• Title/Summary/Keyword: high-purity purification

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Novel Modification of Growth Medium Enables Efficient E. coli Expression and Simple Purification of an Endotoxin-Free Recombinant Murine Hsp70 Protein

  • Zachova, Katerinat;Krupka, Michal;Chamrad, Ivo;Belakova, Jana;Horynova, Milada;Weigl, Evzen;Sebela, Marek;Raska, Milan
    • 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.

A New large-scale Pre-purification for Peroxidase from Plant Cell Cultures (식물세포 배양으로부터 Peroxidase 대량 정제를 위한 전처리 공정 개발)

  • 표상현
    • KSBB Journal
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    • v.15 no.4
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    • pp.342-345
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    • 2000
  • A novel pre-purification method was developed for producing peroxidase to guarantee high purity and yield from plant cell cultures in large-scale process. This method was a simple and efficient procedure for the isolation and pre-purification of peroxidase from the biomass consisting of active clay treatment followed by cationic exchange chromatography. The use of active clay in the pre-purification process allows for rapid and efficient separation of peroxidase from interfering compounds and dramatically increases yield and purity of crude peroxidase for purification steps compared to alternative processes. This pre-purification process serves to minimize the buffer usage size and complexity of the HPLC operations for peroxidase purification. This process is readily scalable to a pilot plant and eventually to a production environment where mass production of material are expected to be produced.

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Mineralogical Analysis and Mechano-Chemical Purification of Natural Silica Ore for High Purity Silica Powder

  • Park, Jesik;Lee, Churl Kyoung;Lee, Hyun-Kwon
    • Korean Journal of Materials Research
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    • v.26 no.6
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    • pp.306-310
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    • 2016
  • To produce 4N grade high-purity silica powder from natural ore, the mineralogical characteristics of natural silica ore were investigated and their effects on the purification process were revealed. The Chinese silica mineral ore used was composed of iron and aluminum as main impurities and calcium, magnesium, potassium, sodium, and titanium as trace impurities; these trace impurities generally exist as either single oxides or complex oxides. It was confirmed that liberation and acidic washing of the impurities were highly dependent on the particle size of the ground silica ore and on its mineralogical characteristics such as the distribution and phases of existing impurities. It is suggested that appropriate size reduction of silica ore should be realized for optimized purification according to the origin of the natural silica ore. A single step purification process, the mechano-chemical washing (MCW) process, was proposed and verified in comparison with the conventional multi step washing process.

Microbiological Purification of L-Arabitol from Xylitol Mother Liquor

  • Jiang, Mingguo;Wang, Ben;Yang, Lifang;Lin, Shuangjun;Cheng, Hairong
    • Journal of Microbiology and Biotechnology
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    • v.21 no.1
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    • pp.43-49
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    • 2011
  • As a rare sugar alcohol, L-arabitol can be used in food and can prevent extra fat deposits in the intestinal tract. Commercially, L-arabitol is prepared from pure L-arabinose by hydrogenation, which needs a high temperature and high pressure, leading to a high production cost for Larabitol. Therefore, this study describes a novel L-arabitol production method based on biological purification from the xylitol mother liquor, a cheap and readily available raw material that contains a high concentration of Larabitol. First, a novel Bacillus megaterium strain was screened that can utilize xylitol, sorbitol, and mannitol, yet not L-arabitol. The isolated strain was inoculated into a medium containing the xylitol mother liquor under formulated culture conditions, where a high L-arabitol yield (95%) and high purity (80%) were obtained when the medium was supplemented with 50 g/l of xylitol mother liquor. Upon further purification of the fermentation broth by ion exchange and decolorization, L-arabitol was crystallized with a purity of 98.5%.

