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Adil Denizli - One of the best experts on this subject based on the ideXlab platform.

  • Biologically Modified poly(2-hydroxylethyl methacrylate) Cryogels for Lysozyme Purification
    2019
    Co-Authors: Duygu Çimen, Adil Denizli
    Abstract:

    Heparin immobilized poly(2-hydroxylethyl methacrylate) PHEMA Cryogel was synthesized and applied for lysozyme purification from egg white. Firstly, the PHEMA Cryogel was synthesized by cryopolymerization and then heparin was covalently immobilized on to the PHEMA Cryogel with cyanogen bromide activation. The modification of PHEMA Cryogel structure with heparin was further confirmed by Fourier-transform infrared spectroscopy (FTIR). The surface and inner structure morphologies of PHEMA Cryogels were studied and characterized by the scanning electron microscope (SEM). The surface area of PHEMA Cryogel was found to be 25.2 m2/g. Heparin immobilized PHEMA Cryogels were used in lysozyme adsorption studies to assess the effects of pH, lysozyme concentration, flow rate, temperature and ionic strength. The maximum lysozyme adsorption on the heparin immobilized PHEMA Cryogel was found to be 48.73 mg/g from aqueous solutions under optimized conditions. 1.0 M NaCI solution was used for desorption of lysozyme in a continuous system. The reusability of heparin immobilized PHEMA Cryogels was tested for 10 adsorption-desorption cycles. The Langmuir adsorption model was plotted and found fitted for adsorption studies. The purity of lysozyme from egg white studies was analysed by sodium-dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) using 12% separating gel.

  • Synthesis of hydroxyethyl-methacrylate-(L)-histidine methyl ester Cryogels. Application on the separation of bovine immunoglobulin G
    Analytical Biochemistry, 2017
    Co-Authors: Assem Elkak, Cana Armutcu, Amar Hamade, Nilay Bereli, Adil Denizli
    Abstract:

    In this study Cryogels based 2-hydroxyethyl methacrylate (HEMA) functionalized with N-methacryloyl-L-histidine methyl ester (MAH) were synthesized and used for the adsorption and separation of bovine IgG. Two series of Cryogels functionalized with 5 and 10 mg of MAH as pseudobioaffinity ligand were prepared and characterized by swelling test, FTIR and SEM analysis. The adsorption efficiency of the bovine immunoglobulin into Cryogels is discussed with respect to the following chromatographic parameters: pH, flow rate, initial IgG concentration, adsorption time and ionic strength. Our results show good adsorption of bovine immunoglobulin under mild separation conditions at pH 7.4. The maximum binding capacity was determined (32.4 mg/g of Cryogel) and demonstrates the efficiency of the used Cryogels. This efficacy is clearly seen upon increasing the maximum binding capacity from 23.2 mg (obtained with Cryogels with 5 mg MAH) to 32.4 mg/g (for Cryogel with 10 mg MAH ligand concentration). The purity of separated fractions was evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Together our observations highlights poly (HEMA-MAH) as an efficient adsorbent for bovine immunoglobulins G separation.

  • Dye functionalized Cryogel columns for reversible lysozyme adsorption
    Journal of biomaterials science. Polymer edition, 2015
    Co-Authors: Murat Uygun, Deniz Aktas Uygun, Sinan Akgol, Begüm Akduman, Adil Denizli
    Abstract:

    In this study, poly (methyl methacrylate–glycidyl methacrylate) [poly(MMA-GMA)] Cryogels were prepared by radical cryocopolymerization of MMA with GMA as a functional comonomer. Reactive Green 19 dye was then attached to the Cryogel by nucleophilic substitution reaction, and this dye-attached Cryogel column was used for lysozyme adsorption. Characterization of the Cryogel was performed by Fourier transform infrared spectroscopy, environmental scanning electron microscopy, Brunauer–Emmett–Teller, and energy dispersive X-ray analysis. Pore size of the Cryogels was 15–30 μm and pores were interconnected structure. Attached amount of Reactive Green 19 to Cryogel support was calculated as 106.25 μmol/g Cryogel. Lysozyme adsorption studies were carried out by using a continuous system. It was found that the maximum amount of lysozyme adsorption (32 mg/g Cryogel) obtained from experimental results was found to be approximately same with the calculated Langmuir adsorption capacity (33 mg/g Cryogel). Desorption of...

