Noboru Kubota

Preparation of polyethylene hollow fiber membrane via thermally induced phase separation

High-density polyethylene (HDPE) hollow fiber membranes were prepared via thermally induced phase separation (TIPS). Two kinds of diluents such as diisodecyl phthalate (DIDP) and liquid paraffin (LP) were used in the preparation of membrane. In the case of DIDP, liquid–liquid phase separation occurred, while only the polymer crystallization occurred in the case of LP. In all cases, asymmetric structures with the smaller pores at the outer surface were obtained. Effects of polymer molecular weight, air gap distance, water bath temperature and kinds of diluents on the pore size and the water permeability were investigated. The use of the polymer with the higher molecular weight, the shorter air gap distance and the higher bath temperature were effective to obtain the higher water permeability. The membrane prepared with LP showed the lower water permeability than that with DIDP.

Radiation-induced graft polymerization is the key to develop high-performance functional materials for protein purification

We have described a preparation scheme for immobilizing polymer chains at a uniformly high density onto a microfiltration membrane. Highly efficient protein recovery was demonstrated by the results of the determination of breakthrough and elution curves. The three requirements of high rate, high capacity, and repeated use for the protein recovery were satisfied by ensuring the occurrence of convection, multilayer binding, and hydrophilization, respectively. In addition, easy scale-up to fabrication of a membrane module was verified on a small scale.

Preparation of a hydrophobic porous membrane containing phenyl groups and its protein adsorption performance

A novel porous membrane was prepared by radiation-induced graft polymerization of an epoxy-group-containing vinyl monomer, glycidyl methacrylate, and subsequent addition of phenol and water. A phenyl group was appended to the polymer chain grafted on to a porous polyethylene hollow-fibre membrane. The presence of a diol group together with the phenyl group was required to reduce non-selective adsorption of the protein. A bovine serum albumin (BSA) phosphate buffer solution containing 2 M ammonium sulfate was forced to permeate through the resultant hydrophobic porous membrane, 0.36 mm thick with a phenyl-group density of 1.3 mmol/g. The breakthrough curves of BSA overlapped independent of the residence time in the range 12–1.2 s because of negligible diffusional mass-transfer resistance. A lower phenyl-group density resulted in a higher recovery of BSA after a series of adsorption and elution steps.

Comparison of protein adsorption by anion-exchange interaction onto porous hollow-fiber membrane and gel bead-packed bed

Protein collection performance by anion-exchange interaction was compared between a single porous hollow-fiber membrane and a gel bead-packed bed under identical conditions from a viewpoint of flow resistance and breakthrough behavior. At a space velocity (SV) of 100 h−1, an ethanolamino-group-containing porous hollow-fiber membrane (EA-C fiber) exhibited the same dynamic binding capacity for bovine serum albumin as a bed charged with diethylaminoethyl-group-containing gel beads (DEAE-G bed). Because mass transfer of the protein was enhanced by convective flow of the protein solution through the pores of the EA-C fiber, the EA-C fiber exhibited a higher productivity than the DEAE-G bed over SV = 100 h−1.

Module performance of anion-exchange porous hollow-fiber membranes for high-speed protein recovery

Abstract A laboratory-scale module was fabricated by housing eight anion-exchange porous hollow-fiber membranes with an effective length of 8 cm in parallel to each other in a cartridge. The hollow fibers were prepared by radiation-induced graft polymerization of glycidyl methacrylate and subsequent chemical modifications using diethylamine and ethanolamine. The resultant hollow fibers had diethylamino and ethanolamino group densities of 2.1 and 1.2 mmol/g of the dry hollow fiber, respectively, with inner and outer diameters of 2.8 and 4.4 mm, respectively, in a wet state. A low operating pressure of 0.1 MPa in the module provided a flow-rate of 100 ml/min for a bovine serum albumin solution. The dynamic binding amount of bovine serum albumin for the module was 270 mg, irrespective of the flow-rate, for rates ranging from 10 to 100 ml/min. The dynamic binding amount for the module was equivalent to eight times that of a single hollow fiber with an identical effective length, which indicates a linear relationship between the module and the number of single fibers for dynamic binding amount.

Comparison of Two Convection-Aided Protein Adsorption Methods Using Porous Membranes and Perfusion Beads

We describe results which serve as a guide in the selection of protein recovery techniques using two new chromatographic methods based on anion‐exchange interaction, i.e., membrane and perfusion chromatographies, which involve enhancement of protein transport by convective flow. Bovine serum albumin solution was permeated through a functionalized porous hollow‐fiber membrane and a bed charged with functionalized porous (perfusion) beads of identical volume. The pores of the membrane are surrounded by grafted polymer chains immobilizing anion‐exchange groups, whereas the throughpores of the beads are surrounded by the diffusive pores at the periphery of which anion‐exchange groups are immobilized. An 8‐fold increase in throughput of the protein using the porous membrane, 37 mg of BSA/mL/min, was achieved at a low operating pressure of 0.1 MPa, compared to that using the perfusion bed.

