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Mollerup J.M.,Prepchrom | Breil M.P.,Novo Nordisk AS
AIChE Journal

Solution of a strong electrolyte in a high-density polar fluid gives rise to a dielectric saturation that decreases the orientation polarizability of the solvent molecules in close proximity to the ions wherefore the relative permittivity in this region is determined solely by the atomic and electronic polarization. This causes a substantial decrease in the static permittivity of the solution. By considering the dielectric saturation, a model for the permittivity of an electrolyte solution have been developed and the parameters, the relative permittivities at dielectric saturation in close proximity to the ions, for 17 ions in water at 298.15 K were determined. By scaling these relative permittivities in proportion to the permittivity of the solvent, the model could be extended to calculate the permittivity of solutions of electrolytes in methanol and admixtures of water and ethanol. © 2015 American Institute of Chemical Engineers. Source

Mollerup J.M.,Prepchrom
Journal of Chromatography A

Oligomer forms of proteins are formed by self and cross association or complex formation with ligands. Modelling studies using an ion-exchange adsorbent have demonstrated that the formation of an oligomer form of a target protein can improve a chromatographic separation because the oligomer form displaces the impurities. The results of the investigation show that the process is very robust, that the purity, the yield, and the productivity increase with increasing load and increasing salt concentration in the eluant. An impurity level less than 1. ppm is easy to achieve having a yield of 98%. A universal model for the adsorption equilibria of mono and oligomer forms of proteins on ion-exchange, hydrophobic, and bimodal adsorbents has been developed. © 2011 Elsevier B.V. Source

Johansson K.,Lund University | Frederiksen S.S.,Novo Nordisk AS | Degerman M.,Lund University | Breil M.P.,Novo Nordisk AS | And 2 more authors.
Journal of Chromatography A

The two main chromatographic modes based on hydrophobicity, hydrophobic interaction chromatography (HIC) and reversed-phase chromatography (RPC), are widely used for both analytical and preparative chromatography of proteins in the pharmaceutical industry. Despite the extensive application of these separation methods, and the vast amount of studies performed on HIC and RPC over the decades, the underlying phenomena remain elusive. As part of a systematic study of the influence of mobile phase modulators in hydrophobicity-based chromatography, we have investigated the effects of both KCl and ethanol on the retention of three insulin variants on two HIC adsorbents and two RPC adsorbents. The focus was on the linear adsorption range, separating the modulator effects from the capacity effects, but some complementary experiments at higher load were included to further investigate observed phenomena. The results show that the modulators have the same effect on the two RPC adsorbents in the linear range, indicating that the modulator concentration only affects the activity of the solute in the mobile phase, and not that of the solute-ligand complex, or that of the ligand. Unfortunately, the HIC adsorbents did not show the same behavior. However, the insulin variants displayed a strong tendency toward self-association on both HIC adsorbents; on one in particular. Since this causes peak fronting, the retention is affected, and this could probably explain the lack of congruity. This conclusion was supported by the results from the non-linear range experiments which were indicative of double-layer adsorption on the HIC adsorbents, while the RPC adsorbents gave the anticipated increased tailing at higher load. © 2015 Elsevier B.V. Source

Breil M.P.,Novo Nordisk AS | Mollerup J.M.,Prepchrom
Fluid Phase Equilibria

The scattering of light originates from the fluctuation in the permittivity that gives rise to a turbidity of the fluid. The turbidity of an otherwise homogeneous fluid is, according to Einstein's theory, proportional to the mean-square fluctuation in the relative permittivity. A general expression for the mean-square fluctuation or variance of a fluctuating property is derived. It shows that the turbidity depends on the second derivatives of an appropriately chosen state function - the choice is dependent on which properties that fluctuate. The paper provides a general derivation and outlines the criteria for the choice of state function.Data for light-scattering are modelled using the Clausius-Mossotti equation and an excess Gibbs energy function. The excess Gibbs energy applied consists of three terms: Flory-Huggins (accounting for the size of the components), Debye-Hückel (accounting for the charges of electrolytes) and Kirkwood (accounting for the dipolar nature of proteins). The present investigation shows that the model parameters determined by light-scattering experiments have a well-defined physical significance and light-scattering data can thus be used in parallel with other data to provide information from which model parameters can be estimated. © 2011 Elsevier B.V. Source

The chromatographic retention in hydrophobic and reversed phase chromatography and the solubility of proteins display some common features. The chromatographic retention, as well as the solubility, is modulated by the thermodynamic properties of the solute in the fluid phase. The retention measurements at linear conditions provide information of the solution properties of the protein at infinite dilution, and the solubility measurements produce the supplementary information about the solution properties at the saturation limit. This provides a useful approach to simultaneous correlation of the chromatographic retention and the solubility.The experimental data, used for the correlation, comprise retention measurements of lysozyme on different HIC adsorbents using an aqueous ammonium sulphate eluant, an aqueous ammonium sulphate eluant with an admixture of ethanol, as well as published solubility data.The chromatographic retention data and the corresponding solubility data have been correlated using a chemical potential model derived from Kirkwood's theory of solutions of charged macro-ions and zwitterions in electrolyte solutions. The model correlated the chromatographic retention factor and the solubility data within the precision of the measurements. The model was applied in a pH range from 4 to 11. It was demonstrated experimentally, as well as theoretically, that an admixture of ethanol to the aqueous eluant changes the thermodynamic retention factor on various adsorbents identically when compared to the thermodynamic retention factor in an ethanol free eluant. © 2010 Elsevier B.V. Source

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