Hybrid machine learning model based predictions for properties of poly(2-hydroxyethyl methacrylate)-poly(vinyl alcohol) composite cryogels embedded with bacterial cellulose.

Supermacroporous composite cryogels with enhanced adjustable functionality have received extensive interest in bioseparation, tissue engineering, and drug delivery. However, the variations in their components significantly impactfinal properties. This study presents a two-step hybrid machine learning approach for predicting the properties of innovative poly(2-hydroxyethyl methacrylate)-poly(vinyl alcohol) composite cryogels embedded with bacterial cellulose (pHEMA-PVA-BC) based on their compositions. By considering the ratios of HEMA (1.0-22.0 wt%), PVA (0.2-4.0 wt%), poly(ethylene glycol) diacrylate (1.0-4.5 wt%), BC (0.1-1.5 wt%), and water (68.0-96.0 wt%) as investigational variables, overlay sampling uniform design (OSUD) was employed to construct a high-quality dataset for model development. The random forest (RF) model was used to classify the preparation conditions. Then four models of artificial neural network, RF, gradient boosted regression trees (GBRT), and XGBoost were developed to predict the basic properties of the composite cryogels. The results showed that the RF model achieved an accurate three-class classification of preparation conditions. Among the four models, the GBRT model exhibited the best predictive performance of the basic properties, with the mean absolute percentage error of 16.04 %, 0.85 %, and 2.44 % for permeability, effective porosity, and height of theoretical plate (1.0 cm/min), respectively. Characterization results of the representative pHEMA-PVA-BC composite cryogel showed an effective porosity of 81.01 %, a permeability of 1.20 × 10-12 m2, and a range of height of theoretical plate between 0.40-0.49 cm at flow velocities of 0.5-3.0 cm/min. These indicate that the pHEMA-PVA-BC cryogel was an excellent material with supermacropores, low flow resistance and high mass transfer efficiency. Furthermore, the model output demonstrates that the alteration of the proportions of PVA (0.2-3.5 wt%) and BC (0.1-1.5 wt%) components in composite cryogels resulted in significant changes in the material basic properties. This work represents an attempt to efficiently design and prepare target composite cryogels using machine learning and providing valuable insights for the efficient development of polymers.

Hydrogenation of MTHPA to MHHPA over Ni-based catalysts: Al2O3 coating, Ru incorporation and kinetics.

Because of its excellent performance, methyl hexahydrophthalic anhydride (MHHPA) is a new anhydride-based epoxy resin curing agent after methyl tetrahydrophthalic anhydride (MTHPA). To improve the activity and stability of conventional RANEY® nickel catalysts in the catalytic hydrogenation of MTHPA to MHHPA reaction, RANEY® nickel encapsulated with porous Al2O3 and alumina-supported Ni-Ru bimetallic catalysts were designed and synthesized in this study. The physicochemical properties and surface reactions over the catalysts were characterized by N2 adsorption and desorption, X-ray diffraction (XRD), hydrogen temperature-programmed reduction/desorption (H2-TPR/TPD), X-ray photoelectron spectroscopy (XPS), scanning electronic microscopy (SEM), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR), and in situ diffuse reflectance infrared Fourier transformations spectroscopy (DRIFTS). The kinetic model of MTHPA hydrogenation over NiRu/Al was established and the parameters were estimated using the least-square method. The results showed that the encapsulation of porous Al2O3 on the surface of RANEY® nickel enhanced the stability of the Ni skeleton and the adsorption ability of the reactant molecules, which improved its activity for the hydrogenation reaction. The introduction of Ru improved the dispersion and stability of metallic Ni, which greatly increased the conversion ability towards MTHPA hydrogenation, but it had a trend to cause C[double bond, length as m-dash]C bond transfer at lower temperatures, increasing the hydrogenation difficulties. The kinetic results based on Ni-Ru bimetallic catalyst showed that the MTHPA hydrogenation reaction rate was first-order with respect to MTHPA concentration and 0.5-order with respect to hydrogen partial pressure, and the apparent activation energy of the hydrogenation reaction was 37.02 ± 2.62 kJ mol-1.

Association equilibrium model. I. Influence of pH and salt concentration on ion-exchanger.

