Steady-state and pulsed NMR studies of gelation in aqueous agarose

Steady-state and pulsed NMR techniques have been used to investigate molecular motion in sols and gels of agarose. In passing through the sol–gel transition, the molecular mobility of water molecules is reduced only by a small amount, whereas motion of the polymer chains is greatly attenuated. The results are discused in terms of the network theory of gelation, with references to the role of water in the process and the nature of the “junction zones” between polymer chains. T₂ and line-width measurements are dominated by exchange broadening. The effects of exchange rate and differences in relaxation time between the exchanging sites are discussed. The temperature hysteresis behavior of agarose gels has been investigated and the effects of “ageing” correlated with changes in nuclear relaxation times. The synergistic increase in gel strength obtained on adding locust bean gum (LBG) to agarose has been investigated. The results indicate that LBG does not form double-helix junctions and may decrease rates of gelation by steric effects. At high agarose concentration, the LBG remains mainly in solution in interstitial water, but at low agarose concentration, it is suggested that the LBG can link gel aggregates together into a self-supporting structure, producing a synergistic increase in gel strength. Comparisons have been made between the nature of the agarose–LBG interaction and agarose–cellulose interactions in biological systems.     

Heat-induced reversible gelation of arachin: kinetics, thermodynamics and protein species involved in the process

Arachin forms a heat-reversible gel under certain experimental conditions. The minimal gelling concentration for this system is 7.25%. Above minimal gelling concentration calculation of thermodynamic parameters for gelation of arachin revealed a constant delta Hbonding (-1220 cal.mol-1) where delta Sbonding values varied with an increase in protein concentration (ranging from -4.01 e.u. at 7.5% to -3.48 e.u. at 10.0%). The main steps involved in the gelation phenomenon include thermal denaturation of arachin, partial aggregation of heat-denatured protein molecules, setting of protein solution and maturation of the gel formed. Gel maturation process follows first order kinetics and is characterized by a large positive delta G+(+) (22,030 cal.mol-1). Determination of delta H+(+) and delta S+(+) for this process revealed that mostly delta S+(+) (-62.9 e.u.) contributes to the large positive delta G+(+), thus decreasing the overall rate of gel maturation process. This large negative delta S+(+) value probably arises from a loss of entropy of protein molecules because of their increased involvement in gel network formation. The polymer gel network seems to be primarily contributed by a part of both arachin dodecameric and hexameric species.     

Controlled gelation of xanthan by trivalent chronic ions

Addition of trivalent chromic ions to xanthan solutions gives rise to gel formation. The dynamic shear storage and loss moduli (0.01 – 10 rad/s) of xanthan solutions with polymer concentrations ranging from 1 to 7 mg/ml and Cr³⁺ concentrations ranging from 0 to 50 m have been studied. It is found that the rate of gel formation is strongly dependent on the Cr³⁺ concentration, but to a much smaller extent on the xanthan concentration. The gelation time is less than 1 h for 50 m Cr³⁺ and about 40 h for 2 m Cr³⁺. It is found that the minimum Cr³⁺ concentration needed to give gelation of 1–7 mg/ml xanthan is 1–2 m .     

Cold-Set Whey Protein Gels with Addition of Polysaccharides

Cold-set whey protein (WP) gels with addition of xanthan or guar were evaluated by mechanical properties and scanning electron microscopy. Gels were formed after the addition of different amounts of glucono-δ-lactone to thermally denatured WP solutions, leading to different acidification rates and final pH values. At lower acidification rates and higher final pH, gels showed more discontinuous structure and weaker and less elastic network, which was attributed to a predominance of phase separation during gel formation due to slower gelation kinetics. In contrast, at higher acidification rates and lower final pHs, gelation prevailed over phase separation, favoring the formation of less porous structures, resulting in stronger and more elastic gels. The gels’ fractal dimension (D _f; structure complexity) and lacunarity were also influenced by the simultaneous effects of gelation and phase separation. For systems where phase separation was the prevailing mechanism, greater lacunarity parameters were usually observed, describing the heterogeneity of pore distribution, while the opposite occurred at prevailing gelation conditions. Increase in guar concentration or lower final pH of xanthan gels entailed in D _f reduction, while the increase in xanthan concentration resulted in higher D _f. Such a result suggests that the network contour length was rugged, but this pattern was reduced by the increase of electrostatic interactions among WP and xanthan. Guar addition caused the formation of gel network with smoother surfaces, which could be attributed to the guar–protein excluded volume effects leading to an increase in protein–protein interactions.     

