• Title/Summary/Keyword: Enzyme immobilization

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Fabrication and Response Characteristics of the Light Addressable Potentiometric Sensor for Detecting the Penicillin Concentration (페니실린 농도 검출을 위한 광지시형 전위차 센서의 제작 및 감응특성)

  • Jang, Su-Won;Jung, Young-Hee;Park, Jin-Ho;Kim, Jea-Ho;Kwon, Dae-Hyuk;Lee, Seung-Ha;Kang, Shin-Won
    • Journal of Sensor Science and Technology
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    • v.13 no.5
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    • pp.356-362
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    • 2004
  • In this study, we developed the measuring system based on light addressable potentiometric sensor for the quantitative analysis of penicillin that is very important element in medicine and pharmacy, clinic. It is investigated the response characteristics by enzyme reaction with penicillinase. First, the surface pre-treatment process of the $Si_{3}N_{4}$ was established. The coupling agent was made using self assembled monolayer method and it was confirmed the immobilization process by AFM. Also, as the measuring system, potentiostat, signal processing part etc. was made by Lab VIEW software, it was reduced detecting time as well as simplifying the system. Fabricated device was shown excellent pH response characteristics, 57 mV/ pH in the range of pH $2{\sim}11$. The response characteristics was 60 mV/decade in the range of $0.1{\sim}10{\;}mM$.

Biomimetic sequestration of $CO_2$ and reformation to $CaCO_3$ using bovine carbonic anhydrase immobilized on SBA-15 (생체모방공학을 이용한 bovine carbonic anhydrase를 SBA-15에 고정화하여 이산화탄소분리와 재구성된 $CaCO_3$ 연구)

  • Vinoba, Mari;Kim, Dae-Hoon;Lim, Kyoung-Soo;Jeong, Soon-Kwan;Alagar, Muthukaruppan
    • Proceedings of the KAIS Fall Conference
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    • 2010.11a
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    • pp.495-499
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    • 2010
  • The biocatalytic capture of $CO_2$, and its precipitationas $CaCO_3$, over bovine carbonic anhydrase (BCA) immobilized on a pore-expanded SBA-15 support was investigated. SBA-15 was synthesized using TMB as a pore expander, and the resulting porous silica was characterized by XRD, BET, IR, and FE-SEM analysis. BCA was immobilized on SBA-15 through various approaches, including covalent attachment (BCA-CA), adsorption (BCA-ADS), and cross-linked enzyme aggregation (BCA-CLEA). The immobilization of BCA on SBA-15 was confirmed by the presence of zinc metal in the EDXS analysis. The effects of pH, temperature, storage stability, and reusability on the biocatalytic performance of BCA were characterized by examining para-nitrophenyl acetate (p-NPA) hydrolysis. The $K_{cat}/K_m$ values for p-NPA hydrolysis were 740.05, 660.62, and $680.11M^{-1}s^{-1}$, respectively, where as $K_{cat}/K_m$ for free BCA was $873.76M^{-1}s^{-1}$. The amount of $CaCO_3$ precipitate was measured quantitatively using anion-selective electrode and was found to be 12.41, 11.82, or 11.28 mg $CaCO_3$/mg for BCA-CLEA, BCA-ADS, or BCA-CA, respectively. The present results indicate that the immobilized BCA-CLEA, BCA-ADS, and BCA-CA are green materials, and are tunable, reusable, and promising biocatalysts for $CO_2$ sequestration.

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Simultaneous Determination of Glucose and Ethanol of Takju by Biosensor using Dual Cathode Electrode (Dual Cathode Electrode를 이용한 바이오센서로 탁주 중의 포도당 및 에탄올의 동시 측정)

  • Park, In-Seon;Kim, Jung-Ho;Kim, Tae-Jin;Kim, Nam-Soo;Noh, Bong-Soo
    • Korean Journal of Food Science and Technology
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    • v.28 no.5
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    • pp.974-980
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    • 1996
  • A biosensor was prepared with dual cathode electrode and immobilized enzyme membrane. A nylon net was used for the immobilization of glucose oxidase and alcohol oxidase. The immobilized enzymes were placed on the surface of the electrode which was prepared with one anode and two cathodes as an oxygen electrode. The determination of components by the biosensor was based on the consumption of dissolved oxygen. The optimum condition of this system was 0.1 M potassium phosphate buffer solution, pH 7.5 at $35^{\circ}C$. Glucose and ethanol in takju were simultaneously determined by the biosensor. Comparing with UV-spectrophotometer and gas chromatograph for cross checking, there was a good correlation between the biosensor and the conventional methods. Biosensor with dual cathode electrode required no clarification or pretreatments. It was used for simultaneous determination of glucose and ethanol during the fermentation of takju.

