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Enzyme Immobilization (enzyme + immobilization)
Selected AbstractsElectrospun Nanofibers Modified with Phospholipid Moieties for Enzyme ImmobilizationMACROMOLECULAR RAPID COMMUNICATIONS, Issue 16 2006Xiao-Jun Huang Abstract Summary: PANCMPCs containing phospholipid side moieties were electrospun into nanofibers with a mean diameter of 90 nm. Field emission SEM was used to characterize the morphologies of the nanofibers. These phospholipid-modified nanofibers were explored as supports for enzyme immobilization due to the characteristics of excellent biocompatibility, high surface/volume ratio, and porosity, which were beneficial to the catalytic efficiency and activity of immobilized enzymes. Lipase from Candida rugosa was immobilized on these nanofibers by adsorption. Preliminary results indicated that the properties of the immobilized lipase on these phospholipid-modified nanofibers were greatly promising. Schematic representation of the structure and electrostatic properties of phospholipid-modified nanofibers. [source] Plasma Polymer Surfaces Compatible with a CMOS Process for Direct Covalent Enzyme ImmobilizationPLASMA PROCESSES AND POLYMERS, Issue 1 2009Yongbai Yin Abstract Plasma polymerized surfaces, prepared using a CMOS compatible plasma enhanced chemical vapor polymerization technique, are found to covalently immobilize enzymes without the need for intermediate chemical linker groups. The polymerized surfaces are smooth, strongly adherent to substrates, and have a long shelf life for storage. After incubation with enzymes, a densely packed monolayer is attached. We report the effects of both oxygen etching and annealing post-processing showing that they can be implemented so as not to affect the enzyme binding performance. The fully compatible polymerization method with CMOS device manufacture processes is a potential candidate for integration into nano-CMOS biochemical sensors for direct immobilization of enzymes. [source] Enzyme immobilization on ultrafine cellulose fibers via poly(acrylic acid) electrolyte graftsBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2005Hong Chen Abstract Ultrafine cellulose fiber (diameter 200,400 nm) surfaces were grafted with polyacrylic acid (PAA) via either ceric ion initiated polymerization or methacrylation of cellulose with methacrylate chloride (MACl) and subsequent free-radical polymerization of acrylic acid. PAA grafts by ceric ion initiated polymerization increased with increasing reaction time (2,24 h), monomer (0.3,2.4 M), and initiator (1,10 mM) concentrations, and spanned a broad range from 5.5,850%. PAA grafts on the methacrylated cellulose fibers also increased with increasing molar ratios of MACl to cellulosic hydroxyl groups (MACl/OH, 2,6.4) and monomer acrylic acid (AA) to initiator potassium persulfate (KPS) ratios ([AA]/[KPS], 1.5,6), and were in a much narrower range between 12.8% and 29.4%. The adsorption of lipase (at 1 mg/ml lipase and pH 7) and the activity of adsorbed lipase (pH 8.5, 30°C), in both cases decreased with increasing PAA grafts. The highest adsorption and activity of the lipase on the ceric ion initiated grafted fibers were 1.28 g/g PAA and 4.3 U/mg lipase, respectively, at the lowest grafting level of 5.5% PAA, whereas they were 0.33 g/g PAA and 7.1 U/mg lipase, respectively, at 12.8% PAA grafts on the methacrylated and grafted fibers. The properties of the grafted fibers and the absorption behavior and activity of lipase suggest that the PAA grafts are gel-like by ceric-initiated reaction and brush-like by methacrylation and polymerization. The adsorbed lipase on the ceric ion-initiated grafted surface possessed greatly improved organic solvent stability over the crude lipase. The adsorbed lipases exhibited 0.5 and 0.3 of the initial activity in the second and third assay cycles, respectively. © 2004 Wiley Periodicals, Inc. [source] Enzyme immobilization via silaffin-mediated autoencapsulation in a biosilica supportBIOTECHNOLOGY PROGRESS, Issue 2 2009Wesley D. Marner II Abstract Enzymes and other biomolecules are often immobilized in a matrix to improve their stability or to improve their ability to be reused. Performing a polycondensation reaction in the presence of a biomolecule of interest relies on random entrapment events during polymerization and may not ensure efficient, homogeneous, or complete biomolecule encapsulation. To overcome these limitations, we have developed a method of incorporating autosilification activity into proteins without affecting enzymatic functionality. The unmodified R5 silaffin peptide from Cylindrotheca fusiformis is capable of initiating silica polycondensation in vitro at ambient temperatures and pressures in aqueous solution. In this study, translational fusion proteins between R5 and various functional proteins (phosphodiesterase, organophosphate hydrolase, and green fluorescent protein) were produced in Escherichia coli. Each of the fusion proteins initiated silica polycondensation, and enzymatic activity (or fluorescence) was retained in the resulting silica spheres. Under certain circumstances, the enzymatically-active biosilica displayed improved stability relative to free enzyme at elevated temperatures. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source] Covalent immobilization of ,-galactosidase on carrageenan coated with chitosanJOURNAL OF APPLIED POLYMER SCIENCE, Issue 1 2009Magdy M.M. Elnashar Abstract ,-Galactosidase was covalently immobilized to carrageenan coated with chitosan for the hydrolysis of lactose. The chitosan-carrageenan polyelectrolyte interaction was found to be dependent on the chitosan pH. At pH 4, the chitosan reached its maximum binding of 28.5% (w/w) where the chitosan surface density was 4.8 mg chitosan/cm2 g of carrageenan gel disks, using Muzzarelli method. Glutaraldehyde was used as a mediator to incorporate new functionality, aldehydic carbonyl group, to the bio-polymers for covalent attachment of ,-galactosidase. The enzyme was covalently immobilized to the biopolymer at a concentration of 2.73 mg protein per g of wet gel. FTIR proved the incorporation of the aldehydic carbonyl group to the carrageenan coated with chitosan at 1720 cm,1. The optimum time for enzyme immobilization was found to be 16 h, after which a plateau was reached. The enzyme loading increased from 2.65 U/g (control gel) to 10.92 U/g gel using the covalent technique. The gel's modification has shown to improve the carrageenan gel thermal stability as well as the immobilized enzyme. For example, the carrageenan gel treated with chitosan showed an outstanding thermal stability at 95°C compared with 35°C for the untreated carrageenan gel. Similarly, the immobilization process shifted the enzyme's optimum temperature from 50°C for the free enzyme towards a wider temperature range 45,55 °C indicating that the enzyme structure is strengthened by immobilization. In brief, the newly developed immobilization method is simple; the carrier is cheap, yet effective and can be used for the immobilization of other enzymes. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009 [source] Novel thermally and mechanically stable hydrogel for enzyme immobilization of penicillin G acylase via covalent techniqueJOURNAL OF APPLIED POLYMER SCIENCE, Issue 6 2008Magdy M. M. Elnashar Abstract ,-Carrageenan hydrogel crosslinked with protonated polyethyleneimine (PEI+) and glutaraldehyde (GA) was prepared and evaluated as a novel biocatalytic support for covalent immobilization of penicillin G acylase (PGA). The method of modification of the carrageenan biopolymer is clearly illustrated using a schematic diagram and was verified by FTIR, elemental analysis, DSC, and INSTRON using the compression mode. Results showed that the gels' mechanical strength was greatly enhanced from 3.9 kg/cm2 to 16.8 kg/cm2 with an outstanding improvement in the gels thermal stability. It was proven that, the control gels were completely dissolved at 35°C, whereas the modified gels remained intact at 90°C. The DSC thermogram revealed a shift in the endothermic band of water from 62 to 93°C showing more gel-crosslinking. FTIR revealed the presence of the new functionality, aldehydic carbonyl group, at 1710 cm,1 for covalent PGA immobilization. PGA was successfully immobilized as a model industrial enzyme retaining 71% of its activity. The enzyme loading increased from 2.2 U/g (control gel) to 10 U/g using the covalent technique. The operational stability showed no loss of activity after 20 cycles. The present support could be a good candidate for the immobilization of industrial enzymes rich in amino groups, especially the thermophilic ones. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008 [source] Chitosan-grafted poly(hydroxyethyl methacrylate- co -glycidyl methacrylate) membranes for reversible enzyme immobilizationJOURNAL OF APPLIED POLYMER SCIENCE, Issue 5 2007M. Yakup Ar Abstract Epoxy group-containing poly(hydroxyethyl methacrylate/glycidyl methacrylate), p(HEMA/GMA), membrane was prepared by UV initiated photopolymerization. The membrane was grafted with chitosan (CH) and some of them were chelated with Fe(III) ions. The CH grafted, p(HEMA/GMA), and Fe(III) ions incorporated p(HEMA/GMA)-CH-Fe(III) membranes were used for glucose oxidase (GOD) immobilization via adsorption. The maximum enzyme immobilization capacity of the p(HEMA/GMA)-CH and p(HEMA/GMA)-CH-Fe(III) membranes were 0.89 and 1.36 mg/mL, respectively. The optimal pH value for the immobilized GOD preparations is found to have shifted 0.5 units to more acidic pH 5.0. Optimum temperature for both immobilized preparations was 10°C higher than that of the free enzyme and was significantly broader at higher temperatures. The apparent Km values were found to be 6.9 and 5.8 mM for the adsorbed GOD on p(HEMA/GMA)-CH and p(HEMA/GMA)-CH-Fe(III) membranes, respectively. In addition, all the membranes surfaces were characterized by contact angle measurements. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 3084,3093, 2007 [source] Methods for stabilizing and activating enzymes in ionic liquids,a reviewJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 7 2010Hua Zhao Abstract Ionic liquids (ILs) have evolved as a new type of non-aqueous solvents for biocatalysis, mainly due to their unique and tunable physical properties. A number of recent review papers have described a variety of enzymatic reactions conducted in IL solutions; on the other hand, it is important to systematically analyze methods that have been developed for stabilizing and activating enzymes in ILs. This review discusses the biocatalysis in ILs from two unique aspects (1) factors that impact the enzyme's activity and stability, (2) methods that have been adopted or developed to activate and/or stabilize enzymes in ionic media. Factors that may influence the catalytic performance of enzymes include IL polarity, hydrogen-bond basicity/anion nucleophilicity, IL network, ion kosmotropicity, viscosity, hydrophobicity, the enzyme dissolution, and surfactant effect. To improve the enzyme's activity and stability in ILs, major methods being explored include the enzyme immobilization (on solid support, sol,gel, or CLEA), physical or covalent attachment to PEG, rinsing with n -propanol methods (PREP and EPRP), water-in-IL microemulsions, IL coating, and the design of enzyme-compatible ionic solvents. It is exciting to notice that new ILs are being synthesized to be more compatible with enzymes. To utilize the full potential of ILs, it is necessary to further improve these methods for better enzyme compatibility. This is what has been accomplished in the field of biocatalysis in conventional organic solvents. Copyright © 2010 Society of Chemical Industry [source] Stabilization of penicillin V acylase from Streptomyces lavendulae by covalent immobilizationJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 5 2001Jesús Torres-Bacete Abstract Penicillin,V acylase from the actinomycete Streptomyces lavendulae ATCC 13664 has been immobilized to epoxy-activated acrylic beads (Eupergit C®) by covalent binding. Further linkage of bovine serum albumin after enzyme immobilization was carried out in order to remove the remaining oxirane groups of the support. The obtained immobilized biocatalyst displayed double exponential deactivation kinetics at temperatures below 55,°C, while the native enzyme followed single exponential decay at the same temperatures. We concluded that soluble penicillin acylase was deactivated in one step, whereas the immobilized enzyme showed an enzymatic intermediate state which is highly thermostable. As a consequence of the immobilization process, the enzyme displayed a 10-fold increase in its half-life at 40,°C. At this temperature, the enzymatic intermediate state was progressively destabilized as the pH of the medium was increased. Thus, the optimum pH range for the immobilized enzyme preparation was established as being from 7.0 to 8.0. Higher pH values led to quicker enzyme deactivation. © 2001 Society of Chemical Industry [source] Synthesis of well-defined glycidyl methacrylate based block copolymers with self-activation and self-initiation behaviors via ambient temperature atom transfer radical polymerizationJOURNAL OF POLYMER SCIENCE (IN TWO SECTIONS), Issue 14 2007Ping Jiang Abstract Well-defined glycidyl methacrylate (GMA) based di- and triblock copolymers, with self-activation and self-initiation behaviors by incorporation of 2-(diethylamino) ethyl methacrylate (DEA) blocks, were synthesized via ambient temperature atom transfer radical polymerization (ATRP). The stability of the GMA pendant oxirane rings in tertiary amine environments at ambient temperature was investigated. More importantly, both self-activation behavior in oxirane ring opening addition reaction and self-initiation behavior in post-cure oxirane ring opening crosslinking of these block copolymers were evidenced by 1H NMR studies. The results demonstrated that the reactivity of pendent oxirane rings was strongly dependant on the nucleophilicity and steric hindrance of tertiary amine moieties and temperature. This facilitated the synthesis of well-defined block copolymers of GMA and DEA via sequential monomer addition ATRP, particularly for polymerization of GMA monomer at ambient temperature. Moreover, these one-component GMA based block polymers have novel self-activation and self-initiation properties, rendering some potential applications in both enzyme immobilization and GMA-based thermosetting materials. © 2007 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 