Rheological and structural properties of aqueous alginate during gelation via the Ugi multicomponent condensation reaction

Turbidity, structure, and rheological features during gelation via the Ugi multicomponent condensation reaction of semidilute solutions of alginate have been investigated at different polymer and cross-linker concentrations and reaction temperatures. The gelation time of the system decreased with increasing polymer and cross-linker concentrations, and a temperature rise resulted in a faster gelation. At the gel point, a power law frequency dependence of the dynamic storage modulus (G' proportional, variant omega(n)(')) and loss modulus (G' ' proportional, variant omega(n)(' ')) was observed for all gelling systems with n' = n' ' = n. By varying the cross-linker density at a fixed polymer concentration (2.2 wt %), the power law exponent is consistent with that predicted (0.7) from the percolation model. The value of n decreases with increasing polymer concentration, whereas higher temperatures give rise to higher values of n. The elastic properties of the gels continue to grow over a long time in the postgel region, and at later stages in the gelation process, a solidlike response is observed. The turbidity of the gelling system increases as the gel evolves, and this effect is more pronounced at higher cross-linker concentration. The small-angle neutron scattering results reveal large-scale inhomogeneities of the gels, and this effect is enhanced as the cross-linker density increases. The structural, turbidity, and rheological features were found to change over an extended time after the formation of the incipient gel. It was demonstrated that temperature, polymer, and cross-linker concentrations could be utilized to tune the physical properties of the Ugi gels such as structure, transparency, and viscoelasticity.     

Properties of agar: parameters affecting gel-formation and the agarose-iodine reaction

The effects of concentration of agarose, methyl sulphoxide, and substituted agaroses on the mechanism of gel formation have been evaluated with reference to the “Network theory of gel formation”. Factors affecting formation of the agarose gel-iodine color complex were investigated, and the results suggest that the iodine molecules are incorporated between the agarose helices in the Gel II state of agarose.     

Interpenetrating network formation in agarose–sodium gellan gel composites

Aqueous cold-set gels from mixtures of agarose and sodium gellan have been characterised structurally and mechanically using optical and electron microscopy, turbidity measurements, differential scanning calorimetry, mechanical spectroscopy and compression testing. Consistent with expectations for charged–uncharged polymer combinations at low ionic strength there is no liquid–liquid demixing in sols prior to gelation, and although transmission electron microscopy reveals heterogeneities in gel microstructures at the higher polymer concentrations, these are small in extent, and are unlikely to arise from normal segregative demixing. Overall, ‘molecularly’ interpenetrating networks (IPNs) are indicated, in which the gellan and agarose architectures pass through one another on a distance scale comparable to their pore sizes. At concentrations greater than 2% w/w gellan, where gellan is the first gelling species, and when the agarose concentration is greater than 0.5% w/w, the composite modulus falls below that expected for the agarose alone. At 0.5% w/w agarose, on the other hand, modulus contributions from the components are much closer to additive. These findings are reflected in the results of large deformation compression testing where breaking stresses show similar trends.     

Demonstration of lag phase in the sol-gel transformation of deoxygenated S hemoglobin without temperature alteration.

1). During the sol to gel transformation of deoxygenated sickle hemoglobin, a time-dependent process preceding gel formation (lag phase) was demonstrated that was inversely proportional to a function of the hemoglobin concentration and that occurred without alteration in temperature, pH, or oxygen tension. 2). As determined by the Schachman modification of the capillary viscometer, preparations of oxyhemoglobin S and A and deoxyhemoglobin A were indistinguishable when compared over a wide range of concentrations. Up to the concentration at which gelling occurred, deoxyhemoglobin S exhibited the same viscosity behavior. The viscosity of deoxygenated hemoglobin S within the lower gelling concentration range was normal during the lag phase and became abnormally high only at the time of gelation.     

