Adsorption equilibrium of fructosyltransferase on a weak anion-exchange resin

The adsorption equilibrium of a glycoprotein, fructosyltransferase from Aureobasidium pullulans, on an anion-exchange resin, Sepabeads FP-DA activated with 0.1M NaOH, was investigated. The adsorption isotherms were determined at 20 degrees C in a phosphate-citrate buffer with pH 6.0 using the static method. Sodium chloride was used to adjust the ionic strength in the range from 0.0215 to 0.1215 mol dm(-3) which provided conditions varying from a weak effect of salt concentration on protein binding to its strong suppression. The equilibrium data were very well fitted by means of the steric mass-action model when the ion-exchange capacity of 290 mmol dm(-3) was obtained from independent frontal column experiments. The model fit provided the protein characteristic charge equal to 1.9, equilibrium constant 0.326, and steric factor 1.095 x 10(5).     

Breakthrough Model of Recombinant Human-Like Collagen in Immobilized Metal Affinity Chromatography

The adsorption of recombinant human-like collagen by metal chelate media was investigated in a batch reactor and in a fixed-bed column. The adsorption equilibrium and kinetics had been studied by batch adsorption experiments. Equilibrium parameters and protein diffusivities were estimated by matching the models with the experimental data. Using the parameters of equilibrium and kinetics, various models, such as axial diffusion model, linear driving force model, and constant pattern model, were used to simulate the breakthrough curves on the columns. As a result, the most suitable isotherm was the Langmuir–Freundlich model, and the ionic strength had no effect on the adsorption capacity of chelate media. In addition, the pore diffusion model fitted very well to the kinetic data. The pore diffusivities decreased with increasing the initial protein concentration, however had little change with the ionic strength. The results also indicated that the models predict breakthrough curves reasonably well to the experimental data, especially at low initial protein concentration (0.3 mg ml⁻¹) and low flow rate (34 cm h⁻¹). By the results, we optimized the experimental conditions of a chromatographic process using immobilized metal affinity chromatography to purify recombinant human-like collagen.     

Novel column-based protein refolding strategy using dye-ligand affinity chromatography based on macroporous biomaterial

A novel column-based chromatographic protein refolding strategy was developed using dye-ligand affinity chromatography (DLAC) based on macroporous biomaterial. Chitosan–silica (CS–silica) biomaterial with macroporous surface was used as the supporting matrix for the preparation of the DLAC material. The dye-ligand Cibacron Blue F3GA (CBF) was selected as affinity handle and could be covalently immobilized to form dye-ligand affinity adsorbent (CBF–CS–silica) using the reactivity of NH₂ on CS–silica biomaterial. After the model protein catalase was denatured with 6 mol/L urea, the denaturant could be rapidly removed and catalase could be successfully refolded as facilitated by the adsorption of CBF–CS–silica. The urea denaturation process and the elute condition for the chromatographic refolding were optimized by measuring tryptophan fluorescence and activity of catalase. The refolding performance of the proposed DLAC was compared with dilution refolding. The protein concentration during the proposed chromatographic refolding increased by a factor of 20 without reducing the yield achieved as compared to dilution refolding. The column-based protein refolding strategy based on dye-ligand affinity chromatography with porous biomaterial being matrix possessed potential in chromatographic refolding of protein.     

Effect of charge regulation on steric mass-action equilibrium for the ion-exchange adsorption of proteins

A thermodynamic formalism is developed for incorporating the effects of charge regulation on the ion-exchange adsorption of proteins under mass-overloaded conditions as described by the steric mass-action (SMA) isotherm. To accomplish this, the pH titration behavior of a protein and the associated adsorption equilibrium of the various charged forms of a protein are incorporated into a model which also accounts for the steric hindrance of salt counterions caused by protein adsorption. For the case where the protein is dilute, the new model reduces to the protein adsorption model described recently by the authors which accounts for charge regulation. Similarly, the new model reduces to the steric mass-action isotherm developed by Brooks and Cramer which applies to mass-overloaded conditions for the case where charge regulation is ignored so that the protein has a fixed charge. Calculations using the new model were found to agree with experimental data for the adsorption of bovine serum albumin (BSA) on an anion-exchange column packing when using reasonable physical properties. The new model was also used to develop an improved theoretical criterion for determining the conditions required for an adsorbed species to displace a protein in displacement chromatography when the pH is near the protein pI.     

