• Title/Summary/Keyword: High-purity purification

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Development of Purification Process of Recombinant Human Vascular Endotherial Growth Factor (VEGF) using Fusion Protein (융합 단백질을 이용한 재조합 인간 혈관내피세포 성장인자의 정제공정 개발)

  • Sung, Keehyun;Kim, In Ho
    • Korean Chemical Engineering Research
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    • v.55 no.3
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    • pp.369-378
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    • 2017
  • Vascular endotherial growth factor (VEGF) is a potent mitogen that stimulates vascular permeability and angiogenesis and has a potential in therapeutic applications. An industrial production method that provides high yield as well as purity is needed. Researches for various factors of mild solubilization with combination of ubiquitin fusion protein to increase solubility were carried out as well as by changing pH and denaturant concentration. Usage of pET28-a bacteral expression vector in BL21 (DE3) host cell was capable of producing approximately 14 g/L VEGF fusion protein in 20L fermentor. A purification process consisting of four chromatography steps including refolding and digestion with UBP1 resulted in mild solublization under the conditions of 2M urea and pH 10.0 due to ubiquitin fusion tag protein that increases in solubility of target protein VEGF. High yield of refolding and dimerization could be obtained between two step Ni-affinity chromatography. Multimeric and misfolded proteins and endotoxin were removed by DEAE anion exchange chromatography. Final monomers were removed from dimers by gel filtration chromatography. Characterization analysis of purified dimeric VEGF was performed using SDS-PAGE and RP-HPLC with a purity of 97%.

Purification and Characterization of Recombinant Human Follicle Stimulating Hormone Produced by Chinese Hamster Ovary Cells

  • NA KYU HEUM;KIM SEUNG CHUL;SEO KWANG SEOK;LEE SUNG HEE;KIM WON BAE;LEE KANG CHOON
    • Journal of Microbiology and Biotechnology
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    • v.15 no.2
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    • pp.395-402
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    • 2005
  • Biologically active recombinant human follicle stimulating hormone (rhFSH) was produced in Chinese hamster ovary cells and purified by a series of chromatographic steps. The chromatographic steps included anion-exchange chromatography (DEAE Sepharose F/F, Q Sepharose F/F), hydrophobic interaction chromatography (Source 15 PHE), and hydroxyapatite chromatography (Macro-Prep ceramic hydroxyapatite type I). A distinctive step of the purification process developed was the use of ZnCl$_2$ for the removal of non-glycosylated or lowly-glycosylated FSH and impurities through co-precipitation with Zn$^{2+}$. Purified rhFSH was identified and characterized by several physicochemical and biological methods such as gel electrophoresis, high-performance liquid chromatography, amino acid analysis, carbohydrate analysis, and biological activity. The overall yield of the purification was ~$30\%$. The rhFSH preparation obtained showed high purity (>$99\%$) and high in vivo potency (>16,000 IU/mg). Carbohydrate analysis suggested that the purified rhFSH contained approximately $40\%$ (w/w) carbohydrate with di­or tri-antennary structure on average, which is somewhat more heavily sialylated than commercially available rhFSH. In conclusion, the results of these analyses established an identity of the purified rhFSH with natural FSH from human pituitary glands, and furthermore, the purified rhFSH preparation showed higher in vivo potency and was slightly more heavily sialylated than commercially available rhFSH.

Improvement of the Fractional Precipitation Process for the Purification of (+)-Dihydromyricetin ((+)-Dihydromyricetin 정제를 위한 분별침전공정 개선)

  • Lim, Min-Kyoung;Kim, Jin-Hyun
    • Microbiology and Biotechnology Letters
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    • v.42 no.1
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    • pp.25-31
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    • 2014
  • Fractional precipitation is a simple method for purifying (+)-dihydromyricetin extracted from biomass. However, the fractional precipitation process has been inherently problematic due to the lengthy precipitation time that is required. The fractional precipitation time was shortened and (+)-dihydromyricetin yield was improved by increasing the surface area per working volume (S/V) of the reacting solution through the addition of a cation exchange resin (Amberlite 200, Amberlite IR 120Na, Amberlite IR 120H, or Amberlite IRC 50). Most of the (+)-dihydromyricetin (>90%) could be obtained after about 16 h of fractional precipitation using Amberlite 200. Since high-purity (+)-dihydromyricetin can be obtained at a high yield and the precipitation time can be reduced by increasing the surface area available for precipitation, this improved method is expected to minimize solvent usage and the size and complexity of the high performance liquid chromatography operation required for (+)-dihydromyricetin purification.

High Purity Hydrogen Generator for Fuel Cell Vehicles (연료전지 자동차 탑재형 고순도 수소생산장치)

  • Han, Jaesung;Lee, Seok-Min
    • Journal of Hydrogen and New Energy
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    • v.12 no.4
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    • pp.277-285
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    • 2001
  • We developed a compact, 10 kWe, purifier-integrated reformer which supplies hydrogen for fuel cell vehicles. Our proprietary technologies regarding hydrogen purification by palladium alloy membrane and catalytic combustion by noble metal coated wire-mesh catalyst were combined with the conventional methanol steam reforming technology, resulting in higher conversion, excellent quality of product hydrogen, and better thermal efficiency than any other systems. In this system, steam reforming, hydrogen purification, and catalytic combustion take place all in a single reactor so that the whole system is compact and easy to operate. The module produces $8.2Nm^3/hr$ of 99.999% or higher purity hydrogen with CO impurity less than 10 ppm, which is equivalent to 10 kWe when PEMFC has 45 % efficiency. Thermal efficiency of the module is 81 % and the power density of the module is 1.6 L/kWe. As the results of experiments, cold-start time has been measured about 20 minutes. Response time of hydrogen production to the change of the feed rate has been within 1 minutes.

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Simulation of IgY(Immunoglobulin Yolk) Purification by SMB(Simulated Moving Bed) (SMB(Simulated Moving Bed)를 이용한 IgY(Immunoglobulin Yolk) 분리의 전산모사)

  • Song, Sung-Moon;Kim, In-Ho
    • Korean Chemical Engineering Research
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    • v.49 no.6
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    • pp.798-803
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    • 2011
  • IgY(Immunoglobulin Yolk) is a specific antibody in egg yolk, and it protects human body from virus and antigen. There are a lot of egg yolk components such as lipoprotein and protein. To separate IgY, HPLC(High Performance Liquid Chromatography) and precipitation were used in a batch mode and SMB(Simulated Moving Bed) was adopted for continuous purification of yolk proteins. IgY and other proteins in yolk were separated by using three-zone SMB chromatography. Before performing SMB experiments, batch chromatography and PIM(pulse input method) were performed to find operation parameters and adsorption isotherms. The results of batch chromatography were compared with simulated results using Aspen chromatography. To find the most suitable separation condition in SMB chromatography, simulations in $m_2$-$m_3$ plane on the triangle theory were carried out. $m_2$ = 0.18, $m_3$ = 1.0 and ${\Delta}$t = 419 s are the best conditions for the highest purity of IgY. With this operating parameters(flow rate in three zone and switching time), the purity of raffinate results in 98.39% from Aspen chromatography simulation. Most of the simulation reached steadystate within second recycle.

Recovery of MFB Generated from Dimethyl Terephtalate Production Process (DMT 제조 과정서 발생하는 MFB의 회수에 관한 연구)

  • Kim, Sun Ho;Ryu, Young;Kim, Jong Cheon;Kim, Seok Chan
    • Applied Chemistry for Engineering
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    • v.26 no.5
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    • pp.621-623
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    • 2015
  • This article describes a purification method yielding high purity of MFB produced from DMT production process. Aldehyde functional group of MFB included in side-products were converted to acetal compound via reacting with methanol and further separated. Hydrolysis process of the acetal product was continued under acidic condition and highly pure MFB were obtained with 90% yield. The structure of MFB was analyzed by $^1H$ NMR and $^{13}C$ NMR spectroscopy. Also, the purity of MFB was estimated to be over 99% by GC analysis.

A Study on the Purification of Zn from Pb Splashing Alloy (Pb Splashing 합금으로부터 Zn정련에 관한 연구)

  • 박재욱;김용하;이대열;신형기;김진한;박성수;정원섭
    • Resources Recycling
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    • v.6 no.4
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    • pp.3-10
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    • 1997
  • Electric are furnace dust (EAF dust) generated in steel production based on scrap melting is contained Zn and Fe about 25 and 30 percent by weight, respectively. From a metallurgical point of view, the dust could be regarded as a raw material for Zn and Fe source. To recover the Zn in the metal from EAF dust, many system are proposed such as Arc Plasma Furmace and Pb splasher method. In this study, to recover high purity Zn from Pb splasing alloy, Zn distillation is carried out at the temperature of 1123, 1173, 1223, 1273 K, the gas flow rate of 2.5, 5.0, 8.0 Ni/min and the distilling time of 10, 30, 60, 90 minutes. The main results obtained from this study are as follows:(1) The amount of evaporated Zn and its evaporating rate increased with increasing temperature, but purity of Zn decreased with increasing temperature. Optimum temperature range was found out to be between 1173∼1223K. (2) The amount of evaporated Zn and evaporation rate increased with increasing gas flow rate at a given temperature and distillation time. Gas flow rate has more influence over the amount of evaporated Zn and evaporation rate with increasing temperature.