45: 2947,2958, 2007 [source] Preparation and HPLC applications of rigid macroporous organic polymer monolithsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10-11 2004Frantisek Svec Abstract Rigid porous polymer monoliths are a new class of materials that emerged in the early 1990s. These monolithic materials are typically prepared using a simple molding process carried out within the confines of a closed mold. For example, polymerization of a mixture comprising monomers, free-radical initiator, and porogenic solvent affords macroporous materials with large through-pores that enable applications in a rapid flow-through mode. The versatility of the preparation technique is demonstrated by its use with hydrophobic, hydrophilic, ionizable, and zwitterionic monomers. Several system variables can be used to control the porous properties of the monolith over a broad range and to mediate the hydrodynamic properties of the monolithic devices. A variety of methods such as direct copolymerization of functional monomers, chemical modification of reactive groups, and grafting of pore surface with selected polymer chains is available for the control of surface chemistry. Since all the mobile phase must flow through the monolith, the convection considerably accelerates mass transport within the molded material, and the monolithic devices perform well, even at very high flow rates. The applications of polymeric monolithic materials are demonstrated mostly on the separations in the HPLC mode, although CEC, gas chromatography, enzyme immobilization, molecular recognition, advanced detection systems, and microfluidic devices are also mentioned. [source] Electrospun Nanofibers Modified with Phospholipid Moieties for Enzyme ImmobilizationMACROMOLECULAR RAPID COMMUNICATIONS, Issue 16 2006Xiao-Jun Huang Abstract Summary: PANCMPCs containing phospholipid side moieties were electrospun into nanofibers with a mean diameter of 90 nm. Field emission SEM was used to characterize the morphologies of the nanofibers. These phospholipid-modified nanofibers were explored as supports for enzyme immobilization due to the characteristics of excellent biocompatibility, high surface/volume ratio, and porosity, which were beneficial to the catalytic efficiency and activity of immobilized enzymes. Lipase from Candida rugosa was immobilized on these nanofibers by adsorption. Preliminary results indicated that the properties of the immobilized lipase on these phospholipid-modified nanofibers were greatly promising. Schematic representation of the structure and electrostatic properties of phospholipid-modified nanofibers. [source] Biocatalytic hydrogels by template polymerizationPOLYMERS FOR ADVANCED TECHNOLOGIES, Issue 5 2008H. El-Sherif Abstract Novel ionizable hydrogels were prepared from poly(acrylic acid) and dimethylaminoethyl methacrylate monomer employing template polymerization technique as an alternative to traditional physical and chemical crosslinking. The mode of interaction, as proved by Fourier Transform Infrared Spectroscopy (FTIR), was multiple H-bonding between the tertiary amino group of the monomer and the carboxylic groups of the polymer. The hydrogels represented suitable matrices for enzyme immobilization. The effect of varying the polymer,monomer molar ratio on the swelling kinetics and parameters was investigated. The dynamic swelling isotherm exhibited a Fickian mode of penetrant sorption and a plateau that increases with the amino group content. A polymer complex of molar ratio (polymer:monomer) 0.5:0.8 had a weight swelling ratio of 10 and 7 at pHs 3 and 8, respectively. The proven pH sensitivity together with the amphoteric character of these hydrogels make them good candidates for another bioapplication such as oral delivery systems of therapeutic peptides and proteins. The structural integrity of the hydrogels was proved by their swelling reversibility. , -Galactosidase, as an acidic model enzyme, was immobilized covalently on the synthesized hydrogels. The maximum enzyme velocity (Vmax) was enhanced to 19,µmol/min/mg, for polycomplex of molar ratio 0.5:0.8, compared with 3.2,µmol/min/mg for the free enzyme. Copyright © 2008 John Wiley & Sons, Ltd. [source] Efficient on-chip proteolysis system based on functionalized magnetic silica microspheresPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 14 2007Yan Li Abstract An easily replaceable enzymatic microreactor has been fabricated based on the glass microchip with trypsin-immobilized magnetic silica microspheres (MS microspheres). Magnetic microspheres with small size (,300,nm in diameter) and high magnetic responsivity to magnetic field (68.2,emu/g) were synthesized and modified with tetraethyl orthosilicate (TEOS). Aminopropyltriethoxysilane (APTES) and glutaraldehyde (GA) were then introduced to functionalize the MS microspheres for enzyme immobilization. Trypsin