• Title/Summary/Keyword: Kinetic Parameters

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Cloning of a Novel vpr Gene Encoding a Minor Fibrinolytic Enzyme from Bacillus subtilis SJ4 and the Properties of Vpr

  • Yao, Zhuang;Meng, Yu;Le, Huong Giang;Lee, Se Jin;Jeon, Hye Sung;Yoo, Ji Yeon;Kim, Hyun-Jin;Kim, Jeong Hwan
    • Journal of Microbiology and Biotechnology
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    • v.30 no.11
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    • pp.1720-1728
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    • 2020
  • We have previously characterized AprESJ4, the major fibrinolytic enzyme from Bacillus subtilis SJ4 (Yao et al., 2019). During that study, we observed a 68 kDa protein with fibrinolytic activity. In this study, we cloned the gene (vprSJ4) encoding the 68 kDa protein, a mature Vpr and minor protease secreted by Bacillus species. vprSJ4 encodes a preproenzyme consisting of 810 amino acids (aa) including signal sequence (28 aa) and prosequence (132 aa). The mature enzyme (650 aa) has a predicted molecular weight of 68,467.35. Unlike Vprs from other B. subtilis strains, VprSJ4 has 4 additional amino acids (DEFA) at the C-terminus. vprSJ4 was overexpressed in Escherichia coli. PreproVprSJ4 was localized in inclusion bodies, and subjected to in vitro renaturation and purification by an affinity column. SDS-PAGE and western blot showed that autoprocessing of preproVprSJ4 occurred and 68 kDa and smaller proteins were produced. The optimum pH and temperature of the recombinant VprSJ4 were pH 7.0 and 40℃, respectively. Kinetic parameters of recombinant VprSJ4 were measured by using an artificial substrate, N-succinyl-ala-ala-pro-phe-p-nitroanilide. Coexpression of vprSJ4 and aprESJ4 using pHY300PLK increased the fibrinolytic activity a further 117% when compared with aprESJ4 single expression using the same vector in B. subtilis WB600.

Fundamental Studies on the Equilibrium and Kinetics for the fractional Distillation Reaction of Waste Organic Solvent (폐용제 분별증류 회수 반응의 평형 및 속도론적 기초연구)

  • Noh Hyun-Sook;Kim Dong-Su
    • Resources Recycling
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    • v.11 no.6
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    • pp.38-46
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    • 2002
  • Fundamental investigations were conducted far the recovery process of waste organic solvent by fractional distillation in the aspects of equilibrium and kinetics. Mixture of toluene and xylene, which were both being used in the largest amount as industrial organic solvent, was taken as the artificial waste organic solvent and their distillation behaviors were studied. The purity of recovered solvent was investigated by Cir Chromatography and shown to be in the range of 94~98%. Based upon equilibrium calculations, the changes in the Gibbs free energy, standard enthalpy, and standard entropy for distillation reaction have been estimated. The standard enthalpy changes for toluene and xylene were shown to be 44.833 and 47.044 kJ $mol^{-1}$ respectively, which were similar to their molar heats of evaporation. The activation energies of distillation fur toluene and xylene obtained from kinetic studies were 3.281 and 2.699 kJ $mol^{-1}$ and they were about one tenths of the standard enthalpy changes of distillation reaction. The highness of the purity of recovered organic solvents suggested the possibility that the recovered waste organic sol-vent could partly replace the original solvent.

Production of Chitosna Oligosaccharides Using Chitin-Immobilized Enzyme (키틴 고정화 효소를 이용한 키토산 올리고당의 생산)