Jamming and gelation of dense beta-casein micelle suspensions

The rheology of dense suspensions of beta-casein micelles is investigated at pH 6. For a given temperature, the viscosity increases dramatically at a critical concentration (Cc) of about 100 g/L due to jamming of the micelles. For a given concentration close to and above Cc, the viscosity of dense suspensions decreases strongly with increasing temperature because Cc increases. The suspensions show weak shear thickening followed by strong shear thinning. At lower pH, that is, closer to the isoelectric point, spontaneous gelation is observed, which is favored by lowering the temperature and addition of sodium polyphosphate. The gelation process is studied at pH 5.5 by rheology and light scattering.     

Gel-forming structures and stages of red algal galactans of different sulfation levels

Sol-gel transition processes of algal galactans were studied using cryofixation method in combination with freeze-drying and scanning electron microscopy (SEM) techniques. The structures formed in successive stages of gelling process upon cooling were rapidly frozen at defined temperature points and viewed by SEM. It was established that in the case of both types of gelling galactans investigated, a fine honeycomb-like network exists for a wide range of solution temperatures. The formation and structure of this network depends on the structural type, gelling stage, and concentration of the galactan in solution. The honeycomb suprastructures exist also in carrageenan and agarose sols (at temperatures considerably exceeding the gelling temperatures). An additional helical network formed showed different behaviour in the case of carrageenan and agar-type polysaccharides. In the gel-formation process, tightening of the network takes place in both types of galactan gels; the honeycomb structures persist in carrageenan (furcellaran) but not in agarose gels.     

Molecular mechanisms of Fe2+-induced beta-lactoglobulin cold gelation

To get more insight into the mechanisms of cold gelation of beta-lactoglobulin (beta-lg), macroscopic and molecular structural changes during Fe(2+)-induced gelation of beta-lg were investigated using Fourier transform-infrared (FTIR) spectroscopy and rheological methods. The FTIR spectroscopy results show that, upon the preheating treatment (first step of gel process), native globular proteins are denatured and aggregated molecules are found in solution. The spectra are similar to those of gels obtained in the second step of the process upon incorporation of Fe, which suggests that aggregated molecules formed during the preheating treatment constitute the structural basis of the aggregation. However, the rheological data show that the aggregation is achieved via two molecular mechanisms, both of which are modulated by the iron concentration. At 30 mM of iron, gel formation is essentially controlled by van der Waals interactions, while at 10 mM of iron, hydrophobic interactions predominate. At the two concentrations, disulfide bonds contribute to gel consolidation, the effect being more pronounced at 10 mM of iron. These mechanisms lead to the formation of gels of different microstructures. At the highest iron concentration, a strong and rapid decrease in the repulsion forces is produced, resulting in random aggregation. At the lowest iron concentration, the iron diminishes the superficial charge of both molecules and aggregated molecules, facilitating the interaction among hydrophobic regions and leading to the growth of the aggregation in the preferential direction and to filamentous gel formation. This study provides a comprehensive view of the different modes of gelation.     

“Melt-in-the-mouth” gels from mixtures of xanthan and konjac glucomannan under acidic conditions: A rheological and calorimetric study of the mechanism of synergistic gelation

