• Title/Summary/Keyword: Ion-Exchange Chromatography

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Properties of Glucoamylase Isozymes Produced by Aspergillus sp. (Aspergillus sp.가 생산하는 Glucoamylase Isozymes의 성질)

  • Park, Inshik;Youngho Chung
    • Microbiology and Biotechnology Letters
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    • v.16 no.4
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    • pp.320-326
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    • 1988
  • Glucoamylase (EC 3.2.1.3) of Aspergillus sp. isolated from soil was partially purified by Sephacryl S-200 gel filtration and DEAE-Sephacel ion exchange chromatography, The glucoamylase activity was separated into two isozymes after DEAE-Sephacel ion exchange chromatrography. The optimum pH and temperature for both glucoamylase isozymes (GI, GII) were identical; pH 4.5 and temperature, $65^{\circ}C$. The molecular weights of GI and GII Isozymes were estimated to be 105,000, which were measured by gel filtration on Sephacryl S-200. Both isozymes were stable at pH ranges of 2 to 7, and up to 6$0^{\circ}C$. Glycerol was effective to stabilize the both isozymes. The activation energies of GI and GII isozymes were 10.63 and 10.33 kcal/mole, respectively. The enzyme activities of both isozymes were completely inactivated by addition of 0.1% Hg$^{++}$. In kinetic studies, the Km values of GI isozyme for soluble starch, dextrin, and glycogen were 0.62%, 0.32%, and 1.02%, respectively. For GII isozyme, they became 0.66%, 0.23%. and 0.14% for the substrates.

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Temperature Effect on the Retention Behavior of Sugars in Ion Exchange Chromatography (이온 교환 크로마토그래피에서 온도가 당의 체류 특성에 미치는 영향)

  • Kim, Jin-Il;Lee, Chong-Ho;Koo, Yoon-Mo
    • Korean Chemical Engineering Research
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    • v.43 no.6
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    • pp.722-727
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    • 2005
  • Dow99Ca350 (Dowex monosphere 99Ca/350 separation resin), MFG-220, and Finex CS-10GC are ion-exchange resins, and primarily used to separate sugars, and all of these resins have poly styrene DVB backbone, and sulfonyl group. These resins are already used to separate sugars continuously at sugar industry at constant temperature. These resins are used in experiments for understanding temperature effect on retention or adsorption behavior. Using Dow99Ca350, swelling test, porosity test, pulse test, and frontal analysis at various temperatures were performed. In the cases of MFG-220, and Finex CS-10GC, the effect of temperature variation was verified by pulse test. The experimental results are shown that Dow99Ca350, MFG-220, and Finex CS-10GC, which are commercial resins for sugar separation, are stable to temperature variation because the maximum change of retention time of fructose, and glucose are 1.76, and 3.37% respectively.

Optimization of Extraction and Purification of Phytic Acid from Defatted Rice Bran (탈지미강으로부터 Phytic Acid의 추출과 정제의 최적화)

  • Choi, Moon Sil;Han, Bok Kyung;Choi, Hyuk Joon;Park, Young-Seo
    • Food Engineering Progress
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    • v.15 no.3
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    • pp.276-281
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    • 2011
  • The optimum condition for the extraction and purification processes of phytic acid from defatted rice bran was examined. The phytic acid was efficiently extracted when the defatted rice bran was treated with 10 volumes of 0.5% HCl for 1 hr. For the neutralization of acid-treated extract, 0.5% NaOH was the most acceptable. To purify phytic acid, Diaion HP20 resin was used to remove impurities from the extract. The flow-through was then loaded onto ion exchange columns packed with various resins and among them, Amberlite IRA-416 resin showed highest recovery yield. When the phytic acid was absorbed onto Amberlite IRA-416 resin and then eluted with 0.5% NaOH, 89% of applied phytic acid was eluted. Most proteins were removed from the purified phytic acid and total protein content of the phytic acid was 0.14%(w/w).

Recombinant Expression and Purification of Functional XorII, a Restriction Endonuclease from Xanthomonas oryzae pv. oryzae

  • Hwang, Dong-Kyu;Cho, Jae-Yong;Chae, Young-Kee
    • Journal of Microbiology
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    • v.45 no.2
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    • pp.175-178
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    • 2007
  • An endonuclease from Xanthomonas oryzae pathovar oryzae KACC 10331, XorII, was recombinantly produced in Escherichia coli using a T7 system. XorII was purified using a combination of ion exchange and immobilized metal affinity chromatography (IMAC). An optimized washing protocol was carried out on an IMAC in order to obtain a high purity product. The final amount of purified XorII was approximately 2.5 mg/L of LB medium. The purified recombinant XorII was functional and showed the same cleavage pattern as PvuI. The enzyme activity tested the highest at $25^{\circ}C$ in 50 mM NaCl, 10 mM Tris-HCl, 10 mM $MgCl_{2}$, and 1 mM dithiothreitol at a pH of 7.9.

Occurrence of ELISA Inhibitors, and Inhibitor-free Determination of Cyclobutane Pyrimidine Dimers in Japanese Cypress Leaves

  • Hashimoto, Tohru;Nakagawa, Noriko;Okano, Michiaki;Nikaido, Osamu
    • Journal of Photoscience
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    • v.9 no.2
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    • pp.448-450
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    • 2002
  • DNA samples extracted from Japanese cypress leaf tissues contain isopropyl alcohol-precipitable, high molecular weight compounds, which interfere ELISA for cyclobutane pyrimidine dimers (CPD). Removal of the compounds is achieved by DEAE ion-exchange column chromatography and improves the ELISA responses of the DNA. When extracting DNA repeatedly from the same leaf tissues, the DNA samples show CPD responses which increase with the order in sequential extraction, and hence for a reliable detennination of DNA lesion a thorough extraction of DNA is required. Clearing these two problems it was demonstrated that CPD level was slightly higher in the leaves of trees growing under full sunlight than in those growing under UBV -cut sunlight.

