Protein—Protein Interactions in Aqueous Ammonium Sulfate Solutions. Lysozyme and Bovine Serum Albumin (BSA)

Osmotic pressures have been measured to determine lysozyme—lysozyme,BSA—BSA, and lysosyme—BSA interactions for protein concentrations to 100 g-L^–1in an aqueous solution of ammonium sulfate at ambient temperature, as a functionof ionic strength and pH. Osmotic second virial coefficients for lysozyme, forBSA, and for a mixture of BSA and lysozyme were calculated from theosmotic-pressure data for protein concentrations to 40 g-L^–1. The osmotic second virialcoefficient of lysozyme is slightly negative and becomes more negative withrising ionic strength and pH. The osmotic second virial coefficient for BSA isslightly positive, increasing with ionic strength and pH. The osmotic second virialcross coefficient of the mixture lies between the coefficients for lysozyme andBSA, indicating that the attractive forces for a lysozyme—BSA pair areintermediate between those for the lysozyme—lysozyme and BSA—BSA pairs. For proteinconcentrations less than 100 g-L^–1, experimental osmotic-pressure data comparefavorably with results from an adhesive hard-sphere model, which has previouslybeen shown to fit osmotic compressibilities of lysozyme solutions.     

Release of lysozyme from the branched polyelectrolyte-lysozyme complexation

On the basis of the discretely charged sphere model of lysozyme, the release behavior of lysozyme from the branched polyelectrolyte-lysozyme complexation is investigated by adding salt and changing the pH values of the solution. It is found that, with the increase of the salt ionic strength of the solution, the lysozymes are gradually released from the oppositely charged polyelectrolyte as a result of the screening of electrostatic attraction between the two ionic species by adding the salt. Interestingly, there exists a critical salt ionic strength at which all proteins are released from the branched polyelectrolyte, and the polyelectrolyte-protein complexation is broken completely. Beyond the critical value, the increase of the salt ionic strength causes self-association of the proteins released from the branched polyelectrolyte-protein complexation. The self-association of the protein is detrimental in biological systems. By calculating the second virial coefficient, we found that the optimal salt content for the dispersion of proteins coincides with the critical ionic strength, because the second virial coefficient reaches its maximum at the critical ionic strength. Similarly, increasing the pH value of the solution can also release the lysozymes from the polyelectrolyte, because the increase of pH value of the solution changes the charge distribution and net charge of the lysozyme, weakens the attraction between lysozymes mediated by polyelectrolyte, and finally leads to the dissolution of the complexation of branched polyelectrolyte with lysozymes in strong alkaline solution. In addition, by exploring the effect of architecture of the polyelectrolyte on the release behavior of proteins, we found that it is more difficult to release proteins from the branched polyelectrolyte than from the linear polyelectrolyte.     

Molecular properties of poly(carboxybetaine) in solutions with different ionic strengths and pH values

Viscometry and dynamic and static light scattering are employed to study the molecular properties of water-soluble poly(carboxybetaine), that is, poly(2-(diallyl(methyl)ammonium) acetate). It is shown that, in solutions with pH 1, the polymer behaves as a polyelectrolyte. In media with pH 6 and 13, an increase in the concentration of sodium chloride increases the intrinsic viscosity of the polymer and the hydrodynamic radius of its macromolecules, thereby indicating the antipolyelectrolyte effect typical of polymer zwitterions. In water and 0.1 M NaOH, the second virial coefficient of the polymer is close to zero, while exponent ν, which relates the sizes of macromolecules to their molecular masses, is 0.5. In 1 M NaCl, the second virial coefficient becomes positive, while exponent increases to 0.58. The Kuhn segment lengths of poly(carboxybetaine) molecules are 6.3 and 6.6 nm in water and 1 M NaCl, respectively. An increase in the hydrodynamic radius of macromolecules with the ionic strength of the solution is due to the shielding of attraction between zwitterions belonging to polybetaine monomer units located far apart along a macromolecular chain.     