The effect of acidification on a typical commercial xanthan and on pyruvate-free xanthan (PFX), alone and in gelling mixtures with konjac glucomannan (KGM), has been studied by differential scanning calorimetry (DSC) and small-deformation oscillatory measurements of storage modulus (G′) and loss modulus (G″). For both xanthan samples, progressive reduction in pH caused a progressive increase in temperature of the disorder–order transition in DSC, and a progressive reduction in gelation temperature with KGM. This inverse correlation is interpreted as showing that synergistic gelation involves disruption of the xanthan 5-fold helix, probably by attachment of KGM to the cellulosic backbone of the xanthan molecule (as proposed previously by a research group in the Institute of Food Research, Norwich, UK). Higher transition temperature accompanied by lower gelation temperature for PFX in comparison with commercial xanthan at neutral pH is explained in the same way. However, an additional postulate from the Norwich group, that attachment of KGM (or galactomannans) can occur only when the xanthan molecule is disordered, is inconsistent with the observation that gelation of acidified mixtures of KGM with PFX can occur at temperatures more than 60 °C below completion of conformational ordering of the PFX component (as characterised by DSC). Increase in G′ on cooling for mixtures of commercial xanthan with KGM at pH values of 4.5 and 4.25 occurred in two discrete steps, the first following the temperature-course observed for the same mixtures at neutral pH and the second occurring over the lower temperatures observed for mixtures of KGM with PFX at the same values of pH. These two “waves” of gel formation are attributed to interaction of KGM with, respectively, xanthan sequences that had retained a high content of pyruvate substituents, and sequences depleted in pyruvate by acid hydrolysis. At pH values of 4.0 and lower, gelation of mixtures of KGM with commercial xanthan followed essentially the same temperature-course as for mixtures with PFX, indicating extensive loss of pyruvate under these more strongly acidic conditions. Mixtures prepared at pH values in the range 4.0–3.5 gave comparable moduli at room temperature (20 °C) to those obtained at neutral pH, but showed substantial softening on heating to body temperature, suggesting possible applications in replacement of gelatin in products where “melt-in-the-mouth” characteristics are important for acceptability to the consumer.     

Transient electric birefringence study of highly dilute agarose solutions

In order to characterize the first step of agarose gelation, highly dilute solutions (2·10^?3 to 0.5 g/L) have been studied by means of the transient electric birefringence technique. The field-free decay curves of the birefringence are well described by a stretched-exponential B(t) ≈ exp(?t/τ)^β; the value of the exponent β is close to 0.5 whatever the agarose concentration. The suspended particles observed by electron microscopy present a rod-like shape with a constant diameter (～50 Å), without any branching; they are polydisperse with a distribution of lengths approximately exponential. The mean length of these fibers, deduced from their mean rotational diffusion coefficient, is proportional to the 0.37 power of the agarose concentration in the solution. Furthermore, these particles possess a strong permanent electrical dipole confirming the side-to-side arrangement of helices into bundles; this dipole is roughly proportional to the particle length, indicating a self-similarity in the unidirectional growth of the agarose fibers, even when approaching the gelling concentration.     

Gelation of xanthan with trivalent metal ions

In-situ gelation of semidilute xanthan solutions with trivalent chromium, aluminum or iron ions was studied by rheology and UV-spectroscopy. Measurements of the elastic modulus of xanthan gel cylinders prepared by dialysis against the complexing ion at pH values from 2 to 6 indicate that monomeric species of the ion are ineffective, whereas dimeric or higher oligomeric species are effective in crosslinking the polysaccharide. When chromium was used as the crosslinking species, the dependence of the gelation rate on the ionic concentration followed a power law with a coefficient of 1·7. The gelation time and the gelation rate were found to extrapolate to zero at 1 m Cr for 2·5 mg/ml xanthan. The limiting concentration of xanthan needed for gelation with 5 m Cr(III) at 20°C was estimated as 0·35 mg/ml. This critical xanthan concentration is close to the overlap concentration c^* estimated from the experimentally determined intrinsic viscosity [η] using c^* = 1·4/[η]. An apparent activation energy for crosslinking of xanthan was calculated as E_a = 42 kJ/mol and E_a = 108 kJ/mol for Cr and Al ions, respectively. The fractal dimensionality of xanthan-Cr at the sol-gel transition was estimated as 1·3 applying the Chambon-Winter criterion for gelation, thus indicating that this gelation criterion is applicable also to stiff-chain polysaccharides such as xanthan.     

Kinetic studies on photolysis-induced gelation of sickle cell hemoglobin suggest a new mechanism.

