Volumetric interpretation of protein adsorption: Partition coefficients, interphase volumes, and free energies of adsorption to hydrophobic surfaces

Hyeran Noh, Erwin A. Vogler

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Abstract

The solution-depletion method of measuring protein adsorption is implemented using SDS gel electrophoresis as a separation and quantification tool. Experimental method is demonstrated using lysozyme (15 kDa), α-amylase (51 kDa), human serum albumin (66 kDa), prothrombin (72 kDa), immunoglobulin G (160 kDa), and fibrinogen (341 kDa) adsorption from aqueous-buffer solution to hydrophobic octyl-sepharose and silanized-glass particles. Interpretive mass-balance equations are derived from a model premised on the idea that protein reversibly partitions from bulk solution into a three-dimensional (3D) interphase volume separating the physical-adsorbent surface from bulk solution. Theory both anticipated and accommodated adsorption of all proteins to the two test surfaces, suggesting that the underlying model is descriptive of the essential physical chemistry of protein adsorption. Application of mass balance equations to experimental data quantify partition coefficients P, interphase volumes VI, and the number of hypothetical layers M occupied by protein adsorbed within VI. Partition coefficients quantify protein-adsorption avidity through the equilibrium ratio of interphase and bulk-solution-phase w/v (mg/mL) concentrations WI and WB, respectively, such that P ≡ WI / WB. Proteins are found to be weak biosurfactants with 45 < P < 520 and commensurately low apparent free-energy-of-adsorption - 6 RT < (Δ Gfrac(ads, phobic)0 = - RT ln P) < - 4 RT . These measurements corroborate independent estimates obtained from interfacial energetics of adsorption (tensiometry) and are in agreement with thermochemical measurements for related proteins by hydrophobic-interaction chromatography. Proteins with molecular weight MW < 100 kDa occupy a single layer at surface saturation whereas the larger proteins IgG and fibrinogen required two layers.

Original languageEnglish (US)
Pages (from-to)5780-5793
Number of pages14
JournalBiomaterials
Volume27
Issue number34
DOIs
StatePublished - Dec 1 2006

All Science Journal Classification (ASJC) codes

  • Bioengineering
  • Ceramics and Composites
  • Biophysics
  • Biomaterials
  • Mechanics of Materials

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