Self-consistent field modeling of non-ionic surfactants at the silica-water interface: incorporating molecular detail

We have constructed a model to predict the properties of non-ionic (alkyl-ethylene oxide) (C(n)E(m)) surfactants, both in aqueous solutions and near a silica surface, based upon the self-consistent field theory using the Scheutjens-Fleer discretisation scheme. The system has the pH and the ionic strength as additional control parameters. At high ionic strength, the solvent quality for the surfactant head groups is affected, which changes both the bulk and the adsorption behavior of the surfactant. For example, with increasing ionic strength, the CMC drops and the aggregation increases. Surfactants adsorb above the critical surface association concentration (CSAC). The CSAC is a function of the surfactant and the surface properties. Therefore, the CSAC varies with both the ionic strength and the pH. We predict that with increasing ionic strength, the CSAC will first slightly increase but then drop substantially. The charge on the surface is pH dependent, and as the head groups bind through H-bonding to the silanol groups, the CSAC increases with increasing pH. We focus on adsorption/desorption transitions for the surfactants and compare these to the experimental data. Both the equilibrium predictions and the consequences for the kinetics of adsorption follow experimental findings. Our results show that molecularly realistic models can reveal a much richer interfacial behavior than anticipated from more generic models.     

Characterization of non-linear adsorption properties of dextran-based polyelectrolyte displacers in ion-exchange systems

Experimental studies were carried out on the non-linear adsorption properties of dextran-based polyelectrolytes in anion- and cation-exchange chromatographic systems. By monitoring both the induced salt gradients and sequential breakthrough fronts, parameters were determined for use in a Steric Mass Action (SMA) model of non-linear ion-exchange chromatography. These parameters include: total ion capacity of the columns, characteristic charge, steric factor, equilibrium constant, and maximum adsorptive capacity for each of the polyelectrolytes. In addition the number of functional groups were determined by elemental analysis. The values of the SMA parameters were found to be independent of salt and polyelectrolyte bulk phase compositions. Parameters were also determined for a variety of proteins. Experimental isotherms for the polyelectrolytes and proteins were compared with those simulated by the SMA model. Finally, the implications of polyelectrolyte adsorption properties with respect to their ability to act as efficient displacers in ion-exchange displacement systems are discussed.     

Adsorption Effects Observed in Gel Permeation Chromatography of Thiouracil

Abstract

In gel permeation chromatography (GPC), several compounds deviate from the molecular volume/elution count relationship which is prepared using satured hydrocarbons. In this paper, this problem is investigated in detail using thiouracil in aqueous solution as a model chromatographic adsorbate. The concentration dependences of elution counts and peak heights prove the adsorption of thiouracil on Sephadex G-25 when water is the solvent. Thus to investigate further the mechanisms of adsorption responsable for the chromatographic behaviour, thiouracil-Sephadex interac—tions were investigated by studying equilibrium adsorption. Isotherms of type IV of BDDT classification were found which are typically associated with a weak adsorption such as physisorption, on a porous solid. The effect of water structure perturbants, ionic strength and pH on this adsorption was consistent with the-hypothesis that with water as a solvent both aromatic adsorption and electrostatic interaction are the determinants of the affinity of this gel for a thiouracil compound. This may be particularly useful since results of equilibrium adsorption isotherms are frequently used to develop liquid chromatographic theories.     