Evidence of hydration forces between proteins

Proteins are fundamental molecules in biology that are also involved in a wide range of industrial and biotechnological processes. Consequently, many works in the literature have been devoted to the study of protein–protein and protein–surface interactions in aqueous solutions. The results have been usually interpreted within the frame of the classical Derjaguin–Landau–Verwey–Overbeek (DLVO) theory for colloidal systems. However, against the DLVO predictions, striking evidence of repulsive forces between proteins at high salt concentrations has been observed in different works based on the analysis of the second virial coefficient or on the direct measurement of protein interaction with an atomic force microscope. Hydration forces due to the adsorption of hydrated cations onto the negatively charged protein surfaces have been invoked to rationalize this anomalous repulsion. The hydration forces between proteins provide protein-covered particles with a non-DLVO colloidal stability at high salt concentrations, as different studies in the literature has proven. This review summarizes the most relevant results published so far on the presence of hydration forces between proteins and protein-coated colloidal particles.     

Analysis of Light Scattering Data on the Calcium Ion Sensitivity of Caseinate Solution Thermodynamics: Relationship to Emulsion Flocculation

We describe the quantitative interrelation between the thermodynamic parameters of caseinate submicelles in the presence of calcium ions (0-14 mM) in aqueous medium and the capacity of the protein to induce depletion flocculation in oil-in-water emulsions at pH 7.0 and ionic strength 0.05 mol dm(-3). Measurements have been made by static and dynamic multiangle laser light scattering of the weight-average molecular weight, the radius of gyration, the hydrodynamic radius, and the second virial coefficient of caseinate submicelles in aqueous solution. Successive thermodynamic approximations with and without consideration of correlations between caseinate submicelles have been used to calculate the osmotic pressure in caseinate aqueous solutions and the free energy of the depletion interaction between droplets in oil-in-water emulsions stabilized by caseinate. Numerical results from both thermodynamic approximations are in reasonably good agreement with experiment, predicting a pronounced decrease in the strength of the depletion attraction at concentrations of Ca(2+) in the range 4-8 mM (with a minimum value at 8 mM). This correlates well with the great enhancement of stability of these emulsions with respect to flocculation in comparison with systems having no added ionic calcium and emulsions with lower (2 mM) or higher (10 mM) Ca(2+) contents. Nevertheless, the allowance for interactive correlations between caseinate submicelles seems to lead to a better prediction of emulsion flocculation on a qualitative level over the whole range of Ca(2+) concentrations studied (2-14 mM). The calculated pronounced decrease in depletion interaction strength is attributable to marked changes in weight-average molecular weight and mean size of aggregates, and to more positive values of the second virial coefficient of caseinate submicelles with increasing Ca(2+) content. Finally, we discuss the part played by the electrical charge on the protein in determining the overall strength of the flocculation-inducing attractive interactions between droplets. Copyright 2001 Academic Press.     

Modification of Surface Forces by Metal Ion Adsorption

Abstract

Sorption of ions may lead to variations in interparticle forces and, thus, changes in the stability of colloidal particles. Chemical interactions between metal ions and colloidal particles modify the molecular structure of the surface, the surface charge, and the electrical potential between colloidal particles. These modifications to the surface and to the electrical double layer due to metal ion sorption are reflected in the interaction force between a particle and another surface, which is measured in this study by atomic force microscopy (AFM). Specifically, AFM is used to investigate the sorption of copper ions from aqueous solutions by silica particles. The influence of metal ion concentration and solution ionic strength on surface forces is studied under transient conditions. Results show that as the metal ion concentration is decreased, charge reversal occurs and a longer period of time is required for the system to reach equilibrium. The ionic strength has no significant effect on sorption kinetics. Furthermore, neither metal concentration nor ionic strength exhibits any effect on sorption equilibria, indicating that for the experimental conditions used in this study, the surface sites of the silica particle are fully occupied by copper ions.     

Ion binding in sulfonate-containing polyelectrolytes

Emf measurements have been used to study the activity and the extent of sodium ion binding to several polyelectrolytes, poly(styrenesulfonate) (PSS), poly(2-acrylamido-2-methylpropanesulfonate) (PAMS), poly(3-methacryloyloxypropane-1-sulfonate) (PMOS), and copoly[(N-t-butylacrylamide)-(2-acrylamido-2-methylpropanesulfonate)] (NB–AMS) (mole ratio 3.8:1). The activity coefficient of sodium ion in Na-PAMS and Na-PMOS without added salt is found to be in the range from 0.1 to 0.3 and is insensitive to changes in polymer concentration. However, it increases with increasing concentration of the added salt. The extent of sodium ion binding at a given sodium ion concentration, as estimated from the activity data for some sodium and tetrabutylammonium (TBA) salts, decreases in the order Na-PAMS ≈? Na-PMOS > TBA–PAMS > TBA–(NB–AMS). This indicates that a significant portion of the binding is attributed to the binding of TBA⁺ ions. Also compared are the results of ion binding in Na-PAMS and Na-PSS as a function of ionic strength. At low ionic strength (<1.0M), the order of binding strength is Na-PAMS > Na-PSS, while the order is reversed at high ionic strength (>1.0M). This finding is in good agreement with data obtained by dilatometry and viscometry.     

