scanned in a DSC to above melting endotherm of sorbitol but well below the onset temperature of melting of sucrose at three different temperatures 110, 130 and 150 degrees C. The heat scanning modes used were with or without isothermal holding. The dissolution of sucrose in the sorbitol liquid melt was manifested by an increase in the glass transition temperature of the melt and corresponding decrease in endothermic melting enthalpy of sucrose. At given experimental conditions, as high as 25 and 85% of sucrose dissolved in the sorbitol melt during 1 h of isothermal holding at 110 and 150 degrees C, respectively. Optical microscopic observation also clearly showed the reduction in the size of sucrose crystals in sorbitol melt during the isothermal holding at those temperatures.     

The glass transition temperature of mixtures of trehalose and hydroxyethyl starch

Although mixtures of HES and sugars are used to preserve cells during freezing or drying, little is known about the glass transition of HES, or how mixtures of HES and sugars vitrify. These difficulties may be due to the polydispersity between HES samples or differences in preparation techniques, as well as problems in measuring the glass transition temperature (T(g)) using differential scanning calorimetry (DSC). In this report, we examine the T(g) of mixtures of HES and trehalose sugar with <1% moisture content using DSC measurements. By extrapolating these measurements to pure HES using the Gordon-Taylor and Fox equations, we were able to estimate the T(g) of our HES sample at 44 degrees C. These results were additionally confirmed by using mixtures of glucose-HES which yielded a similar extrapolated T(g) value. Our approach to estimating the glass transition temperature of HES may be useful in other cases where glass transitions are not easily identified.     

Disentangling alpha from beta mechanical relaxations in the rubber-to-glass transition of high-sugar-chitosan mixtures

The occurrence of molecular motions in addition to those of the glass-transition region (alpha mechanism) were investigated in chitosan and a branched derivative substituted with alkyl chains having eight carbon atoms. Once hydrophobic interactions of the alkyl groups in aqueous solution were demonstrated, polymers were mixed with glucose syrup at high levels of solids. The real (G') and imaginary (G") components of the complex dynamic modulus in high-solid mixtures were measured between 0.1 and 100 rad s(-1) in the temperature range from -55 to 50 degrees C. The method of reduced variables gave superposed curves of G' and G", which unveiled an anomaly in the dispersion of the alkylated derivative both in terms of higher modulus values and dominant elastic component of the polymeric network, as compared with the glass-transition region of chitosan. It was proposed that the new mechanical feature was due to beta mechanism, and master curves of viscoelastic functions and relaxation processes were constructed to rationalize it.     

The glass transition temperatures of sugar mixtures

We measured the glass transition temperatures of mono-, di-, and trisaccharide mixtures using differential scanning calorimeter (DSC) and analyzed these temperatures using the Gordon-Taylor equation. We found that the glass transition temperatures of monosaccharide-monosaccharide and disaccharide-disaccharide mixtures could be described by the conventional Gordon-Taylor equation. However, the glass transition temperatures of monosaccharide-disaccharide and monosaccharide-trisaccharide mixtures deviated from the conventional Gordon-Taylor equation and the amount of deviation in the monosaccharide-trisaccharide mixtures was larger than those in the monosaccharide-disaccharide mixtures. From these results, we conclude that the size and shape of the sugars play an important role in the glass transition temperature of the mixtures.     

Heteronuclear coupling constants of hydroxyl protons in a water solution of oligosaccharides: trehalose and sucrose

Relatively few details are known about the conformational preferences of hydroxyl groups in carbohydrates in water solution, though these would be informative about solvation and H-bonding. We show that highly concentrated solutions of sucrose and trehalose exhibit surprisingly well-resolved ¹H NMR spectra in a deuterium oxide–water solvent mixture at subzero temperatures. Measurement conditions are suitable to extract nearly all homonuclear and, for the first time, heteronuclear coupling constants of OH groups of carbohydrates in their natural abundance. For ^2,3J_HO,C coupling constants new, powerful variants of HETLOC and HECADE techniques were applied. The present data do not support the presence of persistent H-bonds in these two cryogenic disaccharides.     