An association equilibrium model is presented in this work to illustrate the charged state of an ion-exchange adsorbent in electrolytic solution. This semi-empirical model considers the adsorption equilibrium of hydrogen ions and small-molecular salt ions with adsorbents, and it can be used to describe the effects of pH and salt concentration on the zeta potential, associated hydrogen ions and ionic capacity of adsorbents. The association equilibrium parameters of four commercial adsorbents were obtained by experimental data fitting. The model fitted the experimental data well, and their coefficients of determination (R2) of four adsorbents ranged from 0.924 to 0.994. The ratio coefficients of the association reaction with hydrogen ions ranged from 0.15 to 0.44 and those with salt counter-ions were all one. These data demonstrated that association reactions followed stoichiometric law, but that ionizable groups on ion-exchangers could not freely ionize as small molecule ions in solution. In this way, the performance of ion-exchange adsorbents can be characterized based on the zeta potential and dissociated hydrogen ions, and the results from this model were consistent with that from the manufacturer. Furthermore, this model could easily be expanded for multi-component systems.

Chromatographic separation of phenyllactic acid from crude broth using cryogels with dual functional groups.

Phenyllactic acid (PLA) is an important organic acid with wide antimicrobial activities against gram-positive and gram-negative bacteria and some fungi. This interesting compound can be synthesized by the microbial fermentation or the bioconversion using phenylpyruvic acid (PPA) as the key substrate and microorganisms as the whole-cell biocatalysts. However, the isolation of high-purity PLA with a high recovery from the crude fermentation or conversion broth is a challenging task. In this work, the separation of PLA from the crude conversion broth prepared by employing Lactobacillus buchneri cells as the whole-cell catalysts was achieved by the chromatography using the poly(hydroxyethyl methacrylate) (pHEMA)-based cryogel with a combination of anion-exchange and hydrophobic benzyl groups. The static adsorption behaviors of PLA under different salt concentrations and the adsorption capacities of PLA on the cryogel were measured experimentally. The chromatographic performance of PLA from the crude conversion broth was compared with that from the clarified broth. The results showed that the pHEMA-based cryogel has a high capacity of PLA, i.e., 14.64 mg mL-1 cryogel, and the adsorption of PLA was influenced by the salt concentration. By using deionized water as running buffer, PLA with a high purity of 97.6% was obtained with one step elution using 0.3 M NaCl as the elution solution with the recovery at the range of 80.2-90.8% from crude feedstock without any pretreatment at various flow velocities. These values were close to those obtained for the clarified broth, i.e., the purity of 98.4% and the recovery of 92.3% under the same chromatography conditions at 1 cm min-1. The cryogel was then applied to separate PLA from clarified feedstock, high purity (>96.7%) and recovery (>91.4%) of PLA were found with 20 cycles, which verified the selectivity and robustness of prepared pHEMA-VBTAC cryogel. Therefore, the chromatography using pHEMA-based cryogel with the dual functional groups is an effective approach for the isolation of PLA directly from the crude bioconversion broth and thus could be interesting in the separation and production of high-purity PLA in industry.

Star-shaped polyester-based elastomers as an implantable delivery system for insulin: Development, pharmacokinetics, pharmacodynamics, and biocompatibility.

Elastomers are largely developed for biomedical applications; however, little is reported on them although they are an effective and controllable delivery system for proteins. In the present study, we investigated the pharmacokinetics, biosecurity, and hypoglycemic effect of an insulin-loaded elastomer formulation in diabetic rats. Cylindrical insulin-loaded elastomers were fabricated using a UV cross-linking process based on methyl-acrylic-star-poly(ε-caprolactone-co-D,L-lactide) cyclic ester and methyl-bi-acrylic-poly(ε-caprolactone-b-polyethylene glycol-b-ε-caprolactone) (CLPEGCLMA). The encapsulated insulin was well protected during the formulation. An in vitro pharmacokinetic study revealed that the rate of insulin release from the elastomers was affected by the hydrophilicity/hydrophobicity of the system and controlled by the CLPEGCLMA (hydrophilic prepolymer) composition. It was observed that insulin release followed the Higuchi model. In addition, the more hydrophilic elastomers showed higher degradation rates in vivo. Furthermore, in the pharmacodynamic study, all the elastomers, except those that contained star-poly(ε-caprolactone-co-D,L-lactide) (number-average molecular weight, Mn), polyethylene glycol (PEG) (kMn), ε-caprolactone/PEG (mol/mol), and CLPEGCLMA (weight, %) at a ratio of 3432:10:20:30, respectively, decreased blood glucose concentration and maintained it at a stable level. It was observed that the hypoglycemic effect of the drug-loaded elastomers was directly proportional to the rate of in vitro insulin release; however, emaciation was not observed. Moreover, elastomers play a positive role in biosecurity. Therefore, the elastomers might be effective carriers for the delivery of peptide drugs in the form of implants.

