Biporous polymeric microspheres coupled with mercaptopyridine for rapid chromatographic purification of plasmid DNA

A biporous absorbent coupled with mercaptopyridine was synthesized for the purification of plasmid DNA. Analyses by scanning electron microscopy and mercury intrusion porosimetry revealed that the matrix contained two families of pores, i.e., micropores and superpores. The superpores provided not only convective flow channels for the mobile phase, but also a large surface for biomacromolecules binding. So the chromatographic process can be operated at high flow rate with high column efficiency and low backpressure as identified on a 2-mL column. When 10 mL crude feed-stock containing 3 mg of plasmid (5.4 kb pcDNA3) was loaded at a flow rate as high as 20 cm/min, the separation was finished in 10 min, and the plasmid was completely recovered with undetectable impurities of nucleic acids and proteins. The productivity was determined to be 9.0 g/L h, comparable to the pDNA productivity obtained using the commercial column. These results indicate that the biporous medium is promising for high-throughput purification of plasmid DNA. (c) 2007 Wiley Periodicals, Inc.