Protein Adsorption and Elution Performances of Porous Hollow-Fiber Membranes Containing Various Hydrophobic Ligands

Phenyl and butyl groups as hydrophobic ligands for protein binding were appended to the poly(glycidyl methacrylate) chain grafted onto a pore surface of a porous polyethylene hollow‐fiber membrane with a pore size of 0.2 μm. A hydrophobic ligand density of the modified membranes from 0.5 to 2.5 mmol/g was obtained, while pure water flux of the hollow fibers was 80% that of the original hollow fiber. Favorable kinetics, wherein an increasing permeation rate provides an increasing binding rate of bovine serum albumin (BSA), were observed in the permeation mode because of the negligible diffusional mass‐transfer resistance of the protein to the hydrophobic ligand during convective transport. Equilibrium binding capacity of BSA in a phosphate buffer (0.07 M, pH 7.4) containing 2 M (NH4) 2SO4 was 30 mg/g of the modified hollow fibers. A −NH(CH2) 3CH3‐group‐containing membrane exhibited electrostatic interaction as well as hydrophobic interaction with BSA. A −OC6H5‐group‐containing membrane exhibited the highest elution percentage of 83% by permeating an (NH4) 2SO4‐free buffer.

Control of phenyl-group site introduced on the graft chain for hydrophobic interaction chromatography

Abstract A phenyl group was appended to the epoxy-group-containing polymer chain grafted onto the pore surface of a porous hollow-fiber membrane for hydrophobic interaction chromatography. For introduction of the phenyl group at a density sufficient for protein collection, glycidyl methacrylate (GMA) grafted membrane was immersed in 1 M phenol aqueous solution whose pH ranged from 9 to 13. A bovine serum albumin (BSA) buffer solution containing 2 M (NH 4 ) 2 SO 4 was forced to permeate through the resultant porous hydrophobic membrane (Ph fiber). The Ph fiber prepared at a lower pH provided a higher elution percentage of BSA when elution was performed with an (NH 4 ) 2 SO 4 -free buffer. Introduction of the phenyl group at a high density onto the upper part of the shrinking graft layer at a higher pH was demonstrated based on the determination of water breakthrough pressure of the dried Ph fiber, resulting in a lower elution percentage of BSA.

Repeated use of a hydrophobic ligand-containing porous membrane for protein recovery

Abstract A porous hollow-fiber membrane containing a phenyl group as a hydrophobic ligand was prepared by radiation-induced graft polymerization of glycidyl methacrylate, followed by successive ring-opening reactions with phenol and water. Bovine serum albumin (BSA) was bound to the ligand during permeation of a BSA solution in phosphate buffer containing 2M (NH4)2SO4 through the pores of the hollow fiber. Subsequent elution with an (NH4)2SO4-free buffer exhibited an elution percentage of 82%. Repeated cycles of adsorption and elution caused the accumulation of BSA on the pore surface, resulting in a decrease in the binding capacity of BSA with increasing number of cycles. In contrast, by permeating 1 M NaOH after each elution, the binding capacity of BSA was maintained even after ten cycles. This alkaline regeneration was found to be effective in ensuring repeated use of the phenyl-group-containing porous membrane for recovery of proteins.

High-throughput hydrolysis of starch during permeation across α-amylase-immobilized porous hollow-fiber membranes

Abstract Two kinds of supporting porous membranes, ethanolamine (EA) and phenol (Ph) fibers, for immobilization of α -amylase were prepared by radiation-induced graft polymerization of an epoxy-group-containing monomer, glycidyl methacrylate, onto a porous hollow-fiber membrane, and subsequent ring-opening with EA and Ph, respectively. An α -amylase solution was forced to permeate radially outward through the pores of the EA and Ph fibers. α -Amylase was captured at a density of 0.15 and 6.6xa0g/L of the membrane by the graft chain containing 2-hydroxyethylamino and phenyl groups, respectively. A permeation pressure of 0.10xa0MPa provided a space velocity of 780 and 1500xa0h −1 for the α -amylase-immobilized EA and Ph fibers, respectively. Quantitative hydrolysis of starch during permeation of a 20xa0g/L starch solution in the buffer across the α -amylase-immobilized Ph fiber was attained up to a space velocity of about 2000xa0h −1 ; this was achieved because of negligible diffusional mass-transfer resistance of the starch to the α -amylase due to convective flow, whereas an enzyme reaction-controlled system was observed for the α -amylase-immobilized EA fiber.