Use of dynamic mechanical analysis (DMA) to determine critical transition temperatures in frozen biomaterials intended for lyophilization

Dynamic mechanical analysis is widely used to determine glass transitions in solid state materials. However, here we demonstrate the application of DMA for the determination of glass transitions (Tg’) in the frozen liquid state by means of a steel sample pocket. The use of the pocket allows frozen material to be analysed and glass transition events demonstrated. In addition, it allows weak glass transitions to be detected clearly in some complex formulations where they can be obscured by eutectic and other strong thermal events when other methods such as DSC or DTA are used. Classical excipients (trehalose, lactose, dextran) were analysed and shown to give reproducible Tg’ values, though with values slightly higher than those obtained by DSC. Finally, several complex real biological materials, typical of those encountered when freeze drying biological and biopharmaceutical materials, were analysed and the potential value of DMA demonstrated to determine the relevant glass transition temperatures for use in cryobiology and freeze drying.     

Oxidative and nitrosative stress in frozen-thawed pig spermatozoa. II: Effect of the addition of saccharides to freezing medium on sperm function

Saccharides have bioprotective properties, with a high capacity to preserve biological proteins and membranes during sperm cryopreservation. The aim of this study was to evaluate how replacing the lactose of cryopreservation media by sucrose (SUC) or trehalose (TRE) at concentrations of 0.2 M (SUC-1 and TRE-1) and 0.25 M (SUC-2 and TRE-2) affects frozen/thawed pig spermatozoa. The media used were composed of medium A (saccharide/egg yolk) and B (saccharide/egg yolk/glycerol), their osmolality being determined prior to freezing. Cell viability, membrane lipid disorder, acrosome integrity, mitochondrial membrane potential (MMP), lipid peroxidation, thiol group oxidation, total reactive oxygen species (ROS), peroxynitrite and superoxide anion (O₂^●-) were determined through flow cytometry; total motility (TM), progressive motility (PM) and kinetic parameters motility were determined immediately after thawing (T0) and again 30 (T30) and 60 (T60) minutes later. The SUC-2 and TRE-2 groups maintained viability significantly and presented fewer lipid membrane disorders, respectively, both with a significant increase in MMP. The production of O₂^●- and peroxynitrite was lower in the TRE-2 groups compared to the control (P < 0.05). Total motility at T0 was greater in the TRE-2 group (P < 0.05). Sperm kinetics was not affected by the treatment. The use of saccharides SUC and TRE at a concentration of 0.25 M improves sperm quality, so that both non-penetrating cryoprotectants can be utilized in pig sperm freezing media.     

Correlation between bioprotective effectiveness and dynamic properties of trehalose-water, maltose-water and sucrose-water mixtures

The aim of the present work is to link the bioprotective effectiveness to the dynamic properties of a class of homologous disaccharides, that is, trehalose, maltose and sucrose, and their mixtures in water. The findings obtained by elastic neutron scattering point out a harmonic–anharmonic transition for all the three disaccharide mixtures. Using a new operative definition of ‘fragility’, the different degrees of ‘strength’ of the investigated systems are determined. The links existing between the degree of fragility and the cryptoprotective action are also discussed.     

Effect of electrostatic interactions on phase stability of cubic phases of membranes of monoolein/dioleoylphosphatidic acid mixtures.

