What are Cryogels? | Monolith | Ion-exchange | Affinity Chromatography: Cryogels were reported for the first time about five decades ago. Cryogels are a super-macroporous polymeric monolithic material with controlled porosities synthesized in partially frozen aqueous media and continuously interconnected pores after preparation. The monomer/crosslinker solution is frozen at low temperature (≈−20 °C) where crystals form, and two phases are segregated; one with a high concentration of monomer/crosslinker solution (unfrozen liquid microphase) and the ice phase (frozen part). Water is widely used as a solvent for production of cryogel due to its environmentally friendly properties as well as being cheaper than other solvents such as dimethyl sulfoxide. Therefore, in this system, the ice crystals act as porogens and create the porous structure. After allowing sufficient time for cryogelation, cryogels are thawed at room temperature. The ice crystals melt, and the result of polymerization will be permanent super macroporous structures even in the dry state; the geometry of the ice crystals determines the final pore size. In the literature, several biomedical and biotechnological potentials of these materials have now been reported [1-15].
Monolithic cryogels has been found to be an efficient stationary phase for capturing different therapeutic biomolecules with pore diameters in the range of 10-100 µm. The structural features of these adsorbents allow the separation and purification of biomolecules by convective flow as compared to the diffusional limitation observed in the conventional particle based resins. As a result of their simple preparation and enhanced mass transfer properties, monolithic cryogels present an attractive alternative to currently used bead-based adsorbent media. But due to large pores, cryogels offers lower surface area which is an issue and needs to be modified for capturing smaller molecules. Several researchers have reported that cryogels backbone can be modified with different functional groups. Such functionalities have been introduced either directly during the preparation of the monolith backbone [16,17] or by surface grafting [1,2,3,4,5,18,19], where the epoxy groups are first introduced onto the monolith and subsequently modified into any desired functionality. Gu et al. adopted the former approach and synthesized a series of monoliths using functionalized copolymers with strong cation-exchange properties in a single step [20,21]. Singh et al. selected the surface grafting approach and synthesized weak anion-exchangers (WAX) by appending DEAE groups onto the epoxy-functionalized backbone, and reported dynamic binding capacities of up to 27 mg/mL [3,4]. Even Savina et al. have also adopted the surface grafting approach and synthesized monoliths by introducing ionizable groups onto the backbone, and reported protein-binding capacities of up to 6-12 mg/mL .
- N. S. Bibi, N. K. Singh, R. N. Dsouza, M. Aasim, M. Fernandez-Lahore, Synthesis and performance of megaporous immobilized metal-ion affinity cryogels for recombinant protein capture and purification, J. Chromatogr. A 1272 (2013) 145–149. Kudos: https://goo.gl/h2qyMP
- N.K. Singh, R.N. DSouza, N.S. Bibi, M. Fernández-Lahore, Direct Capture of His6-Tagged Proteins Using Megaporous Cryogels Developed for Metal-Ion Affinity Chromatography, in: S. Reichelt (Ed.) Affinity Chromatography: Methods and Protocols, Springer New York, New York, NY, 2015, pp. 201-212. Kudos: https://goo.gl/iLHbTg
- N. K. Singh, R. N. Dsouza, M. Grasselli, M. Fernandez-Lahore, High capacity cryogel-type adsorbents for protein purification, J. Chromatogr. A 1355 (2014) 143–148. Kudos: https://goo.gl/AHkQnN
- N. Singh, R.N. Dsouza, M. Grasselli, M. Fernandez-Lahore, Gamma ray-mediated functionalization of monolithic cryogels for macrobiomolecule purification, N. Biotechnol., 2014, 31, S127-S127. Kudos: https://goo.gl/s4KAFN
- N.K. Singh, R.N. Dsouza, V. Yelemane, N. Nentwig, M. Grasselli, M. Fernandez-Lahore, pDNA Capture using Grafted Adsorbents“, J. Chem. Technol. Biotechnol., 93 (2018) pp. 1975-1979. doi:10.1002/jctb.5671.
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- C. P. Bisjak, R. Bakry, C. W. Huck, G. K. Bonn, Amino-functionalized monolithic poly(glycidyl methacrylate-co-divinylbenzene) ion-exchange stationary phases for the separation of oligonucleotides, Chromatographia 62 (2005) s31–s36.
- W. Wieder, C. P. Bisjak, C. W. Huck, R. Bakry, G. K. Bonn, Monolithic poly(glycidyl methacrylate-co-divinylbenzene) capillary columns functionalized to strong anion exchangers for nucleotide and oligonucleotide separation, J. Sep. Sci. 29 (2006) 2478–2484.
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- Y. Li, B. Gu, H. Dennis Tolley, M. L. Lee, Preparation of polymeric monoliths by copolymerization of acrylate monomers with amine functionalities for anion-exchange capillary liquid chromatography of proteins, J. Chromatogr. A 1216 (2009) 5525–5532.
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I. N. Savina, I. Y. Galaev, B. Mattiasson, Ion-exchange macroporous hydrophilic gel monolith with grafted polymer brushes, J. Mol. Recognit. 19 (2006) 313–321.