  • 전유진;박표잠;변희국;송병권;김세권
    • KSBB Journal
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    • v.13 no.2
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    • pp.147-154
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    • 1998
  • Enzymatic hydrolysis using an immobilized enzyme was carried out to produce chitosan oligosaccharides (COSs) from chitosan effectively. Chitosanase was immobilized on eight different carriers by physical adsorption. The enzyme immobilized on chitin had higher activity than those immobilized on the other carriers in spite of its lower adsorption. The activity of chitin-immobilized enzyme was more than 90% of the original activity. Optimal temperature of the immobilized enzyme increased by about $15^{\circ}C$ and its thermostability was excellent in relatively wide range of temperature. But its effects of pH did not improve compared to the free enzyme. The immobilized enzyme produced 153 mg/g chitosan of the reducing sugar for 3hrs of hydrolytic incubation time. The total content of higher oligomers, tetramer to hexamer, among amount of total COSs obtained for 2hrs was more than 90%. In kinetic parameters for both enzymes, immobilized enzyme showed lower affinity for substrate and reaction rate than free enzyme, however, no reduction of the rate for high substrate concentrations. Consequently, chitin-immobilized could effectively hydrolyse chitosan and produce the higher COSs without activity decrease in comparison with the free enzyme.

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Computational Fluid Dynamics(CFD) Simulation for a Pilot-scale Selective Non-catalytic Reduction(SNCR) Process Using Urea Solution (요소용액을 이용한 파일럿규모 SNCR 공정에 대한 CFD 모델링 및 모사)

  • Nguyen, Thanh D.B.;Kang, Tae-Ho;Lim, Young-Il;Kim, Seong-Joon;Eom, Won-Hyeon;Yoo, Kyung-Seun
    • Korean Chemical Engineering Research
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    • v.46 no.5
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    • pp.922-930
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    • 2008
  • The selective non-catalytic reduction(SNCR) performance is sensitive to the process parameters such as flow velocity, reaction temperature and mixing of reagent(ammonia or urea) with the flue gases. Therefore, the knowledge of the velocity field, temperature field and species concentration distribution is crucial for the design and operation of an effective SNCR injection system. In this work, a full-scale two-dimensional computational fluid dynamics(CFD)-based reacting model involving a droplet model is built and validated with the data obtained from a pilot-scale urea-based SNCR reactor installed with a 150 kW LPG burner. The kinetic mechanism with seven reactions for nitrogen oxides($NO_x$) reduction by urea-water solution is used to predict $NO_x$ reduction and ammonia slip. Using the turbulent reacting flow CFD model involving the discrete droplet phase, the CFD simulation results show maximum 20% difference from the experimental data for NO reduction. For $NH_3$ slip, the simulation results have a similar tendency with the experimental data with regard to the temperature and the normalized stoichiometric ratio(NSR).

Partial Purification and General Properties of Yeast Acetolactate Synthase (효모 Acetolactate Synthase의 부분 정제와 일반 특성 연구)

  • Koh, Eun-Hie;Song, Soo-Mee;Kim, Sun-Young
    • Journal of the Korean Chemical Society
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    • v.39 no.6
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    • pp.459-465
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    • 1995
  • Acetolactate Synthase (ALS) was partially purified from the yeast and its basic biochemical studies were carried out. Yeast was grown in the minimum media containing 0.5% glucose, 51 mM $K_2HPO_4$, 22 mM $KH_2PO_4$, 8 mM $(NH_4)2SO_4,\;0.4\;m M\;MgSO_4$ for 18 hours at 37 $^{\circ}C$. The cell was ruptured in the buffer (20 mM phosphate buffer pH 7.0, 0.1 mM TPP, 0.5 mM DTT, 1 ${\mu}M$ FAD, and 1 mM MgCl_2$) following an overnight suspension. The supernatant fraction was collected from $10,000{\times}g$ and the enzyme was further purified by ammonium sulfate fractionation, DEAE-Sephacel chromatography and leucine-agarose chromatography. The enzyme activity was measured under the various conditions by the function of protein concentration, time, temperature, pH, and substrate. The optimum temperature was found to be 50$^{\circ}C$, optimum pH 8.0∼8.5. The kinetic parameters, $K_m\;and\;V_{max}$ were 8.4 mM and 17.9 nmol/mg/min respectively. Stability of the enzyme was studied with ethylene glycol and glycerol added to the enzyme solution. Both ethylene glycol and glycerol improved the enzyme stability up to 50%. The study of feedback inhibition showed that valine was a strong inhibitor while leucine was a weak inhibitor.