Investigation of salt properties with electro-acoustic measurements and their effect on dynamic binding capacity in hydrophobic interaction chromatography

The pH dependence in hydrophobic interaction chromatography (HIC) is usually discussed exclusively in terms of protein dependence and there are no clear defined trends. Many of the deviations from an ideal solution are caused solely by the high salt concentration, as protein concentration is usually negligible. So pH dependency in hydrophobic interaction chromatography could also be the result of pH dependent changes of ion properties from the salt solution. The possibility that pH dependent ion hydration or ion association in highly concentrated salt solutions may influence the dynamic protein binding capacity onto HIC resins was investigated. In buffer solutions commonly used in HIC e.g. sodium chloride, ammonium sulphate and sodium citrate pH dependent maxima in the electro-acoustic signals were found. These maxima are related to an increase of the ion sizes by hydration or ion association. At low ionic strength the maxima are in the range between 4.5 and 6 and they increased in concentrated electrolyte solutions to values between 6 and 8. The range of these maxima is in the same region as dynamic protein binding capacity maxima often observed in HIC. For a qualitative interpretation of this phenomenon of increased protein stabilization by volume exclusion effect extended scaling theory can be used. This theory predicts a maximum of protein stabilization if the ratio of salt ion diameter to water is 1.8. According to the hypothesis raised here, if the pH dependent ratio of salt ion diameter to water approaches this value the transport of the protein in the pore system is less restricted and an increase in binding capacity can be produced.     

Hofmeister effects in the restabilization of IgG--latex particles: testing Ruckenstein's theory

The hydration interaction is responsible for the colloidal stability observed in protein-coated particles at high ionic strengths. The origin of this non-DLVO interaction is related not only to the local structure of the water molecules located at the surface but also to the structure of those molecules involved in the hydration of the ions that surround the colloidal particles. Ruckenstein and co-workers have recently developed a new theory based on the coupling of double-layer and hydration interactions. Its validity was contrasted by their fitting of experimental data obtained with IgG-latex particles restabilized at high salt concentration. The theory details the important role played by the counterions in the stability at high salt concentrations by proposing an ion pair reaction forming surface dipoles. These surface dipoles are responsible of repulsive interactions between two approaching surfaces. This paper checks the theory with recent data where some ions associated with the Hofmeister series (NO(3)(-), SCN(-) and Ca(2+)) restabilize the same kind of IgG-latex systems by means of hydration forces. Surprisingly, these ions induce stability acting even as co-ions, likely by modifying the water structure at the surface, but not forming surface ion pairs. Therefore, this experimental evidence would question Ruckenstein's theory based on the surface dipole formation for explaining the observed restabilization phenomena.     

Performance of the diffusive gradients in thin films technique for measurement of trace metals in low ionic strength freshwaters

A series of experiments were undertaken to investigate the effect of ionic strength and the concentration of free sodium ions in the resin gel on the performance of the diffusive gradients in thin films (DGT) technique. When the free sodium ion concentration in the resin gel was estimated by the time-dependent release into solution, it agreed with a previous estimate. However, equilibration with different volumes of water gave a higher value, suggesting that inherent averaging in the time-dependent release method underestimates the free concentration. DGT measurements of Cu and Cd were made over a wide range of ionic strengths (from 3 μmol l⁻¹ to 0.8 mol l⁻¹). For all the ionic strengths above 100 μmol l⁻¹ there was no significant difference between measurements made by DGT and measurements made directly on the solution using atomic absorption spectroscopy. Below 100 μmol l⁻¹ results were erratic. They did not comply with a theory that predicts high results for DGT based on enhancement of the diffusion coefficient of trace metal cations by counter diffusion of sodium ions. When Cd in solutions with a range of ionic strengths was measured by DGT there was no difference whether the resin gels were in Na or Ca form. Rather than counter diffusion of Na ions, it is suggested that the spurious behaviour at low ionic strength is due to interactions of the trace metals with the diffusion gel when there are insufficient excess cations present.     

