40 CFR 60.601 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-07-01

... means the water, solvent, or other chemical bath into which the polymer or prepolymer (partially reacted..., transportation, collection, concentration, and purification of organic solvents. It may include enclosures, hoods...

40 CFR 60.601 - Definitions.

Code of Federal Regulations, 2013 CFR

2013-07-01

... means the water, solvent, or other chemical bath into which the polymer or prepolymer (partially reacted..., transportation, collection, concentration, and purification of organic solvents. It may include enclosures, hoods...

40 CFR 60.601 - Definitions.

Code of Federal Regulations, 2012 CFR

2012-07-01

... means the water, solvent, or other chemical bath into which the polymer or prepolymer (partially reacted..., transportation, collection, concentration, and purification of organic solvents. It may include enclosures, hoods...

40 CFR 60.601 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-07-01

... means the water, solvent, or other chemical bath into which the polymer or prepolymer (partially reacted..., transportation, collection, concentration, and purification of organic solvents. It may include enclosures, hoods...

Purification of semiconducting single-walled carbon nanotubes by spiral counter-current chromatography.

PubMed

Knight, Martha; Lazo-Portugal, Rodrigo; Ahn, Saeyoung Nate; Stefansson, Steingrimur

2017-02-03

Over the last decade man-made carbon nanostructures have shown great promise in electronic applications, but they are produced as very heterogeneous mixtures with different properties so the achievement of a significant commercial application has been elusive. The dimensions of single-wall carbon nanotubes are generally a nanometer wide, up to hundreds of microns long and the carbon nanotubes have anisotropic structures. They are processed to have shorter lengths but they need to be sorted by diameter and chirality. Thus counter-current chromatography methods developed for large molecules are applied to separate these compounds. A modified mixer-settler spiral CCC rotor made with 3 D printed disks was used with a polyethylene glycol-dextran 2-phase solvent system and a surfactant gradient to purify the major species in a commercial preparation. We isolated the semi-conducting single walled carbon nanotube chiral species identified by UV spectral analysis. The further development of spiral counter-current chromatography instrumentation and methods will enable the scalable purification of carbon nanotubes useful for the next generation electronics. Copyright © 2016 Elsevier B.V. All rights reserved.

Purification of Carbon Nanotubes: Alternative Methods

NASA Technical Reports Server (NTRS)

Files, Bradley; Scott, Carl; Gorelik, Olga; Nikolaev, Pasha; Hulse, Lou; Arepalli, Sivaram

2000-01-01

Traditional carbon nanotube purification process involves nitric acid refluxing and cross flow filtration using surfactant TritonX. This is believed to result in damage to nanotubes and surfactant residue on nanotube surface. Alternative purification procedures involving solvent extraction, thermal zone refining and nitric acid refiuxing are used in the current study. The effect of duration and type of solvent to dissolve impurities including fullerenes and P ACs (polyaromatic compounds) are monitored by nuclear magnetic reasonance, high performance liquid chromatography, and thermogravimetric analysis. Thermal zone refining yielded sample areas rich in nanotubes as seen by scanning electric microscopy. Refluxing in boiling nitric acid seem to improve the nanotube content. Different procedural steps are needed to purify samples produced by laser process compared to arc process. These alternative methods of nanotube purification will be presented along with results from supporting analytical techniques.

Enhanced fluidity liquid chromatography of inulin fructans using ternary solvent strength and selectivity gradients.

PubMed

Bennett, Raffeal; Olesik, Susan V

2018-01-25

The value of exploring selectivity and solvent strength ternary gradients in enhanced fluidity liquid chromatography (EFLC) is demonstrated for the separation of inulin-type fructans from chicory. Commercial binary pump systems for supercritical fluid chromatography only allow for the implementation of ternary solvent strength gradients which can be restrictive for the separation of polar polymeric analytes. In this work, a custom system was designed to extend the capability of EFLC to allow tuning of selectivity or solvent strength in ternary gradients. Gradient profiles were evaluated using the Berridge function (RF 1 ), normalized resolution product (NRP), and gradient peak capacity (P c ). Selectivity gradients provided the separation of more analytes over time. The RF 1 function showed favor to selectivity gradients with comparable P c to that of solvent strength gradients. NRP did not strongly correlate with P c or RF 1 score. EFLC with the hydrophilic interaction chromatography, HILIC, separation mode was successfully employed to separate up to 47 fructan analytes in less than 25 min using a selectivity gradient. Copyright © 2017 Elsevier B.V. All rights reserved.

[Purification Technology Optimization for Saponins from Ziziphi Spinosae Semen with Macroporous Adsorption Resin by Box-Behnken Design-Response Surface Methodology].

PubMed

Zhao, Hui-ru; Ren, Zao; Liu, Chun-ye

2015-04-01

To compare the purification effect of saponins from Ziziphi Spinosae Semen with different types of macroporous adsorption resin, and to optimize its purification technology. The type of macroporous resins was optimized by static adsorption method. The optimum technological conditions of saponins from Ziziphi Spinosae Semen was screened by single factor test and Box-Behnken Design-Response Surface Methodology. AB-8 macroporous resin had better purification effect of total saponins than other resins, optimum technological parameters were as follows: column height-diameter ratio was 5: 1, the concentration of sample solution was 2. 52 mg/mL, resin adsorption quantity was 8. 915 mg/g, eluted by 3 BV water, flow rate of adsorption and elution was 2 BV/h, elution solvent was 75% ethanol, elution solvent volume was 5 BV. AB-8 macroporous resin has a good purification effect on jujuboside A. The optimized technology is stable and feasible.

Process for the removal of impurities from combustion fullerenes

DOEpatents

Alford, J. Michael; Bolskar, Robert

2005-08-02

The invention generally relates to purification of carbon nanomaterials, particularly fullerenes, by removal of PAHs and other hydrocarbon impurities. The inventive process involves extracting a sample containing carbon nanomaterials with a solvent in which the PAHs are substantially soluble but in which the carbon nanomaterials are not substantially soluble. The sample can be repeatedly or continuously extracted with one or more solvents to remove a greater amount of impurities. Preferred solvents include ethanol, diethyl ether, and acetone. The invention also provides a process for efficiently separating solvent extractable fullerenes from samples containing fullerenes and PAHs wherein the sample is extracted with a solvent in which both fullerenes and PAHs are substantially soluble and the sample extract then undergoes selective extraction to remove PAHs. Suitable solvents in which both fullerenes and PAHs are soluble include o-xylene, toluene, and o-dichlorobenzene. The purification process is capable of treating quantities of combustion soot in excess of one kilogram and can produce fullerenes or fullerenic soot of suitable purity for many applications.

Preparative isolation and purification of astaxanthin from the microalga Chlorococcum sp. by high-speed counter-current chromatography.

PubMed

Li, H B; Chen, F

2001-08-03

High-speed counter-current chromatography was applied to the isolation and purification of astaxanthin from microalgae. The crude astaxanthin was obtained by extraction with organic solvents after the astaxanthin esters were saponified. Preparative high-speed counter-current chromatography with a two-phase solvent system composed of n-hexane-ethyl acetate-ethanol-water (5:5:6.5:3, v/v) was successfully performed yielding astaxanthin at 97% purity from 250 mg of the crude extract in a one-step separation.

Extraction and Isolation of Antineoplastic Pristimerin from Mortonia greggii (Celastraceae).

PubMed

Mejia-Manzano, Luis Alberto; Barba-Dávila, Bertha A; Gutierrez-Uribe, Janet A; Escalante-Vázquez, Edgardo J; Serna-Saldivar, Sergio O

2015-11-01

The aim of this research was to identify, extract and isolate pristimerin in leaves, stems and roots of the Mexican plant Mortonia greggii (Celastraceae). The principal objective was to determine the best laboratory experimental conditions for the extraction and isolation of this powerful natural anticancer agent from the root tissue. Six experimental factors in solid-liquid pristimerin extraction were analyzed: solvent systems, number of extractions, ratio of plant weight (g)/solvent volume (mL) used, time of extraction, temperature and agitation. A mathematical model was generated for pristimerin purity and yield. Ethanol, first extraction, 0.5 ratio of plant weight/solvent volume (g/mL), 0.5 h, 200 rpm and 49.7°C were optimal conditions for the extraction of this phytochemical. The degree of purification of pristimerin root extract was studied by size-exclusion chromatography (SEC) using Sephadex LH-20 reaching fractions with purification indexes (PI) greater than 2 and recoveries of 28.3%. When fractions with purification indices higher than 1 and less than 2 were accumulated, the recovery of pristimerin increased by about 73.6%. By combining the optimum extracts and SEC purification protocols, an enriched fraction containing 245.6 mg pristimerin was obtained from 100 g of root bark, representing about 14.4%, w/w, pristimerin from the total solids presented in the fraction.

Integrating a post-column makeup pump into preparative supercritical fluid chromatography systems to address stability and recovery issues during purifications.

PubMed

Bajpai, Lakshmikant; Naidu, Harshavardhan; Asokan, Kathiravan; Shaik, Khaja Mohiddin; Kaspady, Mahammed; Arunachalam, Piramanayagam; Wu, Dauh-Rurng; Mathur, Arvind; Sarabu, Ramakanth

2017-08-18

Purification of many pharmaceutical compounds by supercritical fluid chromatography (SFC) has always been challenging because of degradation of compound during the isolation step in the presence of acidic or basic modifiers in the mobile phase. Stability of such acid or base-sensitive compounds could be improved by post-column addition of a solvent containing base or acid modifier as counter ion through a make-up pump respectively to neutralize the compound fraction without affecting the resolution. One such case study has been presented in this work where the stability of a base-sensitive compound was addressed by the addition of acidic co-solvent through the make-up pump. Details of this setup and the investigation of degradation of the in-house base-sensitive compound are discussed in this paper. In addition, poor retentivity and low recovery of many non-polar compounds in SFC eluting under low co-solvent percentage is another major concern. Even though the desired separation could be achieved with low percentage of co-solvent, it's difficult to get the proper recovery after purification due to precipitation of the sample and significant aerosol formation inside the cyclone. We have demonstrated the first-time use of a post-column make-up pump on SFC 350 system to introduce additional solvent prior to cyclone to avoid the precipitation, reduce the aerosol formation and thus improve the recovery of non-polar compounds eluting under less than 10% of co-solvent. Copyright © 2017 Elsevier B.V. All rights reserved.

Extraction of neptunium by trilaurylamine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Patil, S.K.; Swarup, R.; Ramaniah, M.V.

1972-07-01

Trilaurylamine (TLA) is considered as useful solvent for the final purification of plutonium and neptunium. As TLA is considered as an alternate possible extractant for the final purification of plutonium and neptunium at Tarapur Reprocessing Plant under construction, it was considered necessary to study the optimum conditions for the extraction of neptunium using TLA.

A gradient-free method for the purification of infective dengue virus for protein-level investigations.

PubMed

Jensen, Stephanie M; Nguyen, Celina T; Jewett, John C

2016-09-01

Dengue virus (DENV) is a mosquito-transmitted flavivirus that infects approximately 100 million people annually. Multi-day protocols for purification of DENV reduce the infective titer due to viral sensitivity to both temperature and pH. Herein we describe a 5-h protocol for the purification of all DENV serotypes, utilizing traditional gradient-free ultracentrifugation followed by selective virion precipitation. This protocol allows for the separation of DENV from contaminating proteins - including intact C6/36 densovirus, for the production of infective virus at high concentration for protein-level analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

Simply enhancing throughput of free-flow electrophoresis via organic-aqueous environment for purification of weak polarity solute of phenazine-1-carboxylic acid in fermentation of Pseudomonas sp. M18.

PubMed

Yang, Jing-Hua; Shao, Jing; Wang, Hou-Yu; Dong, Jing-Yu; Fan, Liu-Yin; Cao, Cheng-Xi; Xu, Yu-Quan

2012-09-01

Herein, a simple novel free-flow electrophoresis (FFE) method was developed via introduction of organic solvent into the electrolyte system, increasing the solute solubility and throughput of the sample. As a proof of concept, phenazine-1-carboxylic acid (PCA) from Pseudomonas sp. M18 was selected as a model solute for the demonstration on feasibility of novel FFE method on account of its faint solubility in aqueous circumstance. In the developed method, the organic solvent was added into not only the sample buffer to improve the solubility of the solute, but also the background buffer to construct a uniform aqueous-organic circumstance. These factors of organic solvent percentage and types as well as pH value of background buffer were investigated for the purification of PCA in the FFE device via CE. The experiments revealed that the percentage and the types of organic solvent exerted major influence on the purification of PCA. Under the optimized conditions (30 mM phosphate buffer in 60:40 (v/v) water-methanol at an apparent pH 7.0, 3.26 mL/min background flux, 10-min residence time of injected sample, and 400 V), PCA could be continuously purified from its impurities. The flux of sample injection was 10.05 μL/min, and the recovery was up to 93.7%. An 11.9-fold improvement of throughput was found with a carrier buffer containing 40% (v/v) methanol, compared with the pure aqueous phase. The developed procedure is of evident significance for the purification of weak polarity solute via FFE. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Secretory immunoglobulin purification from whey by chromatographic techniques.

PubMed

Matlschweiger, Alexander; Engelmaier, Hannah; Himmler, Gottfried; Hahn, Rainer

2017-08-15

Secretory immunoglobulins (SIg) are a major fraction of the mucosal immune system and represent potential drug candidates. So far, platform technologies for their purification do not exist. SIg from animal whey was used as a model to develop a simple, efficient and potentially generic chromatographic purification process. Several chromatographic stationary phases were tested. A combination of two anion-exchange steps resulted in the highest purity. The key step was the use of a small-porous anion exchanger operated in flow-through mode. Diffusion of SIg into the resin particles was significantly hindered, while the main impurities, IgG and serum albumin, were bound. In this step, initial purity was increased from 66% to 89% with a step yield of 88%. In a second anion-exchange step using giga-porous material, SIg was captured and purified by step or linear gradient elution to obtain fractions with purities >95%. For the step gradient elution step yield of highly pure SIg was 54%. Elution of SIgA and SIgM with a linear gradient resulted in a step yield of 56% and 35%, respectively. Overall yields for both anion exchange steps were 43% for the combination of flow-through and step elution mode. Combination of flow-through and linear gradient elution mode resulted in a yield of 44% for SIgA and 39% for SIgM. The proposed process allows the purification of biologically active SIg from animal whey in preparative scale. For future applications, the process can easily be adopted for purification of recombinant secretory immunoglobulin species. Copyright © 2017 Elsevier B.V. All rights reserved.

Centrifugal partition chromatography: A preparative tool for isolation and purification of xylindein from Chlorociboria aeruginosa.

PubMed

Boonloed, Anukul; Weber, Genevieve L; Ramzy, Kelly M; Dias, Veronica R; Remcho, Vincent T

2016-12-23

A centrifugal partition chromatography (CPC) method was developed for the preparative-scale isolation and purification of xylindein from the wood-staining fungi, Chlorociboria aeruginosa. Xylindein, a blue-green pigment naturally secreted from the hyphae and fruiting bodies of the fungus, has great value in the decorative wood industry and textile coloration. Xylindein has great potential for use as a fluorescent labeling agent as well as in organic semiconductor applications. However, a primary limitation of xylindein is its poor solubility in most common HPLC solvents. Consequently, it is arduous to purify using preparative liquid chromatography or solid-phase extraction (SPE). Support-free, liquid-liquid chromatographic methods, including CPC, where solutes are separated based on their different distribution coefficients (K D ) between two immiscible solvent systems, are promising alternatives for the purification of the compound on a preparative scale. In this work, a new biphasic solvent system suitable for CPC separation of xylindein was developed. Various groups of solvents were assessed for their suitability as xylindein extractants. A new solvent system suitable for CPC separation of xylindein, composed of heptane/THF/MEK/acetonitrile/acetic acid/water, was developed. This solvent system yielded a K D value for xylindein of 1.54±0.04, as determined by HPLC (n=3). The compositions of the upper phase and lower phase of the solvent system were determined by Heteronuclear Single Quantum Correlation (HSQC) NMR and proton NMR. A CPC system, equipped with a fraction collector, was used for the isolation of xylindein from crude extracts. The xylindein fractions isolated by the CPC were then analyzed using HPLC and presented as a fractogram. Based on the CPC fractogram, the purified xylindein fractions were achieved after 30min CPC separation time, yielding 71% extraction efficiency. The developed CPC method allowed for isolation of this naturally sourced xylindein in amounts suitable for further study. Copyright © 2016 Elsevier B.V. All rights reserved.

Economic Methods of Ginger Protease'sextraction and Purification

NASA Astrophysics Data System (ADS)

Qiao, Yuanyuan; Tong, Junfeng; Wei, Siqing; Du, Xinyong; Tang, Xiaozhen

This article reports the ginger protease extraction and purification methods from fresh ginger rhizome. As to ginger protease extraction, we adapt the steps of organic solvent dissolving, ammonium sulfate depositing and freeze-drying, and this method can attain crude enzyme powder 0.6% weight of fresh ginger rhizome. The purification part in this study includes two steps: cellulose ion exchange (DEAE-52) and SP-Sephadex 50 chromatography, which can purify crude ginger protease through ion and molecular weight differences respectively.

Isolation and purification of a granulosis virus from infected larvae of the Indian meal moth, Plodia interpunctella.

PubMed

Tweeten, K A; Bulla, L A; Consigli, R A

1977-09-01

A procedure was developed for purification of a granulosis virus inclusion body produced in vivo in the Indian meal moth, Plodia interpunctella (Hübner). Purification was accomplished by differential centrifugation, treatment with sodium deoxycholate, and velocity sedimentation in sucrose gradients. The adequacy of the procedure was confirmed by mixing experiments in which uninfected, radioactively labeled larvae were mixed with infected, unlabeled larvae. After purification, the virus was shown to be free of host tissue, to retain its physical integrity, and to be highly infectious per os. Preparations of purified virus consisted of homogeneous populations of intact inclusion bodies (210 by 380 nm) whose buoyant density was 1.271 g/cm3 when centrifuged to equilibrium in sucrose gradients. Electron microscopy of thin-sectioned virus or of virus sequentially disrupted on electron microscope grids demonstrated three components: protein matrix, envelope, and nucleocapsid.

Isolation and purification of a granulosis virus from infected larvae of the Indian meal moth, Plodia interpunctella.

PubMed Central

Tweeten, K A; Bulla, L A; Consigli, R A

1977-01-01

A procedure was developed for purification of a granulosis virus inclusion body produced in vivo in the Indian meal moth, Plodia interpunctella (Hübner). Purification was accomplished by differential centrifugation, treatment with sodium deoxycholate, and velocity sedimentation in sucrose gradients. The adequacy of the procedure was confirmed by mixing experiments in which uninfected, radioactively labeled larvae were mixed with infected, unlabeled larvae. After purification, the virus was shown to be free of host tissue, to retain its physical integrity, and to be highly infectious per os. Preparations of purified virus consisted of homogeneous populations of intact inclusion bodies (210 by 380 nm) whose buoyant density was 1.271 g/cm3 when centrifuged to equilibrium in sucrose gradients. Electron microscopy of thin-sectioned virus or of virus sequentially disrupted on electron microscope grids demonstrated three components: protein matrix, envelope, and nucleocapsid. Images PMID:334076

Conical coils counter-current chromatography for preparative isolation and purification of tanshinones from Salvia miltiorrhiza Bunge.

PubMed

Liang, Junling; Meng, Jie; Guo, Mengzhe; Yang, Zhi; Wu, Shihua

2013-05-03

Modern counter-current chromatography (CCC) originated from the helical coil planet centrifuge. Recently, spiral coils were found to possess higher separation efficiency in both the retention of stationary phase and solutes resolution than other CCC coils like the helical and toroidal coils used on type-J CCC and cross-axis CCC. In this work, we built a novel conical coil CCC for the preparative isolation and purification of tanshinones from Salvia miltiorrhiza Bunge. The conical coils were wound on three identical upright tapered holders in head-to-tail and left-handed direction and connected in series. Compared with helical and spiral coil CCC, conical coil CCC not only placed CCC column in a two-dimensional centrifugal field, but also provided a potential centrifugal force gradient both in axial and radial directions. The extra centrifugal gradient made mobile phase move faster and enabled CCC much higher retention of stationary phase and better resolution. As a result, higher efficiency has been obtained with the solvent system of hexane-ethyl acetate-methanol-water (HEMWat) with the volume ratio of 5:5:7:3 by using conical coil CCC apparatus. Four tanshinones, including cryptotanshinone (1), tanshinone I (2), 1,2-dihydrotanshinquinone (3) and tanshinone IIA (4), were well resolved from 500mg to 1g crude samples with high purity. Furthermore, the conical coil CCC can make a much higher solid phase retention, which makes it to be a powerful separation tool with high throughput. This is the first report about conical coil CCC for separation of tanshinones and it may also be an important advancement for natural products isolation. Copyright © 2013 Elsevier B.V. All rights reserved.

URANIUM PURIFICATION PROCESS

DOEpatents

Ruhoff, J.R.; Winters, C.E.

1957-11-12

A process is described for the purification of uranyl nitrate by an extraction process. A solution is formed consisting of uranyl nitrate, together with the associated impurities arising from the HNO/sub 3/ leaching of the ore, in an organic solvent such as ether. If this were back extracted with water to remove the impurities, large quantities of uranyl nitrate will also be extracted and lost. To prevent this, the impure organic solution is extracted with small amounts of saturated aqueous solutions of uranyl nitrate thereby effectively accomplishing the removal of impurities while not allowing any further extraction of the uranyl nitrate from the organic solvent. After the impurities have been removed, the uranium values are extracted with large quantities of water.

Extraction, isolation, and purification of analytes from samples of marine origin--a multivariate task.

PubMed

Liguori, Lucia; Bjørsvik, Hans-René

2012-12-01

The development of a multivariate study for a quantitative analysis of six different polybrominated diphenyl ethers (PBDEs) in tissue of Atlantic Salmo salar L. is reported. An extraction, isolation, and purification process based on an accelerated solvent extraction system was designed, investigated, and optimized by means of statistical experimental design and multivariate data analysis and regression. An accompanying gas chromatography-mass spectrometry analytical method was developed for the identification and quantification of the analytes, BDE 28, BDE 47, BDE 99, BDE 100, BDE 153, and BDE 154. These PBDEs have been used in commercial blends that were used as flame-retardants for a variety of materials, including electronic devices, synthetic polymers and textiles. The present study revealed that an extracting solvent mixture composed of hexane and CH₂Cl₂ (10:90) provided excellent recoveries of all of the six PBDEs studied herein. A somewhat lower polarity in the extracting solvent, hexane and CH₂Cl₂ (40:60) decreased the analyte %-recoveries, which still remain acceptable and satisfactory. The study demonstrates the necessity to perform an intimately investigation of the extraction and purification process in order to achieve quantitative isolation of the analytes from the specific matrix. Copyright © 2012 Elsevier B.V. All rights reserved.

New biphasic solvent system based on cyclopentyl methyl ether for the purification of a non-polar synthetic peptide by pH-zone refining centrifugal partition chromatography.

PubMed

Amarouche, Nassima; Boudesocque, Leslie; Borie, Nicolas; Giraud, Matthieu; Forni, Luciano; Butte, Alessandro; Edwards, Florence; Renault, Jean-Hugues

2014-06-01

A new type 1 ternary biphasic system composed of cyclopentyl methyl ether, dimethylformamide and water was developed, characterized and successfully used for the purification of a lipophilic, protected peptide by pH-zone refining centrifugal partition chromatography. The protected peptide is an 8-mer, key intermediate in bivalirudin (Angiomax®) synthesis and shows a very low solubility in the solvents usually used in liquid chromatography. All ionic groups, except the N-terminal end of the peptide, are protected by a benzyl group. The purification of this peptide was achieved with a purity of about 99.04% and a recovery of 94% using the new ternary biphasic system cyclopentyl methyl ether/dimethylformamide/water (49:40:11, v/v) in the descending pH-zone refining mode with triethylamine (28 mM) as the retainer and methanesulfonic acid (18 mM) as the eluter. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of magnetic- and carbon-based nano-adsorbents application in pre-purification of paclitaxel from needles of Taxus baccata

NASA Astrophysics Data System (ADS)

Naghavi, M. R.; Motamedi, E.; Nasiri, J.; Alizadeh, H.; Fattahi Moghadam, M. R.; Mashouf, A.

2015-01-01

In this investigation, the proficiency of a number of magnetic carbon-based nano-adsorbents is evaluated in pre-purification process of the crude paclitaxel extract obtained from fresh needles of yew tree ( Taxus baccata L.). The effectiveness and removal ability of color and impurities from crude extracts, for three novel candidate nano-adsorbents (i.e., Fe3O4 nanoparticles (Fe3O4Nps), graphite oxide (GO), and their hybrids Fe3O4Nps/GO) are compared with commercial graphite in three different solvents. In general, both HPLC and UV-Vis spectroscopy results demonstrate that in less polar solvent (i.e., dichloromethane), the adsorption is greatly affected by the electrostatic attractions, while in more polar solvents (i.e., acetone and ethanol) π-π electron interactions taking place between adsorbent and adsorbate are the most dominant factors in sorption. Considering decolorization efficiency, purity of taxol, recovery and reusability of adsorbents, Fe3O4Nps/GO (50 g/L) in dichloromethane is selected as the best medium for pre-purification of paclitaxel. Additionally, in kinetic studies the sorption equilibrium can be reached within 120 min, and the experimental data are well fitted by the pseudo-second-order model. The Langmuir sorption isotherm model correlates well with the sorption equilibrium data for the crude extract concentration (500-2,000 mg/L). Our findings display promising applications of Fe3O4Nps/GO, as a cost-effective nano-adsorbent, to provide a suitable vehicle toward improvement of paclitaxel pre-purification.

Retrovirus purification: method that conserves envelope glycoprotein and maximizes infectivity.

PubMed Central

McGrath, M; Witte, O; Pincus, T; Weissman, I L

1978-01-01

A Sepharose 4B chromatographic method for purification of retroviruses is described which was less time consuming, increased purified virus yields, conserved viral glycoprotein, and increased recovery of biological infectivity in comparison with conventional sucrose gradient ultracentrifugation techniques. Images PMID:205680

Synthesis, stabilization, and characterization of metal nanoparticles

NASA Astrophysics Data System (ADS)

White, Gregory Von, II

Wet chemical synthesis techniques offer the ability to control various nanoparticle characteristics including size, shape, dispersibility in both aqueous and organic solvents, and tailored surface chemistries appropriate for different applications. Large quantities of stabilizing ligands or surfactants are often required during synthesis to achieve these nanoparticle characteristics. Unfortunately, excess reaction byproducts, surfactants, and ligands remaining in solution after nanoparticle synthesis can impede application, and therefore post-synthesis purification must be employed. A liquid-liquid solvent/antisolvent pair (typically ethanol/toluene or ethanol/hexane for gold nanoparticles, GNPs) can be used to both purify and size-selectively fractionate hydrophobically modified nanoparticles. Alternatively, carbon dioxide may be used in place of a liquid antisolvent, a "green" approach, enabling both nanoparticle purification and size-selective fractionation while simultaneously eliminating mixed solvent waste and allowing solvent recycle. We have used small-angle neutron scattering (SANS) to investigate the ligand structure and composition response of alkanethiol modified gold and silver nanoparticles at varying anti-solvent conditions (CO2 or ethanol). The ligand lengths and ligand solvation for alkanethiol gold and silver NPs were found to decrease with increased antisolvent concentrations directly impacting their dispersibility in solution. Calculated Flory-Huggins interaction parameters support our SANS study for dodecanethiol dispersibility in the mixed organic solvents. This research has led to a greater understanding of the liquid-liquid precipitation process for metal nanoparticles, and provides critical results for future interaction energy modeling.

Organic solvents in the pharmaceutical industry.

PubMed

Grodowska, Katarzyna; Parczewski, Andrzej

2010-01-01

Organic solvents are commonly used in the pharmaceutical industry as reaction media, in separation and purification of synthesis products and also for cleaning of equipment. This paper presents some aspects of organic solvents utilization in an active pharmaceutical ingredient and a drug product manufacturing process. As residual solvents are not desirable substances in a final product, different methods for their removal may be used, provided they fulfill safety criteria. After the drying process, analyses need to be performed to check if amounts of solvents used at any step of the production do not exceed acceptable limits (taken from ICH Guideline or from pharmacopoeias). Also new solvents like supercritical fluids or ionic liquids are developed to replace "traditional" organic solvents in the pharmaceutical production processes.

Isolation of mitochondria from Saccharomyces cerevisiae using magnetic bead affinity purification

PubMed Central

Liao, Pin-Chao; Boldogh, Istvan R.; Siegmund, Stephanie E.

2018-01-01

Isolated mitochondria are widely used to study the function of the organelle. Typically, mitochondria are prepared using differential centrifugation alone or in conjunction with density gradient ultracentrifugation. However, mitochondria isolated using differential centrifugation contain membrane or organelle contaminants, and further purification of crude mitochondria by density gradient ultracentrifugation requires large amounts of starting material, and is time-consuming. Mitochondria have also been isolated by irreversible binding to antibody-coated magnetic beads. We developed a method to prepare mitochondria from budding yeast that overcomes many of the limitations of other methods. Mitochondria are tagged by insertion of 6 histidines (6xHis) into the TOM70 (Translocase of outer membrane 70) gene at its chromosomal locus, isolated using Ni-NTA (nickel (II) nitrilotriacetic acid) paramagnetic beads and released from the magnetic beads by washing with imidazole. Mitochondria prepared using this method contain fewer contaminants, and are similar in ultrastructure as well as protein import and cytochrome c oxidase complex activity compared to mitochondria isolated by differential centrifugation. Moreover, this isolation method is amenable to small samples, faster than purification by differential and density gradient centrifugation, and more cost-effective than purification using antibody-coated magnetic beads. Importantly, this method can be applied to any cell type where the genetic modification can be introduced by CRISPR or other methods. PMID:29698455

Solvent-free direct reductive amination by catalytic use of an organotin reagent incorporated on an ionic liquid.

PubMed

Pham, Phuoc Dien; Bertus, Philippe; Legoupy, Stéphanie

2009-11-07

An organotin reagent supported on an ionic liquid was used as a highly effective catalyst (down to 0.1 mol%) for the direct reductive amination of aldehydes and ketones using PhSiH3; this solvent-free method facilitates purification of the products, thus minimizing the contamination by tin.

High pressure liquid chromatographic gradient mixer

DOEpatents

Daughton, Christian G.; Sakaji, Richard H.

1985-01-01

A gradient mixer which effects the continuous mixing of any two miscible solvents without excessive decay or dispersion of the resultant isocratic effluent or of a linear or exponential gradient. The two solvents are fed under low or high pressure by means of two high performance liquid chromatographic pumps. The mixer comprises a series of ultra-low dead volume stainless steel tubes and low dead volume chambers. The two solvent streams impinge head-on at high fluxes. This initial nonhomogeneous mixture is then passed through a chamber packed with spirally-wound wires which cause turbulent mixing thereby homogenizing the mixture with minimum "band-broadening".

High-pressure liquid chromatographic gradient mixer

DOEpatents

Daughton, C.G.; Sakaji, R.H.

1982-09-08

A gradient mixer effects the continuous mixing of any two miscible solvents without excessive decay or dispersion of the resultant isocratic effluent or of a linear or exponential gradient. The two solvents are fed under low or high pressure by means of two high performance liquid chromatographic pumps. The mixer comprises a series of ultra-low dead volume stainless steel tubes and low dead volume chambers. The two solvent streams impinge head-on at high fluxes. This initial nonhomogeneous mixture is then passed through a chamber packed with spirally-wound wires which cause turbulent mixing thereby homogenizing the mixture with minimum band-broadening.

Solutocapillary Convection Effects on Polymeric Membrane Morphology

NASA Technical Reports Server (NTRS)

Krantz, William B.; Todd, Paul W.; Kinagurthu, Sanjay

1996-01-01

Macro voids are undesirable large pores in membranes used for purification. They form when membranes are cast as thin films on a smooth surface by evaporating solvent (acetone) from a polymer solution. There are two un-tested hypotheses explaining the growth of macro voids. One states that diffusion of the non-solvent (water) is solely responsible, while the other states that solutocapillary convection is the primary cause of macro void growth. Solutocapillary convection is flow-caused by a concentration induced surface-tension gradient. Macrovoid growth in the former hypothesis is gravity independent, while in the latter it is opposed by gravity. To distinguish between these two hypotheses, experiments were designed to cast membranes in zero-gravity. A semi-automated apparatus was designed and built for casting membranes during the 20 secs of zero-g time available in parabolic aircraft flight such as NASA's KC-135. The phase changes were monitored optically, and membrane morphology was evaluated by scanning electron microscopy (SEM). These studies appear to be the first quantitative studies of membrane casting in micro-gravity which incorporate real-time data acquisition. Morphological studies of membranes cast at 0, 1, and 1.8 g revealed the presence of numerous, sparse and no macrovoids respectively. These results are consistent with the predictions of the solutocapillary hypothesis of macrovoid growth.

Effect of solvent volume ratio and time extraction of glycerol purification

NASA Astrophysics Data System (ADS)

Sinaga, M. S.; Rico, G.; Nababan, A. N.; Manullang, T. A.

2018-02-01

Glycerol as a byproduct of biodiesel production about 10% of the biodiesel weight. Impurities which contained in the glycerol such as catalyst, soap, methanol, water, salt, and matter organic nonglycerol (MONG) on have a significant effect on the glycerol concentration. So, it is necessary to treat the impurities. The purpose of this study is to know the effect of ethylene glycol to glycerol purification process with acidification method using phosphoric acid aspretreatment process. This research was begun with an acid addition to the glycerol to neutralize the base content and to split the soap content into free fatty acid and salt, which easier separated from glycerol. Then the process was continued with extraction by the solvent ethylene glycol using the variable of test volume ratio (v/v) (1:0,5, 1:1, 1:1,5) and the extraction time (20, 40, and 60 minutes). The results showed that the more volume of solvent used, gave less extraction time to produce high purity of glycerol. The highest purity produced in this study amounted to 90.646% is obtained at the ratio of the volume solvent (v/v) 1:1 with extraction time 60 minutes.

Rapid purification of staphylococcal enterotoxin B by high-pressure liquid chromatography.

PubMed Central

Strickler, M P; Neill, R J; Stone, M J; Hunt, R E; Brinkley, W; Gemski, P

1989-01-01

The Staphylococcus aureus enterotoxins represent a group of proteins that cause emesis and diarrhea in humans and other primates. We have developed a rapid two-step high-pressure liquid chromatography (HPLC) procedure for purification of staphylococcal enterotoxin B (SEB). Sterile filtrates (2.5 liters) of strain 10-275 were adsorbed directly onto a reversed-phase column (50 mm by 30 cm Delta Pak; 300 A [30 nm], 15 microns, C18). SEB was obtained by using a unique sequential gradient system. First, an aqueous ammonium acetate to acetonitrile gradient followed by an aqueous trifluoroacetic acid (TFA) wash was used to remove contaminants. A subsequent TFA to acetonitrile-TFA gradient eluted the bound SEB. Further purification was obtained by rechromatography on a cation-exchange column. From 35 to 45% of the SEB in starting filtrates was recovered. Analysis by immunoblotting of samples separated on sodium dodecyl sulfate-polyacrylamide gels indicated that HPLC-purified SEB exhibited immunological and biochemical properties similar to those of the SEB standard. Induction of an emetic response in rhesus monkeys showed that the HPLC-purified toxin also retained biological activity. Images PMID:2745678

Ion-exchange chromatography purification of extracellular vesicles.

PubMed

Kosanović, Maja; Milutinović, Bojana; Goč, Sanja; Mitić, Ninoslav; Janković, Miroslava

2017-08-01

Despite numerous studies, isolating pure preparations of extracellular vesicles (EVs) has proven challenging. Here, we compared ion-exchange chromatography (IEC) to the widely used sucrose density gradient (SDG) centrifugation method for the purification of EVs. EVs in bulk were isolated from pooled normal human amniotic fluid (AF) by differential centrifugation followed by IEC or sucrose density gradient separation. The purity of the isolated EVs was evaluated by electrophoresis and lectin blotting/immuno blotting to monitor the distribution of total proteins, different EVs markers, and selected N-glycans. Our data showed efficient separation of negatively charged EVs from other differently charged molecules, while comparative profiling of EVs using SDG centrifugation confirmed anion-exchange chromatography is advantageous for EV purification. Finally, although this IEC-based method was validated using AF, the approach should be readily applicable to isolation of EVs from other sources as well.

Two-step purification of scutellarin from Erigeron breviscapus (vant.) Hand. Mazz. by high-speed counter-current chromatography.

PubMed

Gao, Min; Gu, Ming; Liu, Chun-Zhao

2006-07-11

Scutellarin, a flavone glycoside, popularly applied for the treatment of cardiopathy, has been purified in two-step purification by high-speed counter-current chromatography (HSCCC) from Erigeron breviscapus (vant.) Hand. Mazz. (Deng-zhan-hua in Chinese), a well-known traditional Chinese medicinal plant for heart disease. Two solvent systems, n-hexane-ethyl acetate-methanol-acetic acid-water (1:6:1.5:1:4, v/v/v/v/v) and ethyl acetate-n-butanol-acetonitrile-0.1% HCl (5:2:5:10, v/v/v/v) were used for the two-step purification. The purity of the collected fraction of scutellarin was 95.6%. This study supplies a new alternative method for purification of scutellarin.

Green and Bio-Based Solvents.

PubMed

Calvo-Flores, Francisco G; Monteagudo-Arrebola, María José; Dobado, José A; Isac-García, Joaquín

2018-04-24

Chemical reactions and many of the procedures of separation and purification employed in industry, research or chemistry teaching utilize solvents massively. In the last decades, with the birth of Green Chemistry, concerns about the employment of solvents and the effects on human health, as well as its environmental impacts and its dependence on non-renewable raw materials for manufacturing most of them, has drawn the attention of the scientific community. In this work, we review the concept of green solvent and the properties and characteristics to be considered green. Additionally, we discuss the different possible routes to prepare many solvents from biomass, as an alternative way to those methods currently applied in the petrochemical industry.

Solvent coarsening around colloids driven by temperature gradients

NASA Astrophysics Data System (ADS)

Roy, Sutapa; Dietrich, Siegfried; Maciolek, Anna

2018-04-01

Using mesoscopic numerical simulations and analytical theory, we investigate the coarsening of the solvent structure around a colloidal particle emerging after a temperature quench of the colloid surface. Qualitative differences in the coarsening mechanisms are found, depending on the composition of the binary liquid mixture forming the solvent and on the adsorption preferences of the colloid. For an adsorptionwise neutral colloid, the phase next to its surface alternates as a function of time. This behavior sets in on the scale of the relaxation time of the solvent and is absent for colloids with strong adsorption preferences. A Janus colloid, with a small temperature difference between its two hemispheres, reveals an asymmetric structure formation and surface enrichment around it, even if the solvent is within its one-phase region and if the temperature of the colloid is above the critical demixing temperature Tc of the solvent. Our phenomenological model turns out to capture recent experimental findings according to which, upon laser illumination of a Janus colloid and due to the ensuing temperature gradient between its two hemispheres, the surrounding binary liquid mixture develops a concentration gradient.

Antibiotics from Pseudomonas reptilivora II. Isolation, Purification, and Properties1

PubMed Central

Del Rio, Luís A.; Gorgé, J. López; Olivares, J.; Mayor, F.

1972-01-01

Under well-established culture conditions, Pseudomonas reptilivora produced several antibiotics that have been purified by solvent extraction, chromatography in Sephadex G-25, electrophoresis, and paper chromatography in different solvent systems. Activity has been monitored at the different steps of isolation and purification by measurement of the inhibition of the growth of Staphylococcus aureus by the cylinder-plate method, as well as by bioautography of chromatograms and electropherograms. Three antibiotics have been isolated and named A, B1, and B2. The B1 and B2 activities were studied in greater detail than A. The B1 substance was crystallized, and its chemical properties were found to coincide with those of YC 73 or fluopsin C described by Egawa et al. and Itoh et al., respectively. Images PMID:4790558

Biodiesel production from ethanolysis of palm oil using deep eutectic solvent (DES) as co-solvent

NASA Astrophysics Data System (ADS)

Manurung, R.; Winarta, A.; Taslim; Indra, L.

2017-06-01

Biodiesel produced from ethanolysis is more renewable and have better properties (higher oxidation stability, lower cloud and pour point) compared to methanolysis, but it has a disadvantage such as complicated purification. To improve ethanolysis process, deep eutectic solvent (DES) can be prepared from choline chloride and glycerol and used as co-solvent in ethanolysis. The deep eutectic solvent is formed from a quaternary ammonium salt (choline chloride) and a hydrogen bond donor (Glycerol), it is a non-toxic, biodegradable solvent compared to a conventional volatile organic solvent such as hexane. The deep eutectic solvent is prepared by mixing choline chloride and glycerol with molar ratio 1:2 at temperature 80 °C, stirring speed 300 rpm for 1 hour. The DES is characterized by its density and viscosity. The ethanolysis is performed at a reaction temperature of 70 °C, ethanol to oil molar ratio of 9:1, potassium hydroxide as catalyst concentration of 1.2 wt. DES as co-solvent with concentration 0.5 to 3 wt. stirring speed 400 rpm, and a reaction time 1 hour. The obtained biodiesel is then characterized by its density, viscosity, and ester content. The oil - ethanol phase condition is observed in the reaction tube. The oil - ethanol phase with DES tends to form meniscus compared to without DES, showed that oil and ethanol become more slightly miscible, which favors the reaction. Using DES as co-solvent in ethanolysis showed increasing in yield and easier purification. The esters properties meet the international standards ASTM D6751, with the highest yield achieved 83,67 with 99,77 conversion at DES concentration 2 . Increasing DES concentration above 2 in ethanolysis decrease the conversion and yield, because of the excessive glycerol in the systems makes the reaction equilibrium moves to the reactant side.

Aqueous Chloride Operations Overview: Plutonium and Americium Purification/Recovery

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gardner, Kyle Shelton; Kimball, David Bryan; Skidmore, Bradley Evan

These are a set of slides intended for an information session as part of recruiting activities at Brigham Young University. It gives an overview of aqueous chloride operations, specifically on plutonium and americium purification/recovery. This presentation details the steps taken perform these processes, from plutonium size reduction, dissolution, solvent extraction, oxalate precipitation, to calcination. For americium recovery, it details the CLEAR (chloride extraction and actinide recovery) Line, oxalate precipitation and calcination.

Size exclusion chromatography-gradients, an alternative approach to polymer gradient chromatography: 2. Separation of poly(meth)acrylates using a size exclusion chromatography-solvent/non-solvent gradient.

PubMed

Schollenberger, Martin; Radke, Wolfgang

2011-10-28

A gradient ranging from methanol to tetrahydrofuran (THF) was applied to a series of poly(methyl methacrylate) (PMMA) standards, using the recently developed concept of SEC-gradients. Contrasting to conventional gradients the samples eluted before the solvent, i.e. within the elution range typical for separations by SEC, however, the high molar mass PMMAs were retarded as compared to experiments on the same column using pure THF as the eluent. The molar mass dependence on retention volume showed a complex behaviour with a nearly molar mass independent elution for high molar masses. This molar mass dependence was explained in terms of solubility and size exclusion effects. The solubility based SEC-gradient was proven to be useful to separate PMMA and poly(n-butyl crylate) (PnBuA) from a poly(t-butyl crylate) (PtBuA) sample. These samples could be separated neither by SEC in THF, due to their very similar hydrodynamic volumes, nor by an SEC-gradient at adsorbing conditions, due to a too low selectivity. The example shows that SEC-gradients can be applied not only in adsorption/desorption mode, but also in precipitation/dissolution mode without risking blocking capillaries or breakthrough peaks. Thus, the new approach is a valuable alternative to conventional gradient chromatography. Copyright © 2011 Elsevier B.V. All rights reserved.

Fractionation, partial characterization and bioactivity of water-soluble polysaccharides and polysaccharide-protein complexes from Pleurotus geesteranus.

PubMed

Zhang, Mei; Zhu, Lin; Cui, Steve W; Wang, Qi; Zhou, Ting; Shen, Hengsheng

2011-01-01

Fractionation and purification of mushroom polysaccharides is a critical process for mushroom clinical application. After a hot-water treatment, the crude Pleurotus geesteranus (PG) was further fractionated into four fractions (PG-1, -2, -3, -4) using gradient precipitation with water and ammonia sulphate. By controlling the initial polymer concentration and ratio of solvents, this process produced PG fractions with high chemical uniformity and narrow Mw distribution without free proteins. Structurally, PG-1 and PG-2 are pure homopolysaccharide mainly composed of glucose; and PG-3 and PG-4 are heteropolysaccharide-protein complexes. PG-2, a high M(w) fraction mainly composed of glucose presented significant cytotoxicity at the concentration of 200 and 100 μg/ml to human breast cancer cells. Here, we report a new mushroom polysaccharides extraction and fractionation method, with which we produced four fractions of PG with PG-2 appearing effective anti-tumour activity. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Experimental studies on islets isolation, purification and function in rats

PubMed Central

Pang, Xinlu; Xue, Wujun; Feng, Xinshun; Tian, Xiaohui; Teng, Yan; Ding, Xiaoming; Pan, Xiaoming; Guo, Qi; He, Xiaoli

2015-01-01

To develop a simple and effective method of islet isolation and purification in rats. Collagenase P was injected into pancreatic duct followed by incubation in water bath to digest the pancreas and isolate islet, then discontinuous gravity gradient purification was used to purify the islet. The purified islets were identified by dithizone staining. The viability of islets was assessed by fluorescence staining of acridine orange (AO) and propidium iodide (PI). The function of purified islets was determined by glucose-stimulated insulin release test and transplantation of rat with streptozocin-induced diabetes. 738±193 islets were recovered after purification. The average purity was 77±13%, the viability of islets was more than 95%. When inspected by glucose stimulation, the secreted insulin concentration was 24.31±5.47 mIU/L when stimulated by low concentration glucose and 37.62±4.29 mIU/L by high concentration glucose. There was significant difference between the two phases (P<0.05). The blood sugar concentration recovered to normal level after two days in the animals with islet transplantation. In conclusion, islets can be procured with good function and shape by using the method of injecting collagenase into pancreatic duct followed by incubation in water bath and purification using discontinuous gravity gradient. PMID:26885021

Literature Review: Crud Formation at the Liquid/Liquid Interface of TBP-Based Solvent-Extraction Processes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Delegard, Calvin H.; Casella, Amanda J.

2016-09-30

This report summarizes the literature reviewed on crud formation at the liquid:liquid interface of solvent extraction processes. The review is focused both on classic PUREX extraction for industrial reprocessing, especially as practiced at the Hanford Site, and for those steps specific to plutonium purification that were used at the Plutonium Reclamation Facility (PRF) within the Plutonium Finishing Plant (PFP) at the Hanford Site.

Measurement of dielectric constant of organic solvents by indigenously developed dielectric probe

NASA Astrophysics Data System (ADS)

Keshari, Ajay Kumar; Rao, J. Prabhakar; Rao, C. V. S. Brahmmananda; Ramakrishnan, R.; Ramanarayanan, R. R.

2018-04-01

The extraction, separation and purification of actinides (uranium and plutonium) from various matrices are an important step in nuclear fuel cycle. One of the separation process adopted in an industrial scale is the liquid-liquid extraction or solvent extraction. Liquid-liquid extraction uses a specific ligand/extractant in conjunction with suitable diluent. Solvent extraction or liquid-liquid extraction, involves the partitioning of the solute between two immiscible phases. In most cases, one of the phases is aqueous, and the other one is an organic solvent. The solvent used in solvent extraction should be selective for the metal of interest, it should have optimum distribution ratio, and the loaded metal from the organic phase should be easily stripped under suitable experimental conditions. Some of the important physical properties which are important for the solvent are density, viscosity, phase separation time, interfacial surface tension and the polarity of the extractant.

Analytical RISM-MP2 free energy gradient method: Application to the Schlenk equilibrium of Grignard reagent

NASA Astrophysics Data System (ADS)

Mori, Toshifumi; Kato, Shigeki

2007-03-01

We present a method to evaluate the analytical gradient of reference interaction site model Møller-Plesset second order free energy with respect to solute nuclear coordinates. It is applied to calculate the geometries and energies in the equilibria of the Grignard reagent (CH 3MgCl) in dimethylether solvent. The Mg-Mg and Mg-Cl distances as well as the binding energies of solvents are largely affected by the dynamical electron correlation. The solvent effect on the Schlenk equilibrium is examined.

Purification of uranium alloys by differential solubility of oxides and production of purified fuel precursors

DOEpatents

McLean, II, William; Miller, Philip E.

1997-01-01

A method for purifying metallic alloys of uranium for use as nuclear reactor fuels in which the metal alloy is first converted to an oxide and then dissolved in nitric acid. Initial removal of metal oxide impurities not soluble in nitric acid is accomplished by filtration or other physical means. Further purification can be accomplished by carbonate leaching of uranyl ions from the partially purified solution or using traditional methods such as solvent extraction.

Purification of uranium alloys by differential solubility of oxides and production of purified fuel precursors

DOEpatents

McLean, W. II; Miller, P.E.

1997-12-16

A method is described for purifying metallic alloys of uranium for use as nuclear reactor fuels in which the metal alloy is first converted to an oxide and then dissolved in nitric acid. Initial removal of metal oxide impurities not soluble in nitric acid is accomplished by filtration or other physical means. Further purification can be accomplished by carbonate leaching of uranyl ions from the partially purified solution or using traditional methods such as solvent extraction. 3 figs.

Off-line coupling of new generation centrifugal partition chromatography device with preparative high pressure liquid chromatography-mass spectrometry triggering fraction collection applied to the recovery of secoiridoid glycosides from Centaurium erythraea Rafn. (Gentianaceae).

PubMed

Mandova, Tsvetelina; Audo, Grégoire; Michel, Sylvie; Grougnet, Raphaël

2017-09-01

A purification sequence including a Gilson CPC 250 PRO device coupled to PrepHPLC hyphenated with a MS triggering fraction collector was applied to isolate secoiridoid glycosides from a complex methanolic extract of Centaurium erythraea. This species is widely used for ethnomedicinal purposes around the Mediterranean Sea. The solvent system ethyle acetate/ethanol/water 7.5/3/5 was determined using shake-flask method targeting swertiamarin, the major secoiridoid of the extract. Optimization of CPC experimental parameters enabled the injection of 4g of extract with a flow rate of 40mL/min at 3000rpm to provide a secoiridoid glycosides enriched fraction. 130mg of this latter was submitted to a second step of purification by preparative HPLC (gradient water/formic acid (19:1) (A) and methanol (B) as follows: 0min, 85% A; 8min, 60% A; 12min, 55% A; 35min, 55% A; 40min, 10% A; 50min, 10% A; 52min, 85% A; 55min, 85% A) to give swertiamarin (36mg, yield 27.7%, purity 98.2%). Other secoiridoid glycosides (sweroside, gentiopicroside, secologanol, secoxyloganin) were also isolated in minor amounts. As these monoterpene derivatives are responsible for several biological activities, their quick recovery with high yield and purity may serve as a model for further scale-up and industrial development. Copyright © 2017 Elsevier B.V. All rights reserved.

In-tube electro-membrane extraction with a sub-microliter organic solvent consumption as an efficient technique for synthetic food dyes determination in foodstuff samples.

PubMed

Bazregar, Mohammad; Rajabi, Maryam; Yamini, Yadollah; Asghari, Alireza; Abdossalami asl, Yousef

2015-09-04

A simple and efficient extraction technique with a sub-microliter organic solvent consumption termed as in-tube electro-membrane extraction (IEME) is introduced. This method is based upon the electro-kinetic migration of ionized compounds by the application of an electrical potential difference. For this purpose, a thin polypropylene (PP) sheet placed inside a tube acts as a support for the membrane solvent, and 30μL of an aqueous acceptor solution is separated by this solvent from 1.2mL of an aqueous donor solution. This method yielded high extraction recoveries (63-81%), and the consumption of the organic solvent used was only 0.5μL. By performing this method, the purification is high, and the utilization of the organic solvent, used as a mediator, is very simple and repeatable. The proposed method was evaluated by extraction of four synthetic food dyes (Amaranth, Ponceau 4R, Allura Red, and Carmoisine) as the model analytes. Optimization of variables affecting the method was carried out in order to achieve the best extraction efficiency. These variables were the type of membrane solvent, applied extraction voltage, extraction time, pH range, and concentration of salt added. Under the optimized conditions, IEME-HPLC-UV provided a good linearity in the range of 1.00-800ngmL(-1), low limits of detection (0.3-1ngmL(-1)), and good extraction repeatabilities (RSDs below 5.2%, n=5). It seems that this design is a proper one for the automation of the method. Also the consumption of the organic solvent in a sub-microliter scale, and its simplicity, high efficiency, and high purification can help one getting closer to the objectives of the green chemistry. Copyright © 2015 Elsevier B.V. All rights reserved.

Energy requirements of the switchable polarity solvent forward osmosis (SPS-FO) water purification process

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wendt, Daniel S.; Orme, Christopher J.; Mines, Gregory L.

A model was developed to estimate the process energy requirements of a switchable polarity solvent forward osmosis (SPS FO) system for water purification from aqueous NaCl feed solution concentrations ranging from 0.5 to 4.0 molal at an operational scale of 480 m3/day (feed stream). The model indicates recovering approximately 90% of the water from a feed solution with NaCl concentration similar to seawater using SPS FO would have total equivalent energy requirements between 2.4 and 4.3 kWh per m 3 of purified water product. The process is predicted to be competitive with current costs for disposal/treatment of produced water frommore » oil and gas drilling operations. As a result, once scaled up the SPS FO process may be a thermally driven desalination process that can compete with the cost of seawater reverse osmosis.« less

Energy requirements of the switchable polarity solvent forward osmosis (SPS-FO) water purification process

DOE PAGES

Wendt, Daniel S.; Orme, Christopher J.; Mines, Gregory L.; ...

2015-08-01

A model was developed to estimate the process energy requirements of a switchable polarity solvent forward osmosis (SPS FO) system for water purification from aqueous NaCl feed solution concentrations ranging from 0.5 to 4.0 molal at an operational scale of 480 m3/day (feed stream). The model indicates recovering approximately 90% of the water from a feed solution with NaCl concentration similar to seawater using SPS FO would have total equivalent energy requirements between 2.4 and 4.3 kWh per m 3 of purified water product. The process is predicted to be competitive with current costs for disposal/treatment of produced water frommore » oil and gas drilling operations. As a result, once scaled up the SPS FO process may be a thermally driven desalination process that can compete with the cost of seawater reverse osmosis.« less

Nickel solvent extraction from cold purification filter cakes of Angouran mine concentrate using LIX984N

NASA Astrophysics Data System (ADS)

Balesini, A. A.; Zakeri, A.; Razavizadeh, H.; Khani, A.

2013-11-01

Cold purification filter cakes generated in the hydrometallurgical processing of Angouran mine zinc concentrate commonly contain significant amounts of Zn, Cd, and Ni ions and thus are valuable resources for metal recovery. In this research, a nickel containing solution that was obtained from sulfuric acid leaching of the filter cake following cadmium and zinc removal was subjected to solvent extraction experiments using 10vol% LIX984N diluted in kerosene. Under optimum experimental conditions (pH 5.3, volume ratio of organic/aqueous (O:A) = 2:1, and contact time = 5 min), more than 97.1% of nickel was extracted. Nickel was stripped from the loaded organic by contacting with a 200 g/L sulfuric acid solution, from which 77.7% of nickel was recovered in a single contact at the optimum conditions (pH 1-1.5, O:A = 5:1, and contact time = 15 min).

[Pilot-scale purification of lipopeptide from marine-derived Bacillus marinus].

PubMed

Gu, Kangbo; Guan, Cheng; Xu, Jiahui; Li, Shulan; Luo, Yuanchan; Shen, Guomin; Zhang, Daojing; Li, Yuanguang

2016-11-25

This research was aimed at establishing the pilot-scale purification technology of lipopeptide from marine-derived Bacillus marinus. We studied lipopeptide surfactivity interferences on scale-up unit technologies including acid precipitation, methanol extraction, solvent precipitation, salting out, extraction, silica gel column chromatography and HZ806 macroporous absorption resin column chromatography. Then, the unit technologies were combined in a certain order, to remove the impurities gradually, and to gain purified lipopeptide finally, with high recovery rate throughout the whole process. The novel pilot-scale purification technology could effectively isolate and purify lipopeptide with 87.51% to 100% purity in hectograms from 1 ton of Bacillus marinus B-9987 fermentation broth with more than 81.73% recovery rate. The first practical hectogram production of highly purified lipopeptide derived from Bacillus marinus was achieved. With this new purification method, using complex media became possible in fermentation process to reduce the fermentation cost and scale-up the purification for lipopeptide production. For practicability and economy, foaming problem resulting from massive water evaporation was avoided in this technology.

Dynamic pH junction high-speed counter-current chromatography coupled with microwave-assisted extraction for online separation and purification of alkaloids from Stephania cepharantha.

PubMed

Yuan, Zhiquan; Xiao, Xiaohua; Li, Gongke

2013-11-22

A simple and efficient dynamic pH junction high-speed counter-current chromatography method was developed and further applied to the online extraction, separation and purification of alkaloids from Stephania cepharantha by coupling with microwave-assisted extraction. Mineral acid and organic base were added into the mobile phase and the sample solution, respectively, leading to the formation of a dynamic pH junction in the column and causing focus of alkaloids. Selective focus of analytes can be achieved on the basis of velocity changes of the pH junction through appropriate selection of solvent systems and optimization of additive concentrations. The extract can be directly introduced into the HSCCC for the online extraction, separation and purification of alkaloids from S. cepharantha. Continuous separation can be easily achieved with the same solvent system. Under the optimum conditions, 6.0 g original sample was extracted with 60 mL of the upper phase of hexane-ethyl acetate-methanol-water (1:1:1:1, v/v/v/v) containing 10% triethylamine under 50 °C and 400 W irradiation power for 10 min, the extracts were directly separated and purified by high-speed counter-current chromatography. A total of 5.7 mg sinomenine, 8.3mg 6,7-di-O-acetylsinococuline, 17.9 mg berbamine, 12.7 mg isotetrandrine and 14.6 mg cepharanthine were obtained with purities of 96.7%, 93.7%, 98.7%, 97.3% and 99.3%, respectively. The online method provides good selectivity to ionizable compounds and improves the separation and purification efficiency of the high-speed counter-current chromatography technique. It has good potential for separation and purification of effective compounds from natural products. Copyright © 2013 Elsevier B.V. All rights reserved.

Purification of Torpedo californica post-synaptic membranes and fractionation of their constituent proteins.