  • immunoglobulin g purification from bovine serum with pseudo specific supermacroporous Cryogels
    Separation and Purification Technology, 2013
    Co-Authors: Mira Daoudattieh, Lokma Uzu, Haida Chaib, Cana Armutcu, Assem Elkak, Adil Denizli
    Abstract:

    Abstract Supermacroporous Cryogels were synthesized and used for the purification of immunoglobulin G (IgG) from bovine serum. Imidazole functional groups were incorporated into basic polymeric backbone to gain pseudo-specificity to Cryogels by using n-vinylimidazole as comonomer. The Cryogels were prepared in aqueous solution of monomers partially frozen inside plastic syringe column by Cryogelation. Poly(2-hydroxyethyl methacrylate-n-vinylimidazole) [poly(HEMA-VIM)] Cryogels were prepared by polymerization of water-soluble functional monomer, n-vinylimidazole, with basic monomer HEMA. The characterization methods including swelling test, Fourier transform infrared spectroscopy (FTIR), elemental analysis, and scanning electron microscopy (SEM) were performed to evaluate physical and chemical properties of Cryogels prepared. Bovine IgG adsorption on plain and composite Cryogels was studied with respect to different parameters such as, pH, IgG concentration, flow rate, ionic strengths, and adsorption time. The best adsorption of bovine IgG was observed at pH 7.4 up to 21.1 mg per unit mass of poly(HEMA-VIM) Cryogel. Elution of IgG adsorbed from the Cryogels was easily achieved with 0.1 M acetate buffer containing 1 M NaCl at pH 4.0. In order to describe the adsorption process, we applied some equilibrium and kinetic adsorption models to the data. The results best fitted to Langmuir model showing monolayer protein adsorption and surface homogeneity of Cryogel. Finally, we evaluated IgG purification from bovine serum under optimal condition determined. The purification efficiency and IgG purity were investigated with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE) study.

  • molecularly imprinted poly hydroxyethyl methacrylate based Cryogel for albumin depletion from human serum
    Colloids and Surfaces B: Biointerfaces, 2013
    Co-Authors: Muge Andac, Igor Yu Galaev, Adil Denizli
    Abstract:

    Macroporous Cryogels imprinted with human serum albumin (HSA) have been prepared by copolymerization of 2-hydroxyethyl methacrylate with a functional co-monomer of N-methacryloyl-L-phenylalanine. The Cryogels were used for the depletion of HSA from human serum. HSA-imprinted Cryogels were prepared with gel fraction yields up to 90%, and their chemical structure, morphology and porosity were characterized by FTIR-spectroscopy, scanning electron microscopy, swelling studies and flow dynamics. Selective binding experiments were performed in the presence of competitive proteins like human transferrin and myoglobin. Albumin-imprinted Cryogel column was optimized for fast protein liquid chromatography. Sodium-dodecyl sulfate polyacrylamide gel electrophoresis was used to show the efficiency of albumin depletion.

Ashok Kumar - One of the best experts on this subject based on the ideXlab platform.

  • decellularized liver matrix modified Cryogel scaffolds as potential hepatocyte carriers in bioartificial liver support systems and implantable liver constructs
    ACS Applied Materials & Interfaces, 2018
    Co-Authors: Apeksha Damania, Arun Kumar Teotia, Haruna Kimura, Masamichi Kamihira, Hiroyuki Ijima, Shiv Kumar Sarin, Anupam Kumar, Ashok Kumar
    Abstract:

    Recent progress in the use of decellularized organ scaffolds as regenerative matrices for tissue engineering holds great promise in addressing the issue of donor organ shortage. Decellularization preserves the mechanical integrity, composition, and microvasculature critical for zonation of hepatocytes in the liver. Earlier studies have reported the possibility of repopulating decellularized matrices with hepatic cell lines or stem cells to improve liver regeneration. In this work, we study the versatility of the decellularized liver matrix as a substrate coating of three-dimensional Cryogel scaffolds. The coated Cryogels were analyzed for their ability to maintain hepatic cell growth and functionality in vitro, which was found to be significantly better than the uncoated Cryogel scaffolds. The decellularized liver matrix-coated Cryogel scaffolds were evaluated for their potential application as a cell-loaded bioreactor for bioartificial liver support and as an implantable liver construct. Extracorporeal c...

