Performance of R-N(R')-R'' functionalised poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) monolithic sorbent for plasmid DNA adsorption.
High-throughput plasmid DNA (pDNA) manufacture is obstructed predominantly by the performance of conventional stationary phases. For this reason, the search for new materials for fast chromatographic separation of pDNA is ongoing. A poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) (GMA-EGDMA) monolithic material was synthesised via a thermal-free radical reaction, functionalised with different amino groups from urea, 2-chloro-N,N-diethylethylamine hydrochloride (DEAE-Cl) and ammonia in order to investigate their plasmid adsorption capacities. Physical characterisation of the monolithic polymer showed a macroporous polymer having a unimodal pore size distribution pivoted at 600 nm. Chromatographic characterisation of the functionalised polymers using pUC19 plasmid isolated from E. coli DH5alpha-pUC19 showed a maximum plasmid adsorption capacity of 18.73 mg pDNA/mL with a dissociation constant (KD) of 0.11 mg/mL for GMA-EGDMA/DEAE-Cl polymer. Studies on ligand leaching and degradation demonstrated the stability of GMA-EGDMA/DEAE-Cl after the functionalised polymers were contacted with 1.0 M NaOH, which is a model reagent for most 'cleaning in place' (CIP) systems. However, it is the economic advantage of an adsorbent material that makes it so attractive for commercial purification purposes. Economic evaluation of the performance of the functionalised polymers on the grounds of polymer cost (PC)/mg pDNA retained endorsed the suitability of GMA-EGDMA/DEAE-Cl polymer.
Danquah
Michael K
MK
BEL (Bioengineering Laboratory), Department of Chemical Engineering, Monash University, Melbourne, Australia. michael.danquah@eng.monash.edu.au
Ho
Jenny
J
Forde
Gareth M
GM
D000327:Adsorption; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods; D004269:DNA, Bacterial / Q000737:chemistry*; D004926:Escherichia coli / Q000737:chemistry; D005026:Ethylene Glycols; D008689:Methacrylates / Q000737:chemistry*; D015394:Molecular Structure; D010316:Particle Size; D010957:Plasmids / Q000737:chemistry*; D016062:Porosity; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013499:Surface Properties; D013997:Time Factors
D004269:DNA, Bacterial; D005026:Ethylene Glycols; D008689:Methacrylates; C526534:poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate)