• Title/Summary/Keyword: 이온-교환 크로마토그래피

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Insect Resistance of Tobacco Plant Expressing CpBV-ELP1 Derived from a Polydnavirus (폴리드나바이러스 유래 CpBV-ELP1 발현 담배의 내충성)

  • Kim, Eunseong;Kim, Yonggyun
    • Korean journal of applied entomology
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    • v.56 no.1
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    • pp.19-28
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    • 2017
  • Polydnaviruses (PDVs) are a group of double-stranded DNA viruses symbiotic to some endoparasitoid wasps. Cotesia plutellae bracovirus (CpBV) is a PDV symbiotic to an endoparasitoid wasp, C. plutellae, parasitizing young larvae of Plutella xylostella. An early expressed gene, CpBV-ELP1, plays an important role in the parasitism by suppressing host cellular immunity by its cytotoxic activity against hemocytes. This study aimed to test its oral toxicity against insect pest by expressing it in a recombinant tobacco plant. A recombinant CpBV-ELP1 protein was produced using a baculovirus expression system and secreted to cell culture medium. The cell cultured media were used to purify CpBV-ELP1 by a sequential array of purification steps: ammonium sulfate fractionation, size exclusion chromatography, and ion exchange chromatography. Purified rCpBV-ELP1 exhibited a significant cytotoxicity against Spodoptera exigua hemocytes. CpBV-ELP1 was highly toxic to the fifth instar larvae of S. exigua by injection to hemocoel. It also showed a significant oral toxicity to fifth instar larvae of S. exigua by a leaf-dipping assay. CpBV-ELP1 was cloned into pBI121 vector under CaMV 35S promoter with opaline synthase terminator. Resulting recombinant vector (pBI121-ELP1) was used to transform Agrobacterium tumefaciens LBA4404. The recombinant bacteria were then used to induce callus of a tobacco (Nicotiana tabacum Xanthi) leaves and subsequent generation (T1) plants were selected. T1 generation tobacco plants expressing CpBV-ELP1 gave significant insecticidal activities against S. exigua larvae. These results suggest that CpBV-ELP1 gene can be used to control insect pests by constructing transgenic crops.

Epimerization of L-Arabinose for Producing L-Ribose (L-리보스 생산을 위한 L-아라비노스의 에피머반응)

  • Jeon, Young Ju;Song, Sung Moon;Lee, Chang Soo;Kim, In Ho
    • Korean Chemical Engineering Research
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    • v.49 no.5
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    • pp.628-632
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    • 2011
  • L-ribose has recently attracted interest as a starting material for antiviral drug. It could be obtained from L-arabinose by epimerization reaction. Epimerization reaction was carried out with molybdenium oxide or molybdic acid catalyst and methanol/water solution. Reaction temperature, methanol percentage, and catalyst kind were selected to find an optimum reaction condition. Ion exhange chromatography was used for separating epimerization reaction mixture, and then HPLC chromatogram of L-ribose fraction obtained to calculate the yield of the reaction. Shodex ion exchange HPLC column(Model SC1011) and Phenomenex Luna $NH_2$ HPLC column were compared to employ a convenient HPLC analysis. It was found that the usage of 20% methanol, $60^{\circ}C$, and 40 g/L molybdic acid gives the best reaction condition with a yield of 21%.

Characterization of Carboxymethyl Cellulase Produced by Cellulomonas sp. CS1-1 on Microcrystalline Cellulose (Cellulomonas sp. CS1-1이 미소결정성 섬유소로부터 생산한 Carboxymethyl Cellulase의 효소적 성질)

  • Park, Jong-Soo;Yoon, Min-Ho;Choi, Woo-Young
    • Korean Journal of Agricultural Science
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    • v.24 no.2
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    • pp.275-282
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    • 1997
  • The prcx.iuction of extracellular 1,4-${\beta}$-glucanase by Cellulomonas sp. CS1-1 on microcrystalline cellulose, sigmacell was maximal after 5-day cultivation as 280 units/mL, which was three times higher than the level produced on carboxymethyl cellulose. A carboxymethyl cellulase containing the carbohydrate of 8.2% was purified from the culture filtrate by successive procedures of column chromatographies. Purification factor was calculated as 22-folds with the specific carboxymethyl cellulase activity of 31.9 units/mg. The molecular weight and isoelectric point of the purified enzyme were 54,000 and pI 5.4, respectively. The optimal pH and temperature were 6.0 and $45^{\circ}C$, and the enzyme was stable between pH 6.5 and 7.5 and below $50^{\circ}C$. The estimated Km and Vmax were 10 mg/mL and $6.25{\mu}mol/min$ for carboxymethyl cellulose and 30.3 mg/mL and $2.85{\mu}mol/min$ for sigmacell, respectively. The enzyme was partially inhibited by $Ag^+$, $Zn^{+{+}}$, $Fe^{+{+}}$ and EDTA, while completely inhibited by $Cd^{+{+}}$ and $Hg^{+{+}}$ at 1 mM concentration.

