• Title/Summary/Keyword: 이온-크로마토그래피

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Characterization of Extracellular Peroxidase from Pleurotus ostreatus (Pleurotus ostreatus에서 분비되는 Peroxidase의 특성)

  • 배성호;신광수;강사욱;하영칠;최선진;김규중;최형태
    • Korean Journal of Microbiology
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    • v.27 no.4
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    • pp.348-356
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    • 1989
  • An extracellular peroxidase found in culture broth of Pleurotus ostreatus was induced by syringic acid. This enzyme was fractionated by DEAE Sephadex A-50 ion exchange chromatography and gel filtration chromatogrphy on Sephadex G-150. The enzyme is a glycoprotein containing 35.7% carbohydrate. The results of SDS-linear polyacrylamide gradient gel electrophoresis and gel filtration indicate that the enzyme is a dimer consisted of identical subunits (Mr=72,400). The absorption spectrum of the enzyme indicates the presence of one mole of iron protoporphyrin IX per one mole of subunit. Isoelectric point of the enzyme is 4.26 and $K_m$ values for $H_2O_2$ is $7.2{\mu}M$. The enzyme showed its optimal activity at pH 3.5-4.0 and at $40^{\circ}C$. The Km values of this enzyme for ferulic acid and sinapic acid are 2.4 and 12.3 times higher than those of horseradish peroxidase, respectively.

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The Effect of Temperature and Flow Rate of Eluent on the Separation of Adjacent Lanthanides (La : Ce, Ce : Pr, Pr : Nd) with Displacement Chromatography (치환크로마토그래피에서 온도와 용리액의 흐름속도가 란탄족 원소들 (La : Ce, Ce : Pr, Pr : Nd) 의 분리에 미치는 영향)

  • Ha, Yeong Gu;Song, Gi Hun
    • Journal of the Korean Chemical Society
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    • v.38 no.9
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    • pp.660-666
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    • 1994
  • The effects of temperature and flow rate of eluent on the separation of adjacent lighter lanthanide pairs (La : Ce, Ce : Pr, Pr: Nd) have been studied with displacement chromatography. Two serial columns packed with Amberlite 120 cation exchange resin are used for loading and separation. The retaining ion is $H^+$ ion and the eluent is 0.012M and 0.015M of EDTA solution. The columns and the eluent are maintained at the temperature of 90$^{\circ}C$ and pressurized to reduce vaporizing in the ion-exchange resin column. The eluated solution is analyzed directly with ICP-AES. The separation factors of the lanthanide pairs, La: Ce, Ce :Pr, and Pr: Nd, are 4.6, 2.8, and 1.9, respectively and are higher than that from theoretical calculation at 25$^{\circ}C$. When the flow rate is reduced from 2.5 ml/min to 1.5 ml/min, the HETP is reduced from 1.60 cm to 0.88 cm. The separation efficency can be improved at lower flow rate of eluent and higher operating temperature. The recoveries of pure lanthanides than 99.9% are 49∼77% from this separation.

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Purification and Characterization of Polyphenol Oxidase from Flammulina velutipes (팽나무버섯 polyphenol oxidase의 정제 및 특성)

  • Pyo, Han-Jong;Son, Dae-Yeul;Lee, Chan
    • Korean Journal of Food Science and Technology
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    • v.34 no.4
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    • pp.552-558
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    • 2002
  • Polyphenol oxidase from Flammulina velutipes was purified and characterized. Purification of polyphenol oxidase was achieved by ammonium sulfate precipitation, Superdex G-200 gel filtration chromatography, Phenyl superose affinity chromatography, Mono-Q anion exchange chromatography and Superdex S-200 gel filtration chromatography on FPLC. After these purification steps specific activity of purified polyphenol oxidase increased to 199.1 units/mg. Polyphenol oxidase from F. velutipes was composed of a single polypeptide with molecular weight of about 40 kDa. Optimum pH and temperature for the enzyme reaction were found to be 6.0 and $25^{\circ}C$, respectively. The activity of the enzyme gradually decreased at acidic pH between 3 and 5, and the enzyme lost its activity at alkaline pH between 8 and 10. This enzyme exhibited high substrate specificity to o-diphenols. Km-values for L-DOPA and caffeic acid were found to be 3.97 mM and 1.78 mM, respectively. 2-mercaptoethanol, L-ascorbic acid, sodium bisulfite, EDTA and $Mg^{2+}$ inhibited the activity of pholyphenol oxidase and $Cu^{2+}$, $Fe^{2+}$, $Zn^{2+}$ and $Ni^{2+}$ increased enzyme activity. The activity of enzyme was well maintained at $-70^{\circ}C$ for over 4 months, and at $-20^{\circ}C$ for 1 months.