5-Aminobenzimidazole as new hydrophobic charge-induction ligand for expanded bed adsorption of bovine IgG.

Expanded bed adsorption (EBA) can capture target proteins directly from unclarified feedstock without prior solid-liquid separation. Hydrophobic charge-induction chromatography (HCIC) is a promising technology for biomolecule separation with high capacity, good selectivity and relatively low cost without the pretreatment of dilution or salt addition. In this work, EBA and HCIC were combined to develop a new separation technology, hydrophobic charge-induction EBA. Two HCIC ligands, 4-mercapto-ethyl-pyridine (MEP) and 5-aminobenzimidazole (ABI), were coupled onto agarose beads containing tungsten carbide to prepare the resins for EBA, named T-MEP and T-ABI, respectively. The static adsorption and dynamic binding behaviors of bovine IgG (bIgG) were investigated. Two resins had similar saturated adsorption capacities and salt-tolerant properties, but T-ABI showed higher dynamic binding capacity than T-MEP, indicating that ABI ligand was more suitable for EBA. The performances in expanded bed were verified. With the protein mixture (2mg/ml bIgG and 10mg/ml bovine serum albumin) as the model feedstock, the effects of loading and elution pH, expansion factor and loading volume on the separation performance of bIgG were evaluated. Finally, T-ABI EBA was used to separate bIgG directly from bovine whey with optimized operation conditions. The purity and recovery of bIgG reached 90.6% and 78.2%, respectively. The purification factor was about 19.3. The results demonstrated that the combination of HCIC and EBA would be a potential platform for antibody capture with less feedstock pretreatments, high efficiency and relatively low cost.

Chromatographic adsorption of serum albumin and antibody proteins in cryogels with benzyl-quaternary amine ligands.

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.

Poly(hydroxyethyl methacrylate)-based composite cryogel with embedded macroporous cellulose beads for the separation of human serum immunoglobulin and albumin.

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.

Isolation of immunoglobulin G from bovine milk whey by poly(hydroxyethyl methacrylate)-based anion-exchange cryogel.

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.

Rapid freezing cryo-polymerization and microchannel liquid-flow focusing for cryogel beads: adsorbent preparation and characterization of supermacroporous bead-packed bed.

Cryogel beads, fabricated by the microchannel liquid-flow focusing and cryo-polymerization method, have micron-scale supermacropores allowing the passage of crude feedstocks, and could be of interest as chromatographic adsorbents in bioseparation applications. In this work, we provide a rapid freezing and continuous formation method for cryogel beads by cryo-polymerization using dry ice particles as the freezing source and microchannel liquid-flow focusing using peristaltic pumps for the fluid supply. Polyacrylamide (pAAm)-based supermacroporous cryogel beads were prepared and grafted with N,N-dimethylaminoethyl methacrylate (DMAEMA), which provided the anion-exchange cryogel beads with tertiary amine functional groups suitable for binding proteins. Properties of the supermacroporous cryogel-bead packed bed, i.e., permeability, bed voidage, protein breakthrough as well as protein adsorption performance by using bovine γ-globulin as model protein, were experimentally investigated. A capillary-based model was employed to characterize the supermacroporous bed performance, and gave a reasonable description of the microstructure and thus an insight into the flow, dispersion and mass transfer behaviors within the cryogel bead-packed bed. The results also showed that by using dry ice as the freezing source, it is easy to reduce the temperature below -55 to -61°C in the bulk solution, causing the rapid formation of ice crystals within the monomer drops, and finally effective cryo-polymerization to form supermacropores within the cryogel beads. By using peristaltic pumps, continuous preparation was achieved and the obtained cryogel beads had favorable properties similar to those prepared using syringe pumps in the microchannel liquid-flow focusing process. This method is thus expected to be interesting in the liter- or even larger-scale preparation of cryogel adsorbents.

[Chromatographic separation of plasmid DNA by anion-exchange cryogel].