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Isolation of a Bacteriocin - Producing Lactobacillus sakei Strain from Kimchi (김치에서 박테리오신을 분비하는 Lactobacillus sakei균주의 분리)

  • 김한택;박재용;이강권;김정환
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.33 no.3
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    • pp.560-565
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    • 2004
  • Bacteriocin producing lactic acid bacteria (LAB) were isolated from Kimchi by using spot-on-the-lawn method. Listeria monocytogenes, Staphylococcus aureus, and Lactobacillus plantarum were used as indicators. One isolate (P3-l) produced a bacteriocin efficiently inhibiting the growth of Listeria monocytogenes. 16S rDNA sequence and sugar utilization test identified that P3-1 was a Lactobacillus sakei strain. Accordingly, the isolate was named as Lactobacillus sakei P3-1. L. sakei P3-1 produced a bacteriocin which efficiently inhibited the growth of Listeria monocytogenes but did not inhibit other Gram positive and negative organisms tested. The bacteriocin was stable against heat, organic solvent, and pH variation and it retained 50% of activity after 10 min heat treatment at 10$0^{\circ}C$. The molecular weight of Sakacin P3-1 was estimated to be 4 kDa by SDS-PAGE.

An Anticoagulant Polysaccharide Isolated from the Alkali Extracts of Coriolus versicolor (구름버섯 알칼리 추출물에서 분리한 항응고성 다당류)

  • Lee, Hyun-Sun;Kweon, Mee-Hyang;Lim, Wang-Jin;Sung, Ha-Chin;Yang, Han-Chul
    • Korean Journal of Food Science and Technology
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    • v.29 no.2
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    • pp.369-375
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    • 1997
  • We have isolated an anticoagulant polysaccharide from the alkali extracts of Coriolus versicolor. The anticoagulant polysaccharide was purified through a gradual ethanol precipitation and three concecutive chromatography of DEAE-Toyopearl 650C, Sephadex G-100, and Sepharose CL-6B by measuring activated partial thromboplastin time (aPTT). The anticoagulant polysaccharide showed the homogenecity on HPLC using a gel permeation column and had about $7.2{\times}10^{5}$ molecular weight. The polysaccharide consisted of fucose, glucose, and galactose in a molar ratio of 1.0:0.2:0.2:0.1, and also compromised 19.32% of sulfate at its constituent sugars. The polysaccharide showed the two typical bands of C-O-S $(823\;cm^{-1})$ and S=O $(1257\;cm^{-1})$ in the IR spectroscopy. The sulfated polysaccharide (CV-40-Va-1) inhibited the blood coagulation via the intrinsic pathway like heparin whose activity produced a concentration dependent effect in aPTT and thrombin time (TT).

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Purification of Egg Immunoglobulin IgY (계란 면역 단백질[IgY]의 정제 연구)

  • Kim, In-Ho;Lee, Yong-Tak;Lee, Chung-Hee;Chung, Bong-Hyun
    • KSBB Journal
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    • v.14 no.6
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    • pp.677-681
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    • 1999
  • Purificationi of egg yolk immunoglobulin(IgY) was performed to understand the property of egg immunoglobulin. IgY differs from mammalian IgY in the molecular size(larger), isoelectric point(more acidic), and binding ability with mammalian complement and protein A(nonbinding ability). IgY is also known as ${\gamma}$-livetin and exists in egg yolk together with other two water-solubel proteins, ${\alpha}$-livetin(chicken serum albumin) and ${\beta}$-livetin(${\alpha}_2$-glycoprotein) and various lipoproteins(Low density lipoprotein, LDL and High density lipoprotein, HDL) which are the major components of egg yolk. The first step of isolation of IgY is to separate the water-solube proteins from lipoproteins. We report a simple method for separation of water soluble proteins using k-carrageenan and sedimentation. k-carrageenan was found to be effective for removal of yolk lipoprotein as a precipitate. IgY remained supernatant, and was isolated by chromatography on columns of DEAE-Sephacel and G 75 gel filtration chromatography.