  • Fabrication of macroporous Cryogels as potential hepatocyte carriers for bioartificial liver support.
    Colloids and surfaces. B Biointerfaces, 2015
    Co-Authors: Era Jain, Apeksha Damania, Shiv Kumar Sarin, Anupam Kumar, Akhilesh Kumar Shakya, Ashok Kumar
    Abstract:

    Two different Cryogels composed of copolymer of acrylonitrile (AN) and N-vinyl-2-pyrrolidone (NVP) (poly(AN-co-NVP)) and interpenetrated polymer networks (IPN) of chitosan and poly(N-isopropylacrylamide) (poly(NiPAAm)-chitosan) were fabricated by gelation at sub-zero temperatures. The two Cryogels possess an interconnected network of macropores of size 20-100 μm and efficient transport properties as determined by physiochemical analysis. Both Cryogels support in vitro growth and function of fibroblasts (COS-7) and human liver hepatocarcinoma cells (HepG2). The Cryogels are hemocompatible as demonstrated by low albumin adsorption and platelet adherence. Furthermore, in vivo implantation of poly(NiPAAm)-chitosan Cryogel in mice shows its biocompatibility with the surrounding tissue. Primary rat hepatocytes grown on poly(NiPAAm)-chitosan Cryogel for 96 h formed cellular aggregates and maintained their functions in terms of, ammonia removal, ureagenesis and drug detoxification. Cryogel-based closed continuous bioreactor systems could maintain HepG2 cells at high density for 7 days. Off-line clinical evaluation of these Cryogel-based bioreactors showed the ability of immobilized cells to detoxify circulating plasma obtained from patients with acute on chronic liver failure (ACLF). Altogether, the presented data suggests Cryogels as a potential bioreactor matrix for bio-artificial liver support system.

  • Disposable polymeric Cryogel bioreactor matrix for therapeutic protein production
    Nature Protocols, 2013
    Co-Authors: Era Jain, Ashok Kumar
    Abstract:

    Low cost and high efficiency make disposable bioreactors feasible for small-scale therapeutic development and initial clinical trials. We have developed a Cryogel-based disposable bioreactor matrix, which has been used for production of protein therapeutics such as urokinase and monoclonal antibodies (mAbs). The protocol discusses the application of a Cryogel bioreactor for mAb production. Cryogels composed of either polyacrylamide (PAAm) coupled to gelatin or semi-interpenetrating PAAm-chitosan are synthesized by free-radical polymerization at −12 °C. Hybridoma cells are immobilized over the Cryogel bioreactor and incubated for 48 h. Medium is circulated thereafter at 0.2 ml min^−1 and bioreactors can be run continuously for 60 d. The Cryogel-based packed-bed bioreactor can be formulated as a monolith or as beads; it also has an efficiency four times what can be obtained using a tissue-culture flask, a high surface-to-volume ratio and effective nutrient transport. After incubation, the bioreactor setup will take about 60 min using a pre-prepared sterilized Cryogel.

  • Tunable hybrid Cryogels functionalized with microparticles as supermacroporous multifunctional biomaterial scaffolds
    Journal of biomaterials science. Polymer edition, 2012
    Co-Authors: Haider Sami, Ashok Kumar
    Abstract:

    This study aims at incorporating microparticles (MPs) into polymeric network of Cryogels to fabricate a multifunctional biomaterial scaffold (Cryogel-microparticle [C-MP] composite) which has microstructured biocompatible pore surface, supermacroporous interconnected pore distribution and capability to simultaneously deliver bioactive molecules. Three types of MPs (i.e. poly(D,L-lactide-co-glycolide), chitosan, and silica MPs) were separately incorporated in three different Cryogels (i.e. chitosan–agarose, chitosan–agarose–gelatin, and chitosan–gelatin), demonstrating the generality of the approach. Factors affecting distribution of MPs within the Cryogel were investigated to achieve uniform distribution of MPs within the pore wall of Cryogel as well as the whole Cryogel monolith to create microstructured pore surface of C-MP composites, which was characterized by scanning electron microscopy, fluorescence microscopy, infrared spectral analysis (FTIR), and water uptake properties. Further, mouse embryonic...

  • Thermoresponsive poly(N-vinylcaprolactam) Cryogels: synthesis and its biophysical evaluation for tissue engineering applications
    Journal of Materials Science: Materials in Medicine, 2010
    Co-Authors: Akshay Srivastava, Ashok Kumar
    Abstract:

    The thermoresponsive poly( N -vinylcaprolactam) (PVCl) based Cryogel network were synthesized and characterized with respect to physical and biological properties. The PVCl Cryogel crosslinked with polyethylene glycol-diacrylate (PEGda) was synthesized in 1% dimethyl sulfoxide containing aqueous medium at −12°C for 12–14 h. The Cryogel synthesized in this manner were highly spongy in nature and can absorb water in its porous network. These polymeric Cryogel networks have good physical morphology as confirmed by scanning electron microscopy. The estimated porosity of these Cryogels was 90% as demonstrated by various methods based on absorption of water and cyclohexane. The median pore diameter and surface area was 30 μm and 2.0253 m^2/g, respectively as confirmed by analysis on mercury porosimeter. These materials can interact with biological system without any cytotoxic effects. Change in temperature influenced the adsorption of fetal bovine serum (FBS) on PVCl scaffold which showed maximum protein adsorption at 37°C, as compared to that at 25°C. Furthermore, the fibroblast cell adhesion studies showed the potential of these PVCl based Cryogels as tissue engineering scaffolds.