Adsorption Studies of Nonionic Surfactants onto Polyvinyltoluene Microlatexes in Aqueous Medium

Negatively charged polyvinyltoluene (PVT) microlatexes of various dimensions were prepared by polymerization of the corresponding vinyltoluene-in-water microemulsion precursors containing sodium dodecyl sulfate and n-pentanol. PVT microlatexes were used as adsorbents for adsorption of nonionic surfactants such as Tween 80, Tween 20, and Triton X-100. The extents of adsorption were known from the differences in the diffusion coefficient of the PVT microlatexes containing a solubilized suitable electro active probe. The surface excess (Gamma), equilibrium constant of adsorption (K(ads)), molecular cross-sectional surface area (sigma), intercalation ratio (kappa), etc., were determined for sodium dodecyl sulfate composition variation and ionic strength effects. A surface reconstruction model was proposed to rationalize the results. Copyright 2000 Academic Press.     

The ionic strength effect on microcystin and natural organic matter surrogate adsorption onto PAC

This work aims to contribute to a better understanding of the ionic strength effect on microcystin and natural organic matter (NOM) surrogate adsorption by analyzing the importance of adsorbate molecular size, and surface concentration. Adsorption kinetics and/or isotherms were performed on PAC Norit SA-UF for four microcystin variants (MC-LR, MC-LY, MC-LW, MC-LF), and three NOM surrogates (salicylic acid (SA), tannic acid (TA), Aldrich humic acid (AHA)) at different solution ionic strengths. Results showed that the ionic strength effect depends upon the adsorbate surface concentration, cation charge (mono or divalent), and adsorbate molecular size. Potassium seemed not to affect the MC-LR adsorption, while calcium enhanced MC-LR kinetics and adsorption capacity. K+ and, particularly, Ca2+ improved the adsorption kinetics of the other microcystin variants. For identical surface concentration and ionic strength, the impact of K+ and Ca2+ on NOM surrogates depended on the adsorbate molecular size: K+ effect was only observed for AHA, whereas Ca2+ caused no effect on SA adsorption, slightly enhanced TA adsorption, and greatly enhanced AHA adsorption. MC-LR isotherms with two salt concentrations (KCl or CaCl2) indicated that, for the studied range of equilibrium surface concentration (5.3-18.7 mg/g), an enhanced adsorption regime prevails, and no transition regime was observed.     

Water vapor molecule adsorption onto 'Ajwa' dates: Analytical investigation via infinite multilayer statistical physics model

A gravimetric technique was used to examine the water activity onto the ‘Ajwa’ dates. The equilibrium adsorption isotherms of water molecules were carried out at three temperatures (between 303 K and 323 K). A theoretical method was developed using statistical physics treatment to describe the experimental data at the ionic scale. The date’s isotherms were analyzed via the infinite multilayer adsorption model (formation of a high number of adsorbed layers) which is established based on the ideal gas law (there are no lateral interactions influences on the adsorption mechanism). The chosen model gave significant interpretation of the adsorption of water on the Ajwa dates based on the physicochemical model’s parameters (the density of binding sites (D_m), the number of water molecules per site (n) and the energetic parameters (a₁) and (a₂)). The physicochemical interpretation of the appropriate model indicated that the adsorption of water on the Ajwa dates occurred via a multi-anchorage process since the n values are lower than 1 for the three tested temperatures. The Ajwa dates adsorption was found typical to an exothermic process by the intermediate of the steric parameter D_m (D_m (303 K) = 0.58 kg/kg? D_m (323 K) = 0.33 kg/kg). Moreover, the energies values |?ε₁| and |?ε₂|, which varied from 27.8 KJ/mol to 51.2 KJ/mol, confirmed that the ‘Ajwa’ dates adsorption was a chemical process presenting covalent bonds between the water molecules and the dates’ sites.     

