• Korean Journal of Fisheries and Aquatic Sciences
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• v.26 no.6
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• pp.529-537
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• 1993

Several methods were examined for purification of docosahexaenoic acid (DHA, 22:6n-3) from skipjack Euthynnus pelamis orbital tissue oil, a marine by-product, and a modified method for isolation of a high purity DHA was proposed. Skipjack orbital tissue contained $55.4\%$ of total lipid(TL), and DHA accounted for $23.7\%$ of the TL. Application of low-temperature crystallization and urea inclusion compound methods to the orbital fatty acid mixture resulted in increases of DHA concentrations to approximately $46\%\;and\;61\%$, respectively. These methods were suitable for large production of DHA with relative low purity because of the simple purification procedure. DHA of approximately $74\%$ in purity was obtained by silver nitrate aqueous solution method, but the method gave a very low recovery($<10\%$). Silver nitrate-impregnated silica column chromatography was suitable for purification of a high purity DHA(purity, $>98\%$ and recovery, $>90\%$) A modified method, silver nitrate-impregnated silica column chromatography combined with low-temperature crystallization(two step purication method) was proposed as the most effective method to obtain DHA with high purity($99.9\%$) from the skipjack orbital oil.

• Journal of Microbiology and Biotechnology
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• v.19 no.7
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• pp.727-733
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• 2009

Heat shock protein 70 kDa (hsp70), a molecular chaperone involved in folding of nascent proteins, has been studied for its ability to activate innate and specific immunity. High purity hsp70 preparation is generally required for immunization experiments, because endotoxins and other immunologically active contaminants may affect immune responses independently of hsp70. We have developed a novel modification of E. coli-expression medium that enabled a simple two-step production and purification method for endotoxin-free recombinant hsp70. During Ni-NTA-based affinity purification of hsp70, a contaminating protein from host E. coli cells, L-glutamine-n-fructose-6-phosphate aminotransferase (GFAT), was identified. By testing various compounds, supplementation of growth medium with a GFAT metabolite,N-acetylglucosamine, was found to reduce GFAT expression and increase the total hsp70 yield five times. The new protocol is based on column purification of His-tagged hsp70 protein produced by E. coli with the modified medium, followed by endotoxin removal by Triton X-114 extraction. This approach yielded hsp70 with high purity and minimal endotoxin contamination, making the final product acceptable for immunization experiments. In summary, a simple modification of growth medium allowed production of recombinant mouse hsp70 in high yield and purity, thus compatible with immunological studies. This protocol may be useful for production of other Histagged proteins expressed in E. coli.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.4
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• pp.574-579
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• 2014

A lysozyme gene from breast of Tibetan sheep was successfully expressed by secretion using a-factor signal sequence in the methylotrophic yeast, Pichia pastoris GS115. An expression yield and specific activity greater than 500 mg/L and 4,000 U/mg was obtained. Results at optimal pH and temperature showed recombinant lysozyme has higher lytic activity at pH 6.5 and $45^{\circ}C$. This study demonstrates the successful expression of recombinant lysozyme using the eukaryotic host organism P. pastoris paving the way for protein engineering. Additionally, this study shows the feasibility of subsequent industrial manufacture of the enzyme with this expression system together with a high purity scheme for easy high-yield purification.

• 한국생물공학회:학술대회논문집
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• 2005.04a
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• pp.351-354
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• 2005

A new isolation and purification method was developed aiming at increasing yield and purity for homoharringtonine. This method was a simple andefficient procedures, for the isolation and purification of homoharringtonine from Cephalotaxus koreana, consisting of solvent extraction, adsorbent treatment, low-pressure chromatography, and high performance liquid chromatography (HPLC). The crude homoharringtonine was efficiently pre-purified adequately to perform HPLC through a combination withadsorbent treatment and low-pressure chromatogaphy. The homoharringtonine can be simply obtained with high purity and yield from crude homoharringtonine by HPLC. Purified homoharringtonine was characterized.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.1
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• pp.13-16
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• 2000

Simple procedures have been devised for purifying recombinant human interleukin-2 (hIL-2), which was expressed in Escberichia coli using sequences of glucagon molecules and enterokinase cleavage site as an N-terminus fusion partner. The insoluble aggregates of recombinant fusion protein produced in E. coli cytoplasm were easily dissolved by simple alkaline pH shift $(8\rightarrow12\rightarrow8)$. Following enterokinase cleavage, the recombinant hIL-2 was finally purified by one-step reversed-phase HPLC with high purity. The ease and high efficiency of this simple purification process seem to mainly result from the role of used glucagon fusion partner, which could be applied to the production of other therapeutically important proteins.

• Journal of the Korean Applied Science and Technology
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• v.24 no.2
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• pp.140-148
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• 2007

As an additional high purification method of p-dioxanone monomer for a high molecular weight polymer, the sweating operation of crystalline layer obtained by layered melt crystallization from p-dioxanone-diethylene glycol system was studied. Purity and yield of p-dioxanone crystal depended mainly on the sweating temperature and sweating time. Increasing sweating time and sweating temperature, the purity of p-dioxanone crystal increase, whereas the yield of that decrease, respectively. Through the optimization of sweating operation, p-dioxanone crystal can be upgraded to very high purity over 99.9 % suited to monomer for polymerization.