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Production of casein phosphopeptides using Streptococcus faecalis var. liquefaciens cell immobilization (Streptococcus faecalis var. liquefaciens 전세포 고정화법을 이용한 Casein Phosphopeptides 생산)

  • Lee, Ki-Sung;Shin, Jae-Yoon;Jang, Yi-Hyun;Kweon, Dae-Hyuk;Park, Ki-Moon;Jin, Yong-Su
    • KSBB Journal
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    • v.23 no.1
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    • pp.59-64
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    • 2008
  • Optimum conditions for production of casein phosphopeptides (CPP) from sodium casenate by immobilized cell culture of Streptococcus faecalis var. liquefaciens were investigated. Immobilized cells were made by mixing 60% sodium alginate solution with an equal volume of culture broth at the end of exponential phase and subsequently dropping the mixture into $CaCl_{2}$ solution. Optimum conditions for CPP production by the immobilized cells were the same as those ($50^{\circ}C$, pH 7.0, and 10% substrate concentration) by the crude enzyme solution from the supernatant of culture broth. Optimum loading volume of the immobilized cells into a batch reactor was 30% (w/v). Using a continuous reactor loaded by the immobilized cells under the identified optimal conditions, we were able to produce CPP continuously up to 30 days with a maximum CPP conversion efficiency of 20%.

An Aptamer-Based Electrochemical Sensor That Can Distinguish Influenza Virus Subtype H1 from H5

  • Lee, Jin-Moo;Kim, JunWon;Ryu, Ilhwan;Woo, Hye-Min;Lee, Tae Gyun;Jung, Woong;Yim, Sanggyu;Jeong, Yong-Joo
    • Journal of Microbiology and Biotechnology
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    • v.27 no.11
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    • pp.2037-2043
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    • 2017
  • The surface protein hemagglutinin (HA) mediates the attachment of influenza virus to host cells containing sialic acid and thus facilitates viral infection. Therefore, HA is considered as a good target for the development of diagnostic tools for influenza virus. Previously, we reported the isolation of single-stranded aptamers that can distinguish influenza subtype H1 from H5. In this study, we describe a method for the selective electrical detection of H1 using the isolated aptamer as a molecular probe. After immobilization of the aptamer on Si wafer, enzyme-linked immunosorbent assay (ELISA) and field emission scanning electron microscopy (FE-SEM) showed that the immobilized aptamer bound specifically to the H1 subtype but not to the H5 subtype. Assessment by cyclic voltammetry (CV) also demonstrated that the immobilized aptamer on the indium thin oxide-coated surface was specifically bound to the H1 subtype only, which was consistent with the ELISA and FE-SEM results. Further measurement of CV using various amounts of H1 subtype provided the detection limit of the immobilized aptamer, which showed that a nanomolar scale of target protein was sufficient to produce the signal. These results indicated that the selected aptamer can be an effective probe for distinguishing the subtypes of influenza viruses by monitoring current changes.

Preparation of Enzyme Electrodes for Biofuel Cells Based on the Immobilization of Glucose Oxidase in Polyion Complex (폴리이온복합체를 이용하여 글루코스 산화효소를 고정화한 바이오전지용 효소전극 제조)

  • Nguyen, Linh Thi My;Li, Nan;Yoon, Hyon Hee
    • Applied Chemistry for Engineering
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    • v.24 no.1
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    • pp.99-103
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    • 2013
  • An emzymatic bioanode for a glucose/oxygen biofuel cell was prepared by the sequential coating of carbon nanotube (CNT), charge transfer complex (CTC) based on tetracyanoquinodimethane (TCNQ) and tetrathiafulvalene (TTF), glucose oxidase (GOx), and polyion complex (mixture of poly-L-lysine hydrobromide and poly (sodium 4-styrenesulfonate)) on a glassy carbon electrode. A biocathode was also prepared by the sequential coating of CNT, bilirubin oxidase (BOD), 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), and polyion complex. The effect of CNT and CTC on the electrochemical performance was investigated. The biofuel cell exhibited a promising performance with maximum power densities of 3.6, 10.1, and $46.5{\mu}W/cm^2$ at 5, 20, and 200 mM of glucose concentration, respectively. The result indicates that the biofuel cell architecture prepared in this study can be used in the development of biofuel cells and biosensors.

Sorbitol Production by Zymomonas mobilis Immobilized in Calcium Alginate Gels and Glutaraldehyde (알지네이트 및 글루타르알데하이드 고정화 Zymomonas mobilis에 의한 쏠비톨 생산)

  • Jung, In-Ho;Choi, Do-Jin;Park, Cheol-Jin;Chun, Uck-Han
    • Korean Journal of Food Science and Technology
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    • v.22 no.7
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    • pp.812-816
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    • 1990
  • This study describes the sorbitol production with permeabilized cells of Zymomonas mobilis immobilized in Ca-alginate. Toluene treated cells lose activity of glucose-fructose oxidoreductase due to the leaking of enzyme from the cells. To prevent this leakage, the permeabilized cells were treated with 0.25% glutaraldehyde by stirring for 1 h at room temperature. A continuous process with glutaraldehyde treated cells was developed and no significant reduction in the degree of conversion occurred during 210 h operation. The productivities were estimated to be about $7.2{\sim}7.5\;g/l-h$ for sorbitol at dilution rate $0.18\;h^{-1}$.