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Studies on Constituents of the Higher Fungi of Korea (XLVI) -Antitumor Components Extracted from the Cultured Mycelia of Lyophyllum decastes-

  • Kim, Hye-Ryoung;Shim, Mi-Ja;Kim, Jung-Woo;Kim, Ha-Won;Lee, Chong-Ock;Choi, Eung-Chil;Kim, Byong-Kak
    • Korean Journal of Pharmacognosy
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    • v.15 no.2
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    • pp.61-73
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    • 1984
  • To investigate antitumor component of Lyophyllum decastes, the aqueous extract of its shake-cultured mycelia was subjected to antitumor test against sarcoma 180 cells implanted in ICR mice. The extract showed an inhibition ratio of 65.4% and was found to consist of a polysaccharide moiety and a protein moiety. After purification with DEAE-Sephadex A-50 ion exchange chromatography, Fraction D showed the highest inhibition ratio of 75.7%. The antitumor constituent was examined for immunoaccelerating activity and was found to increase macrophage accumulation in the peritoneal cavity and plaque forming cells of the spleen cells. It was named lyophyllan after the genus name.

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Purification and characterization of Protease from Kyenegum (계내금(鷄內金) 단백질 분해 효소의 정제와 특성)

  • Kim, Do-Wan;Jo, Hye-Sim;Jeong, Yong-Jin;Kim, Kwang-Soo
    • The Korea Journal of Herbology
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    • v.22 no.4
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    • pp.21-28
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    • 2007
  • Objectives : Kyenegum has been popularly used long as the digestive. The purpose of this study was to investigate the purification and characteristics of protease obtained from Kyenegum crude enzyme. Methods : Kyenegum protease was purified by precipitation with ammonium sulfate followed by SP-Spharose ion exchange chromatography. The molecular weight of Kyenegum protease was estimated by SDS-PAGE electrophoresis. Results : Kyenegum protease was 3,087 units/mg protein specific activity, 14.5 purification fold and 9.8 % recovery. The molecular weight of protease was estimated to be 18 kDa. The isoelectric point was pI 8.97 and values of Km and Vmax of its were 48 mg/mL and 2 units/min, respectively. Conclusion : The result suggests that the protease obtained from Kyenegum has excellent stability of temperature, acid and collagen substrate specificity.

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Characterization of Acid Phosphatase from Carrots (당근 Acid Phosphatase의 특성)

  • Kim, Gi-Nahm
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.23 no.3
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    • pp.490-495
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    • 1994
  • Acid phosphatase (EC3.1.3.2) from carrots was partially purified by ammonium sulfate fractionation (30%-80%), Sephacryl S-200 gel filtration, cm-Sepharose CL-6B and DEAE -Sephacel ion exchange chromatography. The optimum ph and temperature of acid phosphatase from carrots were pH 5.5 and 55$^{\circ}C$, respectively. The enzyme was most stable at ph 6.0 and relatively unstable below pH 4.0 . The activation energy of the enayme was determined to be 10.6kcal/mole. The enzyme utilized p-nitrophenyl phosphate as a substrate among tested possible substrates, whereas it hydrolyzed 5' -IMP and 5'-GMP poorly. The Michaelis -Menten constant(Km) of the enzyme with p-nitrophenyl phosphate as a substrate was identified as 0.55mM. Amongtested metal ions and inhibitors, Al+++ Zn++, Cu++ , fluoride, metavanadate and molybdate ions inhibited the enzyme activity drastically.

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방선균이 생산하는 아미노산 대사길항물질, YS-460의 분리 정제 및 특성

  • 박부길
    • Microbiology and Biotechnology Letters
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    • v.24 no.3
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    • pp.327-330
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    • 1996
  • An amino acid antimetabolite named YS-460 was isolated from the culture filtrate of a newly isolated Actinomycetes identified as Streptomyces sp. Fermentation was carried out in the synthetic medium at 30$\circ$C for 5 days. Purification was done by ion exchange resin, active carbon, silica gel column chromatography and obtained 38 mg of pure active substance per liter of the broth. YS-460, an amino acid like substance, has the molecular formula of C$_{7}$H$_{11}$NO$_{3}$- Its structure determined to be furanomycin by spectral analysis. It is active against some bacteria on a chemically defined medium and reversed competitively by L-isoleucine and non-competitively by L-leucine and L-valine.

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Simultaneous determination of betaine and choline using derivatization by HPLC with UV detection (HPLC-UV검출방법으로 유도체화를 통한 비테인과 콜린의 동시분석)

  • Rhee, Insook;Paeng, Ki-Jung
    • Analytical Science and Technology
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    • v.28 no.2
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    • pp.112-116
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    • 2015
  • Extraction of quaternary ammonium compounds (choline and betaine) from plant samples (spinach) using ion exchange resin (AG1, OH form) is a very simple and inexpensive approach. However, it is very hard to determine amounts of choline and betaine simultaneously using high-performance liquid chromatography-ultraviolet (HPLC-UV) detection. Unlike choline, betaine has low molar absorptivity in UV-visible (UV-Vis) region, which makes it difficult to carry out UV-Vis detection of betaine. The mixture of quaternary ammonium compounds (choline and betaine) was derivatized using 2-bromo acetophenone as a derivatizing agent. As a result, choline did not react with the derivatizing agent, whereas betaine formed a betaine derivative. This betaine derivative exhibited detectable UV absorption with baseline separation between choline and the betaine derivative. Thus, with this method, choline and betaine can be determined simultaneously by using the HPLCUV method through one-step derivatization, which is an easy, sensitive, and reliable method.