Construction of polymer monolithic columns in polypropylene ink-pen tubes for separation of proteins by cation-exchange chromatography.

Abstract

We describe the synthesis of polymer monoliths inside polypropylene tubes from ink pens. These tubes are cheap, chemically stable, and resistant to pressure. UV-initiated grafting with 5wt% benzophenone in methanol for 20min activated the internal surface, thus enabling the covalent binding of ethylene glycol dimethacrylate, also via photografting. The pendant vinyl groups attached a poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) monolith prepared via photopolymerization. These tubes measured 100-110mm long, with 2mm of internal diameter. The parent monoliths were functionalized with Na2 SO3 or iminodiacetate to produce strong and weak cation exchangers, respectively. The columns exhibited permeabilities varying from 2.7 to 3.3 × 10-13 m2 , which enabled the separation of proteins at 500 µL/min and back pressures<2.8MPa. Neither structure collapse nor monolith detachment occurred at flow rates as high as 2.0mL/min, which produced back pressures between 6.9 and 9.0MPa. The retention times of ovalbumin, ribonuclease A, cytochrome C, and lysozyme in salt gradient at pH 7.0 followed the order of increasing isoelectric points, confirming the cation exchange mechanism. Separation and determination of lysozyme in egg white proved the applicability of the columns to the analysis of complex samples.