Modelling gradient elution of bioactive multicomponent systems in non-linear ion-exchange chromatography

A theoretical framework for the ion-exchange behaviour of bioactive substances in non-linear ion-exchange chromatogaphy is described. The aim of the study was the creation of a model basis to support a process design for production-scale ion-exchange chromatography. The theory can be applied to a whole variety of biological substances, such as amino acids, polysaccharides, peptides and proteins and either isocratic or gradient elution can be carried out. The influence of the eluent concentration on the ion-exchange as well as on the characteristic charge was considered. Experimental measurements showed a strong non-linear ion-exchange equilibrium with a transition from a Langmuir-type to a sigmoidal isotherm at higher eluent concentrations. Hereby, the compound binds to the surface though it is not ionic. Therefore, the model considered the possibility of ion-exchange as well as adsorption. A simplified distribution of the counter-ions based on the Gouy-Chapman theory with a discrete distribution of the counter-ions was used. The theory was extended by a selectivity in the double layer to allow specific adsorption. Calculations of adsorption-elution cycles showed, in agreement with the experimental observations, the development of non-linear elution profiles with a desorption fronting. As a result, the column loading and the eluent concentration were varied. The effect of contaminants, in this case sodium ions, was investigated and included in the model. Finally, the model was extended to multicomponent systems to investigate the effect of side components on the retention behaviour. The development of the characteristic elution profiles and the effect of the column loading on the separation are discussed. Calculated concentration profiles along the column at discrete time steps were used to reveal the influence of side components and the underlying separation mechanism. The simulations provided a new insight into the phenomena involved in biochromatography and make convenient design concepts at least doubtful as the separation is in this case mainly determined by the loading step and not by the choice of the elution gradient.     

Purification of complex protein mixtures by ion-exchange displacement chromatography using spacer displacers.

The anion-exchange separation of complex protein mixtures by displacement chromatography using spacer displacers driven by high-affinity final displacers is demonstrated. Guinea pig serum was separated on a medium-resolution adsorbent using a single heterogeneous mixture of carboxymethyldextran displacers to space the protein components. Mouse liver cytosol was separated on a low-resolution adsorbent using six carboxymethyldextran spacer displacers of increasing column affinity. The demonstration of the purification of alkaline phosphatase from E. coli periplasm by displacement chromatography on a high-performance liquid chromatography column is reviewed. The benefits of spacer displacers for separating minor components from complex biological protein mixtures is discussed. A simplified method for preparing carboxymethyldextran displacers is presented.     

Characterization of ion-exchange membrane materials: properties vs structure

This review focuses on the preparation, structure and applications of ion-exchange membranes formed from various materials and exhibiting various functions (electrodialytic, perfluorinated sulphocation-exchange and novel laboratory-tested membranes). A number of experimental techniques for measuring electrotransport properties as well as the general procedure for membrane testing are also described. The review emphasizes the relationships between membrane structures, physical and chemical properties and mechanisms of electrochemical processes that occur in charged membrane materials. The water content in membranes is considered to be a key factor in the ion and water transfer and in polarization processes in electromembrane systems. We suggest the theoretical approach, which makes it possible to model and characterize the electrochemical properties of heterogeneous membranes using several transport-structural parameters. These parameters are extracted from the experimental dependences of specific electroconductivity and diffusion permeability on concentration. The review covers the most significant experimental and theoretical research on ion-exchange membranes that have been carried out in the Membrane Materials Laboratory of the Kuban State University. These results have been discussed at the conferences "Membrane Electrochemistry", Krasnodar, Russia for many years and were published mainly in Russian scientific sources.     

Molecular dynamics simulations of polyelectrolyte adsorption

We have performed molecular dynamics simulations of polyelectrolyte adsorption at oppositely charged surfaces from dilute polyelectrolyte solutions. In our simulations, polyelectrolytes were modeled by chains of charged Lennard-Jones particles with explicit counterions. We have studied the effects of the surface charge density, surface charge distribution, solvent quality for the polymer backbone, strength of the short-range interactions between polymers and substrates on the polymer surface coverage, and the thickness of the adsorbed layer. The polymer surface coverage monotonically increases with increasing surface charge density for almost all studied systems except for the system of hydrophilic polyelectrolytes adsorbing at hydrophilic surfaces. In this case the polymer surface coverage saturates at high surface charge densities. This is due to additional monomer-monomer repulsion between adsorbed polymer chains, which becomes important in dense polymeric layers. These interactions also preclude surface overcharging by hydrophilic polyelectrolytes at high surface charge densities. The thickness of the adsorbed layer shows monotonic dependence on the surface charge density for the systems of hydrophobic polyelectrolytes for both hydrophobic and hydrophilic surfaces. Thickness is a decreasing function of the surface charge density in the case of hydrophilic surfaces while it increases with the surface charge density for hydrophobic substrates. Qualitatively different behavior is observed for the thickness of the adsorbed layer of hydrophilic polyelectrolytes at hydrophilic surfaces. In this case, thickness first decreases with increasing surface charge density, then it begins to increase.     

