ABSTRACT
The preparation and characterization of mixed-mode adsorbents for a typical separation purpose are of great importance in bioseparation areas. In this work, we prepared a new monolithic cryogel with a combination of ion-exchange and hydrophobic functions by employing benzyl-quaternary amine groups. The fundamental cryogel properties, protein equilibrium adsorption isotherm and chromatographic adsorption in the cryogel were measured experimentally. The results showed that, by using bovine serum album as the model protein, the dual functional cryogel has protein binding capability even in salt solution and the buffer with pH close or below the protein isoelectric point due to both the electrostatic and hydrophobic interactions. A capillary-based adsorption model was developed, which provided satisfied insights of the microstructure, axial dispersion, mass transfer as well as protein adsorption characteristics within the cryogel bed. The chromatographic isolation of bioactive proteins from rabbit blood serum was carried out by the cryogel. Immunoglobulin G antibody with a purity of 98.2% and albumin with a purity of 96.8% were obtained, indicating that the cryogel could be an interesting and promising adsorbent in bioseparation areas.
Subject(s)
Acrylic Resins/chemistry , Cryogels/chemistry , Immunoglobulin G/chemistry , Polystyrenes/chemistry , Quaternary Ammonium Compounds/chemistry , Serum Albumin/chemistry , Adsorption , Animals , Cattle , Chromatography, Liquid/methods , Cryogels/chemical synthesis , Hydrophobic and Hydrophilic Interactions , Ligands , Molecular Weight , RabbitsABSTRACT
A novel super-macroporous monolithic composite cryogel was prepared by embedding macroporous cellulose beads into poly(hydroxyethyl methacrylate) cryogel. The cellulose beads were fabricated by using a microchannel liquid-flow focusing and cryopolymerization method, while the composite cryogel was prepared by cryogenic radical polymerization of the hydroxyethyl methacrylate monomer with poly(ethylene glycol) diacrylate as cross-linker together with the cellulose beads. After graft polymerization with (vinylbenzyl)trimethylammonium chloride, the composite cryogel was applied to separate immunoglobulin-G and albumin from human serum. Immunoglobulin-G with a mean purity of 83.2% and albumin with a purity of 98% were obtained, indicating the composite cryogel as a promising chromatographic medium in bioseparation for the isolation of important bioactive proteins like immunoglobulins and albumins.
Subject(s)
Cellulose/chemistry , Cryogels/chemistry , Immunoglobulins/isolation & purification , Polyhydroxyethyl Methacrylate/chemistry , Serum Albumin/isolation & purification , Humans , Immunoglobulins/blood , Microspheres , Particle Size , Porosity , Surface PropertiesABSTRACT
Bovine milk whey contains several bioactive proteins such as α-lactalbumin, ß-lactoglobulin, and immunoglobulin G (IgG). Chromatographic separation of these proteins has received much attention in the past few years. In this work, we provide a chromatographic method for the efficient isolation of IgG from bovine milk whey using a poly(2-hydroxyethyl methacrylate)-based anion-exchange cryogel. The monolithic cryogel was prepared by grafting 2-(dimethylamino) ethyl methacrylate onto the poly(2-hydroxyethyl methacrylate)-based cryogel matrix and then employed to separate IgG under various buffer pH and salt elution conditions. The results showed that the buffer pH and the salt concentration in the step elution have remarkable influences on the purity of IgG, while the IgG recovery depended mainly on the loading volume of whey for a given cryogel bed. High purity IgG (more than 95%) was obtained using the phosphate buffer with pH of 5.8 as the running buffer and the salt solution in as the elution liquid. With suitable loading volume of whey, the maximum IgG recovery of about 94% was observed. The present separation method is thus a potential choice for the isolation of high-purity IgG from bovine milk whey.