The kinetics of deoxyhemoglobin S gelation have been investigated using photolytic dissociation of the carbon monoxide complex to initiate the process. Measurements over a wide range of times, 10(-3)-10(4) show that both the concentration dependence of the tenth-time (i.e., the time required to complete one-tenth the reaction) and the time dependence of the process decrease as gelation speeds up. In slowly gelling samples, where single domains of polymers are formed in the small sample volumes employed with this technique (1-2 x 10(-9) cm3), there is a marked increase in the variability of the tenth-times. These results are explained by a mechanism in which gelation is initiated by homogeneous nucleation of polymers in the bulk solution phase, followed by heterogeneous nucleation on the surface of existing polymers. At the lowest concentrations, homogeneous nucleation is so improbable that stochastic behavior is observed in the small sample volumes, and heterogeneous nucleation is the dominant pathway for polymer formation, thereby accounting for the high time dependence. At the highest concentrations homogeneous nucleation becomes much more probable, and the time dependence decreases. The decrease in concentration dependence of the tenth-time with increasing concentration results from a decrease in size of both the homogeneous and heterogeneous critical nuclei. The model rationalizes the major observations on the kinetics of gelation of deoxyhemoglobin S, and is readily testable by further experiments.     

Microscopy of xanthan/galactomannan mixtures

Polarization microscopy has been used to investigate the structure of 50/50 xanthan/galactomannan (guar gum or locust bean gum) mixtures in aqueous solution, the total concentration ranging from 0.5 to 4%. By the use of polarized light microscopy birefringent areas resulting from the formation of cholesteric mesophases in xanthan gum was clearly seen as has previously been reported by several authors. In xanthan/galactomannan mixtures, we also observed birefringent areas. Moreover, these zones in the blend appeared more anisotropic than with xanthan gum alone. This suggests that xanthan molecules organize themselves as liquid crystalline mesophases in definite enriched xanthan areas resulting from a concentration of xanthan inside these birefringent zones. Upon heating, this anisotropy disappears at a temperature well below the helix-coil transition temperature of xanthan molecules. In fact, this loss of order of the mixed system occurs at the same temperature as the melting temperature of the gel, as assessed by the use of rheological measurements. Since the ordered helical structure of the xanthan molecules still exists beyond the melting temperature while anisotropy disappears, this suggests that the xanthan molecules are no longer concentrated in specific areas but more evenly distributed in the medium. Gel melting would, therefore, be the result of the disappearance of these xanthan enriched areas.     

Effect of hydrophobic modification on rheological and swelling features during chemical gelation of aqueous polysaccharides

Rheological characteristics during chemical gelation with the cross-linker ethylene glycol diglycidyl ether (EGDE) of semidilute aqueous solutions of hydroxyethylcellulose (HEC) and of two hydrophobically modified analogues (HM-1-HEC and HM-2-HEC) are reported. In addition, rheological features of gelling samples (dextran and its hydrophobically modified analogue (HM-dextran)) of a different structure have been examined. Some swelling experiments on these gels in the postgel region are also reported. The gelation time of the hydroxyethylcellulose systems decreased with increasing cross-linker concentration, and incorporation of hydrophobic units of HEC resulted in a slower gelation. The time of gelation for the dextran system was only slightly affected by the incorporation of hydrophobic groups (HM-dextran). At the gel point, a power law frequency dependence of the dynamic storage modulus (G' proportional to omegan') and loss modulus (G' proportional to omegan') was observed for all gelling systems with n' = n' = n. The attachment of hydrophobic moieties on the dextran chains had virtually no impact on the value of n (n = 0.77), and the percolation model describes the incipient dextran gels. By increasing the number of hydrophobic groups of the HEC polymer, the value of n for the corresponding incipient gel drops significantly, and the value of the gel strength parameter increases strongly. Incorporation of hydrophobic units in the HEC chains promotes the formation of stronger incipient gels because of the contribution from the hydrophobic association effect. The frequency dependence of the complex viscosity reveals that all the investigated gels become more solidlike in the postgel domain. Far into the postgel region, the hydrophobicity of HEC plays a minor role for the strength of the gel network, whereas the values of the complex viscosity are significantly higher for HM-dextran than for the corresponding dextran gel. The swelling experiments on HEC, HM-1-HEC, and HM-2-HEC systems disclose that the degree of swelling of the postgels in water is quite different, depending on the relative distance from the gel point at which the cross-linker reaction is quenched. At a given distance from the gel point, the swelling of the HEC gel is less pronounced than for the corresponding hydrophobically modified samples. At this stage, the swelling of the HM-dextran gel is stronger than for the dextran gel.     