Preparation of High-Purity Urokinase Using Single-Step Hydrophobic Interaction Chromatography with p-Aminobenzamidine Ligand

  • Cao, Xue-Jun;Zhou, Jian-Hua;Huang, Zhen-Hui;Wu, Xing-Yan;Hur, Byung-Ki
    • Journal of Microbiology and Biotechnology
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    • v.12 no.2
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    • pp.196-203
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    • 2002
  • A novel process for urokinase purification was studied using p-aminobenzamidine as the ligand and sepharose 4B as the matrix. The adsorption, washing, and elution conditions were optimized by an unusual method. An adsorption buffer containing 2.5 M NaCl and $1\%$ Tween 80 facilitated the adsorption of urokinase on the affinity media and prevented contaminants from binding to the p-aminobenzamidine affinity gel. It was found that $5\%$ Tween 80 removed most of the contaminants from the affinity column. A 0.2 M glycine elution buffer containing 0.5 M NaCl (pH 3.0) was found to have a strong elution ability with a high recovery and purity of urokinase. A crude urokinase material of231 IU/mg protein from human urine was purified to 124,300 IU/mg protein with a purification factor of 538 and yield of $86.7\%$. As a result, a high purity urokinase was obtained with only a single affinity chromatography step. The purification process was successfully scaled-up to a 2-1 chromatography column. The resulting urokinase eluate could be directly lyophilized, thereby complying with Chinese pharmacopoeia (1995 version) standards.

Hydrogen Purification by the Four-Bed Pressure Swing Adsorption Process from Steam Methane Reforming Off-Gas (4탑 PSA 공정의 의한 SMR off-gas로부터 수소 정제)

  • Yang, Se-Il;Park, Ju-Yong;Jang, Seong-Cheol;Kim, Sung-Hyun;Choi, Dae-Ki
    • 한국신재생에너지학회:학술대회논문집
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    • 2008.10a
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    • pp.383-386
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    • 2008
  • The four-bed PSA process using a layered bed of activated carbon and zeolite 5A was studied to produce a high purity hydrogen product from SMR off-gas. At a desired product purity (99.999%+), the recovery increased with decreasing the linear velocity. However, the difference of the increasing of the recovery became smaller with the decreasing of the linear velocity and then was similar from below the linear velocity 3.9 cm/s. When the adsorbents, the feed gas composition, and the operating conditions are given, the residence time is mainly a function for design of the PSA bed size. The minimum residence time exists to obtain the maximum recovery at desired product purity.

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High-Purity Purification of Indole Contained in Coal Tar Fraction - Separation of Close Boiling Mixtures of Indole by Solute Crystallization - (콜타르 유분 중에 함유된 인돌의 고순도 정제 - 용액 결정화에 의한 인돌 유사 비점 혼합물의 분리 -)

  • Kim, Su Jin;Kang, Ho-Cheol;Jeong, Hwa Jin
    • Applied Chemistry for Engineering
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    • v.21 no.2
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    • pp.238-241
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    • 2010
  • The purification of indole from 54.3wt% indole fraction (temperature range of distillate: $250{\sim}255^{\circ}C$) recovered by extraction-distillation combination of coal tar fraction (temperature range of distillate: $240{\sim}265^{\circ}C$) was examined by solute crystallization. The feed consists of eight components such as quinoline, iso-quinoline, indole, quinaldine, 1-methylnaphthalene, 2-methylnaphthalene, biphenyl and phenyl ether. Hexane and an aqueous solution of methanol (50 : 50 vol%) were used as the crystallization solvent and the coolant, respectively. A batch stirred tank of glass material was used as a crystallization apparatus. By increasing the operation temperature and the volume ratio of solvent to feed at initial, the purity of indole increas ed, but yields of indole showed a decreasing tendency. Solute crystallization method using hexane as a solvent was excellent because the purity of 99.3 wt% indole was recovered at the yield of 50% without washing operation.

Methods for High Quality Purification of Alginate and Fucoidan from Marine Brown Algae (해조류로부터 푸코이단/알지네이트의 고순도 분리정제)

  • Kim, Jong-Ki;Yang, Jae-Ho
    • Journal of Marine Bioscience and Biotechnology
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    • v.2 no.4
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    • pp.245-249
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    • 2007
  • High quality of purified alginate and fucoidan is required for the medical uses to prevent the unexpected side effects from the contaminated endotoxin present in the materials. We attempted to establish an efficient and fast mass scale production method for the highly purified poly-G alginate from the sea weeds. In addition, lab scale method was established to obtain the fraction of higher purity from the commercially available fucoidan source (about 85% purity).