PubMed Central

Elliott, J; Blanchard, S G; Wu, W; Miller, J; Strader, C D; Hartig, P; Moore, H P; Racs, J; Raftery, M A

1980-01-01

A rapid methof for preparation of membrane fractions highly enriched in nicotinic acetylcholine receptor from Torpedo californica electroplax is described. The major step in this purification involves sucrose-density-gradient centrifugation in a reorienting rotor. Further purification of these membranes can be achieved by selective extraction of proteins by use of alkaline pH or by treatment with solutions of lithium di-idosalicylate. The alkali-treated membranes retain functional characteristics of the untreated membranes and in addition contain essentially only the four polypeptides (mol.wts. 40000, 50000, 60000 and 65000) characteristic of the receptor purified by affinity chromatography. Dissolution of the purified membranes or of the alkali-treated purified membranes in sodium cholate solution followed by sucrose-density-gradient centrifugation in the same detergent solution yields solubilized receptor preparations comparable with the most highly purified protein obtained by affinity-chromatographic procedures. Images Fig. 1. Fig. 2. Fig. 3. Fig. 5. Fig. 7. PLATE 1 PMID:7387629

[Extraction and purification technologies of total flavonoids from Aconitum tanguticum].

PubMed

Li, Yan-Rong; Yan, Li-Xin; Feng, Wei-Hong; Li, Chun; Wang, Zhi-Min

2014-04-01

To optimize the extraction and purification technologies of total flavonoids from Aconitum tanguticum whole plant. With the content of total flavonoids as index, the optimum extraction conditions for the concentration, volume of alcohol, extracting time and times were selected by orthogonal optimized; Comparing the adsorption quantity (mg/g) and resolution (%), four kinds of macroporous adsorption resins including D101, AB-8, X-5 and XAD-16 were investigated for the enrichment ability of total flavonoids from Aconitum tanguticum; Concentration and pH value of sample, sampling amount, elution solvent and loading and elution velocity for the optimum adsorption resin were determined. The content of total flavonoids in Aconitum tanguticum was about 4.39%; The optimum extraction technique was 70% alcohol reflux extraction for three times,each time for one hour, the ratio of material and liquid was 1:10 (w/v); The optimum purification technology was: using XAD-16 macroporous resin, the initial concentration of total flavonoids of Aconitum tanguticum was 8 mg/mL, the sampling amount was 112 mg/g dry resin, the pH value was 5, the loading velocity was 3 mL/min, the elution solvent was 70% ethanol and the elution velocity was 5 mL/min. Under the optimum conditions, the average content of total flavonoids was raised from 4.39% to 46.19%. The optimum extraction and purification technologies for total flavonoids of Aconitum tanguticum were suitable for industrial production for its simplicity and responsibility.

“Measure Your Gradient”: A New Way to Measure Gradients in High Performance Liquid Chromatography by Mass Spectrometric or Absorbance Detection

PubMed Central

Magee, Megan H.; Manulik, Joseph C.; Barnes, Brian B.; Abate-Pella, Daniel; Hewitt, Joshua T.; Boswell, Paul G.

2014-01-01

The gradient produced by an HPLC is never the same as the one it is programmed to produce, but non-idealities in the gradient can be taken into account if they are measured. Such measurements are routine, yet only one general approach has been described to make them: both HPLC solvents are replaced with water, solvent B is spiked with 0.1% acetone, and the gradient is measured by UV absorbance. Despite the widespread use of this procedure, we found a number of problems and complications with it, mostly stemming from the fact that it measures the gradient under abnormal conditions (e.g. both solvents are water). It is also generally not amenable to MS detection, leaving those with only an MS detector no way to accurately measure their gradients. We describe a new approach called “Measure Your Gradient” that potentially solves these problems. One runs a test mixture containing 20 standards on a standard stationary phase and enters their gradient retention times into open-source software available at www.measureyourgradient.org. The software uses the retention times to back-calculate the gradient that was truly produced by the HPLC. Here we present a preliminary investigation of the new approach. We found that gradients measured this way are comparable to those measured by a more accurate, albeit impractical, version of the conventional approach. The new procedure worked with different gradients, flow rates, column lengths, inner diameters, on two different HPLCs, and with six different batches of the standard stationary phase. PMID:25441073

Membrane-Based Technologies in the Pharmaceutical Industry and Continuous Production of Polymer-Coated Crystals/Particles.

PubMed

Chen, Dengyue; Sirkar, Kamalesh K; Jin, Chi; Singh, Dhananjay; Pfeffer, Robert

2017-01-01

Membrane technologies are of increasing importance in a variety of separation and purification applications involving liquid phases and gaseous mixtures. Although the most widely used applications at this time are in water treatment including desalination, there are many applications in chemical, food, healthcare, paper and petrochemical industries. This brief review is concerned with existing and emerging applications of various membrane technologies in the pharmaceutical and biopharmaceutical industry. The goal of this review article is to identify important membrane processes and techniques which are being used or proposed to be used in the pharmaceutical and biopharmaceutical operations. How novel membrane processes can be useful for delivery of crystalline/particulate drugs is also of interest. Membrane separation technologies are extensively used in downstream processes for bio-pharmaceutical separation and purification operations via microfiltration, ultrafiltration and diafiltration. Also the new technique of membrane chromatography allows efficient purification of monoclonal antibodies. Membrane filtration techniques of reverse osmosis and nanofiltration are being combined with bioreactors and advanced oxidation processes to treat wastewaters from pharmaceutical plants. Nanofiltration with organic solvent-stable membranes can implement solvent exchange and catalyst recovery during organic solvent-based drug synthesis of pharmaceutical compounds/intermediates. Membranes in the form of hollow fibers can be conveniently used to implement crystallization of pharmaceutical compounds. The novel crystallization methods of solid hollow fiber cooling crystallizer (SHFCC) and porous hollow fiber anti-solvent crystallization (PHFAC) are being developed to provide efficient methods for continuous production of polymer-coated drug crystals in the area of drug delivery. This brief review provides a general introduction to various applications of membrane technologies in the pharmaceutical/biopharmaceutical industry with special emphasis on novel membrane techniques for pharmaceutical applications. The method of coating a drug particle with a polymer using the SHFCC method is stable and ready for scale-up for operation over an extended period. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Purification and thermal analysis of perfluoro-n-alkanoic acids.

PubMed

Tsuji, Minami; Inoue, Tohru; Shibata, Osamu

2008-01-15

Purification of perfluoro-n-alkanoic acids (C(n)F(2n+1)COOH, n=7, 9, 11, 13, 15 and 17) was made by repeated recrystallizations from n-hexane/acetone mixed solvent, and their purity was found to be more than 99.5% by GC-MS, NMR, and elemental analysis. The thermal behaviors such as melting point and enthalpy change of fusion were investigated using differential scanning calorimetry (DSC). The melting point monotonously increased with increasing carbon number (n) of the acids, while the enthalpy change showed irregularity at n=14. The crystal structure of these acids was found to be dependent upon solvent used for recrystallization; that is, the acids recrystallized from the above solvent becomes more stable energetically, indicating their higher enthalpy change of fusion than that of the solidified acids from fused ones. The solid state was also found to vary depending upon the thermal history, indicating that a few crystal structures of the solid state are quite similar energetically. The melting points (T(m)) of perfluoro-n-alkanoic acids are higher than those of corresponding n-alkanoic acids, and the difference in T(m) increases with increasing carbon number in the acids.

High temperature-ultra performance liquid chromatography-mass spectrometry for the metabonomic analysis of Zucker rat urine.

PubMed

Gika, Helen G; Theodoridis, Georgios; Extance, Jon; Edge, Anthony M; Wilson, Ian D

2008-08-15

The applicability and potential of using elevated temperatures and sub 2-microm porous particles in chromatography for metabonomics/metabolomics was investigated using, for the first time, solvent temperatures higher than the boiling point of water (up to 180 degrees C) and thermal gradients to reduce the use of organic solvents. Ultra performance liquid chromatography, combined with mass spectrometry, was investigated for the global metabolite profiling of the plasma and urine of normal and Zucker (fa/fa) obese rats (a well established disease animal model). "Isobaric" high temperature chromatography, where the temperature and flow rate follow a gradient program, was developed and evaluated against a conventional organic solvent gradient. LC-MS data were first examined by established chromatographic criteria in order to evaluate the chromatographic performance and next were treated by special peak picking algorithms to allow the application of multivariate statistics. These studies showed that, for urine (but not plasma), chromatography at elevated temperatures provided better results than conventional reversed-phase LC with higher peak capacity and better peak asymmetry. From a systems biology point of view, better group clustering and separation was obtained with a larger number of variables of high importance when using high temperature-ultra performance liquid chromatography (HT-UPLC) compared to conventional solvent gradients.

The Synthesis and Study of Azole Carboxamide Nucleosides as Agents Active Against RNA Viruses.

DTIC Science & Technology

1986-09-15

solvent such as nitromethane gave a nucleoside product , identified as 1-(2,3,5-tri-O-benzoyl-o--D-ribofuranosyl)-l,2,4- triazol-3(2H)-one (20, BL-00307...at 2220 cm . Treatment of 26 with NH4OH/H 202 solution, and purification of the reaction product by chromatography on silica gel furnished 1-(2-deoxy...30) in 74% yield. Treatment of 30 with NH4OH/H202 solution, and purification of the reaction product by chroma- tography on silica gel furnished 1

Purification of organic nonlinear optical materials for bulk crystal growth from melt

NASA Astrophysics Data System (ADS)

Gebre, Tesfaye; Bhat, Kamala N.; Batra, Ashok K.; Lal, Ravindra B.; Aggarwal, Mohan D.; Penn, Benjamin G.; Frazier, Donald O.

2002-10-01

The techniques developed for purification of nonlinear optical organic materials, such as benzil, 2-methyl-4-nitroaniline (MNA), Dicyanovinyl anisole (DIVA) and its derivatives, nitrophenyl prolinol (NPP) and other Schiff's base compounds, include Kugelrohy method, physical vapor transport, zone refining and recrystallization from the solvent are described. Purity of the materials is tested using differential thermal analysis, gas chromatograph/Mass detector, Fourier Transform Infrared spectroscopy and melting point measurements. The purified materials were later used in the growth of single crystal by Bridgman-Stockbarger and Czochralski techniques.

Techno-economic assessment of hybrid extraction and distillation processes for furfural production from lignocellulosic biomass.

PubMed

Nhien, Le Cao; Long, Nguyen Van Duc; Kim, Sangyong; Lee, Moonyong

2017-01-01

Lignocellulosic biomass is one of the most promising alternatives for replacing mineral resources to overcome global warming, which has become the most important environmental issue in recent years. Furfural was listed by the National Renewable Energy Laboratory as one of the top 30 potential chemicals arising from biomass. However, the current production of furfural is energy intensive and uses inefficient technology. Thus, a hybrid purification process that combines extraction and distillation to produce furfural from lignocellulosic biomass was considered and investigated in detail to improve the process efficiency. This effective hybrid process depends on the extracting solvent, which was selected based on a comprehensive procedure that ranged from solvent screening to complete process design. Various solvents were first evaluated in terms of their extraction ability. Then, the most promising solvents were selected to study the separation feasibility. Eventually, processes that used the three best solvents (toluene, benzene, and butyl chloride) were designed and optimized in detail using Aspen Plus. Sustainability analysis was performed to evaluate these processes in terms of their energy requirements, total annual costs (TAC), and carbon dioxide (CO 2 ) emissions. The results showed that butyl chloride was the most suitable solvent for the hybrid furfural process because it could save 44.7% of the TAC while reducing the CO 2 emissions by 45.5% compared to the toluene process. In comparison with the traditional purification process using distillation, this suggested hybrid extraction/distillation process can save up to 19.2% of the TAC and reduce 58.3% total annual CO 2 emissions. Furthermore, a sensitivity analysis of the feed composition and its effect on the performance of the proposed hybrid system was conducted. Butyl chloride was found to be the most suitable solvent for the hybrid extraction/distillation process of furfural production. The proposed hybrid sequence was more favorable than the traditional distillation process when the methanol fraction of the feed stream was <3% and more benefit could be obtained when that fraction decreased.

On-line coupling of counter-current chromatography and macroporous resin chromatography for continuous isolation of arctiin from the fruit of Arctium lappa L.

PubMed

Guo, Mengzhe; Liang, Junling; Wu, Shihua

2010-08-13

In this work, we have developed a novel hybrid two-dimensional counter-current chromatography and liquid chromatography (2D CCC x LC) system for the continuous purification of arctiin from crude extract of Arctium lappa. The first dimensional CCC column has been designed to fractionalize crude complex extract into pure arctiin effluent using a one-component organic/salt-containing system, and the second dimensional LC column has been packed with macroporous resin for on-line adsorption, desalination and desorption of arctiin which was effluent purified from the first CCC dimension. Thus, the crude arctiin mixture has been purified efficiently and conveniently by on-line CCC x LC in spite of the use of a salt-containing solvent system in CCC separation. As a result, high purity (more than 97%) of arctiin has been isolated by repeated injections both using the ethyl acetate-8% sodium chloride aqueous solution and butanol-1% sodium chloride aqueous solution. By contrast with the traditional CCC processes using multi-component organic/aqueous solvent systems, the present on-line CCC x LC process only used a one-component organic solvent and thus the solvent is easier to recover and regenerate. All of used solvents such as ethyl acetate, n-butanol and NaCl aqueous solution are low toxicity and environment-friendly. Moreover, the lower phase of salt-containing aqueous solution used as mobile phase, only contained minor organic solvent, which will save much organic solvent in continuous separation. In summary, our results indicated that the on-line hybrid 2D CCC x LC system using one-component organic/salt-containing aqueous solution is very promising and powerful tool for high-throughput purification of arctiin from fruits of A. lappa. 2010 Elsevier B.V. All rights reserved.

Purification and characterization of two isoenzymes of lipoxygenase from soybeans.

PubMed

Diel, E; Stan, H J

1978-01-01

A chromatographic procedure for the purification of two lipoxygenase isoenzymes (linoleate: O2 oxidoreductase, EC 1.13.11.12.) from soybean is described. The procedure for the purification of isoenzyme L-1 includes optimalized extraction, ammonium sulfate fractionation, heat treatment and gradient elution from a CM-Sephadex C-50 column. The purification of L-2 includes ammonium sulfate fractionation, gelfiltration on Sephadex G-150 and gradient elution from a DEAE-cellulose column. Both isoenzymes L-1 and L-2 appear homogeneous after Disc-PAGE. The isoelectric points are 5.6 for L-1 and 5.8 for L-2. Molecular weights are estimated as 100,000 for L-1 as well as L-2 applying three different methods. Both isoenzymes contain 0.9 mol iron per mol protien. The estimated turn over numbers are 8,200 mol linoleate per mol enzyme and min for L-1 and 3,100 for L-2. Amino acid compositions determined after acid hydrolysis show marked differences between L-1 and L-2, particularly with respect to the amino acids Lys, Phe, Ser, Gly and Leu. L-1 posesses a total of 9 cysteine molecules, 6 of which are present as disulfide bonds. L-2 posesses a total of 8 cysteine molecules with only one disulfide bond.

Efficient synthesis of pure monotosylated beta-cyclodextrin and its dimers.

PubMed

Tripodo, Giuseppe; Wischke, Christian; Neffe, Axel T; Lendlein, Andreas

2013-11-15

6-O-Monotosyl-β-cyclodextrin (mono-Ts-βCD) is one of the most important intermediates in the production of substituted βCD. So far, performing the monotosylation reaction and, in particular, the purification steps was challenging, relied on toxic solvents, and resulted in long and expensive procedures at, importantly, low yields. Here, the reaction of cyclodextrin with p-toluenesulfonyl chloride in aqueous environment is described to obtain a highly pure mono-Ts-βCD, for which a single-step purification with a cation exchange resin was applied. With this synthetic route and purification, yields could be increased from typically <10-15% to 35%, and organic solvents could be avoided. As characterized by FTIR, mass spectrometry, elemental analysis, and NMR, mono-Ts-βCD was obtained with a molar purity of >98mol%. From mono-Ts-βCD, β-cyclodextrin dimers linked by ethylenediamine (bis-Et-βCD) were successfully prepared (yield 93%, purity 96mol%) in a one-step approach using an anion exchange resin to trap leaving groups that typically interfere in the reaction. This synthesis procedure with a direct collection of side-products may be a general strategy applicable for nucleophilic substitution of tosylated cyclodextrins. Copyright © 2013 Elsevier Ltd. All rights reserved.

Ultrasound-assisted extraction and preliminary purification of proanthocyanidins and chlorogenic acid from almond (Prunus dulcis) skin.

PubMed

Ma, Xue; Zhou, Xin-Yu; Qiang, Qian-Qian; Zhang, Zhi-Qi

2014-07-01

An aqueous solution of polyethylene glycol (PEG) as a green solvent was employed for the first time to develop the ultrasound-assisted extraction of proanthocyanidins (PA) and chlorogenic acid (CA) from almond skin. The optimized extraction parameters were determined based on response surface methodology, and corresponded to an ultrasound power of 120 W, a liquid-to-solid ratio of 20:1 (mL/g), and a PEG concentration of 50% (v/v). Under these optimized conditions, the extraction yields of PAs and CA from almond skin were 32.68 ± 0.22 and 16.01 ± 0.19 mg/g, respectively. Compared with organic solvent extraction, PEG solution extraction produced higher yields. Different macroporous resins were compared for their performance in purifying PAs and CA from almond skin extract. Static adsorption/desorption experimental results demonstrated that AB-8 resin exhibits excellent purification performance at pH 4. Under the optimized dynamic adsorption/desorption conditions on the AB-8 column, the total recovery of purification for PAs and CA was 80.67%. The total content of PAs and CA in the preliminarily purified extract was 89.17% (with respective contents of 60.90 and 28.27%). © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Solvent effects in time-dependent self-consistent field methods. II. Variational formulations and analytical gradients

DOE PAGES

Bjorgaard, J. A.; Velizhanin, K. A.; Tretiak, S.

2015-08-06

This study describes variational energy expressions and analytical excited state energy gradients for time-dependent self-consistent field methods with polarizable solvent effects. Linear response, vertical excitation, and state-specific solventmodels are examined. Enforcing a variational ground stateenergy expression in the state-specific model is found to reduce it to the vertical excitation model. Variational excited state energy expressions are then provided for the linear response and vertical excitation models and analytical gradients are formulated. Using semiempiricalmodel chemistry, the variational expressions are verified by numerical and analytical differentiation with respect to a static external electric field. Lastly, analytical gradients are further tested by performingmore » microcanonical excited state molecular dynamics with p-nitroaniline.« less

Energy-Efficient Bioalcohol Recovery by Gel Stripping

NASA Astrophysics Data System (ADS)

Godbole, Rutvik; Ma, Lan; Hedden, Ronald

2014-03-01

Design of energy-efficient processes for recovering butanol and ethanol from dilute fermentations is a key challenge facing the biofuels industry due to the high energy consumption of traditional multi-stage distillation processes. Gel stripping is an alternative purification process by which a dilute alcohol is stripped from the fermentation product by passing it through a packed bed containing particles of a selectively absorbent polymeric gel material. The gel must be selective for the alcohol, while swelling to a reasonable degree in dilute alcohol-water mixtures. To accelerate materials optimization, a combinatorial approach is taken to screen a matrix of copolymer gels having orthogonal gradients in crosslinker concentration and hydrophilicity. Using a combination of swelling in pure solvents, the selectivity and distribution coefficients of alcohols in the gels can be predicted based upon multi-component extensions of Flory-Rehner theory. Predictions can be validated by measuring swelling in water/alcohol mixtures and conducting h HPLC analysis of the external liquid. 95% + removal of butanol from dilute aqueous solutions has been demonstrated, and a mathematical model of the unsteady-state gel stripping process has been developed. NSF CMMI Award 1335082.

Molecular Simulation Evaluation of Macromolecular Transport through Nanofiltration Membranes

NASA Astrophysics Data System (ADS)

Almodovar Arbelo, Noelia; Boudouris, Bryan; Corti, David

A hybrid Monte Carlo and Molecular Dynamics simulation technique was implemented to elucidate the equilibrium behavior and transport properties of a model macromolecule as it navigated across a nanoporous polymer thin film (i.e., a nanofiltration membrane). The model linear homopolymer chosen was one that had interactions that were representative of poly(ethylene oxide) (PEO) due to the known interactions of PEO with solution molecules when a PEO chain is dissolved in an aqueous environment. The structural rearrangements of the PEO chain as it passes through the nanopore under an imposed chemical potential gradient was quantified as a function of solvent quality, polymer chain length, nanopore diameter and shape, and PEO-nanopore wall interactions. Thus, these computational studies provide a more detailed picture of the underlying physical mechanisms that drive macromolecular transport through nanopores, and, in particular, how dimensionally-large macromolecules (i.e., with large radii of gyration) enter and move through dimensionally-small pores (i.e., small radii nanopores). The insights gained from these studies will aid in the development of more cost-effective water purification systems in separation technologies for myriad industrial applications.

Comparing cooling systems for the COBE 2991 cell separator used in the purification of human pancreatic islets of Langerhans.

PubMed

Adewola, A; Mage, R; Hansen, M; Barbaro, B; Mendoza-Elias, J; Harvat, T; Morel, P H; Oberholzer, J; Wang, Y

2010-01-01

Two different approaches of controlled cooling of the COBE 2991 cell-separator for islet purification were evaluated. The first method is the new Geneva COBE cooling system (GCCS), which consists of an electronically controlled liquid nitrogen injection system. The second is the University of Illinois at Chicago cooling system (UICCS), which consists of a specially designed "Cold Room" maintained at 1-8 C. For the GCCS, the mean temperatures of the gradient solutions were measured at the beginning and end of centrifugation were found to be 7 +/-0.7 C and 6.8 +/-0.6 C respectively. For the UICCS, the mean temperature of the gradients at the beginning and end of centrifugation were 4.7 +/-0.53 C and 7.03 C+/-0.91 C respectively. The presented COBE cooling systems can easily be adapted to a COBE 2991 cell-separator and are efficient in maintaining gradient solutions at a defined low temperature during centrifugation.

A comparative study of different strategies for removal of endotoxins from bacteriophage preparations.

PubMed

Van Belleghem, Jonas D; Merabishvili, Maya; Vergauwen, Bjorn; Lavigne, Rob; Vaneechoutte, Mario

2017-01-01

Bacterial endotoxins have high immunogenicity. Phage biology studies as well as therapeutic phage applications necessitate highly purified phage particles. In this study, we compared combinations of seven different endotoxin removal strategies and validated their endotoxin removal efficacy for five different phages (i.e. four Pseudomonas aeruginosa phages and one Staphylococcus aureus phage). These purification strategies included Endotrap HD column purification and/or CsCl density centrifugation in combination with Endotrap purification, followed by organic solvent (1-octanol), detergent (Triton X-100), enzymatic inactivation of the endotoxin using alkaline phosphatase and CIM monolytic anion exchange chromatography. We show that CsCl density purification of the P. aeruginosa phages, at an initial concentration of 10 12 -10 13 pfu/ml, led to the strongest reduction of endotoxins, with an endotoxin removal efficacy of up to 99%, whereas additional purification methods did not result in a complete removal of endotoxins from the phage preparations and only yielded an additional endotoxin removal efficacy of 23 to 99%, sometimes accompanied with strong losses in phage titer. Copyright © 2016 Elsevier B.V. All rights reserved.

Microchip electrospray: improvements in spray and signal stability during gradient elution by an inverted postcolumn makeup flow.

PubMed

Jung, Stephanie; Effelsberg, Uwe; Tallarek, Ulrich

2011-12-01

Dynamic changes in mobile phase composition during high-performance liquid chromatography (HPLC) gradient elution coupled to mass spectrometry (MS) sensitively affect electrospray modes. We investigate the impact of the eluent composition on spray stability and MS response by infusion and injection experiments with a small tetrapeptide in water-acetonitrile mixtures. The employed HPLC/electrospray (ESI)-MS configuration uses a microchip equipped with an enrichment column, a separation column, and a makeup flow (MUF) channel. One nano pump is connected to the separation column, while a second one delivers solvent of exactly inverted composition to the MUF channel. Both solvent streams are united behind the separation column, before the ESI tip, such that the resulting electrosprayed solution always has identical composition during a gradient elution. Analyte peak parameters without and with MUF compensation are determined and discussed with respect to the electrospray mode and eluent composition. The postcolumn MUF significantly improves spray and signal stability over the entire solvent gradient, without compromising the performance of the HPLC separation column. It can also be conveniently implemented on microchip platforms.

Solvent Exchange Leading to Nanobubble Nucleation: A Molecular Dynamics Study

PubMed Central

2017-01-01

The solvent exchange procedure has become the most-used protocol to produce surface nanobubbles, while the molecular mechanisms behind the solvent exchange are far from being fully understood. In this paper, we build a simple model and use molecular dynamics simulations to investigate the dynamic characteristics of solvent exchange for producing nanobubbles. We find that at the first stage of solvent exchange, there exists an interface between interchanging solvents of different gas solubility. This interface moves toward the substrate gradually as the exchange process proceeds. Our simulations reveal directed diffusion of gas molecules against the gas concentration gradient, driven by the solubility gradient of the liquid composition across the moving solvent–solvent interface. It is this directed diffusion that causes gas retention and produces a local gas oversaturation much higher near the substrate than far from it. At the second stage of solvent exchange, the high local gas oversaturation leads to bubble nucleation either on the solid surface or in the bulk solution, which is found to depend on the substrate hydrophobicity and the degree of local gas oversaturation. Our findings suggest that solvent exchange could be developed into a standard procedure to produce oversaturation and used to a variety of nucleation applications other than generating nanobubbles. PMID:28742364

Copper(I)/TEMPO Catalyzed Aerobic Oxidation of Primary Alcohols to Aldehydes with Ambient Air

PubMed Central

Hoover, Jessica M.; Steves, Janelle E.; Stahl, Shannon S.

2012-01-01

This protocol describes a practical laboratory-scale method for aerobic oxidation of primary alcohols to aldehydes, using a chemoselective CuI/TEMPO catalyst system. The catalyst is prepared in situ from commercially available reagents, and the reactions are performed in a common organic solvent (acetonitrile) with ambient air as the oxidant. Three different reaction conditions and three procedures for the isolation and purification of the aldehyde product are presented. The oxidations of eight different alcohols, described here, include representative examples of each reaction condition and purification method. Reaction times vary from 20 min to 24 h, depending on the alcohol, while the purification methods each take about 2 h. The total time necessary for the complete protocol ranges from 3 – 26 h. PMID:22635108

Corona-discharge air-purification system

NASA Technical Reports Server (NTRS)

Wydeven, T. J.; Flamm, D. L.

1979-01-01

Plasma reaction chamber removes trace contaminants from spacecraft, submarines, and other closed environments by oxidizing contaminants to produce carbon dioxide and water. Contaminants are alcohols, esters, hydrogen sulfide, and ammonia. Others are lubricant solvents such as Freons, aromatics, and Ketones. Contaminants are removed from chamber by scrubber.

Detergent-compatible, organic solvent-tolerant alkaline protease from Bacillus circulans MTCC 7942: Purification and characterization.

PubMed

Patil, Ulhas; Mokashe, Narendra; Chaudhari, Ambalal

2016-01-01

Proteases are now recognized as the most indispensable industrial biocatalyst owing to their diverse microbial sources and innovative applications. In the present investigation, a thermostable, organic solvent-tolerant, alkaline serine protease from Bacillus circulans MTCC 7942, was purified and characterized. The protease was purified to 37-fold by a three-step purification scheme with 39% recovery. The optimum pH and temperature for protease was 10 and 60 °C, respectively. The apparent molecular mass of the purified enzyme was 43 kD as revealed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The Km and Vmax values using casein-substrate were 3.1 mg/mL and 1.8 µmol/min, respectively. The protease remained stable in the presence of organic solvents with higher (>3.2) log P value (cyclohexane, n-octane, n-hexadecane, n-decane, and n-dodecane), as compared to organic solvents with lower (<3.2) log P value (acetone, butanol, benzene, chloroform, toluene). Remarkably, the protease showed profound stability even in the presence of organic solvents with less log P values (glycerol, dimethyl sulfate [DMSO], p-xylene), indicating the possibility of nonaqueous enzymatic applications. Also, protease activity was improved in the presence of metal ions (Ca(2+), Mg(2+), Mn(2+)); enhanced by biosurfactants; hardly affected by bleaching agents, oxidizing agents, and chemical surfactants; and stable in commercial detergents. In addition, a protease-detergent formulation effectively washed out egg and blood stains as compared to detergent alone. The protease was suitable for various commercial applications like processing of gelatinous film and as a compatible additive to detergent formulation with its operative utility in hard water.

Isolation and purification of rabbit mesenchymal stem cells using an optimized protocol.

PubMed

Lin, Chunbo; Shen, Maorong; Chen, Weiping; Li, Xiaofeng; Luo, Daoming; Cai, Jinhong; Yang, Yuan

2015-11-01

Mesenchymal stem cells were first isolated and grown in vitro by Friedenstein over 40 yr ago; however, their isolation remains challenging as they lack unique markers for identification and are present in very small quantities in mesenchymal tissues and bone marrow. Using whole marrow samples, common methods for mesenchymal stem cell isolation are the adhesion method and density gradient fractionation. The whole marrow sample adhesion method still results in the nonspecific isolation of mononuclear cells, and activation and/or potential loss of target cells. Density gradient fractionation methods are complicated, and may result in contamination with toxic substances that affect cell viability. In the present study, we developed an optimized protocol for the isolation and purification of mesenchymal stem cells based on the principles of hypotonic lysis and natural sedimentation.

Selection of operating parameters on the basis of hydrodynamics in centrifugal partition chromatography for the purification of nybomycin derivatives.