Thermosensitive Poloxamer 407/Poly(Acrylic Acid) Hydrogels with Potential Application as Injectable Drug Delivery System

Thermosensitive hydrogels are of great interest for in situ gelling drug delivery. The thermosensitive vehicle with a gelation temperature in a range of 30–36°C would be convenient to be injected as liquid and transform into gel after injection. To prepare novel hydrogels gelling near body temperature, the gelation temperature of poloxamer 407 (PX) were tailored by mixing PX with poly(acrylic acid) (PAA). The gelation behaviors of PX/PAA systems as well as the interaction mechanism were investigated by tube inversion, viscoelastic, shear viscosity, DSC, SEM, and FTIR studies. The gelation temperature of the plain PX solutions at high concentration of 18, 20, and 22% (w/w) gelled at temperature below 28°C, which is out of the suitable temperature range. Mixing PX with PAA to obtain 18 and 20% (w/w) PX with 1% (w/w) PAA increased the gelation temperature to the desired temperature range of 30–36°C. The intermolecular entanglements and hydrogen bonds between PX and PAA may be responsible for the modulation of the gelation features of PX. The mixtures behaved low viscosity liquid at room temperature with shear thinning behavior enabling their injectability and rapidly gelled at body temperature. The gel strength increased, while the pore size decreased with increasing PX concentration. Metronidazole, an antibiotic used for periodontitis, was incorporated into the matrices, and the drug did not hinder their gelling ability. The gels showed the sustained drug release characteristic. The thermosensitive PX/PAA hydrogel could be a promising injectable in situ gelling system for periodontal drug delivery.     

Thermally induced protein gelation: gelation and rheological characterization of highly concentrated ovalbumin and soybean protein gels

In order to optimize the use of proteins as functional ingredients in foods, one needs more insight into the effects of environmental conditions (pH, ionic strength, and temperature) on the functional properties of protein. This paper summarizes the results of an extensive study on heat-induced gelation of ovalbumin (egg-white protein) and soybean protein in the concentration range from 10 to 35 g/100 g. It was the aim of the study to relate the rheological properties of thermally induced protein gels to the microstructure of the gel and the physicochemical properties of the constituent protein. The gelling behavior of the protein was quantified with rheological techniques, and the physical properties of the gels were determined, at small and large deformations. From the swelling/dissolving behavior of the gels in various media, the nature of the crosslinks was determined qualitatively. The microstructure of the gels was determined with electron microscopy. Nmr-spectroscopy was applied in order to elucidate changes in conformation during heating. It was found that the formation of a continuous covalently crosslinked network is not a prerequisite for thermally-induced protein gelation. The properties of a gel strongly depend on the pH at which the gel is formed. When heat-set at high pH(pH～10), a homogeneous, strong, and almost transparent gel is formed, consisting of flexible crosslinked protein gels. Heat-setting at low pH (pH 5) leads to the formation of a heterogeneous and weak gel, which easily exudes water. This gel consists of crosslinked aggregated protein. The ionic strength of the solvent in which the protein is dissolved and heat-set has a much lower effect on gel properties.     

Influence of alginate properties and gel reinforcement on fermentation characteristics of immobilized yeast cells

The effect of alginate composition, gel concentration, gelation method, cell loading and surface area on fermentation characteristics of immobilized yeast cells have been investigated. Molecular weight and G/M ratio had only little effect on fermentation velocity and gel strength, while increasing the alginate concentration caused a sizeable decrease in fermentation velocity and an increase in gel strength. The internally gelled immobilizates generally showed a higher fermentation velocity for the same gel strength and no decrease in gel strength was seen during fermentation. With high initial cell loadings, the fermentation velocity per g of immobilizate was higher, but the productivity per cell was lower than with low initial cell loadings. The difference decreased with time. Specific surface area (surface/volume) was shown to be an important factor for the observed productivity per gram of immobilizate, with high S/V ratios giving the highest productivity. Gel shape had no influence on fermentation velocity for a given S/V ratio. Gelation behaviour of externally gelled beads was determined by estimating the amount of cells liberated during gel formation through measurement of invertase activity (yeast-bound) in the gelling solution. A method for reinforcement of internally gelled alginate slabs with a nylon mesh was developed and utilized for production of a continuous fermentation reactor with reinforced gels.