Plasmid DNA (pDNA) is used as an important vector for gene therapy, and its wide application is restricted by the purity and yield. To obtain high-purity pDNA, a chromatographic method based on anion-exchange supermacroporous cryogel was explored. The anion-exchange cryogel was prepared by grafting diethylaminoethyl-dextran to the epoxide groups of polyacrylamide-based matrix and pUC19 plasmid was used as a target to test the method. The plasmid was transferred into Escherichia coli DH5alpha, cultivated, harvested and lysed. The obtained culture was centrifuged and the supernatant was used as the plasmid feedstock, which was loaded into the anion-exchange cryogel bed for chromatographic separation. By optimizing the pH of running buffer and the elution conditions, high-purity pDNA was obtained by elution with 0.5 mol/L sodium chloride solution at pH 6.6. Compared to the traditional methods for purification of pDNA, animal source enzymes and toxic reagents were not involved in the present separation process, ensuring the safety of both the purification operations and the obtained pDNA.

Microchannel liquid-flow focusing and cryo-polymerization preparation of supermacroporous cryogel beads for bioseparation.

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 × 10⁻¹° 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.

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.

Modeling of protein breakthrough performance in cryogel columns by taking into account the overall axial dispersion.

A model considering the overall axial dispersion for describing protein adsorption and breakthrough in monolithic cryogel beds has been developed. The microstructure of cryogels was characterized by tortuous capillaries with a normal diameter distribution but a constant pore wall thickness. The axial dispersion within cryogel columns was described by using the overall axial dispersion coefficient, which can be easily obtained by matching the experimental breakthrough curves without adsorption or measuring residence time distributions (RTDs). Experimental breakthrough curves of lysozyme within a metal-chelated affinity cryogel by Persson et al. (Biotechnol. Bioeng. 2004, 88, 224-236) and a cation-exchange cryogel by Yao et al. (J. Chromatogr. A 2007, 1157, 246-251) were employed as examples to test the model. The results showed that by using the axial dispersion coefficient and assuming uniform radial concentration profile at a given cross-section of the cryogel along the bed height, the model can describe the detailed behaviors of the in-bed overall axial dispersion, the in-pore mass transfer, as well as the protein adsorption and breakthrough. For a known overall axial dispersion coefficient, the lumped parameter of the mass transfer coefficient between the bulk liquid and the capillary wall can be determined by fitting the protein breakthrough curve at a known chromatographic condition. Once this parameter is determined, the model can be used to predict the protein breakthrough profiles under different conditions based on the basic physical parameters of the cryogel bed and the properties of the fluid and protein. The effective capillary diameters employed in the model are close to the actual pore sizes observed from the images by SEM. The model predictions of lysozyme breakthrough profiles at various flow rates are also in good agreement with the experimental data in both the metal-chelated affinity and cation-exchange cryogel columns.

Isolation of ATP from a yeast fermentation broth using a cryogel column at high flow velocities.

This communication presents an effective method for isolating adenosine triphosphate (ATP) from a yeast fermentation broth using an anion-exchange supermacroporous cryogel column at high flow velocities. The breakthrough and elution behaviors of pure ATP in the cryogel bed were investigated at flow velocities of 2, 5, and 10 cm/min and the ATP binding capacities were determined. Then the ATP-containing yeast fermentation broth was employed as the test feedstock and various chromatographic runs were conducted to isolate ATP by the cryogel at different high flow velocities. The ATP samples obtained were analyzed quantitatively by HPLC. The results showed that even at a flow velocity of 5 or 10 cm/min, a product purity of 97.4 or 98.0% can be achieved, illustrating the potential of the present method for separation of high-purity ATP directly from fermentation feedstock at high flow velocities.

Chromatographic separation of cytidine triphosphate from fermentation broth of yeast using anion-exchange cryogel.

A novel separation method was developed to isolate directly cytidine triphosphate (CTP) from fermentation broth of yeast using anion-exchange supermacroporous cryogel. The anion-exchange cryogel with tertiary amine groups was prepared by graft polymerization. The breakthrough characteristics and elution performance of pure CTP in the cryogel bed were investigated experimentally and the CTP binding capacity was determined. Then the separation experiments of CTP from crude fermentation broth of yeast using the cryogel column were carried out using deionized water and 0.01 M HCl as washing buffer, respectively. The chromatographic behavior was monitored and analyzed. The purity and concentration of the obtained CTP in these processes were determined quantitatively by HPLC. The maximal purity of CTP obtained at the condition of 0.01 M HCl as washing buffer and 0.5 M NaCl in 0.01 M HCl as elution buffer reached 93%.