PubMed

Adelmann, S; Baldhoff, T; Koepcke, B; Schembecker, G

2013-01-25

The selection of solvent systems in centrifugal partition chromatography (CPC) is the most critical point in setting up a separation. Therefore, lots of research was done on the topic in the last decades. But the selection of suitable operating parameters (mobile phase flow rate, rotational speed and mode of operation) with respect to hydrodynamics and pressure drop limit in CPC is still mainly driven by experience of the chromatographer. In this work we used hydrodynamic analysis for the prediction of most suitable operating parameters. After selection of different solvent systems with respect to partition coefficients for the target compound the hydrodynamics were visualized. Based on flow pattern and retention the operating parameters were selected for the purification runs of nybomycin derivatives that were carried out with a 200 ml FCPC(®) rotor. The results have proven that the selection of optimized operating parameters by analysis of hydrodynamics only is possible. As the hydrodynamics are predictable by the physical properties of the solvent system the optimized operating parameters can be estimated, too. Additionally, we found that dispersion and especially retention are improved if the less viscous phase is mobile. Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.

Activated-charcoal filters: water treatment, pollution control, and industrial applications. January 1970-July 1988 (citations from the US Patent data base). Report for January 1970-July 1988

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

This bibliography contains citations of selected patents concerning activated-charcoal filters and their applications in water treatment, pollution control, and industrial processes. Filtering methods and equipment for air and water purification, industrial distillation and extraction, industrial leaching, and filtration of toxic gases and pollutants are described. Applications include drinking water purification, filtering beverages, production of polymer materials, solvent and metal recovery, swimming pool filtration, waste conversion, automobile fuel and exhaust systems, and footwear deodorizing. (Contains 129 citations fully indexed and including a title list.)

Histamine monolith versatility to purify supercoiled plasmid deoxyribonucleic acid from Escherichia coli lysate.

PubMed

Sousa, A; Almeida, A M; Černigoj, U; Sousa, F; Queiroz, J A

2014-08-15

Preparation of high quantities of supercoiled plasmid DNA of pharmaceutical grade purity is a research area where intensive investigation is being performed. From this standpoint, several downstream methods have been proposed, among them the monolithic chromatographic strategies owing to excellent mass transfer properties of monolithic supports and their high binding capacity for large biomolecules. The present study explores the physicochemical properties of histamine ligand in a supercoiled plasmid DNA purification process from an Escherichia coli clarified lysate, where the emphasis is given to the elution strategy that allows higher selectivity and efficient removal of other impurities besides the open circular isoform. The combination of high NaCl concentration and acidic pH allowed the elimination of 89% of RNA during the preparative loading of the lysate sample. The results of the purification strategy with ascending sodium chloride gradient revealed that 97% of supercoiled plasmid DNA was recovered with a purity degree of 99%. In addition, using a combined purification strategy with ascending sodium chloride (capture step) and then descending ammonium sulfate (polishing step) gradient, it was achieved a lower supercoiled plasmid DNA recovery yield of 79% with a purity degree of 92%, although the dynamic binding capacity under these conditions was higher than in the previous strategy. A significant reduction of host contents, such as proteins, RNA and genomic DNA, was obtained in both purification strategies. Accordingly, histamine is a useful and versatile ligand that allows the desirable supercoiled plasmid purification with high yield and purity level. Copyright © 2014. Published by Elsevier B.V.

Analysis of non-phthalates plasticizers on porous graphitic carbon by supercritical fluid chromatography using evaporative light scattering detection.

PubMed

Vaccher, Claude; Decaudin, Bertrand; Sautou, Valérie; Lecoeur, Marie

2014-09-12

The analysis of several plasticizers, widely used in the production of medical devices, was investigated on porous graphitic carbon (PGC) stationary phase in supercritical fluid chromatography (SFC) with an evaporative light scattering detector (ELSD). Due to strong interaction of compounds with the PGC support, solvents of strong eluotropic strength were added to the CO2 supercritical fluid. The effect of alkyl chain (pentane, hexane, heptane) and chlorinated (CH2Cl2, CHCl3, CCl4) solvents was studied on the retention and on the ELSD detection of plasticizers. A co-solvent mixture composed of CHCl3/heptane, eluted under gradient mode, allowed a significant improvement of the ELSD response compared to the use of each solvent individually. Then, a central composite design (CCD) was implemented to optimize both the separation and the detection of plasticizers. The parameters involved were the outlet pressure, the gradient slope, the co-solvent composition and the drift tube temperature of the ELSD. After optimization, baseline separation of plasticizers was achieved in 7min and best signal-to-noise ratios were obtained with outlet pressure and drift tube temperature of ELSD set at 200bar and 31°C, respectively. The co-solvent mixture was also composed of CHCl3/heptane (35/65 v/v) and a gradient from 15 to 60% of co-solvent in 2.2min was employed. The results demonstrated that CCD is a powerful tool for the optimization of SFC/ELSD method and the response surface model analysis can provide statistical understandings of the significant factors required to achieve optimal separation and ELSD sensitivity. Copyright © 2014 Elsevier B.V. All rights reserved.

Deep Eutectic Solvents Modified Molecular Imprinted Polymers for Optimized Purification of Chlorogenic Acid from Honeysuckle.

PubMed

Li, Guizhen; Wang, Wei; Wang, Qian; Zhu, Tao

2016-02-01

Deep eutectic solvents (DES) were synthesized with choline chloride (ChCl), and DES modified molecular imprinted polymers (DES-MIPs), DES modified non-imprinted polymers (DES-NIPs, without template), MIPs and NIPs were prepared in an identical procedure. Fourier transform infrared spectrometer (FT-IR) and field emission scanning electron microscopy (FE-SEM) were used to characterize the obtained polymers. Rebinding experiment and solid-phase extraction (SPE) were used to prove the high selectivity adsorption properties of the polymers. Box-Behnken design (BBD) with three factors was used to optimize the extraction condition of chlorogenic acid (CA) from honeysuckles. The optimum extraction conditions were found to be ultrasonic time optimized (20 min), the volume fraction of ethanol (60%) and ratio of liquid to material (15 mL g(-1)). Under these conditions, the mean extraction yield of CA was 12.57 mg g(-1), which was in good agreement with the predicted BBD model value. Purification of hawthorn extract was achieved by SPE process, and SPE recoveries of CA were 72.56, 64.79, 69.34 and 60.08% by DES-MIPs, DES-NIPs, MIPs and NIPs, respectively. The results showed DES-MIPs had potential for promising functional adsorption material for the purification of bioactive compounds. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Purification of biodiesel by choline chloride based deep eutectic solvent

NASA Astrophysics Data System (ADS)

Niawanti, Helda; Zullaikah, Siti; Rachimoellah, M.

2017-05-01

Purification is a crucial step in biodiesel production to meet the biodiesel standard. This study purified biodiesel using choline chloride based deep eutectic solvent (DES). DES was used to reduce unreacted oil and unsaponifiable matter in rice bran oil based biodiesel. The objective of this work was to study the effect of extraction time using DES on the content and yield of fatty acid methyl ester (FAME). Rice bran used in this work contains 16.49 % of oil with initial free fatty acids (FFA) of 44.75 %. Acid catalyzed methanolysis was employed to convert rice bran oil (RBO) into biodiesel under following operation conditions: T = 60 °C, t = 8 h, molar ratio of oil to methanol = 1/10, H2SO4 = 1% w/w of oil. Rice bran oil based biodiesel obtained contain 89.05 % of FAME with very low FFA content (0.05 %). DES was made from a mixture of choline chloride and ethylene glycol with molar ratio of 1/2. Molar ratio of crude biodiesel to DES were 1/2 and 1/4. Extraction time was varied from 15 minutes to 240 minutes at 30 °C. The highest FAME content was obtained after purification for 240 min. at molar ratio crude biodiesel to DES 1/4 was 96.60 %. This work shows that DES has potential to purify biodiesel from non-edible raw material, such as RBO.

Effective Method of Purification of Betulin from Birch Bark: The Importance of Its Purity for Scientific and Medicinal Use

PubMed Central

Šiman, Pavel; Filipová, Alžběta; Tichá, Alena; Niang, Mohamed; Bezrouk, Aleš; Havelek, Radim

2016-01-01

A new and relatively simple method for purification of betulin from birch bark extract was developed in this study. Its five purification steps are based on the differential solubility of extract components in various solvents and their crystallization and/or precipitation, on their affinity for Ca(OH)2 in ethanol, and on the affinity of some impurities for silica gel in chloroform. In addition, all used solvents can be simply recycled. Betulin of more than 99% purity can be prepared by this method with minimal costs. Various observations including crystallization of betulin, changes in crystals during heating, and attempt of localization of betulin in outer birch bark are also described in this work. The original extract, fraction without betulinic acid and lupeol, amorphous fraction of pure betulin, final crystalline fraction of pure betulin and commercial betulin as a standard were employed to determine the antiproliferative/cytotoxic effect. We used WST-1 tetrazolium-based assays with triple negative breast cancer cell line BT-549. The decrease in cell survival showed clear relationship with the purity of the samples, being most pronounced using our final product of pure crystalline betulin. WST-1 proliferation/cytotoxicity test using triple negative breast cancer cell line BT-549 clearly showed the importance of purity of betulin for biological experiments and, apparently, for its medicinal use. PMID:27152419

EVALUATION OF THE POLYAD FB AIR PURIFICATION AND SOLVENT RECOVERY PROCESS FOR STYRENE REMOVAL

EPA Science Inventory

The report gives results of a study evaluating the Polyad fluidized-bed (FB) process for controlling styrene emissions at a representative fiberglass shower stall and bath tub manufacturing plan*t. he process was evaluated using a transport able unit supplied by Weatherly, Inc., ...

Purification of Piscirickettsia salmonis and partial characterisation of antigens

USGS Publications Warehouse

Barnes, M.N.; Landolt, M.L.; Powell, D.B.; Winton, J.R.

1998-01-01

Piscirickettsia salmonis is the etiological agent of salmonid rickettsial septicemia, an economically significant disease affecting the salmon aquaculture industry. As with other rickettsial pathogens, antigenic analysis of P. salmonis has been limited by the inherent difficulties of purifying an intracellular organism away from host cell material. In this report, we describe the use of diatrizoate meglumine and diatrizoate sodium (DMDS) density gradient centrifugation to purify P. salmonis grown in chinook salmon embryo (CHSE-214) cells. Plaque assay titers and total protein assays confirmed that viable P. salmonis was consistently concentrated in a visible band within the DMDS density gradient at a density of 1.15 to 1.16 g ml-1. Recovery of purified, viable organisms from DMDS density gradients varied from 0.6 to 3%. Preparations of uninfected CHSE-214 cells, CHSE-214 cells infected with P. salmonis, and gradient-purified P. salmonis were compared using sodium dodecyl sulfate polyacrylamide gel electrophoresis to assess the degree of purification and to identify P. salmonis-specific proteins. Although gradient-purified P. salmonis preparations were not completely free of host cell material, 8 bacterial proteins were identified. Polyclonal rabbit antiserum was used in an immunoblot of proteins from purified P. salmonis to identify 3 major and 5 minor antigens. The major antigens of 56, 30 and 20 kDa were potential candidates for experimental vaccines and development of novel diagnostic assays.

Tall fescue seed extraction and partial purification of ergot alkaloids

PubMed Central

Ji, Huihua; Fannin, F.; Klotz, J.; Bush, Lowell

2014-01-01

Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because synthetically produced ergovaline is difficult to obtain, we developed a seed extraction and partial purification protocol for ergovaline/ergovalinine that provided a biologically active product. Tall fescue seed was ground and packed into several different sized columns for liquid extraction. Smaller particle size and increased extraction time increased efficiency of extraction. Our largest column was a 114 × 52 × 61 cm (W × L × D) stainless steel tub. Approximately 150 kg of seed could be extracted in this tub. The extraction was done with 80% ethanol. When the solvent front migrated to bottom of the column, flow was stopped and seed was allowed to steep for at least 48 h. Light was excluded from the solvent from the beginning of this step to the end of the purification process. Following elution, ethanol was removed from the eluate by evaporation at room temperature and the resulting syrup was freeze-dried. About 80% recovery of alkaloids was achieved with 18-fold increase in concentration of ergovaline. Initial purification of the dried product was accomplished by extracting with hexane/water (6:1, v/v). The aqueous fraction was extracted with chloroform, the aqueous layer discarded, after which the chloroform was removed with a resulting 20-fold increase of ergovaline. About 65% of the ergovaline was recovered from the chloroform residue for an overall recovery of 50%. The resultant partially purified ergovaline had biological activities in in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline. PMID:25566528

Tall fescue seed extraction and partial purification of ergot alkaloids

NASA Astrophysics Data System (ADS)

Bush, Lowell

2014-12-01

Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because synthetically produced ergovaline is difficult to obtain, we developed a seed extraction and partial purification protocol for ergovaline/ergovalinine that provided a biologically active product. Tall fescue seed was ground and packed into several different sized columns for liquid extraction. Smaller particle size and increased extraction time increased efficiency of extraction. Our largest column was a 114 × 52 × 61 cm (W×L×D) stainless steel tub. Approximately 150 kg of seed could be extracted in this tub. The extraction was done with 80% ethanol. When the solvent front migrated to bottom of the column, flow was stopped and seed was allowed to steep for at least 48 h. Light was excluded from the solvent from the beginning of this step to the end of the purification process. Following elution, ethanol was removed from the eluate by evaporation at room temperature. Resulting syrup was freeze-dried. About 80% recovery of alkaloids was achieved with 18-fold increase in concentration of ergovaline. Initial purification of the dried product was accomplished by extracting with hexane/water (6:1, v/v) and the hexane fraction was discarded. The aqueous fraction was extracted with chloroform, the aqueous layer discarded, after which the chloroform was removed with a resulting 20-fold increase of ergovaline. About 65% of the ergovaline was recovered from the chloroform residue for an overall recovery of 50%. The resultant partially purified ergovaline had biological activities in in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline.

Optimal mixing rate in linear solvent strength gradient liquid chromatography. Balanced mixing program.

PubMed

Blumberg, Leonid M; Desmet, Gert

2016-12-09

The mixing rate (R ϕ ) is the temporal rate of increase in the solvent strength in gradient LC. The optimal R ϕ (R ϕ ,Opt ) is the one at which a required peak capacity of gradient LC analysis is obtained in the shortest time. The balanced mixing program is a one where, for better separation of early eluting solutes, the mixing ramp is preceded by a balanced isocratic hold of the duration depending on R ϕ . The improvement in the separation of the earlier eluites due to the balanced programming has been evaluated. The value of R ϕ ,Opt depends on the solvent composition range covered by the mixing ramp and on the column pressure conditions. The R ϕ ,Opt for a column operating at maximum instrumental pressure is different from R ϕ ,Opt for a column operating below the instrumental pressure limit. On the other hand, it has been shown that the difference in the R ϕ ,Opt values under different conditions is not very large so that a single default R ϕ previously recommended for gradient analyses without the isocratic hold also yields a good approximation to the shortest analysis time for all conditions in the balanced analyses. With or without the initial balance isocratic hold, the recommended default R ϕ is about 5%/t 0 (5% increase in the solvent strength per each t 0 -long increment in time) for small-molecule samples, and about an order of magnitude slower (0.5%/t 0 ) for protein samples. A discussion illustrating the use of the optimization criteria employed here for the techniques other than LSS gradient LC is included. Copyright © 2016 Elsevier B.V. All rights reserved.

Microwave-Assisted γ-Valerolactone Production for Biomass Lignin Extraction: A Cascade Protocol.

PubMed

Tabasso, Silvia; Grillo, Giorgio; Carnaroglio, Diego; Calcio Gaudino, Emanuela; Cravotto, Giancarlo

2016-03-26

The general need to slow the depletion of fossil resources and reduce carbon footprints has led to tremendous effort being invested in creating "greener" industrial processes and developing alternative means to produce fuels and synthesize platform chemicals. This work aims to design a microwave-assisted cascade process for a full biomass valorisation cycle. GVL (γ-valerolactone), a renewable green solvent, has been used in aqueous acidic solution to achieve complete biomass lignin extraction. After lignin precipitation, the levulinic acid (LA)-rich organic fraction was hydrogenated, which regenerated the starting solvent for further biomass delignification. This process does not requires a purification step because GVL plays the dual role of solvent and product, while the reagent (LA) is a product of biomass delignification. In summary, this bio-refinery approach to lignin extraction is a cascade protocol in which the solvent loss is integrated into the conversion cycle, leading to simplified methods for biomass valorisation.

Solvent screening for a hard-to-dissolve molecular crystal.

PubMed

Maiti, A; Pagoria, P F; Gash, A E; Han, T Y; Orme, C A; Gee, R H; Fried, L E

2008-09-01

Materials with a high-degree of inter- and intra-molecular hydrogen bonding generally have limited solubility in conventional organic solvents. This presents a problem for the dissolution, manipulation and purification of these materials. Using a state-of-the-art density-functional-theory based quantum chemical solvation model we systematically evaluated solvents for a known hydrogen-bonded molecular crystal. This, coupled with direct solubility measurements, uncovered a class of ionic liquids involving fluoride anions that possess more than two orders of magnitude higher solvation power as compared with the best conventional solvents. The crystal structure of one such ionic liquid, determined by X-ray diffraction spectroscopy, indicates that F- ions are stabilized through H-bonded chains with water. The presence of coordinating water in such ionic liquids seems to facilitate the dissolution process by keeping the chemical activity of the F- ions in check.

Step-wise supercritical extraction of carbonaceous residua

DOEpatents

Warzinski, Robert P.

1987-01-01

A method of fractionating a mixture containing high boiling carbonaceous material and normally solid mineral matter includes processing with a plurality of different supercritical solvents. The mixture is treated with a first solvent of high critical temperature and solvent capacity to extract a large fraction as solute. The solute is released as liquid from solvent and successively treated with other supercritical solvents of different critical values to extract fractions of differing properties. Fractionation can be supplemented by solute reflux over a temperature gradient, pressure let down in steps and extractions at varying temperature and pressure values.

Purification of bacteriophage M13 by anion exchange chromatography.

PubMed

Monjezi, Razieh; Tey, Beng Ti; Sieo, Chin Chin; Tan, Wen Siang

2010-07-01

M13 is a non-lytic filamentous bacteriophage (phage). It has been used widely in phage display technology for displaying foreign peptides, and also for studying macromolecule structures and interactions. Traditionally, this phage has been purified by cesium chloride (CsCl) density gradient ultracentrifugation which is highly laborious and time consuming. In the present study, a simple, rapid and efficient method for the purification of M13 based on anion exchange chromatography was established. A pre-packed SepFast Super Q column connected to a fast protein liquid chromatography (FPLC) system was employed to capture released phages in clarified Escherichia coli fermented broth. An average yield of 74% was obtained from a packed bed mode elution using citrate buffer (pH 4), containing 1.5 M NaCl at 1 ml/min flow rate. The purification process was shortened substantially to less than 2 h from 18 h in the conventional ultracentrifugation method. SDS-PAGE revealed that the purity of particles was comparable to that of CsCl gradient density ultracentrifugation method. Plaque forming assay showed that the purified phages were still infectious. Copyright 2010 Elsevier B.V. All rights reserved.

Tandem dye-ligand chromatography and biospecific elution applied to the purification of glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides.

PubMed Central

Hey, Y; Dean, P D

1983-01-01

1. A total of 65 immobilized triazine dyes were screened for their ability to purify the dual-nucleotide-specific glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides. From this screen a 'negative' (Matrex Gel Purple A) and a 'positive' (Matrex Gel Orange B) adsorbent were found to be the best in terms of overall purification and yield and were therefore combined to give the best purification. 2. Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides was purified approx. 56-fold in a two-step tandem chromatographic system using Matrex Gel Purple A followed by Matrex Gel Orange B chromatography to a specific activity of 228 units/mg of protein in a final yield of 73%. 3. A study of the elution characteristics of glucose-6-phosphate dehydrogenase bound to Matrex Gel Orange B by KCl (pulse and gradient) and biospecific eluents (pulse) was carried out. NADP+, NADPH and adenosine 2',5'-bisphosphate were found to be the only effective biospecific eluents. A pulse of 50 microM-NADP+ (1/2 column vol.) was found to give a better purification than a 0-1 M-KCl gradient and therefore was the preferred method of elution. 4. Presaturation of the enzyme with various nucleotides was carried out to determine the effect on the subsequent binding of glucose-6-phosphate dehydrogenase to Matrex Gel Orange B. The results of these and biospecific-elution studies lead us to propose two possible schemes to explain the mechanism of the dye-protein interaction. 5. Reusability, capacity of the adsorbent and effect of varying the ligand concentration were also studied in the purification of glucose-6-phosphate dehydrogenase on Matrex Gel Orange B. Images Fig. 1. PMID:6847623

Tandem dye-ligand chromatography and biospecific elution applied to the purification of glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides.

PubMed

Hey, Y; Dean, P D

1983-02-01

1. A total of 65 immobilized triazine dyes were screened for their ability to purify the dual-nucleotide-specific glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides. From this screen a 'negative' (Matrex Gel Purple A) and a 'positive' (Matrex Gel Orange B) adsorbent were found to be the best in terms of overall purification and yield and were therefore combined to give the best purification. 2. Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides was purified approx. 56-fold in a two-step tandem chromatographic system using Matrex Gel Purple A followed by Matrex Gel Orange B chromatography to a specific activity of 228 units/mg of protein in a final yield of 73%. 3. A study of the elution characteristics of glucose-6-phosphate dehydrogenase bound to Matrex Gel Orange B by KCl (pulse and gradient) and biospecific eluents (pulse) was carried out. NADP+, NADPH and adenosine 2',5'-bisphosphate were found to be the only effective biospecific eluents. A pulse of 50 microM-NADP+ (1/2 column vol.) was found to give a better purification than a 0-1 M-KCl gradient and therefore was the preferred method of elution. 4. Presaturation of the enzyme with various nucleotides was carried out to determine the effect on the subsequent binding of glucose-6-phosphate dehydrogenase to Matrex Gel Orange B. The results of these and biospecific-elution studies lead us to propose two possible schemes to explain the mechanism of the dye-protein interaction. 5. Reusability, capacity of the adsorbent and effect of varying the ligand concentration were also studied in the purification of glucose-6-phosphate dehydrogenase on Matrex Gel Orange B.

Activated charcoal filters: Water treatment, pollution control, and industrial applications. (Latest citations from the Patent Bibliographic database with exemplary claims. ) Published Search

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

The bibliography contains citations of selected patents concerning activated charcoal filters and their applications in water treatment, pollution control, and industrial processes. Filtering methods and equipment for air and water purification, industrial distillation and extraction, industrial leaching, and filtration of toxic materials and contaminants are described. Applications include drinking water purification, filtering beverages, production of polymer materials, solvent and metal recovery, waste conversion, automotive fuel and exhaust systems, swimming pool filtration, tobacco smoke filters, kitchen ventilators, medical filtration treatment, and odor absorbing materials. (Contains 250 citations and includes a subject term index and title list.)

Journal of Chemical Education: Software.

ERIC Educational Resources Information Center

Journal of Chemical Education, 1988

1988-01-01

Describes a chemistry software program that emulates a modern binary gradient HPLC system with reversed phase column behavior. Allows for solvent selection, adjustment of gradient program, column selection, detectory selection, handling of computer sample data, and sample preparation. (MVL)

A low-cost gradient system for high-performance liquid chromatography. Quantitation of complex pharmaceutical raw materials.

PubMed

Erni, F; Frei, R W

1976-09-29

A device is described that makes use of an eight-port motor valve to generate step gradients on the low-pressure side of a piston pump with a low dead volume. Such a gradient device with an automatic control unit, which also permits repetition of previous steps, can be built for about half the cost of a gradient system with two pumps. Applications of this gradient unit to the separation of complex mixtures of glycosides and alkaloids are discussed and compared with separations systems using two high-pressure pumps. The gradients that are used on reversed-phase material with solvent mixtures of water and completely miscible organic solvents are suitable for quantitative routine control of pharmaceutical products. The reproducibility of retention data is excellent over several months and, with the use of loop injectors, major components can be determined quantitatively with a reproducibility of better than 2% (relative standard deviation). The step gradient selector valve can also be used as an introduction system for very large sample volumes. Up to 11 can be injected and samples with concentrations of less than 1 ppb can be determined with good reproducibilities.

Production, purification and characterization of halophilic organic solvent tolerant protease from marine crustacean shell wastes and its efficacy on deproteinization.

PubMed

Maruthiah, Thirumalai; Somanath, Beena; Jasmin, Jebamonydhas Vijila; Immanuel, Grasian; Palavesam, Arunachalam

2016-12-01

The quantum of marine fish wastes produced by fish processing industries has necessitated to search new methods for its disposal. Hence, this study is focused on production and purification of halophilic organic solvent tolerant protease (HOSP) from marine Alcaligenes faecalis APCMST-MKW6 using marine shell wastes as substrate. The candidate bacterium was isolated from the marine sediment of Manakudi coast and identified as A. faecalis APCMST-MKW6. The purified protease showed 16.39-fold purity, 70.34 U/mg specific activity with 21.67 % yield. The molecular weight of the purified alkaline protease was 49 kDa. This purified protease registered maximum activity at pH 9 and it was stable between pH 8-9 after 1.30 h of incubation. The optimum temperature registered was 60 °C and it was stable between 50 and 60 °C even after 1.30 h of incubation. This enzyme also showed maximum activity at 20 % NaCl concentration. Further, manganese chloride, magnesium chloride, calcium chloride and barium chloride influenced this enzyme activity remarkably and it was also found to be enhanced by many of the tested surfactants and solvents. The candidate bacterium effectively deproteinized the shrimp shell waste compared to the other tested crustaceans shell wastes and also attained maximum antioxidant activity.

Preparative separation of six rhynchophylla alkaloids from Uncaria macrophylla wall by pH-zone refining counter-current chromatography.

PubMed

Zhang, Qinghai; Lin, Changhu; Duan, Wenjuan; Wang, Xiao; Luo, Aiqin

2013-12-12

pH-Zone refining counter-current chromatography was successfully applied to the preparative isolation and purification of six alkaloids from the ethanol extracts of Uncaria macrophylla Wall. Because of the low content of alkaloids (about 0.2%, w/w) in U. macrophylla Wall, the target compounds were enriched by pH-zone refining counter-current chromatography using a two-phase solvent system composed of petroleum ether-ethyl acetate-isopropanol-water (2:6:3:9, v/v), adding 10 mM triethylamine in organic stationary phase and 5 mM hydrochloric acid in aqueous mobile phase. Then pH-zone refining counter-current chromatography using the other two-phase solvent system was used for final purification. Six target compounds were finally isolated and purified by following two-phase solvent system composed of methyl tert-butyl ether (MTBE)-acetonitrile-water (4:0.5:5, v/v), adding triethylamine (TEA) (10 mM) to the organic phase and HCl (5 mM) to aqueous mobile phase. The separation of 2.8 g enriched total alkaloids yielded 36 mg hirsutine, 48 mg hirsuteine, 82 mg uncarine C, 73 mg uncarine E, 163 mg rhynchophylline, and 149 mg corynoxeine, all with purities above 96% as verified by HPLC Their structures were identified by electrospray ionization-mass spectrometry (ESI-MS) and 1H-NMR spectroscopy.

Computational solvent system screening for the separation of tocopherols with centrifugal partition chromatography using deep eutectic solvent-based biphasic systems.

PubMed

Bezold, Franziska; Weinberger, Maria E; Minceva, Mirjana

2017-03-31

Tocopherols are a class of molecules with vitamin E activity. Among those, α-tocopherol is the most important vitamin E source in the human diet. The purification of tocopherols involving biphasic liquid systems can be challenging since these vitamins are poorly soluble in water. Deep eutectic solvents (DES) can be used to form water-free biphasic systems and have already proven applicable for centrifugal partition chromatography separations. In this work, a computational solvent system screening was performed using the predictive thermodynamic model COSMO-RS. Liquid-liquid equilibria of solvent systems composed of alkanes, alcohols and DES, as well as partition coefficients of α-tocopherol, β-tocopherol, γ-tocopherol, and σ-tocopherol in these biphasic solvent systems were calculated. From the results the best suited biphasic solvent system, namely heptane/ethanol/choline chloride-1,4-butanediol, was chosen and a batch injection of a tocopherol mixture, mainly consisting of α- and γ-tocopherol, was performed using a centrifugal partition chromatography set up (SCPE 250-BIO). A separation factor of 1.74 was achieved for α- and γ-tocopherol. Copyright © 2017 Elsevier B.V. All rights reserved.

Microwave-assisted extraction and purification of chlorogenic acid from by-products of Eucommia Ulmoides Oliver and its potential anti-tumor activity.

PubMed

Shao, P; Zhang, J F; Chen, X X; Sun, P L

2015-08-01

An efficient method for the rapid extraction, separation and purification of chlorogenic acid (CGA) from by-products of Eucommia Ulmoides Oliver (E. ulmoides) by microwave-assisted extraction (MAE) coupled with high-speed counter-current chromatography (HSCCC) was developed. The optimal MAE parameters were evaluated by response surface methodology (RSM), and they were extraction time of 12 min, microwave power of 420 W, ethanol concentration of 75 %, solvent/sample ratio of 30:1 (mL/g), yield of CGA reached 3.59 %. The crude extract was separated and purified directly by HSCCC using ethyl acetate-butyl alcohol-water (3:1:4, v/v) as the two-phase solvent system. The 14.5 mg of CGA with the purity of 98.7 % was obtained in one-step separation from 400 mg of crude extract. The chemical structure of CGA was verified with IR, ESI-MS analysis. Meanwhile, the purified CGA extract was evaluated by MTT assay and results indicate that CGA extract exhibited potential anti-tumor activity for AGS gastric cancer cell.

Extraction and Determination of Quercetin from Ginkgo biloba by DESs-Based Polymer Monolithic Cartridge.