was stably immobilized onto the MS microspheres through the reaction of primary amines of the proteins with aldehyde groups on the MS microspheres. The trypsin-immobilized MS microspheres were then locally packed into the microchannel by the application of a strong field magnet to form an on-chip enzymatic microreactor. The digestion efficiency and reproducibility of the microreactor were demonstrated by using cytochrome c (Cyt-C) as a model protein. When compared with an incubation time of 12,h by free trypsin in the conventional digestion approach, proteins can be digested by the on-chip microreactor in several minutes. This microreactor was also successfully applied to the analysis of an RPLC fraction of the rat liver extract. This opens a route for its further application in top-down proteomic analysis. [source] Directed self-immobilization of alkaline phosphatase on micro-patterned substrates via genetically fused metal-binding peptideBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2009Turgay Kacar Abstract Current biotechnological applications such as biosensors, protein arrays, and microchips require oriented immobilization of enzymes. The characteristics of recognition, self-assembly and ease of genetic manipulation make inorganic binding peptides an ideal molecular tool for site-specific enzyme immobilization. Herein, we demonstrate the utilization of gold binding peptide (GBP1) as a molecular linker genetically fused to alkaline phosphatase (AP) and immobilized on gold substrate. Multiple tandem repeats (n,=,5, 6, 7, 9) of gold binding peptide were fused to N-terminus of AP (nGBP1-AP) and the enzymes were expressed in E. coli cells. The binding and enzymatic activities of the bi-functional fusion constructs were analyzed using quartz crystal microbalance spectroscopy and biochemical assays. Among the multiple-repeat constructs, 5GBP1-AP displayed the best bi-functional activity and, therefore, was chosen for self-immobilization studies. Adsorption and assembly properties of the fusion enzyme, 5GBP1-AP, were studied via surface plasmon resonance spectroscopy and atomic force microscopy. We demonstrated self-immobilization of the bi-functional enzyme on micro-patterned substrates where genetically linked 5GBP1-AP displayed higher enzymatic activity per area compared to that of AP. Our results demonstrate the promising use of inorganic binding peptides as site-specific molecular linkers for oriented enzyme immobilization with retained activity. Directed assembly of proteins on solids using genetically fused specific inorganic-binding peptides has a potential utility in a wide range of biosensing and bioconversion processes. Biotechnol. Bioeng. 2009;103: 696,705. © 2009 Wiley Periodicals, Inc. [source] Stabilization of glucose oxidase in alginate microspheres with photoreactive diazoresin nanofilm coatingsBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2005Rohit Srivastava Abstract The nanoassembly and photo-crosslinking of diazo-resin (DAR) coatings on small alginate microspheres for stable enzyme entrapment is described. Multilayer nanofilms of DAR with poly(styrene sulfonate) (PSS) were used in an effort to stabilize the encapsulation of glucose oxidase enzyme for biosensor applications. The activity and physical encapsulation of the trapped enzyme were measured over 24 weeks to compare the effectiveness of nanofilm coatings and crosslinking for stabilization. Uncoated spheres exhibited rapid loss of activity, retaining only 20% of initial activity after one week, and a dramatic reduction in effective activity over 24 weeks, whereas the uncrosslinked and crosslinked {DAR/PSS}-coated spheres retained more than 50% of their initial activity after 4 weeks, which remained stable even after 24 weeks for the two and three bilayer films. Nanofilms comprising more polyelectrolyte layers maintained higher overall activity compared to films of the same composition but fewer layers, and crosslinking the films increased retention of activity over uncrosslinked films after 24 weeks. These findings demonstrate that enzyme immobilization and stabilization can be achieved by using simple modifications to the layer-by-layer self-assembly technique. © 2005 Wiley Periodicals, Inc. [source] Water-in-ionic liquid microemulsion-based organogels as novel matrices for enzyme immobilizationBIOTECHNOLOGY JOURNAL, Issue 8 2010Ioannis V. Pavlidis Abstract The use of water-in-ionic liquid microemulsion-based organogels (w/IL MBGs) as novel supports for the immobilization of lipase B from Candida antarctica and lipase from Chromobacterium viscosum was investigated. These novel lipase-containing w/IL MBGs can be effectively used as solid phase biocatalysts in various polar and non-polar organic solvents or ILs, exhibiting up to 4.4-fold higher esterification activity compared to water-in-oil microemulsion-based organogels. The