Igor Yu Galaev - One of the best experts on this subject based on the ideXlab platform.

  • molecularly imprinted poly hydroxyethyl methacrylate based Cryogel for albumin depletion from human serum
    Colloids and Surfaces B: Biointerfaces, 2013
    Co-Authors: Muge Andac, Igor Yu Galaev, Adil Denizli
    Abstract:

    Macroporous Cryogels imprinted with human serum albumin (HSA) have been prepared by copolymerization of 2-hydroxyethyl methacrylate with a functional co-monomer of N-methacryloyl-L-phenylalanine. The Cryogels were used for the depletion of HSA from human serum. HSA-imprinted Cryogels were prepared with gel fraction yields up to 90%, and their chemical structure, morphology and porosity were characterized by FTIR-spectroscopy, scanning electron microscopy, swelling studies and flow dynamics. Selective binding experiments were performed in the presence of competitive proteins like human transferrin and myoglobin. Albumin-imprinted Cryogel column was optimized for fast protein liquid chromatography. Sodium-dodecyl sulfate polyacrylamide gel electrophoresis was used to show the efficiency of albumin depletion.

  • an improved capillary model for describing the microstructure characteristics fluid hydrodynamics and breakthrough performance of proteins in Cryogel beds
    Journal of Chromatography A, 2011
    Co-Authors: Igor Yu Galaev, Bo Mattiasson, Junxian Yun, Gry Ravn Jespersen, Harald Kirsebom, Pererik Gustavsson
    Abstract:

    A capillary-based model modified for characterization of monolithic Cryogels is presented with key parameters like the pore size distribution, the tortuosity and the skeleton thickness employed for describing the porous structure characteristics of a Cryogel matrix. Laminar flow, liquid dispersion and mass transfer in each capillary are considered and the model is solved numerically by the finite difference method. As examples, two poly(hydroxyethyl methacrylate) (pHEMA) based Cryogel beds have been prepared by radical cryo-copolymerization of monomers and used to test the model. The axial dispersion behaviors, the pressure drop vs. flow rate performance as well as the non-adsorption breakthrough curves of different proteins, i.e., lysozyme, bovine serum albumin (BSA) and concanavalin A (Con A), at various flow velocities in the Cryogel beds are measured experimentally. The lumped parameters in the model are determined by matching the model prediction with the experimental data. The results showed that for a given Cryogel column, by using the model based on the physical properties of the Cryogel (i.e., diameter, length, porosity, and permeability) together with the protein breakthrough curves one can obtain a reasonable estimate and detailed characterization of the porous structure properties of Cryogel matrix, particularly regarding the number of capillaries, the capillary tortuousness, the pore size distribution and the skeleton thickness. The model is also effective with regards to predicting the flow performance and the non-adsorption breakthrough profiles of proteins at different flow velocities. It is thus expected to be applicable for characterizing the properties of Cryogels and predicting the chromatographic performance under a given set of operating conditions.

  • molecularly imprinted phema based Cryogel for depletion of hemoglobin from human blood
    Macromolecular Chemistry and Physics, 2010
    Co-Authors: Ali Derazshamshir, Igor Yu Galaev, Ridva Say, Gozde Baydemir, Muge Andac, Adil Denizli
    Abstract:

    A new generation of molecularly imprinted macroporous hydrogels, so-called Cryogels, were prepared for depletion of hemoglobin from human blood. Hb-PHEMAH Cryogels were prepared with gel fraction yields up to 90%, and its morphology and porosity were characterized by FTIR, SEM, swelling studies, flow dynamics and surface area measurements. Selective binding experiments were performed in the presence of competitive PHEMAH Cryogel exhibited a high binding capacity and selectivity for Hb in the presence of Myb and BSA. The selectivity of Hb-PHEMAH Cryogel for Hb was confirmed by HPLC. Hb depletion from blood hemolysate was also studied using SDS-PAGE. Hb-PHE-MAH Cryogel can be reused many times with no apparent decrease in Hb adsorption capacity. (Less)

  • tissue responses to novel tissue engineering biodegradable Cryogel scaffolds an animal model
    Journal of Biomedical Materials Research Part A, 2009
    Co-Authors: Nimet Bölgen, Fatima M Plieva, Igor Yu Galaev, Ibrahim Vargel, Petek Korkusuz, Elif Guzel, Bo Matiasson, Erhan Pişkin
    Abstract:

    Biodegradable macroporous Cryogels with highly open and interconnected pore structures were produced from dextran modified with oligo L-lactide bearing hydroxyethylmethacrylate (HEMA) end groups in moderately frozen solutions. Tissue responses to these novel scaffolds were evaluated in rats after dorsal subcutaneous implantation, iliac submuscular implantation, auricular implantation, or in calvarial defect model. In no case, either necrosis or foreign body reaction was observed during histological studies. The Cryogel scaffolds integrated with the surrounding tissue and the formation of a new tissue were accompanied with significant ingrowth of connective tissue cells and new blood vessels into the Cryogel. The tissue responses were significantly lower in auricular and calvarial implantations when compared with the subcutanous and the submuscular implantations. The degradation of the scaffold was slower in bone comparing to soft tissues. The biodegradable Cryogels are highly biocompatible and combine extraordinary properties including having soft and elastic nature, open porous structure, and very rapid and controllable swelling. Therefore, the Cryogels could be promising candidates for further clinical applications in tissue regeneration. (C) 2008 Wiley Periodicals, Inc. J Biomed Mater Res 91 A: 60-68, 2009 (Less)

  • monolithic Cryogels with open porous structure and unique double continuous macroporous networks
    Soft Matter, 2008
    Co-Authors: Fatima M Plieva, Pete Ekstrom, Igor Yu Galaev, O Mattiasso
    Abstract:

    A novel approach to create macroporous Cryogels (or gels synthesized at sub-zero temperatures) with increased mechanical strength is presented. The Cryogels with unique double-continuous macroporous networks were prepared via sequential freezing–thawing, when the new Cryogel was synthesized inside the interconnected macropores of the preformed or primary Cryogel. The prepared double-continuous macroporous Cryogels (DM Cryogels) have continuously interlacing networks of primary Cryogel with the newly synthesized Cryogels visualized by confocal laser scanning microscopy and scanning electron microscopy. The open structures of the DM Cryogels with up to 150-μm-sized interconnected pores and tissue-like elasticity make them attractive as three-dimensional cell scaffolds for tissue-engineering applications. In particular, a method of preparation of macroporous Cryogels with open porous structure and/or gradient porosity, increased mechanical strength and increased content of functional groups is shown.

Era Jain - One of the best experts on this subject based on the ideXlab platform.

  • Fabrication of macroporous Cryogels as potential hepatocyte carriers for bioartificial liver support.
    Colloids and surfaces. B Biointerfaces, 2015
    Co-Authors: Era Jain, Apeksha Damania, Shiv Kumar Sarin, Anupam Kumar, Akhilesh Kumar Shakya, Ashok Kumar
    Abstract:

    Two different Cryogels composed of copolymer of acrylonitrile (AN) and N-vinyl-2-pyrrolidone (NVP) (poly(AN-co-NVP)) and interpenetrated polymer networks (IPN) of chitosan and poly(N-isopropylacrylamide) (poly(NiPAAm)-chitosan) were fabricated by gelation at sub-zero temperatures. The two Cryogels possess an interconnected network of macropores of size 20-100 μm and efficient transport properties as determined by physiochemical analysis. Both Cryogels support in vitro growth and function of fibroblasts (COS-7) and human liver hepatocarcinoma cells (HepG2). The Cryogels are hemocompatible as demonstrated by low albumin adsorption and platelet adherence. Furthermore, in vivo implantation of poly(NiPAAm)-chitosan Cryogel in mice shows its biocompatibility with the surrounding tissue. Primary rat hepatocytes grown on poly(NiPAAm)-chitosan Cryogel for 96 h formed cellular aggregates and maintained their functions in terms of, ammonia removal, ureagenesis and drug detoxification. Cryogel-based closed continuous bioreactor systems could maintain HepG2 cells at high density for 7 days. Off-line clinical evaluation of these Cryogel-based bioreactors showed the ability of immobilized cells to detoxify circulating plasma obtained from patients with acute on chronic liver failure (ACLF). Altogether, the presented data suggests Cryogels as a potential bioreactor matrix for bio-artificial liver support system.