PubMed

Wang, Xiaoqin; Li, Guizhen; Ho Row, Kyung

2017-09-01

Deep eutectic solvents (DES) were formed from choline chloride (ChCl). DES-modified polymer monolithic (DES-M), template molecular polymer monolithic and non-DES-M without a molecular template were synthesized in identical process. These polymer materials were characterized by field emission scanning electron microscopy and Fourier transform infrared spectroscopy. The significant selective adsorption properties of the polymers were assessed by an absorption capacity experiment and solid-phase extraction (SPE). The optimized extraction procedure was as follows: ultrasonic time (30 min), optimal solvent (ethanol) and liquid to material ratio (20 mL g-1). Under this condition, the amount of quercetin extracted from Ginkgo biloba was 290.8 mg g-1. The purification of G. biloba was achieved by the SPE process. Based on the results, DESs-based monolithic cartridges can be used for simple and efficient extraction and as a pre-concentration technique for the purification of bioactive compounds or drugs in aqueous environments with high affinity and selectivity. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Tannase-mediated biotransformation assisted separation and purification of theaflavin and epigallocatechin by high speed counter current chromatography and preparative high performance liquid chromatography: A comparative study.

PubMed

Xia, Guobin; Lin, Chunfang; Liu, Songbai

2016-09-01

A large scale isolation and purification of theaflavin (TF) and epigallocatechin (EGC) has been successfully developed by tannase-mediated biotransformation combining high-speed countercurrent chromatography. After tannase hydrolysis of a commercially available theaflavins extract (TE), the content of TF and EGC in tannase-mediated biotransformation product (TBP) achieved approximately 3 times enrichment. SEM studies revealed smooth tannase biotransformation and the possibility of recovery of the tannase. A single 1.5 hours' HSCCC separation for TF and EGC employing a two-phase solvent system could simultaneously produce 180.8 mg of 97.3% purity TF and 87.5 mg of 97.3% purity EGC. However, a preparative HPLC separation of maximum injection volume containing 120 mg TBP prepared 11.2 mg TF of 94.9% purity and 7.7 mg EGC of 89.9% purity. HSCCC separation demonstrated significant advantages over Prep HPLC in terms of sample loading size, separation time, environmental friendly solvent systems, and the production. © 2016 Wiley Periodicals, Inc.

Eco-Contribution for the Production of N-Arylnitrones: Solvent-Free and Assisted by Microwaves

PubMed Central

Reyes, Leonor; Corona, Sandra; Arroyo, Gabriel; Delgado, Francisco; Miranda, René

2010-01-01

A simple green approach for the production of benzylideneaniline oxides is offered. This contribution was performed via the condensation of phenylhydroxylamine with several aryl aldehydes, in the absence of both catalyst and solvent, and using microwave irradiation as the activating reaction mode. In addition, good yields of the products were achieved in a short time. It is also worth noting that the work-up procedure is simple and the products do not require further purification. Finally, an interesting comparison without the use of microwave irradiation is also discussed. PMID:20640169

Poly(ethylene oxide) functionalization

DOEpatents

Pratt, Russell Clayton

2014-04-08

A simple procedure is provided by which the hydroxyl termini of poly(ethylene oxide) can be appended with functional groups to a useful extent by reaction and precipitation. The polymer is dissolved in warmed toluene, treated with an excess of organic base and somewhat less of an excess of a reactive acylating reagent, reacted for several hours, then precipitated in isopropanol so that the product can be isolated as a solid, and salt byproducts are washed away. This procedure enables functionalization of the polymer while not requiring laborious purification steps such as solvent-solvent extraction or dialysis to remove undesirable side products.

Partial Purification of a Legume Nodulation Factor Present in Coconut Water 1

PubMed Central

Schaffer, A. G.; Alexander, M.

1967-01-01

The nodulation of adventitious roots growing from segments of bean hypocotyl tissue was used as a bioassay for the material present in coconut water which stimulated nodulation. The active material in coconut water is acidic, but it was not possible to extract it from an acid solution with organic solvents. A purification of approximately 70-fold (on a dry wt basis) was obtained using activated charcoal, but at least 10 different compounds were present in the active fractions. A purified fraction of coconut water, which is stimulatory to the growth of carrot root explants, was active in the nodulation assay at a concentration of 2 μg/ml. This represents a 4000-fold purification of the diffusible fraction of coconut water. The charcoal fractionation procedure can be applied to the active material present in extracts of bean leaves. PMID:16656538

Application of an efficient strategy based on liquid-liquid extraction, high-speed counter-current chromatography, and preparative HPLC for the rapid enrichment, separation, and purification of four anthraquinones from Rheum tanguticum.

PubMed

Chen, Tao; Liu, Yongling; Zou, Denglang; Chen, Chen; You, Jinmao; Zhou, Guoying; Sun, Jing; Li, Yulin

2014-01-01

This study presents an efficient strategy based on liquid-liquid extraction, high-speed counter-current chromatography, and preparative HPLC for the rapid enrichment, separation, and purification of four anthraquinones from Rheum tanguticum. A new solvent system composed of petroleum ether/ethyl acetate/water (4:2:1, v/v/v) was developed for the liquid-liquid extraction of the crude extract from R. tanguticum. As a result, emodin, aloe-emodin, physcion, and chrysophanol were greatly enriched in the organic layer. In addition, an efficient method was successfully established to separate and purify the above anthraquinones by high-speed counter-current chromatography and preparative HPLC. This study supplies a new alternative method for the rapid enrichment, separation, and purification of emodin, aloe-emodin, physcione, and chrysophanol. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Preparative isolation, fast centrifugal partition chromatography purification and biological activity of cajaflavanone from Derris ferruginea stems.

PubMed

Morel, Sylvie; Landreau, Anne; Nguyen, Van Hung; Derbré, Séverine; Grellier, Philippe; Pape, Patrice Le; Pagniez, Fabrice; Litaudon, Marc; Richomme, Pascal

2012-01-01

The Derris genus is known to contain flavonoid derivatives, including prenylated flavanones and isoflavonoids such as rotenoids, which are generally associated with significant biological activity. To develop an efficient preparative isolation procedure for bioactive cajaflavanone. Fast centrifugal partition chromatography (FCPC) was optimised to purify cajaflavanone from Derris ferruginea stems in a single step as compared to fractionation from the cyclohexane extract by successive conventional solid-liquid chromatography procedures. The purification yield, purity, time and solvent consumption per procedure are described. The anti-fungal, anti-bacterial, anti-leishmanial, anti-plasmodial, anti-oxidant activities and the inhibition of advanced glycation end-products (AGEs) by cajaflavanone accumulation are described. FCPC enabled cajaflavanone purification in a single separation step, yielding sufficient quantities to perform in vitro biological screening. Interestingly, cajaflavanone had an inhibitory effect on the formation of AGEs, without displaying any in vitro anti-oxidant activity. A simple and efficient procedure, in comparison with other preparative methods, for bioactive cajaflavone purification has been developed using FCPC. Copyright © 2011 John Wiley & Sons, Ltd.

Gram-scale purification of aconitine and identification of lappaconitine in Aconitum karacolicum.

PubMed

Tarbe, M; de Pomyers, H; Mugnier, L; Bertin, D; Ibragimov, T; Gigmes, D; Mabrouk, K

2017-07-01

Aconitum karacolicum from northern Kyrgyzstan (Alatau area) contains about 0.8-1% aconitine as well as other aconite derivatives that have already been identified. In this paper, we compare several methods for the further purification of an Aconitum karacolicum extract initially containing 80% of aconitine. Reverse-phase flash chromatography, reverse-phase semi-preparative HPLC, centrifugal partition chromatography (CPC) and recrystallization techniques were evaluated regarding first their efficiency to get the highest purity of aconitine (over 96%) and secondly their applicability in a semi-industrial scale purification process (in our case, 150g of plant extract). Even if the CPC technique shows the highest purification yield (63%), the recrystallization remains the method of choice to purify a large amount of aconitine as i) it can be easily carried out in safe conditions; ii) an aprotic solvent is used, avoiding aconitine degradation. Moreover, this study led us to the identification of lappaconitine in Aconitum karacolicum, a well-known alkaloid never found in this Aconitum species. Copyright © 2017 Elsevier B.V. All rights reserved.

Toxoplasma gondii tissue cyst purification using Percoll gradients

PubMed Central

Watts, Elizabeth A.; Dhara, Animesh; Sinai, Anthony P.

2017-01-01

The protozoan parasite Toxoplasma gondii is capable of infecting all warm blooded animals and humans. Infectious, transmissible forms of the parasite include oocysts produced by the sexual cycle within the definitive feline host and tissue cysts that form Toxoplasma in the CNS and muscle during the asexual cycle within all chronically infected warm-blooded hosts. These tissue cysts are populated with slow growing bradyzoites which have been until recently thought to be dormant entities in the context of immune sufficiency. Reactivation to active growth during immune suppression is of critical clinical importance. Yet we know little about tissue cysts or the bradyzoites they house as the diversity of tissue cysts cannot be replicated in cell culture systems. Our optimization of tissue cyst purification from the brains of infected mice using Percoll gradients provides an efficient means to recover in vivo derived tissue cysts that can be applied to imaging, cell-biologic, biochemical, transcriptomic and proteomic analyses. PMID:28510363

Efficient purification of paclitaxel from yews using high-performance displacement chromatography technique.

PubMed

Watchueng, Jean; Kamnaing, Pierre; Gao, Jin-Ming; Kiyota, Taira; Yeboah, Faustinus; Konishi, Yasuo

2011-05-20

Paclitaxel was purified using high-performance displacement chromatography (HPDC) technique, but not by the mechanism of HPDC. On small scale, paclitaxel was extracted with methanol from dry needles of Taxus canadensis and was enriched by extracting with chloroform after removing water-soluble hydrophilic components and hexane-soluble hydrophobic components. Then, 93-99% purity of paclitaxel was obtained using the HPDC technique. On large scale, taxanes were enriched by solvent partitioning between acetic acid/MeOH/H(2)O and hexane and extracted with CH(2)Cl(2). Taxanes except paclitaxel were further removed by extracting with methanol-water-trifluoroacetic acid (1.0:98.9:0.1, v/v/v). Applying HPDC technique to water-insoluble substances is problematic as this method requires a highly aqueous solvent system. In order to overcome this incompatibility, a system was set up where paclitaxel, although in low concentration, was extracted by methanol-water-trifluoroacetic acid (10.0:89.9:0.1, v/v/v). Recycling the extracting solvent to ensure minimal volume, the extracted paclitaxel was adsorbed on a C(18) trap column. A C(18) column of 4.6mm internal diameter was then connected to the trap column. The HPDC technique was thus carried out using an isocratic acetonitrile-water-trifluoroacetic acid (30.0:69.9:0.1, v/v/v) mobile phase consisting of a displacer cetylpyridinium trifluoroacetate (3mg/mL). Paclitaxel was co-eluted with the displacer and spontaneously crystallized. The crystal (114mg) showed 99.4% purity and only 10% of paclitaxel in the starting crude extract was lost during the enrichment/purification processes. This large scale purification method was successfully applied to purify paclitaxel from Chinese yew in small scale, suggesting general applicability of the method. This is the first report of purifying a water-insoluble natural product using HPDC technique. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

A microfluidic study of liquid-liquid extraction mediated by carbon dioxide.

PubMed

Lestari, Gabriella; Salari, Alinaghi; Abolhasani, Milad; Kumacheva, Eugenia

2016-07-05

Liquid-liquid extraction is an important separation and purification method; however, it faces a challenge in reducing the energy consumption and the environmental impact of solvent (extractant) recovery. The reversible chemical reactions of switchable solvents (nitrogenous bases) with carbon dioxide (CO2) can be implemented in reactive liquid-liquid extraction to significantly reduce the cost and energy requirements of solvent recovery. The development of new effective switchable solvents reacting with CO2 and the optimization of extraction conditions rely on the ability to evaluate and screen the performance of switchable solvents in extraction processes. We report a microfluidic strategy for time- and labour-efficient studies of CO2-mediated solvent extraction. The platform utilizes a liquid segment containing an aqueous extractant droplet and a droplet of a solution of a switchable solvent in a non-polar liquid, with gaseous CO2 supplied to the segment from both sides. Following the reaction of the switchable solvent with CO2, the solvent becomes hydrophilic and transfers from the non-polar solvent to the aqueous droplet. By monitoring the time-dependent variation in droplet volumes, we determined the efficiency and extraction time for the CO2-mediated extraction of different nitrogenous bases in a broad experimental parameter space. The platform enables a significant reduction in the amount of switchable solvents used in these studies, provides accurate temporal characterization of the liquid-liquid extraction process, and offers the capability of high-throughput screening of switchable solvents.

Purification of infectious adenovirus in two hours by ultracentrifugation and tangential flow filtration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ugai, Hideyo; Yamasaki, Takahito; Hirose, Megumi

2005-06-17

Adenoviruses are excellent vectors for gene transfer and are used extensively for high-level expression of the products of transgenes in living cells. The development of simple and rapid methods for the purification of stable infectious recombinant adenoviruses (rAds) remains a challenge. We report here a method for the purification of infectious adenovirus type 5 (Ad5) that involves ultracentrifugation on a cesium chloride gradient at 604,000g for 15 min at 4 deg C and tangential flow filtration. The entire procedure requires less than two hours and infectious Ad5 can be recovered at levels higher than 64% of the number of plaque-formingmore » units (pfu) in the initial crude preparation of viruses. We have obtained titers of infectious purified Ad5 of 1.35 x 10{sup 10} pfu/ml and a ratio of particle titer to infectious titer of seven. The method described here allows the rapid purification of rAds for studies of gene function in vivo and in vitro, as well as the rapid purification of Ad5.« less

Rapid step-gradient purification of mitochondrial DNA.

PubMed

Welter, C; Meese, E; Blin, N

1988-01-01

A convenient modification of the step gradient (CsCl/ethidium bomide) procedure is described. This rapid method allows isolation of covalently closed circular DNA separated from contaminating proteins, RNA and chromosomal DNA in ca. 5 h. Large scale preparations can be performed for circular DNA from eukaryotic organelles (mitochondria). The protocol uses organelle pelleting/NaCl-sarcosyl incubation steps for mitochondria followed by a CsCl step gradient and exhibits yields equal to the conventional procedures. It results in DNA sufficiently pure to be used for restriction endonuclease analysis, subcloning, 5'-end labeling, gel retention assays, and various types of hybridization.

Determination of sulfonamides and trimethoprim using high temperature HPLC with simultaneous temperature and solvent gradient.

PubMed

Giegold, Sascha; Teutenberg, Thorsten; Tuerk, Jochen; Kiffmeyer, Thekla; Wenclawiak, Bernd

2008-10-01

A fast HPLC method for the analysis of eight selected sulfonamides (SA) and trimethoprim has been developed with the use of high temperature HPLC. The separation could be achieved in less than 1.5 min on a 50 mm sub 2 microm column with simultaneous solvent and temperature gradient programming. Due to the lower viscosity of the mobile phase and the increased mass transfer at higher temperatures, the separation could be performed on a conventional HPLC system obtaining peak widths at half height between 0.6 and 1.3 s.

Single fiber lignin distributions based on the density gradient column method

Treesearch

Brian Boyer; Alan W. Rudie

2007-01-01

The density gradient column method was used to determine the effects of uniform and non-uniform pulping processes on variation in individual fiber lignin concentrations of the resulting pulps. A density gradient column uses solvents of different densities and a mixing process to produce a column of liquid with a smooth transition from higher density at the bottom to...

Regenerated silica gel as stationary phase on vacuum column chromatography to purify temulawak's extracts

NASA Astrophysics Data System (ADS)

Cahyono, Bambang; Maduwu, Ratna Dewi; Widayat, Suzery, Meiny

2015-12-01

Commercial silica gel only used once by many researchers and affected high cost for purification process, also less support the green chemistry program. This research focused in regeneration silica gel that used purification of temulawak's extracts (Curcuma xanthorrhiza Roxb) by vacuum column chromatography. Sample extracts (contains 10.1195±0.5971% of curcuminoids) was purified by vacuum column chromatography (pressure: 45 kPa, column: 100mm on length and 16mm on diameter). Ethanol 96% and acetone were compared as eluent. The amount of solvent and yield of curcuminoids used as indicator purification. The silica gel was regenerated with heating in 600°C for 8 hours The silica gels were analyzed by IR spectroscopy and X-ray diffraction. Furthermore, regenerated silica gel was used as the stationary phase in vacuum column chromatography under the same conditions with the previous purification. All the purification experiments were performed in three repetitions. Based on regression equation, y=0.132x+0.0011 (r2=0.9997) the yield of curcuminoids on purified products using ethanol as the eluent was improved 4.26% (to 14.3724±0.5749%) and by acetone was improved 3,03% (to 13.1450 ±0.6318%). The IR spectrum of both silica gel showed the same vibration profile and also there were three crystallinity peaks missing on its X-ray diffraction. Regenerated silica gel has the same performance with new silica gel in purification of temulawak's extract: by ethanol has increased 4.08% (14.1947±0.7415%) and 2.93% (13.0447±0.4822) by acetone. In addition, all purification products showed similar TLC profiles. Purification using regenerated silica gel as the adsorbent on vacuum column chromatography has exactly same potential with the new silica gel.

Preparative high-speed counter-current chromatography for purification of shikonin from the Chinese medicinal plant Lithospermum erythrorhizon.

PubMed

Lu, Hai-Tao; Jiang, Yue; Chen, Feng

2004-01-09

The bioactive compound shikonin was successfully isolated and purified from the crude extract of the traditional Chinese medicinal plant Lithospermum erythrorhizon Sieb. et Zucc. by preparative high-speed counter-current chromatography (HSCCC). The preparative HSCCC was performed using a two-phase solvent system composed of n-hexane-ethylacetate-ethanol-water (16:14:14:5 (v/v)). A total amount of 19.6 mg of shikonin at 98.9% purity was obtained from 52 mg of the crude extract (containing 38.9% shikonin) with 96.9% recovery. The preparative isolation and purification of shikonin by HSCCC was completed in 200 min in a one-step separation.

Activated charcoal filters: Water treatment, pollution control, and industrial applications. October 1970-October 1989 (Citations from the US Patent data base). Report for October 1970-October 1989

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

This bibliography contains citations of selected patents concerning activated charcoal filters and their applications in water treatment, pollution control, and industrial processes. Filtering methods and equipment for air and water purification, industrial distillation and extraction, industrial leaching, and filtration of toxic materials and contaminants are described. Applications include drinking water purification, filtering beverages, production of polymer materials, solvent and metal recovery, waste conversion, automotive fuel and exhaust systems, swimming-pool filtration, tobacco-smoke filters, kitchen ventilators, medical filtration treatment, and odor-absorbing materials. (This updated bibliography contains 173 citations, 12 of which are new entries to the previous edition.)

Activated-charcoal filters: Water treatment, pollution control, and industrial applications. January 1970-August 1989 (Citations from the US Patent data base). Report for January 1970-August 1989

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

This bibliography contains citations of selected patents concerning activated charcoal filters and their applications in water treatment, pollution control, and industrial processes. Filtering methods and equipment for air and water purification, industrial distillation and extraction, industrial leaching, and filtration of toxic materials and contaminants are described. Applications include drinking-water purification, filtering beverages, production of polymer materials, solvent and metal recovery, waste conversion, automotive fuel and exhaust systems, swimming-pool filtration, tobacco-smoke filters, kitchen ventilators, medical-filtration treatment, and odor absorbing materials. (This updated bibliography contains 161 citations, 32 of which are new entries to the previous edition.)

Incorporating high-pressure electroosmotic pump and a nano-flow gradient generator into a miniaturized liquid chromatographic system for peptide analysis.

PubMed

Chen, Apeng; Lynch, Kyle B; Wang, Xiaochun; Lu, Joann J; Gu, Congying; Liu, Shaorong

2014-09-24

We integrate a high-pressure electroosmotic pump (EOP), a nanoflow gradient generator, and a capillary column into a miniaturized liquid chromatographic system that can be directly coupled with a mass spectrometer for proteomic analysis. We have recently developed a low-cost high-pressure EOP capable of generating pressure of tens of thousands psi, ideal for uses in miniaturized HPLC. The pump worked smoothly when it was used for isocratic elutions. When it was used for gradient elutions, generating reproducible gradient profiles was challenging; because the pump rate fluctuated when the pump was used to pump high-content organic solvents. This presents an issue for separating proteins/peptides since high-content organic solvents are often utilized. In this work, we solve this problem by incorporating our high-pressure EOP with a nano-flow gradient generator so that the EOP needs only to pump an aqueous solution. With this combination, we develop a capillary-based nano-HPLC system capable of performing nano-flow gradient elution; the pump rate is stable, and the gradient profiles are reproducible and can be conveniently tuned. To demonstrate its utility, we couple it with either a UV absorbance detector or a mass spectrometer for peptide separations. Copyright © 2014. Published by Elsevier B.V.

[Simultaneous determination of five cold medicine ingredients in paracetamol triprolidine hydrochloride and pseudoephedrine hydrochloride tablets by pH/organic solvent double-gradient high performance liquid chromatography].

PubMed

Xuan, Xueyi; Huang, Lina; Pan, Xiaoling; Li, Ning

2013-02-01

A pH/organic solvent double-gradient mode in reversed-phase high performance liquid chromatography (HPLC) has been established as a new approach to the simultaneous determination of acetaminophen, caffeine, salicylamide, pseudoephedrine hydrochloride and triprolidine hydrochloride in paracetamol triprolidine hydrochloride and pseudoephedrine hydrochloride tablets. Through the optimization of the organic solvent gradient mode and pH/organic solvent double-gradient mode, the optimum double-gradient HPLC system of the five cold medicine ingredients has been built. The determination was carried out on a Diamonsiol C18 column (250 mm x 4.6 mm, 5 microm). The mobile phase consisted of methanol, 0.05 mol/L ammonium acetate solution and 0.08 mol/L acetic acid solution. The column temperature was set at 30 degrees C. The flow rate was 1.0 mL/min. The sample was measured at multiple wavelengths: 0-6 min, 280 nm; 6-7 min, 257 nm; 7-14 min, 280 nm; 14 min, 233 nm. The separation of the five cold medicine ingredients in the tablets was achieved in 25.5 min. The linear ranges of acetaminophen, pseudoephedrine hydrochloride, caffeine, salicylamide and triprolidine hydrochloride were 0.055 -0.998 g/L, 0.053-0.946 g/L, 0.007-0.129 g/L, 0.035-0.622 g/L and 0.002-0.039 g/L, respectively, with their correlation coefficients greater than 0.999 0. The detection limits (S/N = 3) were 0.09, 6, 0.02, 0.128 and 0.02 mg/L, respectively. Their mean recoveries were 97.9%-102.8%. The advantage of the method is the simultaneous determination of acidic, neutral and basic compounds. It also can improve the column efficiency of the analyte, compress the half-peak width and reduce the trailing. The optimized and validated method can be used for the simultaneous determination of the five cold medicine ingredients in the tablets.

Interaction chromatography for characterization and large-scale fractionation of chemically heterogeneous copolymers

NASA Astrophysics Data System (ADS)

Han, Junwon

The remarkable development of polymer synthesis techniques to make complex polymers with controlled chain architectures has inevitably demanded the advancement of polymer characterization tools to analyze the molecular dispersity in polymeric materials beyond size exclusion chromatography (SEC). In particular, man-made synthetic copolymers that consist of more than one monomer type are disperse mixtures of polymer chains that have distributions in terms of both chemical heterogeneity and chain length (molar mass). While the molecular weight distribution has been quite reliably estimated by the SEC, it is still challenging to properly characterize the chemical composition distribution in the copolymers. Here, I have developed and applied adsorption-based interaction chromatography (IC) techniques as a promising tool to characterize and fractionate polystyrene-based block, random and branched copolymers in terms of their chemical heterogeneity. The first part of this thesis is focused on the adsorption-desorption based purification of PS-b-PMMA diblock copolymers using nanoporous silica. The liquid chromatography analysis and large scale purification are discussed for the PS-b-PMMA block copolymers that have been synthesized by sequential anionic polymerization. SEC and IC are compared to critically analyze the contents of PS homopolymers in the as-synthesized block copolymers. In addition, I have developed an IC technique to provide faster and more reliable information on the chemical heterogeneity in the as-synthesized block copolymers. Finally, a large scale (multi-gram) separation technique is developed to obtain "homopolymer-free" block copolymers via a simple chromatographic filtration technique. By taking advantage of the large specific surface area of nanoporous silica (≈300m 2/g), large scale purification of neat PS-b-PMMA has successfully been achieved by controlling adsorption and desorption of the block copolymers on the silica gel surface using a gravity column. The second part of this thesis is focused on the liquid chromatography analysis and fractionation of RAFT-polymerized PS-b -PMMA diblock copolymers and AFM studies. In this study, PS- b-PMMA block copolymers were synthesized by a RAFT free radical polymerization process---the PMMA block with a phenyldithiobenzoate end group was synthesized first. The contents of unreacted PS and PMMA homopolymers in as-synthesized PS-b-PMMA block copolymers were quantitatively analyzed by solvent gradient interaction chromatography (SGIC) technique employing bare silica and C18-bonded silica columns, respectively. In addition, by 2-dimensional large-scale IC fractionation method, atomic force microscopy (AFM) study of these fractionated samples revealed various morphologies with respect to the chemical composition of each fraction. The third part of this thesis is to analyze random copolymers with tunable monomer sequence distributions using interaction chromatography. Here, IC was used for characterizing the composition and monomer sequence distribution in statistical copolymers of poly(styrene-co-4-bromostyrene) (PBrxS). The PBrS copolymers were synthesized by the bromination of monodisperse polystyrenes; the degree of bromination (x) and the sequence distribution were adjusted by varying the bromination time and the solvent quality, respectively. Both normal-phase (bare silica) and reversed-phase (C18-bonded silica) columns were used at different combinations of solvents and non-solvents to monitor the content of the 4-bromostyrene units in the copolymer and their average monomer sequence distribution. The fourth part of this thesis is to analyze and fractionate highly branched polymers such as dendronized polymers and star-shaped homo and copolymers. I have developed an interaction chromatography technique to separate polymers with nonlinear chain architecture. Specifically, the IC technique has been used to separate dendronized polymers and PS-based highly branched copolymers and to ultimately obtain well-defined dendronized or branched copolymers with a low polydispersity. The effects of excess arm-polymers on (1) the micellar self-assembly of dendronized polymers and (2) the regularity of the pore morphology in the low-k applications by the sol-gel process have been studied.

Lateral transport of solutes in microfluidic channels using electrochemically generated gradients in redox-active surfactants.

PubMed

Liu, Xiaoyang; Abbott, Nicholas L

2011-04-15

We report principles for a continuous flow process that can separate solutes based on a driving force for selective transport that is generated by a lateral concentration gradient of a redox-active surfactant across a microfluidic channel. Microfluidic channels fabricated with gold electrodes lining each vertical wall were used to electrochemically generate concentration gradients of the redox-active surfactant 11-ferrocenylundecyl-trimethylammonium bromide (FTMA) in a direction perpendicular to the flow. The interactions of three solutes (a hydrophobic dye, 1-phenylazo-2-naphthylamine (yellow AB), an amphiphilic molecule, 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl)-1-hexadecanoyl-sn-glycero-3-phosphocholine (BODIPY C(5)-HPC), and an organic salt, 1-methylpyridinium-3-sulfonate (MPS)) with the lateral gradients in surfactant/micelle concentration were shown to drive the formation of solute-specific concentration gradients. Two distinct physical mechanisms were identified to lead to the solute concentration gradients: solubilization of solutes by micelles and differential adsorption of the solutes onto the walls of the microchannels in the presence of the surfactant concentration gradient. These two mechanisms were used to demonstrate delipidation of a mixture of BODIPY C(5)-HPC (lipid) and MPS and purification of BODIPY C(5)-HPC from a mixture of BODIPY C(5)-HPC and yellow AB. Overall, the results of this study demonstrate that lateral concentration gradients of redox-active surfactants formed within microfluidic channels can be used to transport solutes across the microfluidic channels in a solute-dependent manner. The approach employs electrical potentials (<1 V) that are sufficiently small to avoid electrolysis of water, can be performed in solutions having high ionic strength (>0.1M), and offers the basis of continuous processes for the purification or separation of solutes in microscale systems. © 2011 American Chemical Society

Continuous Quadrupole Magnetic Separation of Islets during Digestion Improves Purified Porcine Islet Viability.

PubMed

Weegman, Bradley P; Kumar Sajja, Venkata Sunil; Suszynski, Thomas M; Rizzari, Michael D; Scott Iii, William E; Kitzmann, Jennifer P; Mueller, Kate R; Hanley, Thomas R; Kennedy, David J; Todd, Paul W; Balamurugan, Appakalai N; Hering, Bernhard J; Papas, Klearchos K

2016-01-01

Islet transplantation (ITx) is an emerging and promising therapy for patients with uncontrolled type 1 diabetes. The islet isolation and purification processes require exposure to extended cold ischemia, warm-enzymatic digestion, mechanical agitation, and use of damaging chemicals for density gradient separation (DG), all of which reduce viable islet yield. In this paper, we describe initial proof-of-concept studies exploring quadrupole magnetic separation (QMS) of islets as an alternative to DG to reduce exposure to these harsh conditions. Three porcine pancreata were split into two parts, the splenic lobe (SPL) and the combined connecting/duodenal lobes (CDL), for paired digestions and purifications. Islets in the SPL were preferentially labeled using magnetic microparticles (MMPs) that lodge within the islet microvasculature when infused into the pancreas and were continuously separated from the exocrine tissue by QMS during the collection phase of the digestion process. Unlabeled islets from the CDL were purified by conventional DG. Islets purified by QMS exhibited significantly improved viability (measured by oxygen consumption rate per DNA, p < 0.03) and better morphology relative to control islets. Islet purification by QMS can reduce the detrimental effects of prolonged exposure to toxic enzymes and density gradient solutions and substantially improve islet viability after isolation.