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Preparation of High Purity Ammonium Dinitramide and Its Liquid Mono-propellant (암모늄 디나이트라마이드염의 합성 및 액상연료화 연구)

  • Kim, Wooram;Park, Mijeong;Kim, Sohee;Jeon, Jong-Ki;Jo, Youngmin
    • Applied Chemistry for Engineering
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    • v.30 no.5
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    • pp.591-596
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    • 2019
  • A recently developed propellant, ammonium dinitramide (ADN, $NH_4N(NO_2)_2$ is stable and safe at an ambient condition. However, it requires high purity for practical applications. A very little quantity of foreign impurities in ADN may cause clogging of thruster nozzles and catalyst poisoning for the use of a liquid propellant. Thus, several purification processes for precipitated ADN particles such as repetition extraction, activated carbon adsorption and low-temperature extraction were presented in this study. The purifying methods helped to improve the chemical purity as evaluated by FT-IR and UV-Vis spectroscopy in addition to ion chromatography (IC) analyses. Among the purification processes, adsorption was found to be the best, showing a final purity of 99.8% based on relative quantification by IC. Thermal analysis revealed an exothermic temperature of $148^{\circ}C$ for the synthesized liquid monopropellant, but rose to $188^{\circ}C$ when urea was added.

Electrical Properties of Transparent Conductive Films of Single-Walled Carbon Nanotubes with Their Purities

  • Lee, Seung-Ho;Goak, Jeung-Choon;Lee, Chung-Yeol;Lee, Nae-Sung
    • Proceedings of the Korean Institute of Electrical and Electronic Material Engineers Conference
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    • 2010.06a
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    • pp.56-56
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    • 2010
  • Single-walled carbon nanotubes (SWCNTs) have attracted much attention as a promising material for transparent conducting films (TCFs), due to their superior electrical conductivity, high mechanical strength, and complete flexibility as well as their one-dimensional morphological features of extremely high length-to-diameter ratios. This study investigated three kinds of SWCNTs with different purities: as-produced SWCNTs (AP-SWCNTs), thermally purified SWCNTs (TH-SWCNTs), thermally and acid purified SWCNTs (TA-SWCNTs). The purity of each SWCNT sample was assessed by considering absorption peaks in the semiconducting ($S_{22}$) and metallic ($M_{11}$) tubes with UV-Vis NIR spectroscopy and a metal content with thermogravimetric analysis (TGA). The purity increased as proceeding the purification stages from the AP-SWCNTs through the thermal purification to the acid purification. The samples containing different contents of SWCNTs were dispersed in water using sodium dodecyl benzensulfate (SDBS). Aqueous suspensions of different purities of SWCNTs were prepared to have similar absorbances in UV-Vis absorption measurements so that one can make the TCFs possess similar optical transmittances irrespective of the SWCNT purity. Transparent conductive SWCNT networks were formed by spraying an SWCNT suspension onto a poly(ethyleneterephthalate) (PET) substrate. As expected, the TCFs fabricated with AP-SWCNTs showed very high sheet resistances. Interestingly, the TH-SWCNTs gave lower sheet resistances to the TFCs than the TA-SWCNTs although the latter was of higher purity in the SWCNT content than the former. The TA-SWCNTs would be shortened in length and be more bundled by the acid purification, relative to the TH-SWCNTs. For both purified (TH, TA) samples, the subsequent nitric acid ($HNO_3$) treatment greatly lowered the sheet resistances of the TCFs, but almost eliminated the difference of sheet resistances between them. This seems to be because the electrical conductivity increased not only due to further removal of surfactants but also due to p-type doping upon the acid treatment. The doping effect was likely to overwhelm the effect of surfactant removal. Although the nitric acid treatment resulted in the similar. electrical properties to the two samples, the TCFs of TH-SWCNTs showed much lower sheet resistances than those of the TA-SWCNTs prior to the acid treatment.

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