To elucidate effects of electrostatic interactions resulting from surface charges on structures and phase stability of cubic phases of lipid membranes, membranes of 1-monoolein (MO) and dioleoylphosphatidic acid (DOPA) (DOPA/MO membrane) mixtures have been investigated by small-angle x-ray scattering method. As increasing DOPA concentration in the DOPA/MO membrane at 30 wt% lipid concentration, a phase transition from Q(224) to Q(229) phase occurred at 0.6 mol% DOPA, and at and above 25 mol% DOPA, DOPA/MO membranes were in the L(alpha) phase. As NaCl concentration in the bulk phase increased, for 10% DOPA/90% MO membrane in excess water, a Q(229) to Q(224) phase transition occurred at 60 mM NaCl, and then a Q(224) to H(II) phase transition occurred at 1.2 M NaCl. Similarly, for 30% DOPA/70% MO membrane in excess water, at low NaCl concentrations it was in the L(alpha) phase, but at and above 0.50 M NaCl it was in the Q(224) phase, and then at 0.65 M NaCl a Q(224) to H(II) phase transition occurred. These results indicate that the electrostatic interactions in the membrane interface make the Q(229) phase more stable than the Q(224) phase, and that, at larger electrostatic interactions, the L(alpha) phase is more stable than the cubic phases (Q(224) and Q(229)). We have found that the addition of tetradecane to the MO membrane induced a Q(224)-to-H(II) phase transition and also that to the 30% DOPA/70% MO membrane induced an L(alpha)-to-H(II) phase transition. By using these membranes, the effect of the electrostatic interactions resulting from the membrane surface charge (DOPA) on the spontaneous curvature of the monolayer membrane has been investigated. The increase in DOPA concentration in the DOPA/MO membrane reduced the absolute value of spontaneous curvature of the membrane. In the 30% DOPA/70% MO membrane, the absolute value of spontaneous curvature of the membrane increased with an increase in NaCl concentration. On the basis of these new results, the phase stability of DOPA/MO membranes can be reasonably explained by the spontaneous curvature of the monolayer membrane and a curvature elastic energy of the membrane.     

Structure/function relationships of several biopolymers as related to invertase stability in dehydrated systems

Structure/function relationships of different biopolymers (alginate, dextran, or beta-cyclodextrin) were analyzed as single excipients or combined with trehalose in relation to their efficiency as enzyme stabilizers in freeze-dried formulations and compared to trehalose. Particularly, a novel synthesized polymer beta-cyclodextrin-branched alginate (beta-CD-A) was employed as excipient. During freeze-drying, the polymers or their mixtures did not confer better protection to invertase compared to trehalose. Beta-CD-A (with or without trehalose), beta-cyclodextrin (beta-CD), or dextran with trehalose were the best protective agents during thermal treatment, while beta-CD and alginate showed a negative effect on invertase activity preservation. The beta-CD linked alginate combined the physical stability provided by alginate with the stabilization of hydrophobic regions of the enzyme provided by cyclodextrin. Beta-CD-A was effective even at conditions at which trehalose lost its protective effect. A relatively simple covalent combination of two biopolymers significantly affected their functionalities and, consequently, their interactions with proteins, modifying enzyme stability patterns.     

海藻糖对载药脂质体在干燥-再水化过程中的保护作用

包载抗肿瘤药物阿糖胞苷及阿霉素的脂质体,在真空干燥-再水化后,其内含物大部分漏出(保存率在20％以下)。在有足够数量海藻糖存在时,其内含物保存率可达80％以上。干燥状态下贮存,明显地比水化状态脂质体具有更良好的稳定性,本文所用方法对载药脂质体在临床中的应用有一定意义。     

Protein inactivation in amorphous sucrose and trehalose matrices: effects of phase separation and crystallization