Continuous Quadrupole Magnetic Separation of Islets during Digestion Improves Purified Porcine Islet Viability

PubMed Central

Kumar Sajja, Venkata Sunil; Rizzari, Michael D.; Scott III, William E.; Kitzmann, Jennifer P.; Kennedy, David J.; Todd, Paul W.; Balamurugan, Appakalai N.; Hering, Bernhard J.

2016-01-01

Islet transplantation (ITx) is an emerging and promising therapy for patients with uncontrolled type 1 diabetes. The islet isolation and purification processes require exposure to extended cold ischemia, warm-enzymatic digestion, mechanical agitation, and use of damaging chemicals for density gradient separation (DG), all of which reduce viable islet yield. In this paper, we describe initial proof-of-concept studies exploring quadrupole magnetic separation (QMS) of islets as an alternative to DG to reduce exposure to these harsh conditions. Three porcine pancreata were split into two parts, the splenic lobe (SPL) and the combined connecting/duodenal lobes (CDL), for paired digestions and purifications. Islets in the SPL were preferentially labeled using magnetic microparticles (MMPs) that lodge within the islet microvasculature when infused into the pancreas and were continuously separated from the exocrine tissue by QMS during the collection phase of the digestion process. Unlabeled islets from the CDL were purified by conventional DG. Islets purified by QMS exhibited significantly improved viability (measured by oxygen consumption rate per DNA, p < 0.03) and better morphology relative to control islets. Islet purification by QMS can reduce the detrimental effects of prolonged exposure to toxic enzymes and density gradient solutions and substantially improve islet viability after isolation. PMID:27843954

Prayon process for wet acid purification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Davister, A.; Peeterbroeck, M.

Described is a process developed in Belgium which enables the upgrading technical phosphoric acid to feed and food grades. After laboratory and pilot tests, Prayon developed and patented a solvent extraction process using a mixture of di-isopropyl ether and tributyl phosphate as solvent. The purified phosphoric acid obtained complies with the quality requirements of the market and can be used for metal treatments, in the manufacture of pure phosphates, for cattle feed, by the fermentation industry, for beverages, etc. Among the advantages of this process are its simplicity of operation, its low power consumption, and minimal environmental pollution. Extensive technologicalmore » data are given.« less

Simultaneous micronization and purification of bioactive fraction by supercritical antisolvent technology

PubMed Central

Hiendrawan, Stevanus; Veriansyah, Bambang; Widjojokusumo, Edward; Tjandrawinata, Raymond R.

2017-01-01

Simultaneous micronization and purification of DLBS3233 bioactive fraction, a combination of two Indonesian herbals Lagerstroemia speciosa and Cinnamomum burmannii has been successfully performed via supercritical anti-solvent (SAS) technology. The objective of the present study was to investigate the effectiveness of SAS technology to micronize and reduce coumarin content of DLBS3233. The effects of four SAS process parameters, i.e. pressure, temperature, concentration and solution flow rate on particle formation were investigated. In SAS process, DLBS3233 was dissolved in dimethylformamide (DMF) as the liquid solvent. The solution was then pumped through a nozzle into a chamber simultaneously with supercritical carbon dioxide (SC-CO2) which acts as the anti-solvent, resulting in DLBS3233 precipitation. Physicochemical properties of unprocessed DLBS3233 and SAS-processed DLBS3233 particles were analyzed using scanning electron microscopy (SEM) and high pressure liquid chromatography (HPLC). Total polyphenol content (TPC) was also analyzed. Particles with mean particle size ranging from 0.107±0.028 μm to 0.298±0.138 μm were obtained by varying the process parameters. SAS-processed DLBS3233 particles showed no coumarin content in all experiments studied in this work. Results of TPC analysis revealed no significant change in SAS-processed DLBS3233 particles compared to unprocessed DLBS3233. Nano-sized DLBS3233 particles with no coumarin content have been successfully produced using SAS process. This study demonstrates the ability of SAS for processing herbal medicine in single step process. PMID:28516056

Accelerated solvent extraction (ASE) for purification and extraction of silicone passive samplers used for the monitoring of organic pollutants.

PubMed

Brockmeyer, Berit; Kraus, Uta R; Theobald, Norbert

2015-12-01

Silicone passive samplers have gained an increasing attention as single-phased, practical and robust samplers for monitoring of organic contaminants in the aquatic environment in recent years. However, analytical challenges arise in routine application during the extraction of analytes as silicone oligomers are co-extracted and interfere severely during chemical analyses (e.g. gas chromatographic techniques). In this study, we present a fast, practical pre-cleaning method for silicone passive samplers applying accelerated solvent extraction (ASE) for the removal of silicone oligomers prior to the water deployment (hexane/dichloromethane, 100 °C, 70 min). ASE was also shown to be a very fast (10 min) and efficient extraction method for non-polar contaminants (non-exposed PRC recoveries 66-101 %) sampled by the silicone membrane. For both applications, temperature, extraction time and the solvent used for ASE have been optimized. Purification of the ASE extract was carried out by silica gel and high-pressure liquid size exclusion chromatography (HPLC-SEC). The silicone oligomer content was checked by total reflection X-ray fluorescence spectroscopy (TXRF) in order to confirm the absence of the silicone oligomers prior to analysis of passive sampler extracts. The established method was applied on real silicone samplers from the North- and Baltic Sea and showed no matrix effects during analysis of organic pollutants. Internal laboratory standard recoveries were in the same range for laboratory, transport and exposed samplers (85-126 %).

Purification and Characterization of Haloalkaline, Organic Solvent Stable Xylanase from Newly Isolated Halophilic Bacterium-OKH

PubMed Central

Sanghvi, Gaurav; Jivrajani, Mehul; Patel, Nirav; Jivrajani, Heta; Bhaskara, Govinal Badiger; Patel, Shivani

2014-01-01

A novel, alkali-tolerant halophilic bacterium-OKH with an ability to produce extracellular halophilic, alkali-tolerant, organic solvent stable, and moderately thermostable xylanase was isolated from salt salterns of Mithapur region, Gujarat, India. Identification of the bacterium was done based upon biochemical tests and 16S rRNA sequence. Maximum xylanase production was achieved at pH 9.0 and 37°C temperature in the medium containing 15% NaCl and 1% (w/v) corn cobs. Sugarcane bagasse and wheat straw also induce xylanase production when used as carbon source. The enzyme was active over a range of 0–25% sodium chloride examined in culture broth. The optimum xylanase activity was observed at 5% sodium chloride. Xylanase was purified with 25.81%-fold purification and 17.1% yield. Kinetic properties such as Km and Vmax were 4.2 mg/mL and 0.31 μmol/min/mL, respectively. The enzyme was stable at pH 6.0 and 50°C with 60% activity after 8 hours of incubation. Enzyme activity was enhanced by Ca2+, Mn2+, and Mg2+ but strongly inhibited by heavy metals such as Hg2+, Fe3+, Ni2+, and Zn2+. Xylanase was found to be stable in organic solvents like glutaraldehyde and isopropanol. The purified enzyme hydrolysed lignocellulosic substrates. Xylanase, purified from the halophilic bacterium-OKH, has potential biotechnological applications. PMID:27350996

Simultaneous micronization and purification of bioactive fraction by supercritical antisolvent technology.

PubMed

Hiendrawan, Stevanus; Veriansyah, Bambang; Widjojokusumo, Edward; Tjandrawinata, Raymond R

2017-01-01

Simultaneous micronization and purification of DLBS3233 bioactive fraction, a combination of two Indonesian herbals Lagerstroemia speciosa and Cinnamomum burmannii has been successfully performed via supercritical anti-solvent (SAS) technology. The objective of the present study was to investigate the effectiveness of SAS technology to micronize and reduce coumarin content of DLBS3233. The effects of four SAS process parameters, i.e. pressure, temperature, concentration and solution flow rate on particle formation were investigated. In SAS process, DLBS3233 was dissolved in dimethylformamide (DMF) as the liquid solvent. The solution was then pumped through a nozzle into a chamber simultaneously with supercritical carbon dioxide (SC-CO2) which acts as the anti-solvent, resulting in DLBS3233 precipitation. Physicochemical properties of unprocessed DLBS3233 and SAS-processed DLBS3233 particles were analyzed using scanning electron microscopy (SEM) and high pressure liquid chromatography (HPLC). Total polyphenol content (TPC) was also analyzed. Particles with mean particle size ranging from 0.107±0.028 μ m to 0.298±0.138 μ m were obtained by varying the process parameters. SAS-processed DLBS3233 particles showed no coumarin content in all experiments studied in this work. Results of TPC analysis revealed no significant change in SAS-processed DLBS3233 particles compared to unprocessed DLBS3233. Nano-sized DLBS3233 particles with no coumarin content have been successfully produced using SAS process. This study demonstrates the ability of SAS for processing herbal medicine in single step process.

Development of Solvent Extraction Approach to Recycle Enriched Molybdenum Material

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tkac, Peter; Brown, M. Alex; Sen, Sujat

2016-06-01

Argonne National Laboratory, in cooperation with Oak Ridge National Laboratory and NorthStar Medical Technologies, LLC, is developing a recycling process for a solution containing valuable Mo-100 or Mo-98 enriched material. Previously, Argonne had developed a recycle process using a precipitation technique. However, this process is labor intensive and can lead to production of large volumes of highly corrosive waste. This report discusses an alternative process to recover enriched Mo in the form of ammonium heptamolybdate by using solvent extraction. Small-scale experiments determined the optimal conditions for effective extraction of high Mo concentrations. Methods were developed for removal of ammonium chloridemore » from the molybdenum product of the solvent extraction process. In large-scale experiments, very good purification from potassium and other elements was observed with very high recovery yields (~98%).« less

Integrated Method for Purification and Single-Particle Characterization of Lentiviral Vector Systems by Size Exclusion Chromatography and Tunable Resistive Pulse Sensing.

PubMed

Heider, Susanne; Muzard, Julien; Zaruba, Marianne; Metzner, Christoph

2017-07-01

Elements derived from lentiviral particles such as viral vectors or virus-like particles are commonly used for biotechnological and biomedical applications, for example in mammalian protein expression, gene delivery or therapy, and vaccine development. Preparations of high purity are necessary in most cases, especially for clinical applications. For purification, a wide range of methods are available, from density gradient centrifugation to affinity chromatography. In this study we have employed size exclusion columns specifically designed for the easy purification of extracellular vesicles including exosomes. In addition to viral marker protein and total protein analysis, a well-established single-particle characterization technology, termed tunable resistive pulse sensing, was employed to analyze fractions of highest particle load and purity and characterize the preparations by size and surface charge/electrophoretic mobility. With this study, we propose an integrated platform combining size exclusion chromatography and tunable resistive pulse sensing for monitoring production and purification of viral particles.

Development, upscaling and validation of the purification process for human-cl rhFVIII (Nuwiq®), a new generation recombinant factor VIII produced in a human cell-line.

PubMed

Winge, Stefan; Yderland, Louise; Kannicht, Christoph; Hermans, Pim; Adema, Simon; Schmidt, Torben; Gilljam, Gustav; Linhult, Martin; Tiemeyer, Maya; Belyanskaya, Larisa; Walter, Olaf

2015-11-01

Human-cl rhFVIII (Nuwiq®), a new generation recombinant factor VIII (rFVIII), is the first rFVIII produced in a human cell-line approved by the European Medicines Agency. To describe the development, upscaling and process validation for industrial-scale human-cl rhFVIII purification. The purification process involves one centrifugation, two filtration, five chromatography columns and two dedicated pathogen clearance steps (solvent/detergent treatment and 20 nm nanofiltration). The key purification step uses an affinity resin (VIIISelect) with high specificity for FVIII, removing essentially all host-cell proteins with >80% product recovery. The production-scale multi-step purification process efficiently removes process- and product-related impurities and results in a high-purity rhFVIII product, with an overall yield of ∼50%. Specific activity of the final product was >9000 IU/mg, and the ratio between active FVIII and total FVIII protein present was >0.9. The entire production process is free of animal-derived products. Leaching of potential harmful compounds from chromatography resins and all pathogens tested were below the limit of quantification in the final product. Human-cl rhFVIII can be produced at 500 L bioreactor scale, maintaining high purity and recoveries. The innovative purification process ensures a high-purity and high-quality human-cl rhFVIII product with a high pathogen safety margin. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Simple Microfluidic Device For Studying Chemotaxis In Response To Dual Gradients

PubMed Central

Moussavi-Haramic, S. F.; Pezzi, H. M.; Huttenlocher, A.; Beebe, D. J.

2016-01-01

Chemotaxis is a fundamental biological process where complex chemotactic gradients are integrated and prioritized to guide cell migration toward specific locations. To understand the mechanisms of gradient dependent cell migration, it is important to develop in vitro models that recapitulate key attributes of the chemotactic cues present in vivo. Current in vitro tools for studying cell migration are not amenable to easily study the response of neutrophils to dual gradients. Many of these systems require external pumps and complex setups to establish and maintain the gradients. Here we report a simple yet innovative microfluidic device for studying cell migration in the presence of dual chemotactic gradients through a 3-dimensional substrate. The device is tested and validated by studying the migration of the neutrophil-like cell line PLB-985 to gradients of fMLP. Furthermore, the device is expanded and used with heparinised whole blood, whereupon neutrophils were observed to migrate from whole blood towards gradients of fMLP eliminating the need for any neutrophil purification or capture steps. PMID:25893484

ENHANCED CONCENTRATION AND ISOLATION OF CYCLOSPORA CAYETANENSIS OOCYSTS FROM HUMAN FECAL SAMPLES

EPA Science Inventory

Cyclospora cayetanensis is the causative agent of cyclosporiasis, an emerging infections disease. A new method for the purification of Cycloposra cayetanensis oocysts from fecal matter has been developed, using a modified detachment solution and a Renocal-sucrose gradient sedimen...

CONDUCT AN INVESTIGATION OF CIGUATERA POISON

DTIC Science & Technology

Research on ciguatera toxin resulted in a satisfactory method for extracting the toxin from fish muscle. Partial purification was also accomplished...by precipitation and silicic acid adsorption. The most precise fractionation of ciguatera toxin is accomplished on a silicic acid column developed...WILL LEAD TO HIGHLY PURIFIED SAMPLES OF CIGUATERA TOXIN. Paper chromatography was examined using fourteen different solvent systems, but none proved

Process chemistry of americium-241

DOE Office of Scientific and Technical Information (OSTI.GOV)

Navratil, J.D.

1983-01-01

Americium-241, one of the most useful actinide isotopes, is produced as a by-product of plutonium scrap recovery operations. Rocky Flats has supplied high purity americium oxide to the US Department of Energy's Isotope Pool since 1962. Over the years, the evolving separation and purification processes have included such diverse operations as ion exchange, aqueous precipitation, and both molten-salt and organic-solvent extraction.

Preparative Purification of Polyphenols from Aronia melanocarpa (Chokeberry) with Cellular Antioxidant and Antiproliferative Activity.

PubMed

Gao, Ningxuan; Wang, Yuehua; Jiao, Xinyao; Chou, Shurui; Li, Enhui; Li, Bin

2018-01-10

The aim of this study was the purification process of polyphenols from Aronia melanocarpa (chokeberry), and the purification parameters were optimised by adsorption and desorption tests. By comparing adsorption and desorption ability of polyphenols from chokeberry on six kinds of macroporous resin, XAD-7 resin was selected. Experiments prove that the best purification parameters of static adsorption and desorption were sample pH = 4.0 with 4 h of adsorption; and desorption solvent is 95% ethanol (pH = 7.0) with 2 h of desorption. The best dynamic parameters were 9.3 bed volume (BV) of sample loading amount at a feeding flow rate of 2 BV/h, and washing the column with 5.8 BV of water, followed by subsequent elution with an eluent volume of 5.0 mL at an elution flow rate of 2 BV/h. Next the antioxidant and antiproliferative activity of polyphenols from chokeberry, blueberries, haskap berries was studied on HepG2 human liver cancer cells. The results show that polyphenol from chokeberry has a strong antioxidant effect. Taking into account the content of polyphenols in fruit, polyphenols from chokeberry represent a very valuable natural antioxidant source with antiproliferative products.

Reaction of N,N-Dimethyltryptamine with Dichloromethane Under Common Experimental Conditions.

PubMed

Dunlap, Lee E; Olson, David E

2018-05-31

A large number of clinically used drugs and experimental pharmaceuticals possess the N , N -dimethyltryptamine (DMT) structural core. Previous reports have described the reaction of this motif with dichloromethane (DCM), a common laboratory solvent used during extraction and purification, leading to the formation of an undesired quaternary ammonium salt byproduct. However, the kinetics of this reaction under various conditions have not been thoroughly described. Here, we report a series of experiments designed to simulate the exposure of DMT to DCM that would take place during extraction from plant material, biphasic aqueous work-up, or column chromatography purification. We find that the quaternary ammonium salt byproduct forms at an exceedingly slow rate, only accumulates to a significant extent upon prolonged exposure of DMT to DCM, and is readily extracted into water. Our results suggest that DMT can be exposed to DCM under conditions where contact times are limited (<30 min) with minimal risk of degradation and that this byproduct is not observed following aqueous extraction. However, alternative solvents should be considered when the experimental conditions require longer contact times. Our work has important implications for preparing a wide-range of pharmaceuticals bearing the DMT structural motif in high yields and purities.

High-performance liquid chromatography with fluorescence detection for the rapid analysis of pheophytins and pyropheophytins in virgin olive oil.

PubMed

Li, Xueqi; Woodman, Michael; Wang, Selina C

2015-08-01

Pheophytins and pyropheophytin are degradation products of chlorophyll pigments, and their ratios can be used as a sensitive indicator of stress during the manufacturing and storage of olive oil. They increase over time depending on the storage condition and if the oil is exposed to heat treatments during the refining process. The traditional analysis method includes solvent- and time-consuming steps of solid-phase extraction followed by analysis by high-performance liquid chromatography with ultraviolet detection. We developed an improved dilute/fluorescence method where multi-step sample preparation was replaced by a simple isopropanol dilution before the high-performance liquid chromatography injection. A quaternary solvent gradient method was used to include a fourth strong solvent wash on a quaternary gradient pump, which avoided the need to premix any solvents and greatly reduced the oil residues on the column from previous analysis. This new method not only reduces analysis cost and time but shows reliability, repeatability, and improved sensitivity, especially important for low-level samples. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Modifier mass transfer kinetic effect in the performance of solvent gradient simulated moving bed (SG-SMB) process

NASA Astrophysics Data System (ADS)

Câmara, L. D. T.

2015-09-01

The solvent-gradient simulated moving bed process (SG-SMB) is the new tendency in the performance improvement if compared to the traditional isocratic solvent conditions. In such SG-SMB separation process the modulation of the solvent strength leads to significant increase in the purities and productivity followed by reduction in the solvent consumption. A stepwise modelling approach was utilized in the representation of the interconnected chromatographic columns of the system combined with lumped mass transfer models between the solid and liquid phase. The influence of the solvent modifier was considered applying the Abel model which takes into account the effect of modifier volume fraction over the partition coefficient. The modelling and simulations were carried out and compared to the experimental SG-SMB separation of the amino acids phenylalanine and tryptophan. A lumped mass transfer kinetic model was applied for both the modifier (ethanol) as well as the solutes. The simulation results showed that such simple and global mass transfer models are enough to represent all the mass transfer effect between the solid adsorbent and the liquid phase. The separation performance can be improved reducing the interaction or the mass transfer kinetic effect between the solid adsorbent phase and the modifier. The simulations showed great agreement fitting the experimental data of the amino acids concentrations both at the extract as well as at the raffinate.

Optimal conditions for elution of hepatitis B antigen after absorption onto colloidal silica.

PubMed Central

Pillot, J; Goueffon, S; Keros, R G

1976-01-01

Hepatitis B surface antigen (HBSAg) adsorbed from sera onto colloidal silica could be completely eluted through the use of 0.25% sodium deoxycholate in 0.01 M borax, pH 9.3, at 56 degrees C. The HBSAg recovered in the eluate represented 100% of that present in the original serum, and it was contaminated by only trace amounts of serum proteins (in decreasing amounts: beta-lipoprotein, immunoglobulin G, albumin). This preliminary step greatly facilitates purification of large amounts of HBSAg and provides small volumes of highly concentrated material for subsequent purification by density gradient centrifugation. PMID:9423

Hormone purification by isoelectric focusing in space

NASA Technical Reports Server (NTRS)

Bier, M.

1982-01-01

The performance of a ground-prototype of an apparatus for recycling isoelectric focusing was evaluated in an effort to provide technology for large scale purification of peptide hormones, proteins, and other biologicals. Special emphasis was given to the effects of gravity on the function of the apparatus and to the determination of potential advantages deriveable from its use in a microgravity environment. A theoretical model of isoelectric focusing sing chemically defined buffer systems for the establishment of the pH gradients was developed. The model was transformed to a form suitable for computer simulations and was used extensively for the design of experimental buffers.

Purification, crystallization and preliminary X-ray diffraction analysis of royal palm tree (Roystonea regia) peroxidase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Watanabe, Leandra; Nascimento, Alessandro S.; Zamorano, Laura S.

2007-09-01

The purification, crystallization, X-ray diffraction data acquisition and molecular-replacement results of royal palm tree (R. regia) peroxidase are described. Royal palm tree peroxidase (RPTP), which was isolated from Roystonea regia leaves, has an unusually high stability that makes it a promising candidate for diverse applications in industry and analytical chemistry [Caramyshev et al. (2005 ▶), Biomacromolecules, 6, 1360–1366]. Here, the purification and crystallization of this plant peroxidase and its X-ray diffraction data collection are described. RPTP crystals were obtained by the hanging-drop vapour-diffusion method and diffraction data were collected to a resolution of 2.8 Å. The crystals belong to themore » trigonal space group P3{sub 1}21, with unit-cell parameters a = b = 116.83, c = 92.24 Å, and contain one protein molecule per asymmetric unit. The V{sub M} value and solvent content are 4.07 Å{sup 3} Da{sup −1} and 69.8%, respectively.« less

Purification of Houttuynia cordata Thunb. Essential Oil Using Macroporous Resin Followed by Microemulsion Encapsulation to Improve Its Safety and Antiviral Activity.

PubMed

Pang, Jianmei; Dong, Wujun; Li, Yuhuan; Xia, Xuejun; Liu, Zhihua; Hao, Huazhen; Jiang, Lingmin; Liu, Yuling

2017-02-15

Essential oil extracted from Houttuynia cordata Thunb. ( H. cordata ) is widely used in traditional Chinese medicine due to its excellent biological activities. However, impurities and deficient preparations of the essential oil limit its safety and effectiveness. Herein, we proposed a strategy to prepare H. cordata essential oil (HEO) safely and effectively by combining the solvent extraction and the macroporous resin purification flexibly, and then encapsulating it using microemulsion. The extraction and purification process were optimized by orthogonal experimental design and adsorption-desorption tests, respectively. The average houttuynin content in pure HEO was then validated at 44.3% ± 2.01%, which presented a great potential for industrial application. Subsequently, pure HEO-loaded microemulsion was prepared by high-pressure homogenization and was then fully characterized. Results showed that the pure HEO-loaded microemulsion was successfully prepared with an average particle size of 179.1 nm and a high encapsulation rate of 94.7%. Furthermore, safety evaluation tests and in vitro antiviral testing indicated that the safety and activity of HEO were significantly improved after purification using D101 resin and were further improved by microemulsion encapsulation. These results demonstrated that the purification of HEO by macroporous resin followed by microemulsion encapsulation would be a promising approach for industrial application of HEO for the antiviral therapies.

Preparative purification of polyethylene glycol derivatives with polystyrene-divinylbenzene beads as chromatographic packing.

PubMed

Yu, Pengzhan; Li, Xingqi; Li, Xiunan; Lu, Xiuling; Ma, Guanghui; Su, Zhiguo

2007-10-15

A clear and powerful chromatographic approach to purify polyethylene glycol derivatives at a preparative scale was reported, which was based on the polystyrene-divinylbenzene beads with ethanol/water as eluants. The validity of this method was verified with the reaction mixture of mPEG-Glu and mPEG propionaldehyde diethylacetal (ALD-PEG) as the model. The target products were one-step achieved with the purity of >99% on the polymer resins column at gram scale. The method developed was free from such disadvantages as utility of toxic solvent and narrow application scope, which was combined with conventional approaches. The method developed provided an appealing and attractive alternative methods for purification of PEG derivatives at a preparative scale.

Data in support of three phase partitioning of zingibain, a milk-clotting enzyme from Zingiber officinale Roscoe rhizomes

PubMed Central

Gagaoua, Mohammed; Hafid, Kahina; Hoggas, Naouel

2016-01-01

This paper describes data related to a research article titled “Three Phase Partitioning of zingibain, a milk-clotting enzyme from Zingiber officinale Roscoe rhizomes” (Gagaoua et al., 2015) [1]. Zingibain (EC 3.4.22.67), is a coagulant cysteine protease and a meat tenderizer agent that have been reported to produce satisfactory final products in dairy and meat technology, respectively. Zingibains were exclusively purified using chromatographic techniques with very low yield purification. This paper includes data of the effect of temperature, usual salts and organic solvents on the efficiency of the three phase partitioning (TPP) system. Also it includes data of the kinetic activity characterization of the purified zingibain using TPP purification approach. PMID:26909379

Dynamics of water in sulfonated poly(phenylene) membranes

NASA Astrophysics Data System (ADS)

Osti, Naresh; Etampawala, Thusitha; Shrestha, Umesh; Perahia, Dvora; Cornelius, Christopher

2011-03-01

The dynamics of water in networks formed by highly rigid ionic polymers, sulfonated poly(phenylene) as observed by quasi elastic neutron scattering (QENS) is presented. These rigid ionic polymers have potential as effective ion exchange membranes with impact on a large number of applications from water purification to clean energy, where its rigidity distinguishes it from other ionic polymers. Its transport characteristics are affected by its rigidness as well as by direct interactions with the solvent. Our QENS studies as a function of sulfonation levels, temperature and solvent content have shown that on the time scale of the measurement, the polymers are rigid. While macroscopically all samples swell, and transport water, the water molecules appear locally rather confined. Water however remind non-frozen to subzero temperatures. The results will be discussed in view of theoretical models including continues diffusion and hopping of solvent molecules.

PURIFICATION OF ENTEROCYTOZOON BIENEUSI SPORES FROM STOOL SPECIMENS BY GRADIENT AND CELL SORTING TECHNIQUES. (R828042)

EPA Science Inventory

The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

Method and apparatus for converting and removing organosulfur and other oxidizable compounds from distillate fuels, and compositions obtained thereby

DOEpatents

D'Alessandro, Robert N.; Tarabocchia, John; Jones, Jerald Andrew; Bonde, Steven E.; Leininger, Stefan

2010-10-26

The present disclosure is directed to a multi-stage system and a process utilizing said system with the design of reducing the sulfur-content in a liquid comprising hydrocarbons and organosulfur compounds. The process comprising at least one of the following states: (1) an oxidation stage; (2) an extraction state; (3) a raffinate washing stage; (4) a raffinate polishing stage; (5) a solvent recovery stage; (6) a solvent purification stage; and (7) a hydrocarbon recovery stage. The process for removing sulfur-containing hydrocarbons from gas oil, which comprises oxidizing gas oil comprising hydrocarbons and organosulfur compounds to obtain a product gas oil.

Purification of influenza deoxyribonucleic acid-based vaccine using agmatine monolith.

PubMed

Bicho, D; Caramelo-Nunes, C; Sousa, A; Sousa, F; Queiroz, J A; Tomaz, C T

2016-02-15

Lately, researchers have made several efforts to improve vaccine production to fight highly contagious respiratory diseases like influenza. One of the most promising options for reducing the impact of this virus is DNA vaccination. However, a large quantity of highly pure plasmid DNA (pDNA) is necessary to attain this goal. The present work describes the production and purification of the plasmid NTC7482-41H-VA2HA expressing influenza virus hemagglutinin using an agmatine monolith. This ligand was chosen to purify supercoiled (sc) pDNA from complex lysates because of its versatile multimodal character. Its natural intervention in several biological systems together with its similarity with the highly studied arginine ligand allowed the development of a simpler and more specific purification process. Agmatine works under two strategies: descending ammonium sulfate gradient and ascending sodium chloride gradient. Furthermore, pH manipulation revealed an important role in pDNA isoforms selectivity. Dynamic binding capacity (DBC) experiments were performed varying different parameters and showed an increase with pDNA concentration, while high flow rate and high pH had the opposite effect. Sc pDNA was purified with high yield and was efficient with respect to cell transfection and cell viability. This monolith showed to be appropriate to purify the plasmid NTC7482-41H-VA2HA, providing a valuable tool for pDNA influenza vaccines preparation. Copyright © 2016. Published by Elsevier B.V.

A novel multimodal chromatography based single step purification process for efficient manufacturing of an E. coli based biotherapeutic protein product.

PubMed

Bhambure, Rahul; Gupta, Darpan; Rathore, Anurag S

2013-11-01

Methionine oxidized, reduced and fMet forms of a native recombinant protein product are often the critical product variants which are associated with proteins expressed as bacterial inclusion bodies in E. coli. Such product variants differ from native protein in their structural and functional aspects, and may lead to loss of biological activity and immunogenic response in patients. This investigation focuses on evaluation of multimodal chromatography for selective removal of these product variants using recombinant human granulocyte colony stimulating factor (GCSF) as the model protein. Unique selectivity in separation of closely related product variants was obtained using combined pH and salt based elution gradients in hydrophobic charge induction chromatography. Simultaneous removal of process related impurities was also achieved in flow-through leading to single step purification process for the GCSF. Results indicate that the product recovery of up to 90.0% can be obtained with purity levels of greater than 99.0%. Binding the target protein at pH

Preparative isolation and purification of seven isoflavones from Belamcanda chinensis.

