title: Volumetric interpretation of protein adsorption: Partition coefficients, interphase volumes, and free energies of adsorption to hydrophobic surfaces.
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 (15kDa), alpha-amylase (51kDa), human serum albumin (66kDa), prothrombin (72kDa), immunoglobulin G (160kDa), and fibrinogen (341kDa) 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 V(I), and the number of hypothetical layers M occupied by protein adsorbed within V(I). Partition coefficients quantify protein-adsorption avidity through the equilibrium ratio of interphase and bulk-solution-phase w/v (mg/mL) concentrations W(I) and W(B), respectively, such that P identical withW(I)/W(B). Proteins are found to be weak biosurfactants with 45<P<520 and commensurately low apparent free-energy-of-adsorption -6RT<(DeltaG(adsphobic)(0)=-RTlnP)<-4RT. 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<100kDa occupy a single layer at surface saturation whereas the larger proteins IgG and fibrinogen required two layers.
authors.affiliation: Department of Materials Science and Engineering, The Pennsylvania State University, University Park, PA 16802, USA.
authors.forename: Hyeran
authors.initials: H
authors.lastname: Noh
authors.forename: Erwin A
authors.initials: EA
authors.lastname: Vogler
mesh_terms: D000327:Adsorption; D004591:Electrophoresis, Polyacrylamide Gel; D019277:Entropy; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008956:Models, Chemical; D011506:Proteins / Q000737:chemistry*; D012996:Solutions / Q000737:chemistry; D013499:Surface Properties
chemical_list: D011506:Proteins; D012996:Solutions