  • Disposable polymeric Cryogel bioreactor matrix for therapeutic protein production
    Nature Protocols, 2013
    Co-Authors: Era Jain, Ashok Kumar
    Abstract:

    Low cost and high efficiency make disposable bioreactors feasible for small-scale therapeutic development and initial clinical trials. We have developed a Cryogel-based disposable bioreactor matrix, which has been used for production of protein therapeutics such as urokinase and monoclonal antibodies (mAbs). The protocol discusses the application of a Cryogel bioreactor for mAb production. Cryogels composed of either polyacrylamide (PAAm) coupled to gelatin or semi-interpenetrating PAAm-chitosan are synthesized by free-radical polymerization at −12 °C. Hybridoma cells are immobilized over the Cryogel bioreactor and incubated for 48 h. Medium is circulated thereafter at 0.2 ml min^−1 and bioreactors can be run continuously for 60 d. The Cryogel-based packed-bed bioreactor can be formulated as a monolith or as beads; it also has an efficiency four times what can be obtained using a tissue-culture flask, a high surface-to-volume ratio and effective nutrient transport. After incubation, the bioreactor setup will take about 60 min using a pre-prepared sterilized Cryogel.

  • macroporous interpenetrating Cryogel network of poly acrylonitrile and gelatin for biomedical applications
    Journal of Materials Science: Materials in Medicine, 2009
    Co-Authors: Era Jain, Akshay Srivastava, Ashok Kumar
    Abstract:

    Cryogels are supermacroporous gel network formed by Cryogelation of appropriate monomers or polymeric precursors at subzero temperature. The beneficial feature of this system is a unique combination of high porosity with adequate mechanical strength and osmotic stability, due to which they are being envisaged as potential scaffold material for various biomedical applications. One of the important aspect of Cryogel is simple approach by which they can be synthesized and use of aqueous solvent for their synthesis which make them suitable for different biological applications. Various modifications of the Cryogels have been sought which involves coupling of various ligands to its surfaces, grafting of polymer chain to Cryogel surface or interpenetrating networks of two or more polymers to form a Cryogel which provides diversity of its applications. In the following work we have synthesized full interpenetrating network of polyacrylonitrile (PAN)-gelatin with varied gelatin concentration. The PAN-gelatin Cryogel interpenetrating network is macroporous in nature and has high percentage swelling equlibirium in the range of 862–1,200 with a flow rate greater than 10 ml/min, which characterizes the interconnectivity of pores and convective flow within the network. PAN-gelatin interpenetrating Cryogel network has good mechanical stability as determined by Young’s modulus which varies from 123 kPa to 819 kPa depending upon the polymer concentration. Moreover they are shown to be biocompatible and support cell growth within the scaffolds.

  • the physical characterization of supermacroporous poly n isopropylacrylamide Cryogel mechanical strength and swelling de swelling kinetics
    Materials Science and Engineering A-structural Materials Properties Microstructure and Processing, 2007
    Co-Authors: Akshay Srivastava, Era Jain, Ashok Kumar
    Abstract:

    Abstract Poly( N -isopropylacrylamide) [poly(NiPAAm)] and poly(acrylamide) [poly(AAm)] Cryogels were synthesized by radical polymerization at −12 °C for 12 h using monomers of N -isopropylacrylamide (NiPAAm) and acrylamide (AAm) with N , N -methylene bisacrylamide (MBAAm) as cross-linking agent, respectively. The Cryogels synthesized in freezing conditions provided spongy, elastic and supermacroporous character as compared to the hydrogels synthesized at ambient temperatures. Our earlier observations revealed that the elastic deformation of Cryogels either by external forces (mechanical deformation) or internal forces (shrinkage-swelling of poly(NiPAAm) Cryogels) led to detachment of affinity bound bioparticles to these gels, which promises great potential in understanding cell interactions on elastic matrices [M.B. Dainiak, A. Kumar, I.Y. Galaev, B. Mattiasson, Proc. Natl. Acad. Sci. U.S.A. 103 (2006) 849–854]. The deformation characteristic of Cryogels as measured by Young's modulus indicates that the modulus of elasticity of poly(NiPAAm) Cryogel (33–65 kPa) is comparatively lower than the Young's modulus for poly(AAm) Cryogel (42–86 kPa). The Young's modulus of both the Cryogels was found to be dependent on monomer concentration in Cryogels and increases with the increase in concentration. Thus, poly(AAm) Cryogel are mechanically more rigid than poly(NiPAAm) Cryogel. Further, the swelling/de-swelling kinetics study on poly(NiPAAm) Cryogel and hydrogel showed, higher swelling ratios for Cryogels in the range of 13–16 as compared to poly(NiPAAm) hydrogels which were in the range of 7–10. However, the extent of de-swelling is more in the case of poly(NiPAAm) hydrogels.

Junxian Yun - One of the best experts on this subject based on the ideXlab platform.