Separation of chlorine in a uranium compound by pyrohydrolysis and steam distillation, and its determination by ion chromatography (열가수분해 및 수증기증류에 의한 우라늄 화합물 중 염소 분리 및 이온크로마토그래피 정량)

  • Kim, Jung-Suk;Lee, Chang-Hun;Park, Soon-Dal;Han, Sun-Ho;Song, Kyu-Seok
    • Analytical Science and Technology
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    • v.23 no.1
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    • pp.45-53
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    • 2010
  • For the determination of chlorine in uranium compound, analytical methods by using a steam distillation and a pyrohydrolysis have been developed. The steam distillation apparatus was composed of steam generator, distilling flask and condenser etc. The samples were prepared with an aliquot of LiCl standard solution and a simulated spent nuclear fuel. A sample aliquot was mixed with a solution containing 0.2 M ferrous ammonium sulfate-0.5 M sulfamic acid 3 mL, phosphoric acid 6 mL and sulfuric acid 15 mL. The chloride was then distilled by steam at the temperature of $140^{\circ}C$ until a volume of $90{\pm}5\;mL$ is collected. The pyrohydrolysis equipment was composed of air introduction system, water supply, quartz reaction tube, combustion tube furnace, combustion boat and absorption vessel. The chloride was separated from powdered sample which is added with $U_3O_8$ accelerator, by pyrohydrolysis at the temperature of $950^{\circ}C$ for 1 hour in a quartz tube with a stream of air of 1 mL/min supplied from the water reservoir at $80^{\circ}C$. The chlorides collected in each absorption solution by two methods was diluted to 100 mL and measured with ion chromatography to determine the recovery yield. For the ion chromatographic determination of chlorine in molten salt retained in a metal ingot, the chlorine was separated by means of pyrohydrolysis after air and dry oxidation, and grinding for the sample.

Ion-Pair Chromatography of Benzoic Acid and Its Derivatives on XAD-2 (XAD-2 지지체를 이용한 벤조산과 그 유도체들의 이온쌍 크로마토그래피에 관한 연구)

  • Kang, Sam-Woo;Ryu, Sam-Gon;Park, Young-Kyu
    • Journal of the Korean Chemical Society
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    • v.28 no.3
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    • pp.176-184
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    • 1984
  • Retention behavior of benzoic acid and its derivatives on XAD-2 in the alcoholic aqueous solution was investigated and separation was attempted. Retention was affected by the concentration and kinds of added organic solvents, the pH of the aqueous solution, the added $R_4N^+$ and the position and kinds of functional group in the sample molecules. Retention of sample acids in acidic conditions was due to mainly molecular adsorption on nonpolar XAD-2 surface and that in basic conditions was due to mainly ion-pair model. In these bases a mixed sample was separated in EtOH 20% aqueous solution at pH 8.50.

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Uncertainties of ionic species in snowpit samples determined with ion chromatography system (이온크로마토그래피 시스템을 이용한 눈 시료의 이온성분 측정자료의 불확도 산출)

  • Hong, Sang-Bum;Hur, Soon-Do;Kim, Sun-Mee;Hong, Sungmin;Chung, Ji-Woong;Kang, Namgoo;Kang, Chang-Hee
    • Analytical Science and Technology
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    • v.25 no.6
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    • pp.350-363
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    • 2012
  • To determine ionic species in snowpit samples using ion chromatography system, we described the performance of ion chromatography(IC) system, cleaning method of bottle, and interference by filtering procedure. The limit of detection, reproducibilities, and accuracies determined with BCR$^{(R)}$-408 were 0.01-0.26 ${\mu}g$/L, 0.4-17.4%, 4.5-12.0% for cations and 0.02-0.26 ${\mu}g/L$, 0.1-27.6%, 1.3-5.6% for anions, respectively. Lab blank test for sample bottle indicated that $CH_3CO_2{^-}$, $HCO_2{^-}$, and $NH_4{^+}$ can be easily contaminated in the lab environment. The positive interferences of $NO_3{^-}$ were partly attributed to the cleaning method of bottle. The filtering of melted snow sample should be carefully applied because it can positively affect the concentration levels of some ionic species. Finally, this method was applied to measure ionic species in snowpit samples from the upward area near NEEM camp and the uncertainties of measurement data of $F^-$ were also estimated.