A NOVEL METAL CHELATE AFFINITY ADSORBENT FOR PROTEIN UPTAKE

1. INTRODUCTION Chitosan is a hydrolyzed derivative of chitin and belongs to a family of linear unbranched polysaccharides which contain large amounts of 1,4-linked-2-amino-2-deoxy-β-D-glucan residues. The presence of free amine groups in chitosan enhances the solubility and reactivity of this polymer. Interest in modifying chitosan by using glutaraldehyde has recently increased. The derivatized polymers have been employed for many applications [1~2], including protein immobilization…     

An in situ FTIR-ATR study of polyacrylate adsorbed onto hematite at high pH and high ionic strength

FTIR-ATR was used to examine in situ the interaction of polyacrylate and hematite at pH 13. Static light scattering and mobility measurements were used to assess solution polyacrylate dimensions and hematite surface charge, respectively. Polyacrylate adsorption occurred only with the addition of electrolyte (e.g., NaCl), and it was found that excess cations, up to approximately 1 M, facilitated adsorption, above which the effect was found to plateau. At pH 13 and at low ionic strength, adsorption of polyacrylate onto hematite is facilitated by cations in solution shielding both the negative acrylate functionality of the polymer and the negative hematite surface. The shielding of the hematite surface continues to increase with increasing salt concentration up to a measured 3 M. Similarly, the shielding of the polymer increased with electrolyte concentration up to approximately 1 M salt, beyond which no further increase in shielding was observed. At this concentration the polymer assumes a finite minimum size in solution that ultimately limits the amount adsorbed. The dimension of the polymer in solution was found to be independent of monovalent cation type. Thus, at high pH and high ionic strength adsorption is determined by the degree of hematite surface charge reduction. The cation-hematite surface interaction was found to be specific, with lithium leading to greater polyacrylate adsorption than sodium, which was followed by cesium. The stronger affinity of lithium for the hematite surface over sodium and cesium is indicative of the inverse lyotropic adsorption series and has been rationalized in the past by the "structure-making-structure-breaking" model. These results provide a useful insight into the likely adsorption mechanism for polyacrylate flocculants at high pH and ionic strength onto residues in the Bayer processing of bauxite.     

Adsorption studies of cationic starch onto quartz surface through an electrode-separated piezoelectric sensor

In the construction of an electrode-separated piezoelectric sensor (ESPS), the quartz surface is in direct contact with the liquid phase. The negatively charged quartz crystal surface can adsorb cationic starch. This adsorption process was in situ monitored from the frequency shift of the ESPS. It was shown that the adsorption of cationic starch onto the quartz surface is reversible with respect the dilution of the bulk phase. The adsorption behavior can be described by Langmuir model. The adsorption density and kinetics parameters were estimated from the frequency responses of the ESPS. The influence of pH and ionic strength on adsorption parameters was investigated. It was shown that the influence of pH on the adsorption rate was slight. With increasing ionic strength, the rate constants for adsorption and desorption increase, but the adsorption equilibrium constant and saturation adsorption density decrease. The adsorption equilibrium constant and adsorption density reach a maximum in buffer of pH 10.     

A NOVEL METAL CHELATE AFFINITY ADSORBENT FOR PROTEIN UPTAKE

In this article,a spherical chitosan gel crosslinked by epichlorohydrin was prepared.It was then loaded with copper ions to produce a metal chelate affinity adsorbent for protein.The uptake of bovine serum albumin(BSA)by the affinity adsorbent was investigated.and the adsorption capacity for BSA as high as 40mg/g-wet beads was observed.The adsorption equilibrium data was well correlated by the Langmuir equation.The adsorption was considerably affected by pH.In additio.The amount of BSA adsorbed onto the beads decreased with the increasing of aqueous phase ionic strength,so adsorbed BAS can be desorbed by adjusting pH orionic strength of the solution.     

Modeling of the salt effects on hydrophobic adsorption equilibrium of protein

A two-state protein model is proposed to describe the salt effects on protein adsorption equilibrium on hydrophobic media. This model assumes that protein molecules exist in two equilibrium states in a salt solution, that is, hydrated and dehydrated states, and only the dehydrated-state protein can bind to hydrophobic ligands. In terms of the two-state protein hypothesis and the steric mass-action theory, protein adsorption equilibrium on hydrophobic media is formulated by a five-parameter equation. The model is demonstrated with the adsorption of bovine serum albumin to Phenyl Sepharose gels as a model system. The effects of salt type (sodium chloride, sodium sulfate and ammonium sulfate) on the model parameters are discussed. Then, the model formulism is simplified in terms of the small magnitude of the protein dehydration equilibrium constant in the model. This simplification has returned the model derived on the basis of the two-state protein hypothesis to its original mechanism of salt effects on the hydrophobic adsorption of protein. This simplified model also creates satisfactory prediction of protein adsorption isotherms.     