  • Strategy of Combining Prefiltration and Chromatography Using Composite Cryogels for Large-Scale Separation of Biotransformation Compounds from Crude High-Cell-Density Broth
    Industrial & Engineering Chemistry Research, 2015
    Co-Authors: Junxian Yun, Jie Liu, Shaochuan Shen, Songhong Zhang, Kejian Yao, Shan-jing Yao
    Abstract:

    The separation of interesting biomolecules or compounds from high-cell-density suspensions has received intensive attention in the biotechnology industry. In recent years, monolithic Cryogels have been suggested as a novel class of chromatographic adsorbents for the direct separation of biomolecules from such crude microbial feedstocks. However, the preparation of large-scale monolithic Cryogels for industrial applications and the direct separation of compounds from high-cell-density broths on an industrial scale are challenging tasks. In this work, a strategy for the separation of biotransformation compounds from high-cell-density broths was developed by combination of the prefiltration of microbial cells using large-scale Cryogels as the filter medium and chromatography followed using large-scale ion-exchange Cryogels as the adsorbents. Composite Cryogel disks (diameter of 143 mm) of poly(2-hydroxyethyl methacrylate) embedded with SiO₂ nanoparticles (pHEMA–SiO₂) and their anion-exchange supports grafted with 2-(dimethylamino)ethyl methacrylate (pHEMA–SiO₂–DMAEMA) were prepared successfully. As an example, the separation of cytidine triphosphate (CTP) from a crude high-cell-density broth containing 233 g/L Saccharomyces cerevisiae cells was carried out by the prefiltration of cells using a Cryogel bed packed with large-scale pHEMA–SiO₂ disks followed by anion-exchange chromatography using a packed bed of the large-scale pHEMA–SiO₂–DMAEMA Cryogel disks. The results showed that about half of the yeast cells have been removed successfully from the crude broth by the prefiltration using pHEMA–SiO₂ Cryogel disks, and CTP with a high purity of 98.2% and recovery of 98.3% were achieved by anion-exchange chromatography using pHEMA–SiO₂–DMAEMA Cryogel disks from the dissolved feedstock filtrated thereafter, indicating that the present strategy is effective and these large-scale composite Cryogels could be potentially applied as interesting filter media and chromatography adsorbents in industrial biotechnology and downstream processes.

  • microchannel liquid flow focusing and cryo polymerization preparation of supermacroporous Cryogel beads for bioseparation
    Journal of Chromatography A, 2012
    Co-Authors: Junxian Yun, Shaochuan Shen, Songhong Zhang, Kejian Yao, Dongqiang Lin, Yantao Guo, Yixin Guan, Shan-jing Yao
    Abstract:

    Abstract Polymeric Cryogels are sponge-like materials with supermacroporous structure, allowing them to be of interest as new chromatographic supports, cell scaffolds and drug carriers in biological and biomedical areas. The matrices of Cryogels are always prepared in the form of monoliths by cryo-polymerization under frozen conditions. However, there are limited investigations on the production of Cryogels in the form of adsorbent beads suitable for bioseparation. In this work, we provide a new approach by combining the microchannel liquid-flow focusing with cryo-polymerization for the preparation of polyacrylamide-based supermacroporous Cryogel beads with a narrow particle size distribution. The present method was achieved by introducing the aqueous phase solution containing monomer, cross-linker and redox initiators, and the water-immiscible organic oil phase containing surfactant simultaneously into a microchannel with a cross-shaped junction, where the aqueous drops with uniform sizes were generated by the liquid shearing and the segmentation due to the steady flow focusing of the immiscible phase streams. These liquid drops were in situ suspended into the freezing bulk oil phase for cryo-polymerization and the Cryogel matrix beads were obtained by thawing after the achievement of polymerization. By grafting the polymer chains containing sulfo binding groups onto these matrix beads, the cation-exchange Cryogel beads for protein separation were produced. The results showed that at the aqueous phase velocities from 0.5 to 2.0 cm/s and the total velocities of the water-immiscible phase from 2.0 to 6.0 cm/s, the obtained Cryogel beads by the present method have narrow size distributions with most of the bead diameters in the range from 800 to 1500 μm with supermacropores in sizes of about 3–50 μm. These beads also have high porosities with the averaged maximum porosity of 96.9% and the mean effective porosity of 86.2%, which are close to those of the polyacrylamide-based Cryogel monoliths. The packed bed using the Cryogel beads with mean diameter of 1248 μm, as an example, has reasonable and acceptable liquid dispersion, but high water permeability (4.29 × 10−10 m2) and high bed voidage (90.2%) owing to the supermacropores within the beads, enhanced the rapid binding and separation of protein from the feedstock even at high flow velocities. The purity of the obtained lysozyme from chicken egg white by one-step chromatography using the packed bed was in the range of about 78–92% at the flow velocities of 0.5–15 cm/min, indicating that the present Cryogel beads could be an effective chromatographic adsorbent for primary bioseparation.