• Bulletin of the Korean Chemical Society
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• v.35 no.9
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• pp.2665-2668
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• 2014

Aptamers are oligonucleotides or peptide molecules that are able to bind to their specific target molecules with high affinity via molecular recognition. In this study, we present development of aptamer-based precipitation assays (or simply aptamoprecipitation) for His-tagged proteins and thrombin to compare their purification efficiency with other conventional affinity precipitation methods. A crosslinking method was employed to immobilize thiol-functionalized aptamers onto the surface of polystyrene resins, enabling them to specifically bind to His-tag and to thrombin, respectively. The resulting aptamer-functionalized resins were successfully applied via a one-step experiment to purification of His-tagged proteins from complex E. coli and to thrombin extraction, exhibiting superior or at least comparable purification results to the conventional immobilized metal affinity precipitation or immunoprecipitation.

• Journal of Microbiology
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• v.35 no.2
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• pp.97-102
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• 1997

Cysteine desulfhydrase (EC 4.4.1.1.) was purified from the culture supernatant of Streptomyces albidoflavus SMF301 by hydroxyapatite, gel filtration and Resource Q ion-exchange chromatography with a purification fold of six identical subunits. The enzyme was stabilized by dithiothreitol and pyridoxal 5'-phosphate during the purification procedures. The optimum pH and temperature were pH 8.6 and 35$^{\circ}C$, respectively. The N-terminal amino acid sequence was identified as A-P-L-P-T-A-D-V-R-S-D-P-G-Y-R-E-W-L-G-E-A-V. The purified cystein desulfhydrase had a high substrate specificity toward cysteine, and exhibited no cystahionine $\gamma$-lyase activity. The $K_m$ value for cysteine was determined to be 0.37 mM.

• Preventive Nutrition and Food Science
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• v.1 no.1
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• pp.106-110
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• 1996

Bromelain(EC 3.4.22.4), the collective name for the proteolytic enzymes found in tissues of the plant family Bromeliaceae(pineapple), has been used as a tenderizing agent in food processing, and as an antiinflammatory agent in pharmaceuticals. In this paper, we describe the simple purification method of bromelain using Korean pineapple fruit. After removing contaminants at 30% saturation of ammonium sulfate, the supernatant obtained was treated again with ammonium sulfate to 80% saturation. Wit the above salt fractionation, partially purified bromelain could be obtained. To get highly purified bromelain, the previous 30% to 80% ammonium sulfate treated precipitate was dialyzed against 25mM sodium acetate buffer(pH 5.0) followed by passing through a CM- cellulose cation exchange column. Fruit bromelain was eluted as a major peak at 0.5~0.8M NaCI gradient. The present method is simpler with high wield than the traditional purification method-acetone treatment and several consecutive chromatographic processes.

• Korean Journal of Ecology and Environment
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• v.35 no.4 s.100
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• pp.285-294
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• 2002

The purpose of this study was to evaluate the relationships between purification characteristics and hydraulic conditions, and to clarify the basic and essential factors required to be considered in the construction and management of artificial wetland system for the improvement of reservoir water quality. The artificial wetland system was composed of a pumping station and six sequential plants beds with five species of macrophytes: Oenanthe javanica, Acorus calamus, Zizania latifolia, Typha angustifolia, and Phragmites australis. The system was operated on free surface-flow system, and operation conditions were $3,444-4,156\; m^3/d$ of inflow rate, 0.5-2.0 hr of HRT, 0.1-0.2 m of water depth, 6.0-9.4 m/d of hydraulic loading, and relatively low nutrients concentration (0.224-2.462 mgN/L, 0.145-0.164 mgP/L) of inflow water. The mean purification efficiencies of TN ranged from 12.1% to 14.3% by showing the highest efficiency at the Phragmites australis bed, and these of TP were 6.3-9.5% by showing the similar ranges of efficiencies among all species. The mean purification efficiencies of SS and Chl-A ranged from 17.4% to 38.5% and from 12.0% to 20.2%, respectively, and the Oenanthe javanica bed showed the highest efficiency with higher concentration of influent than others. The mean purification amount per day of each pollutant were $9.8-4.1\;g{\cdot}m^{-2}{\cdot}d^{-1}$ in BOD, $1.299-2.343\;g{\cdot}m^{-2}{\cdot}d^{-1}$ in TN, $0.085-1.821\;g{\cdot}m^{-2}{\cdot}d^{-1}$ in TP, $17.9-111.6\;g{\cdot}m^{-2}{\cdot}d^{-1}$ in SS and $0.011-0.094\;g{\cdot}m^{-2}{\cdot}d^{-1}$ in Chl-a. The purification amount per day of TN revealed the hi링hest level at the Zizania latifolia bed, and TP showed at the Acrous calamus bed. SS and Chl-a, as particulate materials, revealed the highest purification amount per day at the Oenanthe javanica bed that was high on the whole parameters. It was estimated that the purification amount per day was increased with the high concentration of influent and shoot density of macrophytes, as was shown in the purification efficiency. Correlation coefficients between purification efficiencies and hydraulic conditions (HRT and inflow rate) were 0.016-0.731 of $R^2$ in terms of HRT, and 0.015-0.868 of $R^2$ daily inflow rate. Correlation coefficients of purification amounts per day with hydraulic conditions were 0.173-0.763 of Ra in terms of HRT, and 0.209-0.770 daily inflow rate. Among the correlation coefficients between purification efficiency and hydraulic condition, the percentages of over 0.5 range of $R^2$ were 20% in HRT and in daily inflow rate. However, the percentages of over 0.5 range of correlation coefficients ($R^2$) between purification amount per day and hydraulic conditions were 53% in HRT and 73% in daily inflow rate. The relationships between purificationamount per day and hydraulic condition were more significant than those of purifi-cation efficiency. In this study, high hydraulic conditions (HRT and inflow rate) are not likely to affect significantly the purification efficiency of nutrient. Therefore, the emphasis should be on the purification amounts per day with high hydraulicloadings (HRT and inflow rate) for the improvement of eutrophic reservoir withrelatively low nutrients concentration and large quantity to be treated.