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Synthesis, Potentiometric, Spectral Characterization and Microbial Studies of Transition Metal Complexes with Tridentate Ligand (세자리 리간드의 전이금속 착물에 대한 합성과 전위차 및 분광학적 확인 그리고 미생물학적 연구)

  • Jadhav, S.M.;Munde, A.S.;Shankarwar, S.G.;Patharkar, V.R.;Shelke, V.A.;Chondhekar, T.K.
    • Journal of the Korean Chemical Society
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    • v.54 no.5
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    • pp.515-522
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    • 2010
  • A relation between antimicrobial activities and the formation constants of solid complexes of Cu(II), Ni(II), Co(II), Mn(II) and Fe(III) with tridentate Schiff base ligand, 4-hydroxy-3(1-{2-(benzylideneamino)-phenylimino}-ethyl)-6-methyl-2Hpyran-2-one (HL) derived from o-phenylene diamines, dehydroacetic acid (DHA) and p-chloro benzaldehyde have been studied. The ligand and metal complexes were characterized by elemental analysis, conductivity, magnetic susceptibility, thermal analysis, X-ray diffraction, IR, $^1H$-NMR, UV-vis and mass spectra. From the analytical data, the stiochiometry of the complexes was found to be 1:2 (metal:ligand) with octahedral geometry. The molar conductance values suggest the nonelectrolytic nature of metal complexes. The X-ray diffraction data suggests monoclinic crystal system for Ni(II) and orthorhombic crystal system for Cu(II) and Co(II) complexes. The IR spectral data suggest that the ligand behaves as tridentate ligand with ONN donor atoms sequence towards central metal ion. Thermal behavior (TG/DTA) and kinetic parameters calculated by Coats-Redfern method suggests more ordered activated state in complex formation. The protonation constants of the complexes were determined potentiometrically in THF:water (60:40) medium at $25^{\circ}C$ and ionic strength ${\mu}=0.1\;M$ ($NaClO_4$). Antibacterial activities in vitro were performed against Staphylococcus aureu and Escherichia coli. Antifungal activities were studied against Aspergillus Niger and Trichoderma. The effect of the metal ions and stabilities of complexes on antimicrobial activities are discussed.

Adsorption Equilibrium, Kinetics and Thermodynamics Studies of Malachite Green Using Zeolite (제올라이트를 이용한 말라카이트 그린의 흡착평형, 동력학 및 열역학 연구)

  • Lee, Jong-Jib
    • Clean Technology
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    • v.18 no.1
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    • pp.76-82
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    • 2012
  • The paper includes utlization of zeolite as potential adsorbent to remove a hazardous malachite green from waste water. The adsorption studies were carried out at 298, 308 and 318 K and effects of temperature, contact time, initial concentration on the adsorption were measured. On the basis of adsorption data Langmuir and Freundlich adsorption isotherm model were also confirmed. The equilibrium process was described well by Freundlich isotherm model, showing a selective adsorption by irregular energy of zeolite surface. From determined isotherm constants, zeolite could be employed as effective treatment for removal of malachite green. From kinetic experiments, the adsorption process followed the pseudo second order model, and the adsorption rate constant ($k_2$) decreased with increasing initial concentration of malachite green. Thermodynamic parameters like activation energy, change of free energy, enthalpy, and entropy were also calculated to predict the nature adsorption. The activation energy calculated from Arrhenius equation indicated that the adsorption of malachite green on the zeolite was physical process. The negative free energy change (${\Delta}G^{\circ}$ =-6.47~-9.07 kJ/mol) and the positive enthalpy change (${\Delta}H^{\circ}$ = +32.414 kJ/mol) indicated the spontaneous and endothermic nature of the adsorption in the temperature range 298~318 K.