immobilized lipases retain their activity for several hours at 70°C, while their half life time is up to 25-fold higher compared to that observed in w/IL microemulsions. Fourier-transform infrared spectroscopy data indicate that immobilized lipases adopt a more rigid structure, referring to the structure in aqueous solution, which is in correlation with their enhanced catalytic behavior observed. [source] Amyloid fibrils as a nanoscaffold for enzyme immobilizationBIOTECHNOLOGY PROGRESS, Issue 1 2010Sarah M. Pilkington Abstract Amyloid fibrils are a misfolded state, formed by many proteins when subjected to denaturing conditions. Their constituent amino acids make them ideally suited as a readily functionalized nanoscaffold for enzyme immobilization and their strength, stability, and nanometer size are attractive features for exploitation in the creation of new bionanomaterials. We report successful functionalization of amyloid fibrils by conjugation to glucose oxidase (GOD) using glutaraldehyde. GOD retained activity upon attachment and successful cross-linking was determined using electrophoresis, centrifugation, sucrose gradient centrifugation, and TEM. The resulting functionalized enzyme scaffold was then incorporated into a model poly(vinyl alcohol) (PVOH) film, to create a new bionanomaterial. The antibacterial effect of the functionalized film was then tested on E. coli, the growth of which was inhibited, demonstrating the incorporation of GOD antibacterial activity into the PVOH film. The incorporation of the GOD-functionalized amyloid fibrils into PVOH provides an excellent ,proof of concept' model for the creation of a new bionanomaterial using a functionalized amyloid fibril scaffold. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [source] Production of Low-Lactose Milk by Means of Nonisothermal BioreactorsBIOTECHNOLOGY PROGRESS, Issue 5 2004Valentina Grano The effect of the immobilization time on the activity of immobilized ,-galactosidase from K. lactis was investigated. Six biocatalytic membranes, different only for the time of the enzyme immobilization, were obtained by using nylon membranes grafted with glycidyl methacrylate (GMA) and activated by hexamethylenediamine (HMDA) and glutaraldehyde (Glu), used as spacer and coupling agent, respectively. Comparison between the isothermal and nonisothermal yield of these biocatalytic membranes was carried out in the process of lactose hydrolysis in milk. All of the results, reported as a function of the immobilization time, have evidenced the influence of our variable parameter on the activity of the catalytic membranes. The membrane giving highest yield under isothermal and nonisothermal conditions was that obtained with 2 h of immobilization time. The industrial application of these membranes has been discussed in terms of percentage reduction of the production times. [source] Immobilization of ,-Galactosidase on Fibrous Matrix by Polyethyleneimine for Production of Galacto-Oligosaccharides from LactoseBIOTECHNOLOGY PROGRESS, Issue 2 2002Nedim Albayrak The production of galacto-oligosaccharides (GOS) from lactose by Aspergillus oryzae ,-galactosidase immobilized on cotton cloth was studied. A novel method of enzyme immobilization involving PEI,enzyme aggregate formation and growth of aggregates on individual fibrils of cotton cloth leading to multilayer immobilization of the enzyme was developed. A large amount of enzyme was immobilized (250 mg/g support) with about 90,95% efficiency. A maximum GOS production of 25,26% (w/w) was achieved at near 50% lactose conversion from 400 g/L of lactose at pH 4.5 and 40 °C. Tri- and tetrasaccharides were the major types of GOS formed, accounting for about 70% and 25% of the total GOS produced in the reactions, respectively. Temperature and pH affected not only the reaction rate but also GOS yield to some extend. A reaction pH of 6.0 increased GOS yield by as much as 10% compared with that of pH 4.5 while decreased the reaction rate of immobilized enzyme. The cotton cloth as the support matrix for enzyme immobilization did not affect the GOS formation characteristics of the enzyme under the same reaction conditions, suggesting diffusion limitation was negligible in the packed bed reactor and the enzyme carrier. Increase in the thermal stability of PEI-immobilized enzyme was also observed. The half-life for the immobilized enzyme on cotton cloth was close to 1 year at 40 °C and 21 days at 50 °C. Stable, continuous operation in a plug-flow reactor was demonstrated for about 3 days without any apparent problem. A maximum GOS production of 26% (w/w) of total sugars was attained at 50% lactose conversion with a feed containing 400 g/L of lactose at pH 4.5 and 40 °C. The corresponding reactor productivity was 6 kg/L/h, which is several-hundred-fold higher than those previously reported. [source] | ||||||||||||||||