• Journal of Sensor Science and Technology
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• v.8 no.3
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• pp.247-252
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• 1999

The oxygen electrode of a biosensor needs enzyme immobilized membrane and a dialysis membrane to measure the oxygen concentration that remains after an enzyme reacts with its substrate. Accodingly, a single-layer PVA laminated CTA/PCL membrane was developed as an oxygen biosensor electrode. The enzymes were immobilized on a cellulose triacetate/polycarprolactone membrane using the 1,1'-carbonyl diimidazole(CDI) method, and then laminated with polyvinyl alcohol, aldehyde and acid. The alcohol oxidase and PVA laminated CTA/PCL membrane was tested with various concentration of enzyme substrates using a Yellow Springs Instrument(YSI) oxygen sensor. Under 5-10mmol substrates produced $0.37{\sim}0.83{\mu}A$(r=0.995) currents, and ater 8 weeks the glucose oxidase activity remained at about 56%, while the other activities remained very low. A SEM indicated a smooth surface and tightly attached PVA on the enzyme-immobilized CTA/PCL membranes.

• Microbiology and Biotechnology Letters
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• v.9 no.4
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• pp.197-205
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• 1981

Penicillin amidase of Bacillus megaterium was recovered from the fermentation broth by adsorption on celite and immobilized by entrapping the adsorbed enzyme in acrylamide gel. The operational stability in column reactor was greatly increased by entrappment as compared with that of without entrappment. The optimum pH of the immobilized enzyme was 8.7 with broader activity profile than that of the free enzyme, while the most stable pH range appeared to be between pH 7.5 and 8.0. The optimum temperature was shifted to 5$0^{\circ}C$ from 45$^{\circ}C$ for the soluble enzyme. The values of Km and the inhibition constants for 6-APA( $K_{ia}$ ) and phenylacetic acid ( $K_{ip}$ ), were 4.55 mM, 36.5mM, and 10.5mM, respectively. No significant internal pore diffusion limitation was found since the value of effectiveness factor was 0.95. The operational half life in a column reactor at pH 8.0 was 6.8 days at 4$0^{\circ}C$ and 47 days at 3$0^{\circ}C$, whereas that of without entrappment was only 1 day and 4 days, respectively. The performance of a batch and a column reactor was also discussed with respect to the productivity. The results demonstrated that the entrappment of an adsorbed enzyme for the enhancement of the operational stability of the immobilized enzyme was useful especially when an extracellular enzyme was used.

• Microbiology and Biotechnology Letters
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• v.40 no.2
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• pp.163-167
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• 2012

Since D-xylose is not fermentable in Saccharomyces cerevisiae, its conversion to D-xylulose is required for its application in biotechnological industries using S. cerevisiae. In order to convert D-xylose to D-xylulose by way of an enzyme immobilized system, D-xylose isomerase (XI) of Escherichia coli was fused with 10-arginine tag (R10) at its C-terminus for the simple purification and immobilization process using a cation exchanger. The fusion protein XIR10 was overexpressed in recombinant E. coli and purified to a high purity by a single step of cation exchange chromatography. The purified XIR10 was immobilized to a cation exchanger via the electrostatic interaction with the C-terminal 10-arginine tag. Both the free and immobilized XIR10 exhibited similar XI activities at various pH values and temperatures, indicating that the immobilization to the cation exchanger has a small effect on the enzymatic function of XIR10. Under optimized conditions for the immobilized XIR10, D-xylose was isomerized to D-xylulose with a conversion yield of 25%. Therefore, the results of this study clearly demonstrate that the electrostatic immobilization of XIR10 via the interaction between the 10-arginine tag and a cation exchanger is an applicable form of the conversion of D-xylose to D-xylulose.

• Applied Biological Chemistry
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• v.26 no.4
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• pp.217-221
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• 1983

$\beta$-Galactosidase($\beta$-D-galactoside galactohydrolase, E.C. 3. 2. 1. 23) from E. coli K-12 CSH 36 was immobilized on porous cellulose beads which were previously activated with tannin and p-benzoquinone. Their general properties and applicational possibities were investigated. The most effective, enzyme immobilization was obtained when tannin and p-benzoquinone, pH 11.0, were used together as activation reagents and a period of 6 hours of activation. The optimum pH of $\beta$-galactosidase was 5.5 for free enzyme and 6. 0 for the immobilized enzyme, the optimum temperatures for native and immobilized enzyme were both $50^{\circ}C$. Kms of native $\beta$-galactosidase and immobilized enzyme for ONPG(o-nitrophenyl galactopyranoside) were about $4.0{\times}10^(-4)M$ and $7.5{\times}10^(-4)M$, respectively. In the case of tannin : p-benzoquinone activated cellulose beads, the immobilized enzyme retained over 80% of the initial enzyme activity after 20 runs, which is very promising result far a possible industrial application.