Trehalose is the most effective carbohydrate in preserving the structure and function of biological systems during dehydration and subsequent storage. We have studied the kinetics of protein inactivation in amorphous glucose/sucrose (1:10, w/w) and glucose/trehalose (1:10, w/w) systems, and examined the relationship between protein preservation, phase separation and crystallization during dry storage. The glucose/trehalose system preserved glucose-6-phosphate dehydrogenase better than did the glucose/sucrose system with the same glass transition temperature (T_g). The Williams-Landel-Ferry kinetic analysis indicated that the superiority of the glucose/trehalose system over the glucose/sucrose system was possibly associated with a low free volume and a low free volume expansion at temperatures above the T_g. Phase separation and crystallization during storage were studied using differential scanning calorimetry, and three separate domains were identified in stored samples (i.e., sugar crystals, glucose-rich and disaccharide-rich amorphous domains). Phase separation and crystallization were significantly retarded in the glucose/trehalose system. Our data suggest that the superior stability of the trehalose system is associated with several properties of the trehalose glass, including low free volume, restricted molecular mobility and the ability to resist phase separation and crystallization during storage.     

Dielectric spectroscopy study of myoglobin in glycerol-water mixtures

Due to the interest in protein dynamics, there are numerous dielectric relaxation studies of proteins in water and in glass-forming aqueous solvents such as glycerol-water mixtures. In the regime of low frequencies, the inevitable dc-conductivity of such systems limits the resolution of dynamics that are slow compared with the solvent relaxation. Solutions of myoglobin in glycerol/water mixtures of various compositions are measured by dielectric spectroscopy in the frequency range from 10 mHz to 10 MHz. The resolution of low frequency modes is improved by two approaches: electrical ‘cleaning’ and the analysis of the derivative of the real component of permittivity, which shows no direct signature of dc-conductivity. Effects of internal interfacial polarization are also addressed by measuring the same solvents in confinement as well as mixed with glass beads. We find two processes, the structural relaxation of the solvent and the slower rotational mode of the protein, with no indication at even lower frequencies of a dielectric signature of fluctuations associated with protein dynamics.     

Effect of glycerol on behaviour of amylose and amylopectin films

The effect of water and glycerol on sorption and calorimetric T_gs of amylose and amylopectin films were examined. The mechanical properties of the films were also analysed under varying glycerol content at constant RH and temperature. Based on changes observed in sorption and tensile failure behaviour glycerol was strongly interacted with both starch polymers. Even though water was observed to be more efficient plasticiser than glycerol, glycerol also affected the T_g. But in spite of the observed decrease in T_g under low glycerol contents brittleness of the films increased based on changes in elongation. The increase in brittleness of both polymers was also in agreement with their actual behaviour. At around 20% glycerol great change in the rheological properties occurred. Above 20% glycerol amylose film showed much larger elongation than the low glycerol content films and was still strong but the amylopectin produced a very week and non-flexible film.     

Temperature dependent vibrational modes of glycosidic bond in disaccharide sugars

We studied the temperature dependent vibrational modes of the glycosidic bond in trehalose, sucrose, and maltose at wavenumbers ranging from 1000 to 1200 cm(-1). We found that the slope of temperature dependent Raman shifts of the glycosidic bond in trehalose and sucrose changed at temperatures around 120 degrees C, indicating a bond length or a bond angle (dihedral and torsional angles) change. However, we did not observe any slope change in maltose because the melting temperature of maltose is very close to 120 degrees C. We also found, at temperatures below 120 degrees C, that Raman shifts of the vibrational modes of the glycosidic bond in trehalose showed the strongest temperature dependence among the three disaccharides.     

The glass transition temperatures of amorphous trehalose-water mixtures and the mobility of water: an experimental and in silico study

Isothermal-isobaric molecular dynamics simulations are used to calculate the specific volume of models of trehalose and three amorphous trehalose-water mixtures (2.9%, 4.5% and 5.3% (w/w) water, respectively) as a function of temperature. Plots of specific volume versus temperature exhibit a characteristic change in slope when the amorphous systems change from the glassy to the rubbery state and the intersection of the two regression lines provides an estimate of the glass transition temperature T(g). A comparison of the calculated and experimental T(g) values, as obtained from differential scanning calorimetry, shows that despite the predicted values being systematically higher (about 21-26K), the trend and the incremental differences between the T(g) values have been computed correctly: T(g)(5.3%(w/w))相似文献