One-step isolation of adenosine triphosphate from crude fermentation broth of Saccharomyces cerevisiae by anion-exchange chromatography using supermacroporous cryogel.

Adenosine triphosphate (ATP) is an important high-energy compound widely used in biological and therapeutic fields. It can be produced by phosphorylation of adenosine monophosphate (AMP) with microbial cells in industrial scale and the effective isolation of ATP from microbial fermentation broth is a challenging work. In this work, we develop a novel one-step method to directly separate ATP from fermentation broth of Saccharomyces cerevisiae by anion-exchange chromatography using supermacroporous cryogel. The cryogel bed with tertiary amine groups was prepared by grafting N,N-dimethylaminoethyl methacrylate (DMAEMA) monomer chains onto the matrix of a polyacrylamide-based cryogel in a glass column and its properties of liquid dispersion, water permeability, porosity as well as the ligand density were measured. Chromatographic separation of ATP from the fermentation broth by the cryogel was carried out using deionised water and 0.01 M HCl as running buffer, respectively. The breakthrough characteristics and elution performance in the cryogel bed were revealed and analyzed. The purities of the obtained ATP were analyzed quantitatively by high performance liquid chromatography (HPLC). The maximal purity of ATP by the one-step separation method was 95.5% using 0.01 M HCl as running buffer in this work. The corresponding chromatographic behaviors were investigated and analyzed.

In-situ graft-polymerization preparation of cation-exchange supermacroporous cryogel with sulfo groups in glass columns.

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.

Characterization of a novel continuous supermacroporous monolithic cryogel embedded with nanoparticles for protein chromatography.

A novel continuous supermacroporous monolithic cryogel embedded with nanometer-size particles was prepared by the radical cryogenic co-polymerization of acrylamide (AAm), N,N'-methylene-bis-acrylamide (MBAAm), allyl glycidyl ether (AGE) and the dispersed surfactant-stabilized Fe3O4 nanoparticles under the freezing-temperature variation condition in a glass column. This special separation matrix has interconnected supermacropores with pore size of 10-50 microm, which permit the free-passage of microbial cells or cell debris in the culture fluids and then is interest in downstream processes. The axial liquid dispersion coefficients of the new continuous supermacroporous monolithic bed at different liquid flow rates were obtained by measuring residence time distributions (RTDs) using tracer pulse-response method. The experimental results showed that the axial liquid dispersion within the bed was weak in a wide water flow rate of 0.5-15 cm/min. The axial dispersion coefficient was found to be increased exponentially with the increase of liquid flow rate. Chromatographic process of bovine serum albumin (BSA) in the cryogel monolithic bed was carried out to reveal the protein breakthrough and elution characteristics. Compared with other reported cryogel beds in literature, the protein adsorption capacity of the present cryogel bed was improved due to the embedded nano-sized solid adsorbents in the gel matrix. Microstructure morphology of the embedded nanoparticles in the cryogel and the gel matrix structure were also analyzed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) in this paper.

Predictive modeling of protein adsorption along the bed height by taking into account the axial nonuniform liquid dispersion and particle classification in expanded beds.

Expanded bed adsorption (EBA) is a special chromatography technique with perfect classification of adsorbent particles in the column, thus the performance of protein adsorption in expanded beds is particular, obviously nonuniform and complex along the column. Detailed description of the complex adsorption kinetics of proteins in expanded bed is essential for better analyzing of adsorptive mechanisms, the design of chromatographic processes and the optimization of operation parameters of EBA processes. In this work, a theoretical model for the prediction of protein adsorption kinetics in expanded beds was developed by taking into account the classified distribution of adsorbent particles along the bed height, the nonuniform behaviors of axial liquid dispersion, the axial variation of local bed voidage as well as the axial changes of target component mass transfer. The model was solved using the implicit finite difference scheme combining with the orthogonal collocation method, and then applied to predict the breakthrough behaviors of bovine serum albumin (BSA) on Streamline DEAE and lysozyme on Streamline SP along the bed height in expanded beds under various conditions. In addition, the experiments of front adsorption of BSA on Streamline DEAE at different axial column positions were carried out to reveal the adsorption kinetics of BSA along the bed height in a 20 mm I.D. expanded bed, and the influences of liquid velocity and feed concentration on the breakthrough behaviors were also analyzed. The breakthrough behaviors predicted by the present model were compared with the experimental data obtained in this work and in the literature published. The agreement between the prediction and the experimental breakthrough curves is satisfied.