Steady-state and transient electrical properties of ion-exchange membrane systems in asymmetric arrangements

The steady-state and transient electrical properties of ion-exchange membranes placed between two solutions with different values of the electrolyte concentration, have been simulated using the network simulation method. The ionic transport processes are theoretically described on the basis of the Nernst-Planck and Poisson equations. The system under study is constituted by a cation-exchange membrane in which the fixed-charge is homogeneously distributed in space and two diffusion boundary layers on both sides of the membrane, the electric double layers at the interfaces being included. The steady-state voltage-current characteristic and the profiles of the ionic concentrations and the electric potential, are analysed. Also, the choronopotentiometric response of the system has been discussed and the time evolution of the electric energy consumption evaluated. In particular, the influence of the ratio of the bathing concentrations on the permselectivity and the chronopotentiometric response of the ion-exchange membrane systems, has been established.     

Brownian dynamics simulations of polyelectrolyte adsorption in shear flow

Brownian dynamics simulations are used to study the adsorption of an isolated polyelectrolyte molecule onto an oppositely charged flat surface in the absence and the presence of an imposed shear flow. The polyelectrolyte is modeled as a freely jointed bead-rod chain where excluded volume interactions are incorporated by using a hard-sphere potential. The total charge along the backbone is distributed uniformly among all the beads, and the beads are allowed to interact with one another and the charged surface through screened Coulombic interactions. The simulations are performed by placing the molecule a fixed distance above the surface, and the adsorption behavior is then studied as a function of screening length. In the absence of an imposed flow, the chain is found to lie flat and extended on the adsorbing surface in the limit of weak screening, whereas in the limit of strong screening it desorbs from the surface and attains free-solution behavior. For intermediate screening, only a small portion of the chain adsorbs and it becomes highly extended in the direction normal to the surface. An imposed shear flow tends to orient the chain in the direction of flow and also leads to increased contact of the chain with the surface.     

Preparation and adsorption of refined polyelectrolyte complex nanoparticles

We report on bulk and surface properties of centrifuged nonstoichiometric polyelectrolyte complex (PEC) dispersions. PECs were prepared by mixing poly(diallyldimethylammonium chloride) (PDADMAC) and sodium poly(maleic acid-co-alpha-methylstyrene) (PMA-MS) at the monomolar mixing ratio of 0.6 and polymer concentration >/=1 mmol/l. Centrifugation of initial PEC dispersions revealed three phases: supernatant (SUP), coacervate (COAC), and an insoluble precipitate. Mass, turbidity, particle hydrodynamic radii (R(h)), and the titratable charge amount were determined for those phases. The turbid COAC phase consisted of 200-nm nanoparticles and carried 60% of the polymer mass and 20% of the titratable charge amount of the initial PEC dispersion. The SUP phase showed no turbidity and no such nanoparticles, but carried 80% of the initial titratable charge amount, presumably caused by excess polycations. Furthermore, linear dependences of turbidity and R(h) on COAC concentration was observed. COAC adsorption was studied at polyelectrolyte multilayer (PEM) modified silicon surfaces in dependence on both adsorption time and concentration using attenuated total-reflection-Fourier transform infrared (ATR-FTIR) spectroscopy. The adsorption data were fitted by the simple Langmuir model. Comparison of COAC particles and polystyrene latices revealed similar adsorption features. SEM and AFM measurements resulted in hemispherically shaped adsorbed COAC particles with coverages >/=25%, whose calculated volumes correlated well with those in dispersion obtained by PCS.     

Molecular recognition phenomena in polyelectrolyte systems

The term “molecular recognition” is commonly used to describe various specific interactions, sometimes being not well defined. Let's assume that some species A can separately interact with either species B₁, or B₂, (or B₃, B₄ etc.) forming A·B₁, or A·B₂, (or A·B₃, or A·B₄ etc.) complexes. Here we say “recognition” assuming selective complexation of A with B₁ in the mixture containing also B₂ (B₃, B₄… etc.). At the same time it means discrimination of all other B species except B₁.     