PubMed

Lee, Yeon Sil; Kim, Seon Ha; Kim, Jin Kyu; Lee, Sanghyun; Jung, Sang Hoon; Lim, Soon Sung

2011-01-01

Isoflavonoids from Belamcanda chinensis are known to have a number of physiological benefits including anti-inflammatory, anti-angiogenic and anti-mutagenic properties. However, there have been no reports on the effective isolation and purification of isoflavonoids from B. chinensis. To develop an efficient method for the preparative isolation and purification of isoflavones from B. chinensis by high-speed counter-current chromatography (HSCCC). A two-step HSCCC isolation method was developed using solvent system of n-hexane-ethyl acetate-2-propanol-methanol-water (5:6:2:3.5:6, v/v) and of ethyl acetate-methanol-water (10:2:9, v/v). FLASH purification system (45% methanol, isocratic) was also used for further purification. The purities and chemical structures of the isolated compounds were determined by high-performance liquid chromatography-photodiode array detection (HPLC-PDA), electrospray ionisation-mass spectrometry (ESI-MS), ¹H- and ¹³C-nuclear magnetic resonance spectrometry (NMR) and nuclear overhauser enhancement (NOE). HSCCC was successfully used for the preparative separation and purification of seven isoflavones, including tectoridin (145.4 mg, 97.5%), iridin (77.9 mg, 94.0%), irilin D (42.0 mg, 92.0%), tectorigenin (294.1 mg, 98.6%), iristectorigenin A (86.8 mg, 93.4%), irigenin (141.8 mg, 95.8%) and irisflorentin (73.4 mg, 94.7%) from the rhizomes of B. chinensis. Two isoflavone glycosides and five isoflavone derivatives were successfully isolated and purified from the crude methanol extract of dried rhizomes of the B. chinensis by HSCCC. Copyright © 2011 John Wiley & Sons, Ltd.

Protein Purification and Its Application to Crystallization

DTIC Science & Technology

1988-08-30

separation can be altered by adding various organic modifiers to the separation solvents. The Laboratory for the Structure of Matter is involved in a...SQUID ORGANOPHOSPHOROUS ACID ANHYDRASE 2.2.1 Introduction Cephalopod optic ganglion and hepatopancrease contain an organic phosphorous acid anhydrase...number of other sources including erythrocytes, and various organs (21,22). AChE hydrolyzes the ester linkage in acetylcholine releasing acetate and

Purification of caprolactam from recycled nylon

DOEpatents

Moens, Luc

1999-01-01

A method of removing 1,11-diamino-6-undecanone from the pyrolysis product of nylon comprising: a) pyrolyzing nylon-6 to form a pyrolyzate containing a caprolactam mixture; b) dissolving the caprolactam mixture in a solvent to form a solution; c) passing carbon dioxide gas through the solution to form a precipitate; d) removing the precipitate from the solution; and e) recovering the purified caprolactam from the solution.

Nuclear Weapons. National Nuclear Security Administration’s Plans for Its Uranium Processing Facility Should Better Reflect Funding Estimates and Technology Readiness

DTIC Science & Technology

2010-11-01

metal. Recovery extraction centrifugal contactors A process that uses solvent to extract uranium for purposes of purification. Agile machining A...extraction centrifugal contactors 5 6 Yes 6 No Agile machining 5 5 No 6 No Chip management 5 6 Yes 6 No Special casting 3 6 Yes 6 No Source: GAO

A DIETHER ANALOG OF PHOSPHATIDYL GLYCEROPHOSPHATE IN HALOBACTERIUM CUTIRUBRUM,

DTIC Science & Technology

The major phosphatide in the extremely halophilic bacterium, Halobacterium cutirubrum, was isolated by a combination of solvent fractionation...precipitation through the barium salt, and final purification as the sodium salt. Analytical and degradative data showed the phosphatide to be a...phosphatidyl glycerophosphate with two long-chain ether groups instead of fatty acid ester groups. Both long-chain groups were found to be identical and were

Sulfur tolerant hydrophobic ionic liquid solvent

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luebke, David; Nulwala, Hunaid; Kail, Brian

Exemplary embodiments relate to methods for removal of CO.sub.2 and other acid gases from gaseous fuels prior to combustion. Exemplary methods may be used for CO.sub.2 capture, H.sub.2 purification and natural gas sweetening. Exemplary methods use at least one ionic liquid that has excellent CO.sub.2 solubility, selectivity over hydrogen, low viscosity, and resistance to H.sub.2S.

Solvent-programmed microchip open-channel electrochromatography.

PubMed

Kutter, J P; Jacobson, S C; Matsubara, N; Ramsey, J M

1998-08-01

Open-channel electrochromatography in combination with solvent programming is demonstrated using a microchip device. Channel walls were coated with octadecylsilanes at ambient temperatures, yielding stationary phases for chromatographic separations of neutral dyes. The electroosmotic flow after coating was sufficient to ensure transport of all species and on-chip mixing of isocratic and gradient elution conditions with acetonitrile-buffer mixtures. Chips having different channel depths between 10.2 and 2.9 μm were evaluated for performance, and van Deemter plots were established. Channel depths of about 5 μm were found to be a good compromise between efficiency and ease of operation. Isocratic and gradient elution conditions were easily established and manipulated by computer-controlled application of voltages to the terminals of the microchip. Linear gradients with different slopes, start times, duration times, and start percentages of organic modifier proved to be powerful tools to tune selectivity and analysis time for the separation of a test mixture. Even very steep gradients still produced excellent efficiencies. Together with fast reconditioning times, complete runs could be finished in under 60 s.

Chemotaxis of Molecular Dyes in Polymer Gradients in Solution.

PubMed

Guha, Rajarshi; Mohajerani, Farzad; Collins, Matthew; Ghosh, Subhadip; Sen, Ayusman; Velegol, Darrell

2017-11-08

Chemotaxis provides a mechanism for directing the transport of molecules along chemical gradients. Here, we show the chemotactic migration of dye molecules in response to the gradients of several different neutral polymers. The magnitude of chemotactic response depends on the structure of the monomer, polymer molecular weight and concentration, and the nature of the solvent. The mechanism involves cross-diffusion up the polymer gradient, driven by favorable dye-polymer interaction. Modeling allows us to quantitatively evaluate the strength of the interaction and the effect of the various parameters that govern chemotaxis.

Isolation, production, purification and characterization of an organic-solvent-thermostable alkalophilic cellulase from Bacillus vallismortis RG-07.

PubMed

Gaur, Rajeeva; Tiwari, Soni

2015-03-19

The rising concerns about the scarcity of fossil fuels, the emission of green house gasses and air pollution by incomplete combustion of fossil fuel have also resulted in an increasing focus on the use of cellulases to perform enzymatic hydrolysis of the lignocellulosic materials for the generation of bioethanol. The aim of this study was to isolate a potential thermo-solvent tolerant cellulase producing bacterium from natural resources, and then applied for purification and characterization. The purified enzyme was to be accessible for the bioethanol production as well as industrial exploitation (discuss in our next study). It is the first instance when thermo-solvent tolerant cellulase producing bacterium was isolated from soil sample. The culture was identified as Bacillus vallismortis RG-07 by 16S rDNA sequence analysis. Bacillus vallismortis RG-07 reported maximum cellulase production from sugarcane baggase (4105 U ml(-1)) used as agro-waste carbon source. The cellulase enzyme produced by the Bacillus sp. was purified by (NH4)2SO4 precipitation, ion exchange and gel filtration chromatography, with overall recovery of 28.8%. The molecular weight of purified cellulase was 80 kDa as revealed by SDS-PAGE and activity gel analysis. The optimum temperature and pH for enzyme activity was determined as 65°C and 7.0 and it retained 95 and 75% of activity even at 95°C, and 9.0 respectively. The enzyme activity was enhanced in the presence of organic solvents (30%) n-dodecane, iso-octane, n-decane, xylene, toluene, n-haxane, n-butanol, and cyclohexane, after prolonged incubation (7 days). The enzyme activity was also stimulated by Ca(2+), mercaptoethanol, Tween-60, and Sodium hypochloride whereas strongly inhibited by Hg. Kinetic analysis of purified enzyme showed the Km and Vmax to be 1.923 mg ml(-1) and 769.230 μg ml(-1) min(-1), respectively. The unique property of solvent-thermostable-alkalophilic, nature proves the potential candidature of this isolate for current mainstream biomass conversion into fuel and other industrial process.

Regenerated silica gel as stationary phase on vacuum column chromatography to purify temulawak’s extracts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cahyono, Bambang; Maduwu, Ratna Dewi; Widayat,

Commercial silica gel only used once by many researchers and affected high cost for purification process, also less support the green chemistry program. This research focused in regeneration silica gel that used purification of temulawak’s extracts (Curcuma xanthorrhiza Roxb) by vacuum column chromatography. Sample extracts (contains 10.1195±0.5971% of curcuminoids) was purified by vacuum column chromatography (pressure: 45 kPa, column: 100mm on length and 16mm on diameter). Ethanol 96% and acetone were compared as eluent. The amount of solvent and yield of curcuminoids used as indicator purification. The silica gel was regenerated with heating in 600°C for 8 hours The silica gelsmore » were analyzed by IR spectroscopy and X-ray diffraction. Furthermore, regenerated silica gel was used as the stationary phase in vacuum column chromatography under the same conditions with the previous purification. All the purification experiments were performed in three repetitions. Based on regression equation, y=0.132x+0.0011 (r{sup 2}=0.9997) the yield of curcuminoids on purified products using ethanol as the eluent was improved 4.26% (to 14.3724±0.5749%) and by acetone was improved 3,03% (to 13.1450 ±0.6318%). The IR spectrum of both silica gel showed the same vibration profile and also there were three crystallinity peaks missing on its X-ray diffraction. Regenerated silica gel has the same performance with new silica gel in purification of temulawak’s extract: by ethanol has increased 4.08% (14.1947±0.7415%) and 2.93% (13.0447±0.4822) by acetone. In addition, all purification products showed similar TLC profiles. Purification using regenerated silica gel as the adsorbent on vacuum column chromatography has exactly same potential with the new silica gel.« less

Solvent extraction of gold using ionic liquid based process

NASA Astrophysics Data System (ADS)

Makertihartha, I. G. B. N.; Zunita, Megawati; Rizki, Z.; Dharmawijaya, P. T.

2017-01-01

In decades, many research and mineral processing industries are using solvent extraction technology for metal ions separation. Solvent extraction technique has been used for the purification of precious metals such as Au and Pd, and base metals such as Cu, Zn and Cd. This process uses organic compounds as solvent. Organic solvents have some undesired properties i.e. toxic, volatile, excessive used, flammable, difficult to recycle, low reusability, low Au recovery, together with the problems related to the disposal of spent extractants and diluents, even the costs associated with these processes are relatively expensive. Therefore, a lot of research have boosted into the development of safe and environmentally friendly process for Au separation. Ionic liquids (ILs) are the potential alternative for gold extraction because they possess several desirable properties, such as a the ability to expanse temperature process up to 300°C, good solvent properties for a wide range of metal ions, high selectivity, low vapor pressures, stability up to 200°C, easy preparation, environmentally friendly (commonly called as "green solvent"), and relatively low cost. This review paper is focused in investigate of some ILs that have the potentials as solvent in extraction of Au from mineral/metal alloy at various conditions (pH, temperature, and pressure). Performances of ILs extraction of Au are studied in depth, i.e. structural relationship of ILs with capability to separate Au from metal ions aggregate. Optimal extraction conditon in order to gain high percent of Au in mineral processing is also investigated.

Size and DNA distributions of electrophoretically separated cultured human kidney cells

NASA Technical Reports Server (NTRS)

Kunze, M. E.; Plank, L. D.; Todd, P. W.

1985-01-01

Electrophoretic purification of purifying cultured cells according to function presumes that the size of cycle phase of a cell is not an overriding determinant of its electrophoretic velocity in an electrophoretic separator. The size distributions and DNA distributions of fractions of cells purified by density gradient electrophoresis were determined. No systematic dependence of electrophoretic migration upward in a density gradient column upon either size or DNA content were found. It was found that human leukemia cell populations, which are more uniform function and found in all phases of the cell cycle during exponential growth, separated on a vertical sensity gradient electrophoresis column according to their size, which is shown to be strictly cell cycle dependent.

Evaluating Water Quality in the Lovros River (Greece), Using Biotic Indices based on Invertebrate Communities.

ERIC Educational Resources Information Center

Koussouris, Theodore; And Others

1990-01-01

Presented is a survey of a river including physiochemical measurements and river fauna observations. It is shown that the self-purification gradient of river water quality and the possible ecological disturbances due to pollutants entering the river create an unpredictable pattern of recovery. (CW)

Purification of 1.9-nm-diameter semiconducting single-wall carbon nanotubes by temperature-controlled gel-column chromatography and its application to thin-film transistor devices

NASA Astrophysics Data System (ADS)

Thendie, Boanerges; Omachi, Haruka; Hirotani, Jun; Ohno, Yutaka; Miyata, Yasumitsu; Shinohara, Hisanori

2017-06-01

Large-diameter semiconductor single-wall carbon nanotubes (s-SWCNTs) have superior mobility and conductivity to small-diameter s-SWCNTs. However, the purification of s-SWCNTs with diameters larger than 1.6 nm by gel filtration has been difficult owing to the low selectivity of the conventional purification method in these large-diameter regions. We report a combination of temperature-controlled gel filtration and the gradient elution technique that we developed to enrich a high-purity s-SWCNT with a diameter as large as 1.9 nm. The thin-film transistor (TFT) device using the 1.9-nm-diameter SWCNT shows an average channel mobility of 23.7 cm2 V-1 s-1, which is much higher than those of conventional SWCNT-TFTs with smaller-diameters of 1.5 and 1.4 nm.

Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

PubMed

Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

2016-10-01

Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a cost-effective, rapid, and reliable avenue for the purification of recombinant proteins in heterologous hosts. Copyright © 2016 Elsevier Inc. All rights reserved.

Preparative isolation and purification of hainanmurpanin, meranzin, and phebalosin from leaves of Murraya exotica L. using supercritical fluid extraction combined with consecutive high-speed countercurrent chromatography.

PubMed

Yan, Rongwei; Shen, Jie; Liu, Xiaojing; Zou, Yong; Xu, Xinjun

2018-05-01

The objective of this study was to develop a consecutive preparation method for the isolation and purification of hainanmurpanin, meranzin, and phebalosin from leaves of Murraya exotica L. The process involved supercritical fluid extraction with CO 2 , solvent extraction, and two-step high-speed countercurrent chromatography. Pressure, temperature, and the volume of entrainer were optimized as 27 MPa, 52°C, and 60 mL by response surface methodology in supercritical fluid extraction with CO 2 , and the yield of the crude extracts was 7.91 g from 100 g of leaves. Subsequently, 80% methanol/water was used to extract and condense the three compounds from the crude extracts, and 4.23 g of methanol/water extracts was obtained. Then, a two-step high-speed countercurrent chromatography procedure was developed for the isolation of the three target compounds from methanol/water extracts, including conventional high-speed countercurrent chromatography for further enrichment and consecutive high-speed countercurrent chromatography for purification. The yield of concentrates from high-speed countercurrent chromatography was 2.50 g from 4.23 g of methanol/water extracts. Finally, the consecutive high-speed countercurrent chromatography produced 103.2 mg of hainanmurpanin, 244.7 mg of meranzin, and 255.4 mg of phebalosin with purities up to 97.66, 99.36, and 98.64%, respectively, from 900 mg of high-speed countercurrent chromatography concentrates in one run of three consecutive sample loadings without exchanging a solvent system. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Isolation and purification of orientin and vitexin from Trollius chinensis Bunge by high-speed counter-current chromatography.

PubMed

Yu, Xiao-Xue; Huang, Jie-Yun; Xu, Dan; Xie, Zhi-Yong; Xie, Zhi-Sheng; Xu, Xin-Jun

2014-01-01

Orientin and vitexin are the two main bioactive compounds in Trollius chinensis Bunge. In this study, a rapid method was established for the isolation and purification of orientin and vitexin from T. chinensis Bunge using high-speed counter-current chromatography in one step, with a solvent system of ethyl acetate-ethanol-water (4:1:5, v/v/v). A total of 9.8 mg orientin and 2.1 mg vitexin were obtained from 100 mg of the ethyl acetate extract, with purities of 99.2% and 96.0%, respectively. Their structures were identified by UV, MS and NMR. The method was efficient and convenient, which could be used for the preparative separation of orientin and vitexin from T. chinensis Bunge.

Separation and purification of astaxanthin from Phaffia rhodozyma by preparative high-speed counter-current chromatography.

PubMed

Du, Xiping; Dong, Congcong; Wang, Kai; Jiang, Zedong; Chen, Yanhong; Yang, Yuanfan; Chen, Feng; Ni, Hui

2016-09-01

An effective high-speed counter-current chromatography (HSCCC) method was established for the preparative isolation and purification of astaxanthin from Phaffia rhodozyma. With a two-phase solvent system composed of n-hexane-acetone-ethanol-water (1:1:1:1, v/v/v/v), 100mg crude extract of P. rhodozyma was separated to yield 20.6mg of astaxanthin at 92.0% purity. By further one step silica gel column chromatography, the purity reached 99.0%. The chemical structure of astaxanthin was confirmed by thin layer chromatography (TLC), UV spectroscopy scanning, high performance liquid chromatography with a ZORBAX SB-C18 column and a Waters Nova-pak C18 column, and ESI/MS/MS. Copyright © 2016 Elsevier B.V. All rights reserved.

Enrichment of undifferentiated type a spermatogonia from goat testis using discontinuous percoll density gradient and differential plating.

PubMed

Heidari, Banafsheh; Gifani, Minoo; Shirazi, Abolfazl; Zarnani, Amir-Hassan; Baradaran, Behzad; Naderi, Mohammad Mehdi; Behzadi, Bahareh; Borjian-Boroujeni, Sara; Sarvari, Ali; Lakpour, Niknam; Akhondi, Mohammad Mehdi

2014-04-01

The well documented source for adult multipotent stem cells is Spermatogonial Stem Cells (SSCs). They are the foundation of spermatogenesis in the testis throughout adult life by balancing self-renewal and differentiation. The aim of this study was to assess the effect of percoll density gradient and differential plating on enrichment of undifferentiated type A spermatogonia in dissociated cellular suspension of goat testes. Additionally, we evaluated the separated fractions of the gradients in percoll and samples in differential plating at different times for cell number, viability and purification rate of goat SSCs in culture. Testicular cells were successfully isolated from one month old goat testis using two-step enzymatic digestion and followed by two purification protocols, differential plating with different times of culture (3, 4, 5, and 6 hr) and discontinuous percoll density with different gradients (20, 28, 30, and 32%). The difference of percentage of undifferentiated SSCs (PGP9.5 positive) in each method was compared using ANOVA and comparison between the highest percentage of corresponding value between two methods was carried out by t-test using Sigma Stat (ver. 3.5). The highest PGP9.5 (94.6±0.4) and the lowest c-Kit positive (25.1±0.7) in Percoll method was significantly (p ≤ 0.001) achieved in 32% percoll gradient. While the corresponding rates in differential plating method for the highest PGP9.5 positive cells (81.3±1.1) and lowest c-Kit (17.1±1.4) was achieved after 5 hr culturing (p < 0.001). The enrichment of undifferentiated type A spermatogonia using Percoll was more efficient than differential plating method (p < 0.001). Percoll density gradient and differential plating were efficient and fast methods for enrichment of type A spermatogonial stem cells from goat testes.

Enrichment of Undifferentiated Type A Spermatogonia from Goat Testis Using Discontinuous Percoll Density Gradient and Differential Plating

PubMed Central

Heidari, Banafsheh; Gifani, Minoo; Shirazi, Abolfazl; Zarnani, Amir-Hassan; Baradaran, Behzad; Naderi, Mohammad Mehdi; Behzadi, Bahareh; Borjian-Boroujeni, Sara; Sarvari, Ali; Lakpour, Niknam; Akhondi, Mohammad Mehdi

2014-01-01

Background The well documented source for adult multipotent stem cells is Spermatogonial Stem Cells (SSCs). They are the foundation of spermatogenesis in the testis throughout adult life by balancing self-renewal and differentiation. The aim of this study was to assess the effect of percoll density gradient and differential plating on enrichment of undifferentiated type A spermatogonia in dissociated cellular suspension of goat testes. Additionally, we evaluated the separated fractions of the gradients in percoll and samples in differential plating at different times for cell number, viability and purification rate of goat SSCs in culture. Methods Testicular cells were successfully isolated from one month old goat testis using two-step enzymatic digestion and followed by two purification protocols, differential plating with different times of culture (3, 4, 5, and 6 hr) and discontinuous percoll density with different gradients (20, 28, 30, and 32%). The difference of percentage of undifferentiated SSCs (PGP9.5 positive) in each method was compared using ANOVA and comparison between the highest percentage of corresponding value between two methods was carried out by t-test using Sigma Stat (ver. 3.5). Results The highest PGP9.5 (94.6±0.4) and the lowest c-Kit positive (25.1±0.7) in Percoll method was significantly (p ≤ 0.001) achieved in 32% percoll gradient. While the corresponding rates in differential plating method for the highest PGP9.5 positive cells (81.3±1.1) and lowest c-Kit (17.1±1.4) was achieved after 5 hr culturing (p < 0.001). The enrichment of undifferentiated type A spermatogonia using Percoll was more efficient than differential plating method (p < 0.001). Conclusion Percoll density gradient and differential plating were efficient and fast methods for enrichment of type A spermatogonial stem cells from goat testes. PMID:24834311

Spectroscopic Studies of Nuclear Submarine Atmospheres. I. Desorption and Analysis of Contaminants from Hopcalite

DTIC Science & Technology

A semi-microanalytical method is described for desorbing contaminants from Hopcalite , an oxide catalyst used in the purification of submarine...contaminants are desorbed from the Hopcalite by passing pressurized steam through a column of the catalyst. The various compounds are eluted from the column...appropriate solvent. Infrared spectral analyses of contaminants desorbed from Hopcalite following its use as a catalyst for the oxidation of pure

Purification of caprolactam from recycled nylon

DOEpatents

Moens, L.

1999-07-06

A method is disclosed of removing 1,11-diamino-6-undecanone from the pyrolysis product of nylon comprising: (a) pyrolyzing nylon-6 to form a pyrolyzate containing a caprolactam mixture; (b) dissolving the caprolactam mixture in a solvent to form a solution; (c) passing carbon dioxide gas through the solution to form a precipitate; (d) removing the precipitate from the solution; and (e) recovering the purified caprolactam from the solution. 3 figs.

Purification of optical imaging ligand-Cybesin by high-speed counter-current chromatography

PubMed Central

Ma, Zhiyong; Ma, Ying; Sun, Xilin; Ye, Yunpeng; Shen, Baozhong; Chen, Xiaoyuan; Ito, Yoichiro

2010-01-01

Fluorescent Cybesin (Cypate-Bombesin Peptide Analogue Conjugate) was synthesized from Indocyanine Green (ICG) and the bombesin receptor ligand as a contrast agent for detecting pancreas tumors. However, the LC–MS analysis indicated that the target compound was only a minor component in the reaction mixture. Since preparative HPLC can hardly separate such a small amount of the target compound directly from the original crude reaction mixture without a considerable adsorptive loss onto the solid support, high-speed counter-current chromatography (HSCCC) was used for purification since the method uses no solid support and promises high sample recovery. A suitable two-phase solvent system composed of hexane/ethyl acetate/methanol/methyl t.-butyl ether/acetonitrile/water) at a volume ratio of 1:1:1:4:4:7 was selected based on the partition coefficient of Cybesin (K ≈ 0.9) determined by LC–MS. The separation was performed in two steps using the same solvent system with lower aqueous mobile phase. From 400 mg of the crude reaction mixture the first separation yielded 7.7 mg of fractions containing the target compound at 12.8% purity, and in the second run 1 mg of Cybesin was obtained at purity of 94.0% with a sample recovery rate of over 95% based on the LC–MS Analysis. PMID:20933483

Accurate prediction of retention in hydrophilic interaction chromatography by back calculation of high pressure liquid chromatography gradient profiles.

PubMed

Wang, Nu; Boswell, Paul G

2017-10-20

Gradient retention times are difficult to project from the underlying retention factor (k) vs. solvent composition (φ) relationships. A major reason for this difficulty is that gradients produced by HPLC pumps are imperfect - gradient delay, gradient dispersion, and solvent mis-proportioning are all difficult to account for in calculations. However, we recently showed that a gradient "back-calculation" methodology can measure these imperfections and take them into account. In RPLC, when the back-calculation methodology was used, error in projected gradient retention times is as low as could be expected based on repeatability in the k vs. φ relationships. HILIC, however, presents a new challenge: the selectivity of HILIC columns drift strongly over time. Retention is repeatable in short time, but selectivity frequently drifts over the course of weeks. In this study, we set out to understand if the issue of selectivity drift can be avoid by doing our experiments quickly, and if there any other factors that make it difficult to predict gradient retention times from isocratic k vs. φ relationships when gradient imperfections are taken into account with the back-calculation methodology. While in past reports, the accuracy of retention projections was >5%, the back-calculation methodology brought our error down to ∼1%. This result was 6-43 times more accurate than projections made using ideal gradients and 3-5 times more accurate than the same retention projections made using offset gradients (i.e., gradients that only took gradient delay into account). Still, the error remained higher in our HILIC projections than in RPLC. Based on the shape of the back-calculated gradients, we suspect the higher error is a result of prominent gradient distortion caused by strong, preferential water uptake from the mobile phase into the stationary phase during the gradient - a factor our model did not properly take into account. It appears that, at least with the stationary phase we used, column distortion is an important factor to take into account in retention projection in HILIC that is not usually important in RPLC. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of liquid chromatography mobile phases and buffer salts on phosphorus inductively coupled plasma atomic emission and mass spectrometries utilizing ultrasonic nebulization and membrane desolvation.

PubMed

Carr, John E; Kwok, Kaho; Webster, Gregory K; Carnahan, Jon W

2006-01-23

Atomic spectrometry, specifically inductively coupled plasma atomic emission spectrometry (ICP-AES) and mass spectrometry (ICP-MS) show promise for heteroatom-based detection of pharmaceutical compounds. The combination of ultrasonic nebulization (USN) with membrane desolvation (MD) greatly enhances detection limits with these approaches. Because pharmaceutical analyses often incorporate liquid chromatography, the study herein was performed to examine the effects of solvent composition on the analytical behaviors of these approaches. The target analyte was phosphorus, introduced as phosphomycin. AES response was examined at the 253.7 nm atom line and mass 31 ions were monitored for the MS experiments. With pure aqueous solutions, detection limits of 5 ppb (0.5 ng in 0.1 mL injection volumes) were obtained with ICP-MS. The ICP-AES detection limit was 150 ppb. Solvent compositions were varied from 0 to 80% organic (acetonitrile and methanol) with nine buffers at concentrations typically used in liquid chromatography. In general, solvents and buffers had statistically significant, albeit small, effects on ICP-AES sensitivities. A few exceptions occurred in cases where typical liquid chromatography buffer concentrations produced higher mass loadings on the plasma. Indications are that isocratic separations can be reliably performed. Within reasonable accuracy tolerances, it appears that gradient chromatography can be performed without the need for signal response normalization. Organic solvent and buffer effects were more significant with ICP-MS. Sensitivities varied significantly with different buffers and organic solvent content. In these cases, gradient chromatography will require careful analytical calibration as solvent and buffer content is varied. However, for most buffer and solvent combinations, signal and detection limits are only moderately affected. Isocratic separations and detection are feasible.

Effect of additives on the activity of tannase from Aspergillus awamori MTCC9299.

PubMed

Chhokar, Vinod; Sangwan, Meenakshi; Beniwal, Vikas; Nehra, Kiran; Nehra, Kaur S

2010-04-01

Tannase from Aspergillus awamori MTCC 9299 was purified using ammonium sulfate precipitation followed by ion-exchange chromatography. A purification fold of 19.5 with 13.5% yield was obtained. Temperature of 30 degrees C and pH of 5.5 were found optimum for tannase activity. The effects of metals and organic solvents on the activity of tannase were also studied. Metal ions Mg(+2), Mn(+2), Ca(+2), Na(+), and K(+) stimulated the tannase activity, while Cu(+2), Fe(+3), and Co(+2) acted as inhibitors of the enzyme. The addition of organic solvents like acetic acid, isoamylalcohol, chloroform, isopropyl alcohol, and ethanol completely inhibited the enzyme activity. However, butanol and benzene increased the enzyme activity.

Demonstration of a strategy for product purification by high-gradient magnetic fishing: recovery of superoxide dismutase from unconditioned whey.

PubMed

Meyer, Andrea; Hansen, Dennis B; Gomes, Cláudia S G; Hobley, Timothy J; Thomas, Owen R T; Franzreb, Matthias

2005-01-01

A systematic approach for the design of a bioproduct recovery process employing magnetic supports and the technique of high-gradient magnetic fishing (HGMF) is described. The approach is illustrated for the separation of superoxide dismutase (SOD), an antioxidant protein present in low concentrations (ca. 0.15-0.6 mg L(-1)) in whey. The first part of the process design consisted of ligand screening in which metal chelate supports charged with copper(II) ions were found to be the most suitable. The second stage involved systematic and sequential optimization of conditions for the following steps: product adsorption, support washing, and product elution. Next, the capacity of a novel high-gradient magnetic separator (designed for biotechnological applications) for trapping and holding magnetic supports was determined. Finally, all of the above elements were assembled to deliver a HGMF process for the isolation of SOD from crude sweet whey, which consisted of (i) binding SOD using Cu2+ -charged magnetic metal chelator particles in a batch reactor with whey; (ii) recovery of the "SOD-loaded" supports by high-gradient magnetic separation (HGMS); (iii) washing out loosely bound and entrained proteins and solids; (iv) elution of the target protein; and (v) recovery of the eluted supports from the HGMF rig. Efficient recovery of SOD was demonstrated at approximately 50-fold increased scale (cf magnetic rack studies) in three separate HGMF experiments, and in the best of these (run 3) an SOD yield of >85% and purification factor of approximately 21 were obtained.