• Microbiology and Biotechnology Letters
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• v.13 no.2
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• pp.115-118
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• 1985

Rifamycin B oxidase converts rifamycin B to rifamycin S using oxygen as cosubstrate. Humnicola spp. (ATCC 20620) was treated with acetone and the cell powder was immobilized with cellulose acetate. The properties of the immobilized enzyme was examined. The optimum pHs of the immobilized and the free enzymes were 7.2. The optimum temperature of the immobilized enzyme was at 50-55$^{\circ}C$, which was 5$^{\circ}C$ higher than that of the free enzyme. The activities of the immobilized enzyme appeared less sensistive with respect to the changes of temperature and pH as compared to those of the free enzyme. Twenty percent of the enzyme activity was recovered when the enzyme was immobilized in 3mm beads. The storage stability was good below 4$0^{\circ}C$, but the activity decreased very rapidly above 5$0^{\circ}C$. The physical strength of the beads was good and was suitable as packing material in a three-phase enzyme reactor.

• KSBB Journal
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• v.15 no.1
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• pp.88-95
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• 2000

We have produced new-structured oligosaccharides using immoobilized mixed-enzyme reactor of destransucrase from Leuconostoc mesenteroides B-512FMCM and $\alpha$-amylase from Aspergillus oryzae. The reactors of immobilized mixed-exzyme beads were more efficient for the production of oligosaccharides than that of each immobilized enzyme bead in stirred-tank reactior(STR) or in packed-bed reactor(PCR). In continuous flow reactor, the immobilized mixed-enzyme bead in PBR was more stable than in STR, and 52% of initial yield was maintained for 200 hr. New structured-oligosaccharides (NOS) reduced the change of pH in the culture of Streptococcus mutans. It also showed an inhibitory effect on the growth of Staphylococcus aureus.

• KSBB Journal
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• v.6 no.2
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• pp.181-187
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• 1991

Physiological and morphological characteristics of Anabaena azollae cells immobilized in a synthetic polymer, polyvinyl(PV), were investigated. The cell density of the non-immersed PV foam reached 4.4mg Chl/g dry wt. PV foam. This is 8 times higher than that of PV-immobiliz action in immersed batch system. And MSX-induced ammonia productivity and the photosynthectic oxygen evolution activity are higher than that of free cells after short-term dark storage. Nitrogenase activity and thermostability of photosynthetic activity are also higher than that of free Anabaena cells after immobilization. Total hydrogen production reached to 1.6ml $H_2$ per reactor (total 4mg Chl) after 6 days.

• Microbiology and Biotechnology Letters
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• v.14 no.4
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• pp.285-291
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• 1986

Immobilized Candida lipolytica cells were prepared by entrapping the whole cells in calcium alginate gel. To enhance citric acid productivity, immobilized cells were Incubated with activation medium in fluidized-bed reactors. When the activation was done in batch operation, maximum citric acid productivity appeared in a much shorter time than in continuous operation. Activated immobilized cells were enhanced about 10-fold in citric acid production relative to non-activated immobilized cells. The productivity of citric acid was also influenced by bead size. When Immobilized cells were reacted in a fluidized-bed reactor with the same quantity of cells, the citric acid productivity was increased as the bead size was decreased.

• Microbiology and Biotechnology Letters
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• v.9 no.1
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• pp.35-44
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• 1981

Whole cell penicillin amidase of Escherichia coli was immobilized by entrapment in gelatin followed by extrusion and crosslinking with glutaraldehyde. The immobilized engyme preparation demonstrated the recovery yield of activity up to 70% and good stability during storage and operation. The half life of activity decay during the operation was estimated to be about 50 days. The optimum pH and temperature for both of immobilized and soluble enzyme are 8.5 and 5$0^{\circ}C$, respectively. No significant change was demonstrated in the effect of pH and temperature, but the increase in heat stability at high temperature was observed in the case of the immobilized enzyme. It was found that the plug flow reactor could be operated favorably since the pH drop along the column path due to tile reaction product was minimized by employing substrate solution with moderate buffer strength. The optimal condition of reactor operation was discussed with regard to the effect of substrate concentration and the residence time on the conversion efficiency and productivity.

• Current Research on Agriculture and Life Sciences
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• v.3
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• pp.166-173
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• 1985

Several enzyme immobilization methods has been compared for immobilization of pepsin. Carboxymethyl cellulose and diethylaminoethyl cellulose were activated with Hcl and with NaOH, and were used for immobilization of pepsin. Sepharose-4B was activated cyanogen bromide, and was used for immobilization of pepsin. Porous glass beads were derivatized with 3-aminopropyitrlethoxysilane and with succinicanhydride, and were used for immobilization of pepsin. The results abtained were summarized as follow, 1. 10 mg/gr. dry bead and 15mg/gr. dry bead of pepsin were absorbed to CM-cellulose and DEAE-cellulose, 20 mg/gr. dry bead and 27 mg/gr. dry bead were coupled to CM-cellulose and DEAE-cellulose with glutaraldehyde respectively. Enzyme yields were 22% and 24% of soluble pepsin. 2. 16 mg/gr. dry bead of pepsin was attached to cyanogen bromide activated sepharose-4B, 19mg/gr. dry bead was cross linked to the activated bead with glutaraldehyde. Immobilized enzyme activity was 23% of soluble pepsin. 3. 40 mg/gr. dry bead of pepsin was conjugated to the derivatized glass beads. Immobilized enzyme activity was 45% of soluble pepsin.