Protein sorption and recovery by hydrogels using principles of aqueous two-phase extraction.
Use of the thermodynamic principles of aqueous two-phase extraction (ATPE) to drive protein into a crosslinked gel is developed as a protein isolation and separation technique, and as a protein loading technique for drug delivery applications. A PEG/dextran gel system was chosen as a model system because PEG/dextran systems are widely used in aqueous two-phase extraction and dextran gels (Sephadex(R)) are common chromatographic media. The effects of polymer concentrations and molecular weights, salts, and pH on the partitioning of ovalbumin matched ATPE heuristics and data trends. Gel partition coefficients (Cgel/Csolution) increased with increasing PEG molecular weight and concentration and decreasing dextran concentration (increased gel swelling). The addition of PEG to the buffer solution yielded partition coefficients more than an order of magnitude greater than those obtained in systems with buffer alone, or added salt. A combined salt/PEG system yielded an additional order of magnitude increase. For example, when ovalbumin solution (2.3 mg/mL) was equilibrated with Sephadex(R) G-50 at pH 6.75, the partition coefficients were 0.13 in buffer, 0.11 in buffer with 0.22M KI, 2.3 in 12 wt% PEG-10,000 and 32.0 in 12 wt% PEG-10, 000 with 0.22M KI. The effect of anions and cations as well as ionic strength and pH on the partitioning of ovalbumin also matched ATPE heuristics. Using the heuristics established above, partition coefficients as high as 80 for bovine serum albumin and protein recoveries over 90% were achieved. In addition, the wide range of partition coefficients that were obtained for different proteins suggests the potential of the technique for separating proteins. Also, ovalbumin sorption capacities in dextran were as high as 450 mg/g dry polymer, and the sorption isotherms were linear over a broad protein concentration range.
Gehrke
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Department of Chemical Engineering, Mail Location 0171, University of Cincinnati, Cincinnati, Ohio 45221-0171, USA. Stevin.Gehrke@uc.edu
Vaid
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McBride
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D000327:Adsorption; D000818:Animals; D002417:Cattle; D003911:Dextrans; D005782:Gels; D020100:Hydrogels; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D010047:Ovalbumin / Q000737:chemistry / Q000302:isolation & purification*; D011092:Polyethylene Glycols; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D012492:Salts; D012710:Serum Albumin, Bovine / Q000302:isolation & purification; D012996:Solutions; D055672:Static Electricity
D003911:Dextrans; D005782:Gels; D020100:Hydrogels; D007202:Indicators and Reagents; D011506:Proteins; D012492:Salts; D012996:Solutions; D012710:Serum Albumin, Bovine; D011092:Polyethylene Glycols; D010047:Ovalbumin; C025614:sephadex