Characteristics of Angiotensin Converting Enzyme Inhibitory Peptides from Thermolysin Hydrolysate of Manila clam, Ruditapes philippinarum Proteins (바지락 단백질 Thermolysin 가수분해물의 Angiotensin Converting Enzyme 저해 Peptide의 특성)

  • Lee Tae Gee;Yeum Dong Min;Kim Seon Bong
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.35 no.5
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    • pp.529-533
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    • 2002
  • The peptides inhibiting angiotensin converting enzyme (ACE) were isolated from the hydrolysate of manila clam (Ruditapes philippinamm) proteins prepared with thermolysin. The thermolysin hydrolysate was pretreated with membrane filter (MW cut-off 10,000) to obtain the peptide fraction with ACE inhibition. The crude peptides were applied to a Sephadex LH-20 column and eluted with $30\%$ methanol. The three active fractions (A, B and C) were collected and concentrated, and then applied to a SP-Toyopearl 650S column equilibrated with distilled water and was eluted with a linear gradient of NaCl concentration (0 to 1 M). The four active fractions (A-1, A-2, B-1 and C-1) were collected and concentrated, and then applied to a SuperQ-Toyopearl 650S column equilibrated with distilled water and was eluted with a linear gradient of NaCl concentration (0 to 1 M). The maximum inhibitory activity was observed in the fraction B-1Q showed the IC_{50} values of 0.748 $\mu$g. The abundant amino acids obtained from active fraction B-1Q were leucine, isoleucine, alanine and threonine.

Studies on the Elution Behavior and the Simultaneous Analysis of Some Metal-Dithiocarbamate Chelates by Reversed-Phase High Performance Liquid Chromatography (역상 액체 크로마토그래피에 의한 몇가지 금속-Dithiocarbamate 킬레이트의 용리거동 및 동시분석에 관한 연구)

  • Dai Woon Lee;Yun Je Kim;Hyun Chul Kim;Won Lee
    • Journal of the Korean Chemical Society
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    • v.32 no.3
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    • pp.211-226
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    • 1988
  • Liquid chromatographic behavior of several metal ions in dithiocarbamate(DTC) chelates were investigated by reversed phase high performance liquid chromatography on Novapak $C_{18}$ and ${\mu}$-Bondapak $C_{18}$ columns. The optimum conditions for the separation of DTC-metal chelate were examined with respect to the pH, shaking time, flow rate, extraction solvent, and mobile phase strength. The metal ions in mixtures at trace level, chelated with some dithiocarbamate derivatives were separated successfully on Novapak $C_{18}$ column using acetonitrile/methanol/water or acetonitril/water mixtures as mobile phases. It was found that all DTC metal chelates studied were eluted in an acceptable range of capacity factor values ($0{\leqq}log\;k'{\leqq}1$). Although several foreign metal ions were coexisted, high recovery and good precision were attained ; 97.0-106.7 % for the recovery and 0.98-3.41% for the coefficient of variation. Under the optimum analytical conditions, trace metal ions in the composite water samples were determined sucessfully with in relative error of about {\pm}$6.7 %.

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Analysis of Amino Acid Residues Involved in Activities of Chitin Deacetylase of Aspergillus nidulans (Aspergillus nidulans에서 분리된 키틴 탈아세틸화 효소활성에 영향을 미치는 아미노산 잔기 분석)

  • Kim, Jong-Il;Song, Da-Hyun
    • Korean Journal of Microbiology
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    • v.47 no.4
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    • pp.302-307
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    • 2011
  • Native chitin deacetylase of Aspergillus nidulans was purified to apparent homogeneity by a combination of phenyl-Sepharose and Q-Sepharose column chromatography. In order to analyze the amino acid residues involved in the enzyme activity, the enzyme was chemically modified with chemical agent, which selectively reacted with the specific amino acid residue on the protein. When the enzyme was chemically modified with diethylpyrocarbonate, which specifically reacted with histidine residues on the protein, the activity was eliminated. The chitin deacetylase, chemically modified with 100 ${\mu}M$ modifier at the residue of arginine or tyrosine, has shown to have decreased activities. It was shown that the modification at aspartic acid or glutamic acid did not affect the enzyme activity to a greater extent, which would not implicate that acid amino residues were directly involved in catalytic reaction and would affect on the global structures of the proteins. This results demonstrated that histidine and tyrosine residues of enzyme would participate in an important function of the chitin deacetylase activity.

Production and Characterization of Manganese Peroxidase from the White Rot Fungus Pleurotus ostreatus in Liquid Culture (액체배양한 느타리 버섯균(Pleurotus ostreatus)으로부터 망간퍼옥시데이즈의 생산 및 특성)

  • Lee, Jae-Sung;Ha, Hyo-Cheol
    • Applied Biological Chemistry
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    • v.47 no.1
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    • pp.22-26
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    • 2004
  • The ligninolytic basidiomycete, Pleurotus ostreatus K-2946, was produced a manganese peroxidase (MnP) activity when grown in liquid culture with glucose-yeast-peptone (G-Y-P) medium. However, lignin peroxidase (LiP) was not detected in this culture medium. The purification progress of MnP was purified that included chromatography on Sepharose CL-6B, Superdex 75 prep grade and Mono-Q. MnP purified by column chromatography, was 36400 dalton and a pI of 3.95. The optimal pH and temperature of the purified MnP activity were 5.0 and $55^{\circ}C$. The characteristics of MnP produced was quite similar to those of MnP 3 isoenzyme produced by other strains of P. ostreatus.