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A Study on the Neutron Activation Analysis of Noble Metals in the Ancient Coin (고전(古錢)내 귀금속 원소의 중성자 방사화 분석에 관한 연구)

  • Kwon Soo Chun;Chul Lee;Myung-Zoon Czae;Jong Du Lee;Koo Soon Chung
    • Journal of the Korean Chemical Society
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    • v.37 no.11
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    • pp.961-966
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    • 1993
  • The determination of noble metals such as Ir, Au and Ag in the ancient coins has been studied. For the measurement of the activity of $^{192}Ir,\;^{198}Au\;and\;^{110m}Ag$, radiochemical separations including solvent extraction and ion-exchange chromatography were applied to reduce the interference of high energy ${\gamma}$-ray emitted from various radionuclides with long half-life. As a results, $10^{-11}$ g/g level of Ir could be detected and it was found that the three kinds of the detection limits, i.e., critical, detection, quantitative limit, calculated by the method proposed by Currie, were enhanced. Prior to the re-irradiation with neutron, inactive carrier was added in order to determine the recovery yield of Ir in the radiochemical separation. The average recovery yields of Ir, Au and Ag in the 5 coins were 65.3%, 98.5%, 99.5%, respectively.

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Development of an Enzyme-Linked Immunosorbent Assay for the Iletection of Aflatoxin $B_1$ (Aflatoxin $B_1$의 검출을 위한 효소면역측정법의 개발)

  • 손동화;박애란;서병철;김진철;이인원;남영중;허우덕
    • Microbiology and Biotechnology Letters
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    • v.20 no.2
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    • pp.225-232
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    • 1992
  • In order to develop an enzyme-linked immunosorbent assay(ELISA) for detecting aflatoxin $B_1(AFB_1)$, we produced and purified antibodies, thereafter established and evaluated methods of direct and indirect competitive ELISA. Anti-AFB, antisera, produced by immunizing rabbits with $AFB_1$-1-(0-carboxymethy1)oxime-bovine serum albumin conjugate ($AFB_1$-BSA), were removed of anti-BSA antibodies by quantitative precipitation reaction and further purified by ammonium sulfate precipitation and DEAE-Sephadex A-50 ion exchange chromatography. Purified IgG fractions were used as anti-$AFB_1$ antibodies. The antibodies, whose titer was deterrnined extremely high above $2 \times 10^6$, showed low cross-reactivity of 3~34% against $AFB_1$ analogues such as G2, B2, and GI. From the standard curves of direct and indirect competitive ELISA for AFBI, the detection ranges were found 0.2~20 and 1~10, 000 ng/ml(ppb) respectively. In their sensitivity, stability, simplicity, and rapidity, the direct method was more suitable than the indirect method for practical use.

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Study on the Separation of $^{55}Fe$, $^{90}FSr$$^{94}Nb$ in Radioactive Wastes (방사성 폐기물 내 $^{55}Fe$, $^{90}FSr$$^{94}Nb$의 분리 연구)

  • 이창헌;정기철;임석남;김원호;지광용
    • Proceedings of the Korean Radioactive Waste Society Conference
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    • 2003.11a
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    • pp.54-59
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    • 2003
  • Several radionuclides are considered as an object of the assesment to develop a scaling factor and a periodical verification method which are needed for the evaluation of radionuclide inventory of various radioactive wastes from nuclear power plants in Korea. A selective separation of $^{55}Fe$, $^{90}FSr$$^{94}Nb$ which should be recovered individually for the radiochemical analysis was described in detail. Sorption and desorption behaviours of ion exchange and extraction chromatographic resins for Fe, Sr, Nb and co-existing metal ions were Investigated using an artificial waste solution simulated of chemical composition of real radioactive wastes. Separation conditions available for the sequential recovery of these metal ions from a single sample were optimized to minimize a discharge of radioactive wastes produced through the analytical process and a radiation exposure to analysts. Their recovery yields were measured with reliability.

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Direct Determination of Tellurium in Simulated Nuclear Spent Fuels by Hydride Generation-Inductively Coupled Plasma Atomic Emission Spectrometry (수소화물 생성-유도결합플라스마 원자방출분광법을 이용한 모의사용후 핵연료 중의 텔루르 분석)

  • Choi, Kwang Soon;Lee, Chang Heon;Han, Sun Ho;Joe, Kih Soo;Kim, Won Ho
    • Analytical Science and Technology
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    • v.13 no.6
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    • pp.781-788
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    • 2000
  • Tellurium in simulated nuclear spent fuels (SIMFUEL) has been determined by hydride generation-inductively coupled plasma atomic emission spectrometry (HG-ICP-AES). Parameters such as concentrations of HCl and $NaBH_4$, flow rate of HCl and $NaBH_4$ were optimized and then the effects of U, Mo, Pd, Rh and Ru on the Te intensity were investigated. A thiourea as a masking agent was used to eliminate or minimize such interferences specially caused by palladium. Tellurium was measured by HG-ICP-AES and ICP-MS after separation of tellurium from SIMFUEL with cation exchange chromatography. The relative deviation between direct measurement and separation method was less than 6% based on the data by ICP-MS.

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