Two variables dominating the retention of intact proteins under gradient elution with simultaneous ultrafast high-resolution separation by hydrophobic interaction chromatography

The Analyst ◽  
2015 ◽  
Vol 140 (19) ◽  
pp. 6692-6704 ◽  
Author(s):  
Xindu Geng ◽  
Xiaodan Jia ◽  
Peng Liu ◽  
Fei Wang ◽  
Xiaoming Yang
Keyword(s):  
High Resolution ◽  
Hydrophobic Interaction ◽  
Hydrophobic Interaction Chromatography ◽  
Gradient Elution ◽  
Intact Proteins ◽  
And Migration

The retention of proteins under gradient elution in HIC is dominated by two variables of steady and migration regions.

High resolution of multiple forms of rabbit reticulocyte hexokinase type I by hydrophobic interaction chromatography

1994 ◽  
Vol 676 (1) ◽  
pp. 51-63 ◽  
Author(s):  
Vilberto Stocchi ◽  
Paola Cardoni ◽  
Paola Ceccaroli ◽  
Giovanni Piccoli ◽  
Luigi Cucchiarini ◽  
...  
Keyword(s):  
High Resolution ◽  
Hydrophobic Interaction ◽  
Hydrophobic Interaction Chromatography ◽  
Type I ◽  
Multiple Forms ◽  
Hexokinase Type I

scholarly journals Prediction of the Elution Profiles of Proteins in Mixed Salt Systems in Hydrophobic Interaction Chromatography

2021 ◽  
Author(s):  
Fabian Jirasek
Keyword(s):  
Kinetic Model ◽  
Hydrophobic Interaction ◽  
Hydrophobic Interaction Chromatography ◽  
Taylor Series Expansion ◽  
Gradient Elution ◽  
Ternary Mixtures ◽  
Mixed Salt ◽  
Cooperative Effects ◽  
Adsorption Equilibria ◽  
Salt Mixtures

Hydrophobic interaction chromatography (HIC) is often used for purifying proteins. A mathematical model to describe the complex effects of salts on the adsorption equilibria in HIC has recently been introduced by our group. It describes not only the influence of single salts, but also salt mixtures, in which cooperative effects may occur. The influence of the salts is thereby modeled with a Taylor series expansion in the individual ion molarities. In the present study, the model of the adsorption equilibrium is coupled with a lumped kinetic model of the adsorption kinetics to obtain a model of the elution of proteins in HIC adsorption columns. The column model is tested using experimental data on the adsorption of bovine serum albumin (BSA) and lysozyme (LYS) on the mildly hydrophobic resin Toyopearl PPG-600M at pH 7. The studied salts are ammonium chloride, sodium chloride, ammonium sulfate, and sodium sulfate as well as binary and ternary mixtures of them. The parameters of the lumped kinetic model are protein-specific and were fitted to the elution profiles of the single proteins in presence of single salts. The model was then used to predict the elution profiles of BSA and LYS solutions containing both proteins, for single salts as well as for binary and ternary salt mixtures. Both isocratic and gradient elution were studied. Furthermore, the model was applied to identify the optimal overall ionic strength for the separation of the two proteins by isocratic elution and the optimal linear gradient of the salt concentration in a multicriteria approach where the conflicting goals are high separation yield and low elution volume.

scholarly journals Prediction of the Elution Profiles of Proteins in Mixed Salt Systems in Hydrophobic Interaction Chromatography

2021 ◽  
Author(s):  
Nadia Galeotti ◽  
Eva Hackemann ◽  
Fabian Jirasek ◽  
Hans Hasse
Keyword(s):  
Kinetic Model ◽  
Hydrophobic Interaction ◽  
Hydrophobic Interaction Chromatography ◽  
Taylor Series Expansion ◽  
Gradient Elution ◽  
Ternary Mixtures ◽  
Mixed Salt ◽  
Cooperative Effects ◽  
Adsorption Equilibria ◽  
Salt Mixtures

Hydrophobic interaction chromatography (HIC) is often used for purifying proteins. A mathematical model to describe the complex effects of salts on the adsorption equilibria in HIC has recently been introduced by our group. It describes not only the influence of single salts, but also salt mixtures, in which cooperative effects may occur. The influence of the salts is thereby modeled with a Taylor series expansion in the individual ion molarities. In the present study, the model of the adsorption equilibrium is coupled with a lumped kinetic model of the adsorption kinetics to obtain a model of the elution of proteins in HIC adsorption columns. The column model is tested using experimental data on the adsorption of bovine serum albumin (BSA) and lysozyme (LYS) on the mildly hydrophobic resin Toyopearl PPG-600M at pH 7. The studied salts are ammonium chloride, sodium chloride, ammonium sulfate, and sodium sulfate as well as binary and ternary mixtures of them. The parameters of the lumped kinetic model are protein-specific and were fitted to the elution profiles of the single proteins in presence of single salts. The model was then used to predict the elution profiles of BSA and LYS solutions containing both proteins, for single salts as well as for binary and ternary salt mixtures. Both isocratic and gradient elution were studied. Furthermore, the model was applied to identify the optimal overall ionic strength for the separation of the two proteins by isocratic elution and the optimal linear gradient of the salt concentration in a multicriteria approach where the conflicting goals are high separation yield and low elution volume.

Purification of Glycerol Kinase by "Dye-Ligand" Chromatography and Hydrophobic Interaction Chromatography on Bead-Cellulose Derivatives

1993 ◽  
Vol 58 (2) ◽  
pp. 445-451 ◽  
Author(s):  
Vladimír Žúbor ◽  
Albert Breier ◽  
Marta Horváthová ◽  
Dagmar Hagarová ◽  
Peter Gemeiner ◽  
...  
Keyword(s):  
Hydrophobic Interaction ◽  
Specific Activity ◽  
Hydrophobic Interaction Chromatography ◽  
Glycerol Kinase ◽  
Enzymic Activity ◽  
Cellulose Derivatives ◽  
Long Term Storage ◽  
Bead Cellulose ◽  
Term Storage

The crude extract of cytosole enzymes was obtained from homogenized cells of Saccharomyces cerevisiae by partition. The enzyme was then isolated from the lower aqueous phase displaying higher glycerol kinase activity by dye-ligand chromatography on Cibacron Blue (CB) or Remazol Brilliant Blue R (RB)-derivatized bead-cellulose, ATP being the eluent. The specific activity of glycerol kinase rised more than 10 and 7-times after affinity dye-ligand chromatography and hydrophobic interaction chromatography, respectively. Glycerol kinase obtained by the latter method was purified by CB-bead cellulose. The final preparation maintained its enzymic activity without noticeable losses during a long-term storage at 4 °C in dark.

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