  • an improved capillary model for describing the microstructure characteristics fluid hydrodynamics and breakthrough performance of proteins in Cryogel beds
    Journal of Chromatography A, 2011
    Co-Authors: Igor Yu Galaev, Bo Mattiasson, Junxian Yun, Gry Ravn Jespersen, Harald Kirsebom, Pererik Gustavsson
    Abstract:

    A capillary-based model modified for characterization of monolithic Cryogels is presented with key parameters like the pore size distribution, the tortuosity and the skeleton thickness employed for describing the porous structure characteristics of a Cryogel matrix. Laminar flow, liquid dispersion and mass transfer in each capillary are considered and the model is solved numerically by the finite difference method. As examples, two poly(hydroxyethyl methacrylate) (pHEMA) based Cryogel beds have been prepared by radical cryo-copolymerization of monomers and used to test the model. The axial dispersion behaviors, the pressure drop vs. flow rate performance as well as the non-adsorption breakthrough curves of different proteins, i.e., lysozyme, bovine serum albumin (BSA) and concanavalin A (Con A), at various flow velocities in the Cryogel beds are measured experimentally. The lumped parameters in the model are determined by matching the model prediction with the experimental data. The results showed that for a given Cryogel column, by using the model based on the physical properties of the Cryogel (i.e., diameter, length, porosity, and permeability) together with the protein breakthrough curves one can obtain a reasonable estimate and detailed characterization of the porous structure properties of Cryogel matrix, particularly regarding the number of capillaries, the capillary tortuousness, the pore size distribution and the skeleton thickness. The model is also effective with regards to predicting the flow performance and the non-adsorption breakthrough profiles of proteins at different flow velocities. It is thus expected to be applicable for characterizing the properties of Cryogels and predicting the chromatographic performance under a given set of operating conditions.

  • Preparation of Supermacroporous Composite Cryogel Embedded with SiO2 Nanoparticles
    Chinese Journal of Chemical Engineering, 2010
    Co-Authors: Yuchen Yao, Junxian Yun, Shaochuan Shen, Kejian Yao
    Abstract:

    Abstract Supermacroporous composite Cryogels embedded with SiO 2 nanoparticles were prepared by radical cryogenic copolymerization of the reactive monomer mixture of acrylamide (AAm) and N, N -methylene-bis-acrylamide (MBAAm) containing SiO 2 nanoparticles (mass ratios of nanoparticles to the monomer AAm from 0.01 to 0.08) under the freezing-temperature variation condition in glass columns. The properties of these composite Cryogels were measured. The height equivalent to theoretical plate (HETP) of the Cryogel beds at different liquid flow rates was determined by residence time distribution (RTD) using tracer pulse-response method. The composite Cryogel matrix embedded with the mass fraction of SiO 2 nanoparticles of 0.02 presented the best properties and was employed in the following graft polymerization. Chromatographic process of lysozyme in the composite Cryogel grafted with 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPSA) was carried out to evaluate the protein breakthrough and elution characteristics. The chromatography can be carried out at relatively high superficial velocity, i.e ., 15 cm·min −1 , indicating the satisfactory mechanical strength due to the embedded nanoparticles.

  • in situ graft polymerization preparation of cation exchange supermacroporous Cryogel with sulfo groups in glass columns
    Journal of Chromatography A, 2007
    Co-Authors: Kejian Yao, Junxian Yun, Shaochuan Shen, Fang Chen
    Abstract:

    Graft polymerization of monomer chains with expected functional groups onto the matrix pore surfaces by initiator is an effective approach for introducing ion-exchange groups to Cryogel matrix to get anion- or cation-exchange supermacroporous Cryogels. In this work, a novel cation-exchange Cryogel with sulfo binding groups was prepared by grafting of 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPSA) onto polyacrylamide-based Cryogels in glass columns. The grafting polymerization was achieved in an in-situ manner which was performed by pumping the initiator and the reactive solution of graft monomer with sulfo binding groups directly through a Cryogel bed pre-produced in a glass column under frozen condition. The axial liquid dispersion characteristics within the monolithic Cryogel beds before and after the in-situ polymerization were compared by measuring residence time distributions (RTDs) at various liquid flow rates using tracer pulse-response method. Microstructure morphology of pores within Cryogels was analyzed by scanning electron microscopy (SEM). Chromatography of lysozyme was carried out to reveal the protein breakthrough and elution characteristics in the obtained Cryogel beds.