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Radiochemical separation of 89Zr: a promising radiolabel for immuno-PET

  • Vyas, Chirag K.;Park, Jeong Hoon;Yang, Seung Dae
    • Journal of Radiopharmaceuticals and Molecular Probes
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    • v.2 no.1
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    • pp.43-50
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    • 2016
  • $^{89}Zr$ with the favorable nuclear decay kinetics and chemical properties is an appealing radiometal for its application in immuno-PET using radiolabeled monoclonal antibodies. Rising demand of ultrahigh purity and high-specific activity $^{89}Zr$ has propelled the radiochemist worldwide to develop an overall efficacious method for its promising separation from the target matrix $^{89}Y$. The requirement of elevated radiochemical purity (${\geq}$ 99.99%) has accelerated the efforts since last two decades to achieve higher decontamination and separation factors of carrier free $^{89}Zr$ over $^{89}Y$ using several suitable separation techniques. However, each of the technique has its own pros and cons which prior to its actual medical application needs to be optimized and thoroughly scrutinized to avoid further complications during radiolabelling of the pharmaceuticals. In this short review article we will specifically consider as well focus on the historical development and the recent advances on the radiochemical separation of $^{89}Zr$ from $^{89}Y$ which will be helpful for the separation scientist involved in this area to understand the existing available means and plan the strategy to investigate and develop the novel techniques to overcome the problems involved in the present methods.

Preparation of High Purity α-Alumina from Aluminum Black Dross by Redox Reaction (알루미늄 블랙 드로스로부터 산화 환원반응을 이용한 고순도 알파 알루미나의 제조)

  • Shin, Eui-Sup;An, Eung-Mo;Lee, Su-Jeong;Ohtsuki, Chikara;Kim, Yun-Jong;Cho, Sung-Baek
    • Korean Journal of Materials Research
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    • v.22 no.9
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    • pp.445-449
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    • 2012
  • We investigate the effects of redox reaction on preparation of high purity ${\alpha}$-alumina from selectively ground aluminum dross. Preparation procedure of the ${\alpha}$-alumina from the aluminum dross has four steps: i) selective crushing and grinding, ii) leaching process, iii) redox reaction, and iv) precipitation reaction under controlled pH. Aluminum dross supplied from a smelter was ground to separate metallic aluminum. After the separation, the recovered particles were treated with hydrochloric acid(HCl) to leach aluminum as aluminum chloride solution. Then, the aluminum chloride solution was applied to a redox reaction with hydrogen peroxide($H_2O_2$). The pH value of the solution was controlled by addition of ammonia to obtain aluminum hydroxide and to remove other impurities. Then, the obtained aluminum hydroxide was dried at $60^{\circ}C$ and heat-treated at $1300^{\circ}C$ to form ${\alpha}$-alumina. Aluminum dross was found to contain a complex mixture of aluminum metal, aluminum oxide, aluminum nitride, and spinel compounds. Regardless of introduction of the redox reaction, both of the sintered products are composed mainly of ${\alpha}$-alumina. There were fewer impurities in the solution subject to the redox reaction than there were in the solution that was not subject to the redox reaction. The impurities were precipitated by pH control with ammonia solution, and then removed. We can obtain aluminum hydroxide with high purity through control of pH after the redox reaction. Thus, pH control brings a synthesis of ${\alpha}$-alumina with fewer impurities after the redox reaction. Consequently, high purity ${\alpha}$-alumina from aluminum dross can be fabricated through the process by redox reaction.

A Novel Simple Method to Purify Recombinant Soluble Human Complement Receptor Type 1 (sCR 1) from CHO Cell Culture

  • Wang, Pi-Chao;Hisamune Kato;Takehiro Inoue;Masatoshi Matsumura;Noriyuki Ishii;Yoshinobu Murakami;Tsukasa Seya
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.7 no.2
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    • pp.67-75
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    • 2002
  • The human complement receptor type 1 (CR 1, C3 b/C4b receptor) is a polymorphic membrane glycoprotein expressed on human erythrocytes, peripheral leukocytes, plasma and renal glomerular podocytes, which consists of transmembrane and cytoplasmic domains with 30 repeating homologous protein domains known as short consensus repeats (SCR). CR1 has been used as an inhibitor for inflammatory and immune system for the past several years. Recently; it is reported that CRl was found to suppress the hyper-acute rejection in xeno-transplantation and can be used to cure autoimmune diseases. A soluble form of CRl, called sCRl, is a recombinant CRl by cleaving the transmembrane domain at C-terminus and has been expressed in Chinese Hamster Ovary (CHO) cells. Several purification methods for sCR1 from CHO cells have been reported, but most of them require complicated steps at high cost. Moreover, such methods are mostly performed under the pH condition apt to denaturing sCR1 and causes sCRl losing its activity. We here report a rapid and efficient method to purify sCR1 from CHO cell. The new method consists of a two-stage of cell culture by cultivating cells in serum medium followed by serum-free medium, and a two-stage of column purification by means of heparin and gel filtration column chromatography. By using this novel method, sCR1 can be purified in a simple and effective way with high yield and purity, furthermore, the purified sCR1 was confirmed to retain its activity to suppress the complement activation in vivo and ex vivo.