Polydisperse adsorbability composition of several natural and synthetic organic matrices

The polydisperse composition of nine dissolved organic materials (DOMs) from two river water sources, one ground water source, two biologically treated wastewater sources, and two commercial sources was analyzed based on their adsorbabilities by activated carbon. For each DOM, batch adsorption isotherms measured for both TOC and UV260 were analyzed using an overall isotherm model derived from the IAST-Freundlich expression. By accounting for the heterogeneity of each DOM with a log-normal distribution of the Freundlich parameter (K), its adsorption behavior was characterized with only four parameters (including three fitting ones). The average adsorptive strength (KM) and heterogeneity (sigma) determined for all DOMs, which were defined by the mean value and the standard deviation of the log-normal distribution of the Freundlich K, changed over the ranges 2.5-62.2 and 0.22-0.97 (mg/g)/(mg/l)(1/n), respectively, when the TOC index was used. Among all DOMs studied, a river water DOM at the upper stream was found least heterogeneous: the Freundlich K of its organic constituents varied in the range 10.8-190 (mg/g)/(mg/l)(1/n), as compared to a commercial humic acid that exhibited the broadest Freundlich K distribution of 0.01-1494.3 (mg/g)/(mg/l)(1/n). KM and sigma, along with other two parameters (the Freundlich exponent 1/n and the nonadsorbable organic fraction parameter Cnon/CT0), changed with both indices of TOC and UV260 in a regular manner, indicating that UV-absorbing organic molecules possessed adsorbabilities different from non-UV-absorbing ones. Also based on HPSEC chromatograms measured for solutions before adsorption, the molecular weight composition of all DOMs was also assessed and the molecular size impacts on adsorption characteristics of DOMs were briefly discussed.     

Relationship between interfacial forces measured by colloid-probe atomic force microscopy and protein resistance of poly(ethylene glycol)-grafted poly(L-lysine) adlayers on niobia surfaces

Adsorbed layers of "comb-type" copolymers consisting of PEG chains grafted onto a poly(l-lysine) (PLL) backbone on niobium oxide substrates were studied by colloid-probe AFM in order to characterize the interfacial forces associated with coatings of varying architectures (PEG/PLL ratios and PEG chain lengths) and their relevance to protein resistance. The steric and electrostatic forces measured varied substantially with the architecture of the PLL-g-PEG copolymers. Varying the ionic strength of the buffer solutions enabled discrimination between electrostatic and steric-entropic contributions to the net interfacial force. For high PEG grafting densities the steric component was most prominent, but at low ionic strengths and high grafting densities, a repulsive electrostatic surface force was also observed; its origin was assigned to the niobia charges beneath the copolymer, as insufficient protonated amine groups in the PLL backbone were available for compensation of the oxide surface charges. For lower grafting densities and lower ionic strengths there was a substantial attractive electrostatic contribution arising from interaction of the electrical double layer arising from the protonated amine groups, with that of the silica probe surface (as under low ionic strength conditions, the electrical double layer was thicker than the PEG layer). For these PLL-g-PEG coatings the net interfacial force can thus be a markedly varying superposition of electrostatic and steric-entropic contributions, depending on various factors. The force curves correlate with protein adsorption data, demonstrating the utility of AFM colloid-probe force measurements for quantitative analysis of surface forces and how they determine interfacial interactions with proteins. Such characterization of the net interfacial forces is essential to elucidate the multiple types of interfacial forces relevant to the interactions between PLL-g-PEG coatings and proteins and to advance interpretation of protein adsorption or repellence beyond the oversimplified steric barrier model; in particular, our data demonstrate the importance of an ionic-strength-dependent minimum PEG layer thickness to screen the electrostatic interactions of charged interfaces.     