The osmotic second virial coefficient and the Gibbs–McMillan–Mayer framework

The osmotic second virial coefficient is a key parameter in light scattering, protein crystallisation, self-interaction chromatography, and osmometry. The interpretation of the osmotic second virial coefficient depends on the set of independent variables. This commonly includes the independent variables associated with the Kirkwood–Buff, the McMillan–Mayer, and the Lewis–Randall solution theories. In this paper we analyse the osmotic second virial coefficient using a Gibbs–McMillan–Mayer framework which is similar to the McMillan–Mayer framework with the exception that pressure rather than volume is an independent variable. A Taylor expansion is applied to the osmotic pressure of a solution where one of the solutes is a small molecule, a salt for instance, that equilibrates between the two phases. Other solutes are retained. Solvents are small molecules that equilibrate between the two phases. The independent variables of the solvents are temperature, pressure, and chemical potentials. The derivatives in the Gibbs–McMillan–Mayer framework are transformed into derivatives in the Gibbs framework. This offers the possibility for an interpretation and correlation of the osmotic second virial coefficient using activity coefficient models.     

Perturbation of water structure due to monovalent ions in solution

The ion induced modification to the tetrahedral structure of water is a topic of much current interest. We address this question by interpreting neutron diffraction data from monovalent ionic solutions of NaCl and KCl using a computer assisted structural modeling technique. We investigate the effect that these ions have on the water-water O-O, O-H and H-H radial distribution functions as a function of ionic concentration. It is found that the O-H and H-H functions are only marginally affected by ionic composition, signaling that hydrogen bonding between water molecules remains largely intact, even at the highest concentrations. On the other hand the O-O functions are strongly modified by the ions. In particular the position of the second peak in g(OO)(r), is found to move inwards with increasing salt concentration, in a manner closely analogous to what happens in pure water under pressure. Furthermore by recalculating g(OO)(r) after excluding all the water molecules in the first hydration shell of each ion, we show that this structural perturbation exists outside the first hydration shell of the ions.     

Computer simulation study on the concentration distribution of spherical colloids within confined spaces of well‐defined pores

Aside from the virial expansion and density functional methods, theoretical results on the concentration partitioning behavior for charged colloids within cylindrical pores have not been presented so far. With the increase of relative solute size as well as solute concentration, however, the approximate analytic methods have proven to be unreliable. A suitable Monte Carlo simulation, which is proved as a rigorous technique for concentrated colloids, has been applied in the present study. The concentration profiles within the pore representing the effects of solute concentration as well as solution ionic strength are obtained via a stochastic process, from which the partition coefficient is estimated. Previously developed analyses on the linearized Poisson‐Boltzmann (P‐B) equation are employed for the estimation of long‐range electrostatic interaction. Both the singularity method and the analytical solution with series representation properly determine respective interaction energies between pairs of solute particles and between the solute particle and the pore wall. The effect of solute‐solute and solute‐wall interactions associated with repulsive energy is presented on the partitioning of colloids. Simulation results show that the partition coefficient is evidently enhanced when no particle‐wall interaction exists. Hindered diffusion can be predicted by the simplifying assumption of the centerline approximation analogy, where a dependence on the solute concentration becomes greater as the solution ionic strength decreases.     

Existence of hydration forces in the interaction between apoferritin molecules adsorbed on silica surfaces

The atomic force microscope, together with the colloid probe technique, has become a very useful instrument to measure interaction forces between two surfaces. Its potential has been exploited in this work to study the interaction between protein (apoferritin) layers adsorbed on silica surfaces and to analyze the effect of the medium conditions (pH, salt concentration, salt type) on such interactions. It has been observed that the interaction at low salt concentrations is dominated by electrical double layer (at large distances) and steric forces (at short distances), the latter being due to compression of the protein layers. The DLVO theory fits these experimental data quite well. However, a non-DLVO repulsive interaction, prior to contact of the protein layers, is observed at high salt concentration above the isoelectric point of the protein. This behavior could be explained if the presence of hydration forces in the system is assumed. The inclusion of a hydration term in the DLVO theory (extended DLVO theory) gives rise to a better agreement between the theoretical fits and the experimental results. These results seem to suggest that the hydration forces play a very important role in the stability of the proteins in the physiological media.     