An Experimental Study on the Devolatilization Kinetics of Ashless coal in Fixed and Entrained Conditions (초청정 석탄의 탈휘발 반응률에 관한 실험적 연구)

  • Yu, Da-Yeon;Lee, Byoung-Hwa;Song, Ju-Hun;Lee, Si-Hyun;Jeon, Chung-Hwan
    • Transactions of the Korean Society of Mechanical Engineers B
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    • v.35 no.10
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    • pp.997-1003
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    • 2011
  • In order to investigate devolatilization characteristics for ashless coal with relatively low ash content and high heating value, an experiment was performed in different bed configurations of TGA and DTF(Drop Tube Furnace) at atmospheric pressure condition. The heating rate was $10^{\circ}C$/min up to $950^{\circ}C$ in TGA, while the temperatures of DTF varied from 500 to $1300^{\circ}C$ in step of $200^{\circ}C$. A weight loss and particle temperature were obtained to determine devolatilization kinetics. The kinetic parameters including an activation energy and pre-exponential factor for ashless coal were obtained using Coats-Redfern method in TGA and single step method in DTF. Furthermore, the devolatilization kinetics of the ashless coal were compared with the results of different kinds of conventional coal such as sub-bituminous and bituminous. The results show that the activation energy of devolatilazation for ashless coal is lower than those of others in fixed and entrained conditions.

Structure and Reactivity of Alkylchloroformates. MO Theoretical Interpretations on Halide Exchange Reaction (염화 포름산 알킬의 구조와 반응성. 할로겐화 이온 교환반응에 대한 분자궤도론적 고찰)

  • Lee, Bon Su;Lee, Ik Choon
    • Journal of the Korean Chemical Society
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    • v.18 no.4
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    • pp.223-238
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    • 1974
  • CNDO/2 MO theoretical studies and kinetic studies of halide exchange reactions for alkylchloroformates have been carried out in order to investigate structure-reactivity relationship of alkylchloroformates. From the result of energetics, it was concluded that the most stable configuration of alkylchloroformate is that in which alkyl group and chlorine are trans to each other, and that the hindered rotation about the bond between the carbonyl carbon and alkoxy-oxygen bond is attributed to the ${\pi}-$electron delocalization. It has been found that the large charge separation is due to -M effect of carbonyl and alkoxy oxygens and-I effect of chlorine. The order of rates in solvents studied was $(CH_3)_2 > CO > CH_3CN{\gg}MeOH.$$I^->Br^->Cl^-$ in protic solvent, and of Cl^->Br^- >I^-$ in dipolar aprotic solvents. Alkyl group contribution has the decreasing order of $CH_3-> C_2H-{\gg}i-C_3H_7-.$ The solvent effect has been interpreted on the basis of initial and final state contribution. A transition state model has been suggested, and it has been proposed that the most favorable mechanism is the addition-elimination. From the results of activation parameters and electronic properties, an energy profile has been proposed. Structural factors determining reactivities of alkylchloroformates have been shown to be charge, energy level of ${\alpha}^*LUMO$ to C-Cl bond and ${\alpha}^{\ast} $antibonding strength with respect to C-Cl bond in this MO. Charge and polarizability of nucleophile, and the interaction of these effects with solvent structures are also found to be important.

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Kinetics and Mechanism for Aquation of $[Co(en)_2Cl_2]^+$ in the $Hg^{2+}$ Aqueous Solution ($Hg^{2+}$수용액에서 $[Co(en)_2Cl_2]^+$의 아쿠아 반응속도론과 메카니즘)

  • Byung-Kak Park;Joo-Sang Lim;Sung Nak-Jung;Il-Bong Lee;Kwang-Jin Kim;Sung-Gu Kang
    • Journal of the Korean Chemical Society
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    • v.32 no.4
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    • pp.342-350
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    • 1988
  • An experimental investigation is made to determine the mechanism of the aquation of $[Co(en)_2Cl_2]^+$ in $Hg^{2+}$ aqueous solution. The progress of reaction is followed UV/vis-spectrophotometrically by a measurment of the absorbance at a specific wavelength of Co(III) complex as a function of time. The aquation of cis-$[Co(en)_2Cl_2]^+$ and trans-$[Co(en)_2Cl_2]^+$ has been found to be first order and second order with respect to the concentration of $Hg^{2+}$ catalyst, respectively. It has been found that the reaction rate for aquation of the trans-form is faster than that of cis-form, and that the product of either cis-form or trans-form is always in the mixture ratio of 97 % to 3 %. Plausible reaction mechanism is proposed for the reaction system on the basis of kinetic data and activation parameters. Theoretical rate equation derived from the proposed mechanism is consistent with the observed one.

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