Competitive ion-exchange adsorption of proteins: Competitive isotherms with controlled competitor concentration

Cano, Tony; Offringa, Natalie D.; Willson, Richard C.

doi:10.1016/j.chroma.2005.03.120

Cited by 19 publications

(15 citation statements)

References 30 publications

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“…Some authors have reported that the Langmuir isotherm is unable to describe the adsorption of proteins onto ion exchangers, mainly due to the presence of competing proteins [8]. However, the results presented here demonstrated that, for phycocyanin, the Langmuir model explained the protein adsorption onto the ion exchanger well, in agreement with some other authors, who also used the Langmuir equation to fit their data [3,19].…”

Section: Adsorption Isothermsupporting

confidence: 91%

Separation of phycocyanin from Spirulina platensis using ion exchange chromatography

Silveira

Quines

Burkert

et al. 2008

Bioprocess Biosyst Eng

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This paper presents the evaluation of some important parameters for the purification of phycocyanin using ion exchange chromatography. The influences of pH and temperature on the equilibrium partition coefficient were investigated to establish the best conditions for phycocyanin adsorption. The equilibrium isotherm for the phycocyanin-resin system was also determined. The separation of phycocyanin using the Q-Sepharose ion exchange resin was evaluated in terms of the pH and elution volume that improved the increase in purity and recovery. The highest partition coefficients were obtained in the pH range from 7.5 to 8.0 at 25 degrees C. Under these conditions the equilibrium isotherm for phycocyanin adsorption was well described by the Langmuir model, attaining a Q (m) of 22.7 mg/mL and K (d) of 3.1 x 10(-2) mg/mL. The best conditions for phycocyanin purification using the ion exchange column were at pH 7.5 with an elution volume of 36 mL, obtaining 77.3% recovery and a 3.4-fold increase in purity.

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Section: Adsorption Isothermsupporting

confidence: 91%

Separation of phycocyanin from Spirulina platensis using ion exchange chromatography

Silveira

Quines

Burkert

et al. 2008

Bioprocess Biosyst Eng

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“…On increasing the salt concentration, the value of n H shifted closer to unity, indicating a lower degree of heterogeneity, and suggesting the participation of a reduced set of more homogeneous functional adsorbent sites. Moreover, in what we believe were the first true competitive protein adsorption isotherms (in the strict sense of “isotherm”, with all intensive variables held constant including competitor concentration), at higher competitor concentrations the heterogeneity of the anion-exchange adsorption of a test protein was decreased [7]. We believe this reflects the existence of a heterogeneous population of sites with different affinities, on which different proteins compete with varying success.…”

Section: Resultsmentioning

confidence: 98%

High ionic strength narrows the population of sites participating in protein ion-exchange adsorption: A single-molecule study

Kisley

Chen

Mansur

et al. 2014

Journal of Chromatography A

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The retention and elution of proteins in ion-exchange chromatography is routinely controlled by adjusting the mobile phase salt concentration. It has repeatedly been observed, as judged from adsorption isotherms, that the apparent heterogeneity of adsorption is lower at more-eluting, higher ionic strength. Here, we present an investigation into the mechanism of this phenomenon using a single-molecule, super-resolution imaging technique called motion-blur Points Accumulation for Imaging in Nanoscale Topography (mbPAINT). We observed that the number of functional adsorption sites was smaller at high ionic strength and that these sites had reduced desorption kinetic heterogeneity, and thus narrower predicted elution profiles, for the anion-exchange adsorption of α-lactalbumin on an agarose-supported, clustered-charge ligand stationary phase. Explanations for the narrowing of the functional population such as inter-protein interactions and protein or support structural changes were investigated through kinetic analysis, circular dichroism spectroscopy, and microscopy of agarose microbeads, respectively. The results suggest the reduction of heterogeneity is due to both electrostatic screening between the protein and ligand and tuning the steric availability within the agarose support. Overall, we have shown that single molecule spectroscopy can aid in understanding the influence of ionic strength on the population of functional adsorbent sites participating in the ion-exchange chromatographic separation of proteins.

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“…Since ion‐exchange adsorption of multicomponent proteins began to be concerned in 1980s , dozens of literature has accumulated on the subject of adsorption equilibria of multiple component proteins . There are many factors influencing ion‐exchange adsorption of multicomponent proteins.…”

Section: Introductionmentioning

confidence: 99%

Single and binary adsorption of proteins on ion‐exchange adsorbent: The effectiveness of isothermal models

Liang

Fieg

Shi

et al. 2012

J of Separation Science

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Simultaneous and sequential adsorption equilibria of single and binary adsorption of bovine serum albumin and bovine hemoglobin on Q Sepharose FF were investigated in different buffer constituents and initial conditions. The results in simultaneous adsorption showed that both proteins underwent competitive adsorption onto the adsorbent following greatly by protein-surface interaction. Preferentially adsorbed albumin complied with the universal rule of ion-exchange adsorption whereas buffer had no marked influence on hemoglobin adsorption. Moreover, an increase in initial ratios of proteins was benefit to a growth of adsorption density. In sequential adsorption, hemoglobin had the same adsorption densities as single-component adsorption. It was attributed to the displacement of preadsorbed albumin and multiple layer adsorption of hemoglobin. Three isothermal models (i.e. extended Langmuir, steric mass-action, and statistical thermodynamic (ST) models) were introduced to describe the ion-exchange adsorption of albumin and hemoglobin mixtures. The results suggested that extended Langmuir model gave the lowest deviation in describing preferential adsorption of albumin at a given salt concentration while steric mass-action model could very well describe the salt effect in albumin adsorption. For weaker adsorbed hemoglobin, ST model was the preferred choice. In concert with breakthrough data, the research further revealed the complexity in ion-exchange adsorption of proteins.

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Competitive ion-exchange adsorption of proteins: Competitive isotherms with controlled competitor concentration

Cited by 19 publications

References 30 publications

Separation of phycocyanin from Spirulina platensis using ion exchange chromatography

Separation of phycocyanin from Spirulina platensis using ion exchange chromatography

High ionic strength narrows the population of sites participating in protein ion-exchange adsorption: A single-molecule study

Single and binary adsorption of proteins on ion‐exchange adsorbent: The effectiveness of isothermal models

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