批量法研究牛γ-球蛋白在尼龙亲和膜上的等温吸附行为

以尼龙膜为基质,L－色氨酸（Trp)为配基,合成了一种新的亲和介质用以吸附牛γ-球蛋白（BGG）。用批量法系统考察了温度、离子强度和pH以亲和等温吸附的影响。研究结果表明,BCG与氨基酸之间的亲和相互作用力主要是静电力和疏水相互作用力。在最适条件下吸附遵循Langmuir型吸附,并且亲和吸附量最大而非特异性吸附最小。偏离该条件则会发生在蛋白质在膜上的堆积,蛋白质构型变化及蛋白质与配基间的空间取向的变化,从而使吸附不再遵循Langmuir型吸附。     

Effects of carboxyl groups on the adsorption behavior of low-molecular-weight substances on a stainless steel surface

The adsorption isotherms of various carboxylic acids and several amines on a stainless steel surface were taken as a function of pH and the ionic strength of the solution at 30 degrees C. In particular, the effect of the number of carboxyl groups on the adsorption behavior was investigated. Monocarboxylic acids such as benzoic acid and n-butyric acid were reversibly adsorbed on the stainless steel particles and showed a Langmuir-type adsorption isotherm, i.e., Q=KqmC/(1+KC), where Q and C are, respectively, the amount of adsorbate adsorbed and the equilibrium concentration in the bulk solution, qm, the maximum adsorbed amount, and K is the adsorption equilibrium constant. Carboxylic acids having plural carboxyl groups had much higher affinity to the surface and were adsorbed in both reversible and irreversible modes. The adsorption isotherms for the carboxylic acids having plural carboxyl groups could be expressed by a modified Langmuir-type adsorption isotherm, i.e., Q=q(irrev)+Kq(rev)C/(1+KC), where q(irrev) and q(rev) are, respectively, the maximum amounts adsorbed irreversibly and reversibly. The K and q(irrev) values increased with an increase in the number of carboxyl groups except for isophthalic acid and terephthalic acid. On the basis of the pH dependencies of K, qm, q(irrev), and q(rev) as well as the surface properties of the stainless steel, both reversible and irreversible adsorptions were considered to occur through the electrostatic interaction between negatively charged carboxyl groups and the positively charged sites on the surface. The dependency of the q(irrev) value on ionic strength was discussed on the basis of the differences in their adsorbed state with the interaction forces to the surface and repulsive forces among the adsorbed molecules. The adsorption of amine components was quite weak. The RA-IR and molecular dynamics calculation were done to investigate the adsorption states of phthalic acid, trimellitic acid, and mellitic acid.     

Polyelectrolyte-mediated protein adsorption: fluorescent protein binding to individual polyelectrolyte nanospheres

We have used confocal fluorescence microscopy with single molecule sensitivity to characterize uptake and release of fluorescent protein (mEosFP) molecules by individual spherical polyelectrolyte brush (SPB) nanoparticles that were immobilized on a glass surface. The SPB particles consisted of a solid core particle of 100 nm diameter onto which long polyelectrolyte chains were affixed. They could be loaded with up to 30 000 mEosFP molecules in a solvent of low ionic strength. The concentration dependence of protein loading can be described with a simple bimolecular binding model, characterized by an equilibrium dissociation coefficient of 0.5 microM. Essentially complete release of the bound proteins was observed after increasing the ionic strength by adding 250 mM NaCl to the solvent. Fluorescence emission spectra and time-resolved fluorescence intensity decays were measured on individual, mEosFP-loaded SPB nanoparticles, and also on the dissolved mEosFP before and after adsorption. These results indicate that the mEosFP molecules remained structurally intact in this procedure. Hence, the present investigation demonstrates unambiguously that polyelectrolyte-mediated protein adsorption onto SPB particles presents a viable process for protein immobilization.