The effect of a background electrolyte on the premicellar association and average activity of sodium dodecyl sulfate ions

Premicellar association in a sodium dodecyl sulfate-NaCl-water system is studied by potentiometry using a sodium-selective electrode. The dependences of constants and degrees of dimerization of surfactant molecules on the background electrolyte concentration are determined. The average activity coefficient of surfactant ions is shown to increase with the ionic strength of a solution. The disclosed regularity is explained in terms of the Debye-Hückel theory with allowance made for the dependence of premicellar solution composition on the content of NaCl.__________Translated from Kolloidnyi Zhurnal, Vol. 67, No. 2, 2005, pp. 206–212.Original Russian Text Copyright © 2005 by Podchasskaya, Usyarov.     

Ion and Adsorbing Colloid Flotation of Auramine

Auramine, a cationic dye, was removed from synthetic wastewater by ion flotation of auramine‐sodium lauryl sulfate complex. Over 98% of auramine was removed from the solution in 15 min. A stoichiometric amount of surfactant (1 mol of surfactant to 1 mol of dye) was found to be most effective for auramine removal. The rate of separation and ultimate removal of auramine increased with increasing the rate of air flow and decreased with increasing concentration of NaNO₃. Auramine was also removed by adsorbing colloid flotation technique using ferric hydroxide as the coagulant. Sodium lauryl sulfate was used as the collector, and over 95% of auramine was removed in 10 min. The separation efficiency decreased with increasing ionic strength of the solution. The deleterious effect of neutral salt was compensated somewhat with the aid of aluminum ions as the activator. Both ion flotation and adsorbing colloid flotation are promising approaches for the removal of cationic dye from wastewater.     

Particulate hematite diffusion in sodium polyacrylate solutions. The effect of ionic strength

The diffusion coefficients of hematite particles in polyelectrolyte solution have been investigated using dynamic light scattering. Two apparent diffusion coefficients, a fast and a slow diffusional mode, are observed for the hematite particles in high-molecular-weight sodium polyacrylate solution at pH 10.5. The slow diffusion coefficient (Dslow) shows a decrease with increase in polyelectrolyte concentration. The fast diffusion coefficient (Dfast) shows an increase to a maximum with increasing polyelectrolyte concentration and then a rapid decrease as the polyelectrolyte concentration increases further. With an increase in ionic strength from 10(-4) to 0.1 M NaNO3, the maximum value of Dfast increased in magnitude, while the polyacrylate concentration at which the maximum occurs is seen to increase. The dependence of Dfast on the measurement angle indicates that it is coupled to the fluctuations of the chains. The observed behavior is attributed to the hematite probe particle sensing both macroscopic (viscous) and elastic fluctuations associated with the polyelectrolyte motion.     

Charging and aggregation of positively charged latex particles in the presence of anionic polyelectrolytes

Charging behavior and colloidal stability of amidine latex particles are studied in the presence of poly(sodium styrene sulfonate) (PSS) and KCl. Detailed measurements of electrophoretic mobility, adsorbed layer thickness, and aggregation (or coagulation) rate constant on varying the polymer dose, molecular mass of the polymer, and ionic strength are reported. Polyelectrolyte adsorption leads to the characteristic charge reversal (or overcharging) of the colloidal particles at the isoelectric point (IEP). In accordance with classical Derjaguin-Landau-Verwey-Overbeek (DLVO) theory, uncharged particles tend to aggregate because of van der Waals attraction, whereas charged particles are stabilized by electrical double layer repulsion. Attractive patch-charge interactions originating from the laterally inhomogeneous structure of the adsorbed polymer substantially decrease the suspension stability or even accelerate the aggregation rate beyond diffusion control. These electrostatic non-DLVO forces become progressively important with increasing molecular mass of the polymer and the ionic strength of the solution. At higher polymer dose of typically 10 times the IEP, one observes the formation of a saturated layer of the adsorbed polymer with a thickness of several nanometers. Its thickness increases with increasing molecular mass, whereby the layer becomes increasingly porous. This layer does not seem to be involved in the suspension stabilization, since at such high polymer doses the double layer repulsion has attained sufficient strength to stabilize the suspension.