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Radiation Technology in the Preparation of Polyethylene Oxide Hydrophilic Gels and Immobilization of Proteases for Use in Medical Practice

  • E.I.Vereschagin;Han, Do-Hung;A.W.Troitsky;O.V.Grishin;S.E.Petrov;E.P.Gulyaeva;L.A.Bogdanova;M.V.Korobeinikov;V.L.Auslender
    • Archives of Pharmacal Research
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    • v.24 no.3
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    • pp.229-233
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    • 2001
  • This Paper deals with the development of a technology for making a hydrophilic gel of Polyethylene oxide reception in which radiating ability is employed to cause cross-linking of Polymers in a water solution. The gel of polyethylene oxide was shown to be nontoxic contain 5-50% of polymer and be useful in composite medicinal forms along with biologically active substances including Bac. subtilis proteases. Proteases immobilized in the gel possess high thermal stability and proteolytic activity and are readily applied in medicine. The effect of immobilized proteolytic and glucolytic enzymes of Bac. subtillis (Immozimase) on the warm ischemia-reperfusion (I/R) which can cause hepatic and jejunum injury was also studied. These enzymes were immobilized on water-soluble polymer polyethylene glycol by means of an electron beam. The number of degraanulated mast cells as well as serum ALT after I/R in the group with Immozimase was decreased to almost half as compared with the control group. Pretreatment with Immozimase resulted in significant reduction of hepatic and gut neutrophil accumulation as compared with control animals. It was concluded that Immozimase has a protective effect for hepatic and gut ischemia/reperfusion, and this effect seems to be associated with prevention of leukocyte accumulation .

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Optimization of Culture Medium for Novel Cell-Associated Tannase Production from Bacillus massiliensis Using Response Surface Methodology

  • Belur, Prasanna D.;Goud, Rakesh;Goudar, Dinesh C.
    • Journal of Microbiology and Biotechnology
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    • v.22 no.2
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    • pp.199-206
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    • 2012
  • Naturally immobilized tannase (tannin acyl hydrolase, E.C. 3.1.1.20) has many advantages, as it avoids the expensive and laborious operation of isolation, purification, and immobilization, plus it is highly stable in adverse pH and temperature. However, in the case of cell-associated enzymes, since the enzyme is associated with the biomass, separation of the pure biomass is necessary. However, tannic acid, a known inducer of tannase, forms insoluble complexes with media proteins, making it difficult to separate pure biomass. Therefore, this study optimizes the production of cell-associated tannase using a "protein-tannin complex" free media. An exploratory study was first conducted in shake-flasks to select the inducer, carbon source, and nitrogen sources. As a result it was found that gallic acid induces tannase synthesis, a tryptose broth gives higher biomass, and lactose supplementation is beneficial. The medium was then optimized using response surface methodology based on the full factorial central composite design in a 3 l bioreactor. A $2^3$ factorial design augmented by 7 axial points (${\alpha}$ = 1.682) and 2 replicates at the center point was implemented in 17 experiments. A mathematical model was also developed to show the effect of each medium component and their interactions on the production of cell-associated tannase. The validity of the proposed model was verified, and the optimized medium was shown to produce maximum cell-associated tannase activity of 9.65 U/l, which is 93.8% higher than the activity in the basal medium, after 12 h at pH 5.0, $30^{\circ}C$. The optimum medium consists of 38 g/l lactose, 50 g/l tryptose, and 2.8 g/l gallic acid.

Detection of Food-Grade Hydrogen Peroxide by HRP-Biocomposite Modified Biosensors

  • Chang, Seung-Cheol
    • Journal of Food Hygiene and Safety
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    • v.32 no.6
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    • pp.447-454
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    • 2017
  • A new amperometric biosensor has been developed for the detection of hydrogen peroxide ($H_2O_2$). The sensor was fabricated through the one-step deposition of a biocomposite layer onto a glassy carbon electrode at neutral pH. The biocomposite, as a $H_2O_2$ sensing element, was prepared by the electrochemical deposition of a homogeneous mixture of graphene oxide, aniline, and horseradish peroxidase. The experimental results clearly demonstrated of that the sensor possessed high electrocatalytic activity and responded to $H_2O_2$ with a stable and rapid manners. Scanning electron microscopy, cyclic voltammetry, and amperometry were performed to optimize the characteristics of the sensor and to evaluate its sensing chemistry. The sensor exhibited a linear response to $H_2O_2$ in the range of 10 to $500{\mu}M$ concentrations, and its detection limit was calculated to be $1.3{\mu}M$. The proposed sensing-chemistry strategy and the sensor format were simple, cost-effective, and feasible for analysis of "food-grade $H_2O_2$" in food samples.