Improving selectivity in multimodal chromatography using controlled pH gradient elution

Holstein, Melissa; Nikfetrat, Amir A.M.; Gage, Mark J.; Hirsh, Allen G.; Cramer, Steven M.

doi:10.1016/j.chroma.2012.01.074

Cited by 43 publications

(22 citation statements)

References 30 publications

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“…As mentioned earlier, the results for the ubiquitin–guanidine simulations have been reported in a previous publication and are reproduced here to facilitate comparisons. The density profile of the guanidine is presented in Figure a, whereas the distributions of the guanidinium and the zwitterionic moieties of arginine are shown in Figure b and c, respectively. Red indicates regions of high density ( ρ / ρ 0 ∼ 20) and blue indicates regions of low density ( ρ / ρ 0 ∼ 1).…”

Section: Resultsmentioning

confidence: 99%

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

Parimal

Garde

Cramer

2017

Biotechnology Progress

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The addition of fluid phase modifiers provides significant opportunities for increasing the selectivity of multimodal chromatography. In order to optimize this selectivity, it is important to understand the fundamental interactions between proteins and these modifiers. To this end, molecular dynamics (MD) simulations were first performed to study the interactions of guanidine and arginine with three proteins. The simulation results showed that both guanidine and arginine interacted primarily with the negatively charged regions on the proteins and that these regions could be readily predicted using electrostatic potential maps. Protein surface characterization was then carried out using computationally efficient coarse-grained techniques for a broader set of proteins which exhibited interesting chromatographic retention behavior upon the addition of these modifiers. It was shown that proteins exhibiting an increased retention in the presence of guanidine possessed hydrophobic regions adjacent to negatively charged regions on their surfaces. In contrast, proteins which exhibited a decreased binding in the presence of guanidine did not have hydrophobic regions adjacent to negatively charged patches. These results indicated that the effect of guanidine could be described as a combination of competitive binding, charge neutralization and increased hydrophobic interactions for certain proteins. In contrast, arginine resulted in a significant decrease in protein retention times primarily due to competition for the resin and steric effects, with minimal accompanying increase in hydrophobic interactions. The approach presented in this paper which employs MD simulations to guide the application of coarse-grained approaches is expected to be extremely useful for methods development in downstream bioprocesses. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:435-447, 2017.

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Section: Resultsmentioning

confidence: 99%

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

Parimal

Garde

Cramer

2017

Biotechnology Progress

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“…While it is possible to perform elution from multimodal exchange using a pH gradient , the study reported here focused only on a bind‐and‐elute strategy at constant pH modulated by NaCl salt concentration. Compared to traditional cation exchange resins , the surface responses shown here (Figs.…”

Section: Resultsmentioning

confidence: 99%

“…Multimodal chromatography ligands can provide unique separation capabilities by coupling different chromatography modalities, typically ion exchange (IEX) and hydrophobic interaction chromatography (HIC). The competing orthogonal chromatography modes can lead to unique separation capabilities with nontraditional strategies, salts, and mobile phase modifiers . For mAb bioprocessing, multimodal resins have also demonstrated unique separation capabilities for mAb monomer and aggregates .…”

Section: Introductionmentioning

confidence: 99%

High‐throughput techniques to evaluate the effect of ligand density for impurity separations with multimodal cation exchange resins

Welsh

Bao

Barlow

et al. 2015

Engineering in Life Sciences

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High-throughput techniques to evaluate the effect of ligand density for impurity separations with multimodal cation exchange resins Scale-down, high-throughput screening techniques are well on their way to becoming a commodity in downstream bioprocess development, especially for the rapid development of chromatography process steps. This work used both resin slurry plate and miniature column high-throughput screening methodologies to identify the best resin properties for mAb separations utilizing a multimodal chromatography ligand interaction. A ligand with both cation exchange and hydrophobic interaction properties was studied at several ligand densities and compared to a commercially available multimodal resin with a larger particle size at high ligand density. The resins were screened with mAbs containing distinct process impurities (aggregates and a hydrophobic variant), and optimized conditions provided more than a log of clearance of both types of impurities for the different resins screened. These studies reveal that while a smaller particle size is generally preferable, optimal ligand densities can be different depending on the properties of both the mAb and impurity studied.

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“…Separation of proteins is achieved by the changing pH values of the mobile phase [12][13][14][15][16][17]. By increasing or decreasing the pH value, bound proteins lose their net charges and desorb from the stationary phase when the pH of the mobile phase approaches their respective pIs [12][13][14][15][16][17].…”

Section: Introductionmentioning

confidence: 99%

Modeling of dual gradient elution in ion exchange and mixed-mode chromatography

Lee

Schmidt

Graalfs

et al. 2015

Journal of Chromatography A

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Improving selectivity in multimodal chromatography using controlled pH gradient elution

Cited by 43 publications

References 30 publications

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

High‐throughput techniques to evaluate the effect of ligand density for impurity separations with multimodal cation exchange resins

Modeling of dual gradient elution in ion exchange and mixed-mode chromatography

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