Mathematical correlations for predicting protein retention times in hydrophobic interaction chromatography
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Updated: Sep 28, 2026

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry
Published on: November 29, 2013
M Elena Lienqueo1, Andrea Mahn, Juan A Asenjo
1Centre for Biochemical Engineering and Biotechnology, Millennium Institute for Advanced Studies in Cell Biology and Biotechnology, Department of Chemical Engineering, University of Chile, Beauchef 861, Santiago, Chile.
Abstract:
This paper reports how a novel method for determining the surface hydrophobicity of a protein can be used to predict the dimensionless retention time (DRT) in hydrophobic interaction chromatography. The methodology has three steps. First, it is necessary to know the three-dimensional structure of the target protein. Then, it is necessary to calculate the surface hydrophobicity of the protein, considering that each amino acid has a relative contribution to the surface properties, using an appropriate equation. This will depend largely on the relative scale used to evaluate the hydrophobicity of each amino acid. Forty-two scales were investigated and two gave the best correlation, the Miyazawa-Jernigan and the Cowan-Whittaker scales. Finally, a "quadratic model" (DRT = a + b phi(surface) + c phi2(surface)) is used to predict the retention time of the target protein. The methodology was able to correlate adequately the retention data for monomeric proteins not included in the generation of the model.
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