Synthesis of monodisperse nonporous crosslinked poly(glycidyl methacrylate) particles with metal affinity ligands for protein adsorption

Abstract

Monodisperse nonporous crosslinked poly(glycidyl methacrylate) (PGMA) particles with immobilized metal affinity ligands were prepared for selective recovery of proteins. The PGMA particles, with an average size of 2.2 µm, were prepared by a simple dispersion polymerization of glycidyl methacrylate (GMA) and ethylene glycol dimethacrylate (EGDMA). The particles were characterized by scanning electron microscopy (SEM) and Fourier-transform infrared spectroscopy (FTIR). The epoxy groups of the particles were modified with the metal chelating agent iminodiacetic acid (IDA), which forms metal-IDA chelates at the active sites. After charging with copper ions, the particles were used to recover a model protein, bovine hemoglobin (BHb), in a batchwise manner. The particles had the adsorption capacity of 218.7 mg g −1 with little nonspecific adsorption. The adsorption behavior could be described with the Langmuir equation. The effect of pH on the adsorption was also studied. Regeneration of the metal-chelated particles was easily performed with 50 mmol L −1 ethylenediaminetetraacetic acid (EDTA), followed by washing with water and reloading with Cu 2+ . The particles could be very useful as an affinity separation adsorbent.

Chromatographic Removal of Endotoxins: A Bioprocess Engineer's Perspective

“…Dispersion, emulsion and suspension polymerisations involving glycidyl methacrylate (GMA) and ethylene glycol dimethylacrylate (EDMA) make PGMA [31]. PGMA has large pores, which translate to rapid mass transfer, high adsorption kinetics and negligible lowflow resistance [29].…”

“…PGMA has large pores, which translate to rapid mass transfer, high adsorption kinetics and negligible lowflow resistance [29]. Past researchers have functionalised PGMA with urea, ammonia and DEAE-Cl as well as covalent coupling with imminodiacetic acid-metal (IDAmetal) chelator for the purification of plasmid DNA [29,31]. Adsorption isotherm was determined as Langmuir due to the strong plasmid molecules-ligands interactions.…”

“…PGMA functionalised with IDAmetal chelator has been used in Immobilised Metal Affinity Chromatography (IMAC) for protein adsorption and has shown favourable results. Addition of Ethylenediaminetetraacetic Acid (EDTA) provides for the regeneration of metal-chelated particles with no morphology damage or protein adsorption capacity loss [31]. Chelating agents destroy the bridging effect of calcium ions leading to the prevention of endotoxin aggregation [5].…”

“…For instance, high endotoxin retention is promoted using small pore size based on size-exclusion effects while large pore size reduces ionic interactions for negatively charged proteins [35]. A Monolith (i) PGMA has large pores which translate to rapid mass transfer, high adsorption kinetics and negligible low-flow resistance [29,31] (ii) Addition of Ethylenediaminetetraacetic Acid (EDTA) provides for the regeneration of metal-chelated particles with no morphology damage or protein adsorption capacity loss…”

“…It is a strong pDNA binder and elutes pDNA only at high-salt conditions [34]. However, silica beads are unstable under extreme pH condition [31]. Sephacryl S-500 HR has a high selectivity towards pDNA over RNA through implementation of selective access of molecules into pores and allows passing through of RNA and proteins but bans pDNA [58].…”