Binary adsorption of globular proteins on ion-exchange media

Adsorption equilibrium of binary pairs of lysozyme (LYS), cytochrome c (CYC) and ribonuclease A (RNase) has been measured on different cation-exchange media at various solution conditions. Adsorption patterns largely follow the intrinsic protein–surface interactions, but can differ significantly for different pairs or even for one pair at different solution conditions. LYS/CYC adsorption shows similar behavior on all the adsorbents examined, with competitive adsorption dominated by LYS and the presence of LYS reducing the adsorption of CYC significantly. Simultaneous and sequential measurements for LYS/CYC show that the order of adsorption does not have a significant effect on the adsorption equilibrium. For LYS/RNase, LYS is consistently more strongly adsorbed. For CYC/RNase, both proteins can display significant adsorption, depending on the pH and salt concentration. A model based on colloidal energetics is developed to calculate the binary adsorption isotherms using parameter values obtained from single-component isotherms. The calculated adsorption is in good agreement with experimental results, with significantly better representation than for other commonly used binary isotherms.     

Characterization of BSA-fouling of ion-exchange membrane systems using a subtraction technique for lumped data

Electrical impedance spectroscopy (EIS) techniques were used to gain insight into BSA fouling of the Neosepta CMX and AMX ion-exchange membranes (IEMs). EIS characterizations were made at concentrations above 0.1 M KCl because the conductance of the IEMs was higher than that of bulk solutions of concentration below 0.1 M KCl. Spectra, expressed in terms of dispersions of the conductance and capacitance with frequency, provided an enhanced indication of IEM fouling during separation processes. Bulk conductance measurements of the solution alone, membrane immersed in solution and fouled membrane immersed in solution showed good agreement with general theoretical predictions. Strong dispersions in capacitance were observed below 1 kHz for each of these configurations. Differences in the dispersions arising from fouling were identified by subtracting the impedance of the solution from those of unfouled and fouled IEMs in solution. The conductance and capacitance dispersions of fouled IEMs decreased with the accumulation of the BSA fouling layer on the surface.     

Self-assembly in ternary systems: cross-linked polyelectrolyte,linear polyelectrolyte and surfactant

The competitive interactions in ternary systems consisting of a slightly cross-linked polyelectrolyte hydrogel and the mixture of linear polyelectrolyte and micelle forming surfactant both oppositely charged relative to the polyelectrolyte network were studied. It was shown that the equilibrium in the competitive reactions depends on the linear polyion charge density and the length of the surfactant aliphatic radical. Dependency on these characteristics the interpolyelectrolyte complex formed by cross-linked and linear polyelectrolytes can uptake surfactant ions from water solution transforming into the cross-linked polyelectrolyte-surfactant complex and releasing the linear polyelectrolyte or vice versa. The ternary systems of this kind are perspective to design the novel family of delivery constructs.     

Colligative properties of polyelectrolyte solutions

The solution properties of polyelectrolytes are not well understood despite increasing theoretical and experimental effort, particularly during the last 10 years. Some of the current models for polyelectrolyte solutions are valid only at infinite dilution because they account only for long-range interactions. Nagvekar and Danner [M. Nagvekar, R.P. Danner, An Excess Gibbs Free Energy Model for Polyelectrolyte Solutions, Fluid Phase Equilibria 53 (1989) 219.] have developed an excess Gibbs free energy model for polyelectrolyte solutions. The key feature of the model is to express the excess Gibbs free energy as the sum of the contributions from long-range and short-range interactions. The `limiting laws' of Manning [G.S. Manning, Limiting Laws and Counterion Condensation in Polyelectrolyte Solutions. I. Colligative Properties, J. Chem. Phys. 51 (1969a) 924; G.S. Manning, Limiting Laws and Counterion Condensation in Polyelectrolyte Solutions. II. Self-diffusion of Small Ions, J. Chem. Phys. 51 (1969b) 934.] were used to account for the long-range interactions while a local composition model of the Non-Random Two Liquid (NRTL) type was used for short-range interactions. In this work, we present the expressions for counterion activity coefficients and the motic coefficients in polyelectrolyte solutions with and without added salts at finite concentrations. Data for a number of polyelectrolyte systems with univalent and divalent counterions are analyzed using Nagvekar's proposed model. In the low concentration regime, the osmotic coefficient data are practically independent of concentration, and are in reasonable agreement with the Manning model. With increasing polyelectrolyte concentration, the osmotic coefficient data are a strong function of concentration, and the local composition model is shown to successfully represent the data.     

Dual-beam polarization interferometry resolves mechanistic aspects of polyelectrolyte adsorption

The electrostatically driven binding dynamics of a polyelectrolyte multilayer (PEMU) film was investigated in real-time using dual-beam polarization interferometry (DPI) and independently supported by quartz crystal microbalance with dissipation monitoring (QCM-D) studies. Multilayer assemblies of the polyanions poly[1-[4[(3-carboxy-4-hydroxyphenylazo)benzenesulfonamido]-1,2-ethanediyl sodium salt] (PAZO) and poly(styrene sulfonate) (PSS) were respectively constructed with the polycation poly(ethylenimine) (PEI) on anionic functionalized substrates using the layer-by-layer electrostatic self-assembly method. DPI measurements indicate that polyelectrolyte adsorption occurs in three distinct stages. In the first stage, for approximately 5 s, coil-like segments of polyanion partially tether to the surface of the oppositely charged PEI. In the second stage, these coils unfurl over a period of approximately 10 s to cover the surface resulting in an increase in average density of the film. During the final adsorption step, the surface-bound polyelectrolyte diffuses into the multilayer assembly, exposing the surface to further deposition. This last step occurs over a much longer time period and results in a highly interpenetrated film containing a charge-overcompensated region at the film surface.     

Kinetic aspects of polymer and polyelectrolyte adsorption on surfaces

The behavior of polymers adsorbed on a single surface or bridging two surfaces is fairly well understood when thermodynamic equilibrium conditions prevail. The work of Nobel prize winner, P.G. de Gennes, to whom the papers in this issue of Advances in Colloid and Interface Science are dedicated, has greatly contributed to this field. Useful as this knowledge is, in practise many situations are encountered in which polymer adsorption is not thermodynamically but kinetically controlled. In this paper we discuss various aspects of non-equilibrium polymer and polyelectrolyte adsorption, based on recent experimental findings. The discussion of these aspects is mainly qualitative, since we are still far from a detailed quantitative theory of the kinetics of polymer adsorption. Many of the examples are relevant to the chemistry of papermaking.     

Electroconvection in systems with heterogeneous ion-exchange membranes

The results of studying the surface morphology of heterogeneous cation-(MK-40) and anion-exchange (MA-40) membranes and calculating the structure of electroconvective vortices generated by the electric body force are shown. The body force and its distribution are estimated by taking into account real parameters of the membrane surface morphology. The calculations of vortices were carried out by solving the Navier-Stokes equation with the no-slip boundary condition and the preset body force distribution. It is shown that the body force induced by the flowing current can generate pairs of electroconvective vortices (electroosmosis of the second kind), where the size of induced vortices is comparable with the intermembrane gap in electrodialysis cells.     

Modification of a supported lipid bilayer by polyelectrolyte adsorption

Addition of a weak polyelectrolyte, poly(methacrylic acid) (PMA), to a supported phospholipid bilayer made from 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) depresses the melting temperature and alters the morphology of the bilayer in the gel phase. Ellipsometry measurements show that PMA adsorption lowers the phase transition temperature by 2.4 degrees C. Atomic force microscopy (AFM) showed no visible contrast in the fluid phase (above the melting temperature) but a rich morphology in the gel phase. In the gel phase, adsorption leads to formation of significantly less mobile phospholipid islands and other defects. One consequence of this lower mobility is a decrease in the implied cooperativity number of the phase transition, N, when polymer is added. Additionally, AFM images of the gel-phase bilayer show a highly defected structure that anneals significantly more slowly than in the absence of adsorbed polymer. Tentatively, we suggest that PMA preferentially decorates island and defect edges of the DMPC bilayer.     

Contact ion-exchange interactions in systems involving zeolites

Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 8, pp. 1505–1506, August, 1994.     

Solubilization of phenols in surfactant/polyelectrolyte systems

The properties of the microheterogeneous systems formed by mixtures of cetyltrimethylammonium bromide (CTAB) and an alternating copolymer of maleic acid and styrene, MAS, and their anionic monoesters, MAS-n with n=2, 4, 6, 8, were investigated. The fluorescence of pyrene was used to sense the polarity of the polymer/CTAB aggregates. Measurements of the ratio III/I in pyrene fluorescence spectra indicate that the polymer/CTAB aggregates are more hydrophobic than normal micelles. A series of p-alkyl substituted phenols were employed to probe the solubilization ability of these aggregates. The distribution constant K(S) of phenol, p-methylphenol, p-ethylphenol, and p-propylphenol between water and MAS-n/CTAB aggregates and the corresponding free energy of transfer Deltamicro(0)(t) have been determined using the pseudo-phase model. The results show that the distribution is mainly determined by the phenol structure, and a linear free energy relationship has been found between Deltamicro(0)(t) and the structure of phenols. On the other hand, an increase in the number of methylene groups in the side alkyl chain has no effect on Deltamicro(0)(t). The results are discussed and compared with those obtained for ionic micelles.