COMPUTER-ASSISTED HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY METHOD DEVELOPMENT WITH APPLICATIONS TO THE ISOLATION AND ANALYSIS OF PHYTOPLANKTON PIGMENTS. (R826944)

EPA Science Inventory

We used chromatography modeling software to assist in HPLC method development, with the goal
of enhancing separations through the exclusive use of gradient time and column temperature. We
surveyed nine stationary phases for their utility in pigment purification and natur...

The chemical composition of lymphocystis disease virus of fish.

PubMed

Robin, J; Larivière-Durand, C; Bernard, J

1983-07-01

After concentration with PEG-6000 and purification on a metrizamide gradient, the Lymphocystis Disease Virus (LDV) can be shown as a particle with a chemical composition of 1,6% DNA, 42,3% protein and 17,1% lipid, most of them being phospholipids. Sugars might constitute a major portion of the 39% unidentified components.

Optimization of elution salt concentration in stepwise elution of protein chromatography using linear gradient elution data. Reducing residual protein A by cation-exchange chromatography in monoclonal antibody purification.

PubMed

Ishihara, Takashi; Kadoya, Toshihiko; Endo, Naomi; Yamamoto, Shuichi

2006-05-05

Our simple method for optimization of the elution salt concentration in stepwise elution was applied to the actual protein separation system, which involves several difficulties such as detection of the target. As a model separation system, reducing residual protein A by cation-exchange chromatography in human monoclonal antibody (hMab) purification was chosen. We carried out linear gradient elution experiments and obtained the data for the peak salt concentration of hMab and residual protein A, respectively. An enzyme-linked immunosorbent assay was applied to the measurement of the residual protein A. From these data, we calculated the distribution coefficient of the hMab and the residual protein A as a function of salt concentration. The optimal salt concentration of stepwise elution to reduce the residual protein A from the hMab was determined based on the relationship between the distribution coefficient and the salt concentration. Using the optimized condition, we successfully performed the separation, resulting in high recovery of hMab and the elimination of residual protein A.

Ionic liquid solutions as extractive solvents for value-added compounds from biomass

PubMed Central

Passos, Helena; Freire, Mara G.; Coutinho, João A. P.

2014-01-01

In the past few years, the number of studies regarding the application of ionic liquids (ILs) as alternative solvents to extract value-added compounds from biomass has been growing. Based on an extended compilation and analysis of the data hitherto reported, the main objective of this review is to provide an overview on the use of ILs and their mixtures with molecular solvents for the extraction of value-added compounds present in natural sources. The ILs (or IL solutions) investigated as solvents for the extraction of natural compounds, such as alkaloids, flavonoids, terpenoids, lipids, among others, are outlined. The extraction techniques employed, namely solid–liquid extraction, and microwave-assisted and ultrasound-assisted extractions, are emphasized and discussed in terms of extraction yields and purification factors. Furthermore, the evaluation of the IL chemical structure and the optimization of the process conditions (IL concentration, temperature, biomass–solvent ratio, etc.) are critically addressed. Major conclusions on the role of the ILs towards the extraction mechanisms and improved extraction yields are additionally provided. The isolation and recovery procedures of the value-added compounds are ascertained as well as some scattered strategies already reported for the IL solvent recovery and reusability. Finally, a critical analysis on the economic impact versus the extraction performance of IL-based methodologies was also carried out and is here presented and discussed. PMID:25516718

Ionic liquid solutions as extractive solvents for value-added compounds from biomass.

PubMed

Passos, Helena; Freire, Mara G; Coutinho, João A P

2014-12-01

In the past few years, the number of studies regarding the application of ionic liquids (ILs) as alternative solvents to extract value-added compounds from biomass has been growing. Based on an extended compilation and analysis of the data hitherto reported, the main objective of this review is to provide an overview on the use of ILs and their mixtures with molecular solvents for the extraction of value-added compounds present in natural sources. The ILs (or IL solutions) investigated as solvents for the extraction of natural compounds, such as alkaloids, flavonoids, terpenoids, lipids, among others, are outlined. The extraction techniques employed, namely solid-liquid extraction, and microwave-assisted and ultrasound-assisted extractions, are emphasized and discussed in terms of extraction yields and purification factors. Furthermore, the evaluation of the IL chemical structure and the optimization of the process conditions (IL concentration, temperature, biomass-solvent ratio, etc.) are critically addressed. Major conclusions on the role of the ILs towards the extraction mechanisms and improved extraction yields are additionally provided. The isolation and recovery procedures of the value-added compounds are ascertained as well as some scattered strategies already reported for the IL solvent recovery and reusability. Finally, a critical analysis on the economic impact versus the extraction performance of IL-based methodologies was also carried out and is here presented and discussed.

Process analysis and modeling of a single-step lutein extraction method for wet microalgae.

PubMed

Gong, Mengyue; Wang, Yuruihan; Bassi, Amarjeet

2017-11-01

Lutein is a commercial carotenoid with potential health benefits. Microalgae are alternative sources for the lutein production in comparison to conventional approaches using marigold flowers. In this study, a process analysis of a single-step simultaneous extraction, saponification, and primary purification process for free lutein production from wet microalgae biomass was carried out. The feasibility of binary solvent mixtures for wet biomass extraction was successfully demonstrated, and the extraction kinetics of lutein from chloroplast in microalgae were first evaluated. The effects of types of organic solvent, solvent polarity, cell disruption method, and alkali and solvent usage on lutein yields were examined. A mathematical model based on Fick's second law of diffusion was applied to model the experimental data. The mass transfer coefficients were used to estimate the extraction rates. The extraction rate was found more significantly related with alkali ratio to solvent than to biomass. The best conditions for extraction efficiency were found to be pre-treatment with ultrasonication at 0.5 s working cycle per second, react 0.5 h in 0.27 L/g solvent to biomass ratio, and 1:3 ether/ethanol (v/v) with 1.25 g KOH/L. The entire process can be controlled within 1 h and yield over 8 mg/g lutein, which is more economical for scale-up.

Applications of thermal-gradients method for the optimization of α-amylase crystallization conditions based on dynamic and static light scattering data

NASA Astrophysics Data System (ADS)

Delboni, L. F.; Iulek, J.; Burger, R.; da Silva, A. C. R.; Moreno, A.

2002-02-01

The expression, purification, crystallization, and characterization by X-ray diffraction of α-amylase are described here. Dynamic and static light scattering methods with a temperature controller was used to optimize the crystallization conditions of α-amylase from Bacillus stearothermophilus an important enzyme in many fields of industrial activity. After applying thermal gradients for growing crystals, X-ray cryo-crystallographic methods were employed for the data collection. Crystals grown by these thermal-gradients diffracted up to a maximum resolution of 3.8 Å, which allowed the determination of the unit cell constants as follows: a=61.7 Å, b=86.7 Å, c=92.2 Å and space group C222 (or C222 1).

Screening and identification of genetic loci involved in producing more/denser inclusion bodies in Escherichia coli

PubMed Central

2013-01-01

Background Many proteins and peptides have been used in therapeutic or industrial applications. They are often produced in microbial production hosts by fermentation. Robust protein production in the hosts and efficient downstream purification are two critical factors that could significantly reduce cost for microbial protein production by fermentation. Producing proteins/peptides as inclusion bodies in the hosts has the potential to achieve both high titers in fermentation and cost-effective downstream purification. Manipulation of the host cells such as overexpression/deletion of certain genes could lead to producing more and/or denser inclusion bodies. However, there are limited screening methods to help to identify beneficial genetic changes rendering more protein production and/or denser inclusion bodies. Results We report development and optimization of a simple density gradient method that can be used for distinguishing and sorting E. coli cells with different buoyant densities. We demonstrate utilization of the method to screen genetic libraries to identify a) expression of glyQS loci on plasmid that increased expression of a peptide of interest as well as the buoyant density of inclusion body producing E. coli cells; and b) deletion of a host gltA gene that increased the buoyant density of the inclusion body produced in the E. coli cells. Conclusion A novel density gradient sorting method was developed to screen genetic libraries. Beneficial host genetic changes could be exploited to improve recombinant protein expression as well as downstream protein purification. PMID:23638724

Screening and identification of genetic loci involved in producing more/denser inclusion bodies in Escherichia coli.

PubMed

Pandey, Neeraj; Sachan, Annapurna; Chen, Qi; Ruebling-Jass, Kristin; Bhalla, Ritu; Panguluri, Kiran Kumar; Rouviere, Pierre E; Cheng, Qiong

2013-05-02

Many proteins and peptides have been used in therapeutic or industrial applications. They are often produced in microbial production hosts by fermentation. Robust protein production in the hosts and efficient downstream purification are two critical factors that could significantly reduce cost for microbial protein production by fermentation. Producing proteins/peptides as inclusion bodies in the hosts has the potential to achieve both high titers in fermentation and cost-effective downstream purification. Manipulation of the host cells such as overexpression/deletion of certain genes could lead to producing more and/or denser inclusion bodies. However, there are limited screening methods to help to identify beneficial genetic changes rendering more protein production and/or denser inclusion bodies. We report development and optimization of a simple density gradient method that can be used for distinguishing and sorting E. coli cells with different buoyant densities. We demonstrate utilization of the method to screen genetic libraries to identify a) expression of glyQS loci on plasmid that increased expression of a peptide of interest as well as the buoyant density of inclusion body producing E. coli cells; and b) deletion of a host gltA gene that increased the buoyant density of the inclusion body produced in the E. coli cells. A novel density gradient sorting method was developed to screen genetic libraries. Beneficial host genetic changes could be exploited to improve recombinant protein expression as well as downstream protein purification.

Binding mechanisms for histamine and agmatine ligands in plasmid deoxyribonucleic acid purifications.

PubMed

Sousa, Ângela; Pereira, Patrícia; Sousa, Fani; Queiroz, João A

2014-10-31

Histamine and agmatine amino acid derivatives were immobilized into monolithic disks, in order to combine the specificity and selectivity of the ligand with the high mass transfer and binding capacity offered by monolithic supports, to purify potential plasmid DNA biopharmaceuticals. Different elution strategies were explored by changing the type and salt concentration, as well as the pH, in order to understand the retention pattern of different plasmids isoforms The pVAX1-LacZ supercoiled isoform was isolated from a mixture of pDNA isoforms by using NaCl increasing stepwise gradient and also by ammonium sulfate decreasing stepwise gradient, in both histamine and agmatine monoliths. Acidic pH in the binding buffer mainly strengthened ionic interactions with both ligands in the presence of sodium chloride. Otherwise, for histamine ligand, pH values higher than 7 intensified hydrophobic interactions in the presence of ammonium sulfate. In addition, circular dichroism spectroscopy studies revealed that the binding and elution chromatographic conditions, such as the combination of high ionic strength with extreme pH values can reversibly influence the structural stability of the target nucleic acid. Therefore, ascending sodium chloride gradients with pH manipulation can be preferable chromatographic conditions to be explored in the purification of plasmid DNA biopharmaceuticals, in order to avoid the environmental impact of ammonium sulfate. Copyright © 2014. Published by Elsevier B.V.

Deep Eutectic Solvents as Convenient Media for Synthesis of Novel Coumarinyl Schiff Bases and Their QSAR Studies.

PubMed

Molnar, Maja; Komar, Mario; Brahmbhatt, Harshad; Babić, Jurislav; Jokić, Stela; Rastija, Vesna

2017-09-05

Deep eutectic solvents, as green and environmentally friendly media, were utilized in the synthesis of novel coumarinyl Schiff bases. Novel derivatives were synthesized from 2-((4-methyl-2-oxo-2 H -chromen-7-yl)oxy)acetohydrazide and corresponding aldehyde in choline chloride:malonic acid (1:1) based deep eutectic solvent. In these reactions, deep eutectic solvent acted as a solvent and catalyst as well. Novel Schiff bases were synthesized in high yields (65-75%) with no need for further purification, and their structures were confirmed by mass spectra, ¹H and 13 C NMR. Furthermore, their antioxidant activity was determined and compared to antioxidant activity of previously synthesized derivatives, thus investigating their structure-activity relationship utilizing quantitative structure-activity relationship QSAR studies. Calculation of molecular descriptors has been performed by DRAGON software. The best QSAR model ( R tr = 0.636; R ext = 0.709) obtained with three descriptors ( MATS3m , Mor22u , Hy ) implies that the pairs of atoms higher mass at the path length 3, three-dimensional arrangement of atoms at scattering parameter s = 21 Å - ¹, and higher number of hydrophilic groups (-OH, -NH) enhanced antioxidant activity. Electrostatic potential surface of the most active compounds showed possible regions for donation of electrons to 1,1-diphenyl-2-picryhydrazyl (DPPH) radicals.

Synthesis of Surface-Modified Iron Oxides for the Solvent-Free Recovery of Bacterial Bioactive Compound Prodigiosin and Its Algicidal Activity.

PubMed

Arivizhivendhan, K V; Mahesh, M; Boopathy, R; Patchaimurugan, K; Maharaja, P; Swarnalatha, S; Regina Mary, R; Sekaran, G

2016-09-15

Prodigiosin (PG) is a bioactive compound produced by several bacterial species. Currently, many technologies are being developed for the production of PG by fermentation processes. However, new challenges are being faced with regard to the production of PG in terms of the recovery and purification steps, owing to the labile nature of PG molecules and the cost of the purification steps. Conventional methods have limitations due to high cost, low reusability, and health hazards. Hence, the present investigation was focused on the development of surface-functionalized magnetic iron oxide ([Fe3O4]F) for solvent-free extraction of bioactive PG from the bacterial fermented medium. Fe3O4 was functionalized with diethanolamine and characterized by FT-IR, diffuse reflectance spectroscopy, thermogravimetric analysis, scanning electron microscopy, and confocal microscopy. The various process parameters, such as contact time, temperature, pH, and mass of Fe3O4, were optimized for the extraction of PG using functionalized Fe3O4. Instrumental analyses confirmed that the PG molecules were cross-linked with functional groups on [Fe3O4]F through van der Waals forces of attraction. PG extracted through Fe3O4 or [Fe3O4]F was separated from the fermentation medium by applying an external electromagnetic field and regenerated for successive reuse cycles. The purity of the extracted PG was characterized by high-performance liquid chromatography, FT-IR, and UV-visible spectroscopy. The iron oxide-diethanolamine-PG cross-linked ([Fe3O4]F-PG) composite matrix effectively deactivates harmful fouling by cyanobacterial growth in water-treatment plants. The present investigation provides the possibility of solvent-free extraction of bacterial bioactive PG from a fermented medium using functionalized magnetic iron oxide.

URANIUM PURIFICATION PROCESS

DOEpatents

Winters, C.E.

1957-11-12

A method for the preparation of a diethyl ether solution of uranyl nitrate is described. Previously the preparation of such ether solutions has been difficult and expensive, since crystalline uranyl nitrate hexahydrate dissolves very slowly in ether. An improved method for effecting such dissolution has been found, and it comprises adding molten uranyl nitrate hexahydrate at a temperature of 65 to 105 deg C to the ether while maintaining the temperature of the ether solvent below its boiling point.

A Facile Preparation of Imidazolinium Chlorides

PubMed Central

Kuhn, Kevin M.; Grubbs, Robert H.

2009-01-01

A process for the preparation of symmetric and unsymmetric imidazolinium chlorides that involves reaction of a formamidine with dichloroethane and a base (a) is described. This method makes it possible to obtain numerous imidazolinium chlorides under solvent-free reaction conditions and in excellent yields with purification by simple filtration. Alternatively, symmetric imidazolinium chlorides can be prepared directly in moderate yields from substituted anilines by utilizing half of the formamidine intermediate as sacrificial base (b). PMID:18412354

Aqueous sulfate separation by crystallization of sulfate–water clusters

DOE Office of Scientific and Technical Information (OSTI.GOV)

Custelcean, Radu; Williams, Neil J.; Seipp, Charles A.

An effective approach to separating sulfates from aqueous solutions is based on the crystallization of extended [SO 4(H 2O) 5 2-] n sulfate–water clusters with a bis(guanidinium) ligand. The ligand was generated in situ by hydrazone condensation in water, thus avoiding elaborate syntheses, tedious purifications, and organic solvents. Crystallization of sulfate–water clusters represents an alternative to the now established sulfate separation strategies that involve encapsulating the “naked” anion.

Aqueous sulfate separation by crystallization of sulfate–water clusters

DOE PAGES

Custelcean, Radu; Williams, Neil J.; Seipp, Charles A.

2015-08-07

An effective approach to separating sulfates from aqueous solutions is based on the crystallization of extended [SO 4(H 2O) 5 2-] n sulfate–water clusters with a bis(guanidinium) ligand. The ligand was generated in situ by hydrazone condensation in water, thus avoiding elaborate syntheses, tedious purifications, and organic solvents. Crystallization of sulfate–water clusters represents an alternative to the now established sulfate separation strategies that involve encapsulating the “naked” anion.

Purification of α-glucosidase from mouse intestine by countercurrent chromatography coupled with a reverse micelle solvent system.

PubMed

He, Kai; Zou, Zongyao; Hu, Yinran; Yang, Yong; Xiao, Yubo; Gao, Pincao; Li, Xuegang; Ye, Xiaoli

2016-02-01

Countercurrent chromatography coupled with a reverse micelle solvent was applied to separate α-glucosidase, which is stable at pH 6.0-8.8, 15-50°C. The separation conditions are as follows: stationary phase: pH 4.0 Tris-HCl buffer phase containing 50 mM Tris-HCl and 50 mM KCl; mobile phase A: isooctane containing 50 mM anionic surfactant sodium di(2-ethylhexyl)sulfosuccinate; mobile phase B: 50 mM Tris-HCl buffer containing 500 mM KCl (pH 8.0); In total, 25 mL (23.9 mg) crude enzyme was injected through the injection valve, the enzymatic reaction and sodium dodecylsulfate polyacrylamide gel electrophoresis results imply that the activity of purified α-glucosidase is 6.63-fold higher than that of the crude enzyme. Therefore, countercurrent chromatography coupled with a reverse micelle solvent is capable for protein separation and enrichment. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reduction of Solvent Effect in Reverse Phase Gradient Elution LC-ICP-MS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sullivan, Patrick Allen

2005-12-17

Quantification in liquid chromatography (LC) is becoming very important as more researchers are using LC, not as an analytical tool itself, but as a sample introduction system for other analytical instruments. The ability of LC instrumentation to quickly separate a wide variety of compounds makes it ideal for analysis of complex mixtures. For elemental speciation, LC is joined with inductively coupled plasma mass spectrometry (ICP-MS) to separate and detect metal-containing, organic compounds in complex mixtures, such as biological samples. Often, the solvent gradients required to perform complex separations will cause matrix effects within the plasma. This limits the sensitivity ofmore » the ICP-MS and the quantification methods available for use in such analyses. Traditionally, isotope dilution has been the method of choice for LC-ICP-MS quantification. The use of naturally abundant isotopes of a single element in quantification corrects for most of the effects that LC solvent gradients produce within the plasma. However, not all elements of interest in speciation studies have multiple naturally occurring isotopes; and polyatomic interferences for a given isotope can develop within the plasma, depending on the solvent matrix. This is the case for reverse phase LC separations, where increasing amounts of organic solvent are required. For such separations, an alternative to isotope dilution for quantification would be is needed. To this end, a new method was developed using the Apex-Q desolvation system (ESI, Omaha, NE) to couple LC instrumentation with an ICP-MS device. The desolvation power of the system allowed greater concentrations of methanol to be introduced to the plasma prior to destabilization than with direct methanol injection into the plasma. Studies were performed, using simulated and actual linear methanol gradients, to find analyte-internal standard (AIS) pairs whose ratio remains consistent (deviations {+-} 10%) over methanol concentration ranges of 5%-35% (simulated) and 8%-32% (actual). Quadrupole (low resolution) and sector field (high resolution) ICP-MS instrumentation were utilized in these studies. Once an AIS pair is determined, quantification studies can be performed. First, an analysis is performed by adding both elements of the AIS pair post-column while performing the gradient elution without sample injection. A comparison of the ratio of the measured intensities to the atomic ratio of the two standards is used to determine a correction factor that can be used to account for the matrix effects caused by the mobile phase. Then, organic and/or biological molecules containing one of the two elements in the AIS pair are injected into the LC column. A gradient method is used to vary the methanol-water mixture in the mobile phase and to separate out the compounds in a given sample. A standard solution of the second ion in the AIS pair is added continuously post-column. By comparing the ratio of the measured intensities to the atomic ratio of the eluting compound and internal standard, the concentration of the injected compound can be determined.« less

Thermal Motion and Forced Migration of Colloidal Particles Generate Hydrostatic Pressure in Solvent

PubMed Central

Hammel, H. T.; Scholander, P. F.

1973-01-01

A colloidal solution of ferrite particles in an osmometer has been used to demonstrate that the property that propels water across the semipermeable membrane is the decrease in hydrostatic pressure in the water of the solution. A magnetic field gradient directed so as to force the ferrite particles away from the semipermeable membrane of the osmometer and toward the free surface of the solution enhanced the colloidal osmotic pressure. The enhancement of this pressure was always exactly equal to the augmentation of the pressure as measured by the outward force of the particles, against the area of the free surface. Contrariwise, directing the magnetic field gradient so as to force the ferrite particles away from the free surface and toward the semipermeable membrane diminished the colloidal osmotic pressure of the solution. For a sufficiently forceful field gradient, the initial colloidal osmotic pressure could be negative, followed by an equilibrium pressure approaching zero regardless of the force of the particles against the membrane. Thus, the osmotic pressure of a solution is to be attributed to the pressure in the solvent generated in opposition to the pressure of the solute particles caused by their interaction with the free surface (Brownian motion and/or an external field force), or by their viscous shear when they migrate through the solvent, or both. PMID:16592046

Isolation of purified oocyst walls and sporocysts from Toxoplasma gondii.

PubMed

Everson, William V; Ware, Michael W; Dubey, J P; Lindquist, H D Alan

2002-01-01

Toxoplasma gondii oocysts are environmentally resistant and can infect virtually all warm-blooded hosts, including humans and livestock. Little is known about the biochemical basis for this resistance of oocysts, and mechanism for excystation of T. gondii sporozoites. The objective of the present study was to evaluate different methods (mechanical fragmentation, gradients, flow cytometry) to separate and purify T. gondii oocyst walls and sporocysts. Oocyst walls were successfully separated and purified using iodixanol gradients. Sporocysts were successfully separated and purified using iodixanol and Percoll gradients. Purification was also achieved by flow cytometry. Flow cytometry with fluorescence-activated cell sorting (FACS) yielded analytical quantities of oocyst walls and intact sporocysts. Flow cytometry with FACS also proved useful for quantitation of purity obtained following iodixanol gradient fractionation. Methods reported in this paper will be useful for analytical purposes, such as proteomic analysis of components unique to this life cycle stage, development of detection methods, or excystation studies.

DNA stable-isotope probing (DNA-SIP).

PubMed

Dunford, Eric A; Neufeld, Josh D

2010-08-02

DNA stable-isotope probing (DNA-SIP) is a powerful technique for identifying active microorganisms that assimilate particular carbon substrates and nutrients into cellular biomass. As such, this cultivation-independent technique has been an important methodology for assigning metabolic function to the diverse communities inhabiting a wide range of terrestrial and aquatic environments. Following the incubation of an environmental sample with stable-isotope labelled compounds, extracted nucleic acid is subjected to density gradient ultracentrifugation and subsequent gradient fractionation to separate nucleic acids of differing densities. Purification of DNA from cesium chloride retrieves labelled and unlabelled DNA for subsequent molecular characterization (e.g. fingerprinting, microarrays, clone libraries, metagenomics). This JoVE video protocol provides visual step-by-step explanations of the protocol for density gradient ultracentrifugation, gradient fractionation and recovery of labelled DNA. The protocol also includes sample SIP data and highlights important tips and cautions that must be considered to ensure a successful DNA-SIP analysis.

Semi-automated hydrophobic interaction chromatography column scouting used in the two-step purification of recombinant green fluorescent protein.

PubMed

Stone, Orrin J; Biette, Kelly M; Murphy, Patrick J M

2014-01-01

Hydrophobic interaction chromatography (HIC) most commonly requires experimental determination (i.e., scouting) in order to select an optimal chromatographic medium for purifying a given target protein. Neither a two-step purification of untagged green fluorescent protein (GFP) from crude bacterial lysate using sequential HIC and size exclusion chromatography (SEC), nor HIC column scouting elution profiles of GFP, have been previously reported. Bacterial lysate expressing recombinant GFP was sequentially adsorbed to commercially available HIC columns containing butyl, octyl, and phenyl-based HIC ligands coupled to matrices of varying bead size. The lysate was fractionated using a linear ammonium phosphate salt gradient at constant pH. Collected HIC eluate fractions containing retained GFP were then pooled and further purified using high-resolution preparative SEC. Significant differences in presumptive GFP elution profiles were observed using in-line absorption spectrophotometry (A395) and post-run fluorimetry. SDS-PAGE and western blot demonstrated that fluorometric detection was the more accurate indicator of GFP elution in both HIC and SEC purification steps. Comparison of composite HIC column scouting data indicated that a phenyl ligand coupled to a 34 µm matrix produced the highest degree of target protein capture and separation. Conducting two-step protein purification using the preferred HIC medium followed by SEC resulted in a final, concentrated product with >98% protein purity. In-line absorbance spectrophotometry was not as precise of an indicator of GFP elution as post-run fluorimetry. These findings demonstrate the importance of utilizing a combination of detection methods when evaluating purification strategies. GFP is a well-characterized model protein, used heavily in educational settings and by researchers with limited protein purification experience, and the data and strategies presented here may aid in development other of HIC-compatible protein purification schemes.

Algorithms for Solvents and Spectral Factors of Matrix Polynomials

DTIC Science & Technology

1981-01-01

spectral factors of matrix polynomials LEANG S. SHIEHt, YIH T. TSAYt and NORMAN P. COLEMANt A generalized Newton method , based on the contracted gradient...of a matrix poly- nomial, is derived for solving the right (left) solvents and spectral factors of matrix polynomials. Two methods of selecting initial...estimates for rapid convergence of the newly developed numerical method are proposed. Also, new algorithms for solving complete sets of the right

Theoretical investigation of the weak interaction between graphene and alcohol solvents

NASA Astrophysics Data System (ADS)

Wang, Haining; Chen, Sian; Lu, Shanfu; Xiang, Yan

2017-05-01

The dispersion of graphene in five different alcohol solvents was investigated by evaluating the binding energy between graphene and alcohol molecules using DFT-D method. The calculation showed the most stable binding energy appeared at the distance of ∼3.5 Å between graphene and alcohol molecules and increased linearly as changing the alcohol from methanol to 1-pentanol. The weak interaction was further graphically illustrated using the reduced density gradient method. The theoretical study revealed alcohols with more carbon atoms could be a good starting point for screening suitable solvents for graphene dispersion.

A novel 9 × 9 map-based solvent selection strategy for targeted counter-current chromatography isolation of natural products.

PubMed

Liang, Junling; Meng, Jie; Wu, Dingfang; Guo, Mengzhe; Wu, Shihua

2015-06-26

Counter-current chromatography (CCC) is an efficient liquid-liquid chromatography technique for separation and purification of complex mixtures like natural products extracts and synthetic chemicals. However, CCC is still a challenging process requiring some special technical knowledge especially in the selection of appropriated solvent systems. In this work, we introduced a new 9 × 9 map-based solvent selection strategy for CCC isolation of targets, which permit more than 60 hexane-ethyl acetate-methanol-water (HEMWat) solvent systems as the start candidates for the selection of solvent systems. Among these solvent systems, there are clear linear correlations between partition coefficient (K) and the system numbers. Thus, an appropriate CCC solvent system (i.e., sweet spot for K = 1) may be hit by measurement of k values of the target only in two random solvent systems. Besides this, surprisingly, we found that through two sweet spots, we could get a line ("Sweet line") where there are infinite sweet solvent systems being suitable for CCC separation. In these sweet solvent systems, the target has the same partition coefficient (K) but different solubilities. Thus, the better sweet solvent system with higher sample solubility can be obtained for high capacity CCC preparation. Furthermore, we found that there is a zone ("Sweet zone") where all solvent systems have their own sweet partition coefficients values for the target in range of 0.4 < K< 2.5 or extended range of 0.25 < K < 16. All results were validated by using 14 pure GUESSmix mimic natural products as standards and further confirmed by isolation of several targets including honokiol and magnolol from the extracts of Magnolia officinalis Rehd. Et Wils and tanshinone IIA from Salvia miltiorrhiza Bunge. In practice, it is much easier to get a suitable solvent system only by making a simple screening two to four HEMWat two-phase solvent systems to obtain the sweet line or sweet zone without special knowledge or comprehensive standards as references. This is an important advancement for solvent system selection and also will be very useful for isolation of current natural products including Traditional Chinese Medicines. Copyright © 2015 Elsevier B.V. All rights reserved.

In vitro cell-mediated immunity after thermal injury is not impaired. Density gradient purification of mononuclear cells is associated with spurious (artifactual) immunosuppression.

PubMed Central

Xu, D Z; Deitch, E A; Sittig, K; Qi, L; McDonald, J C

1988-01-01

Mononuclear cells isolated by density gradient centrifugation from the peripheral blood of burn patients, but not healthy volunteers, are contaminated with large numbers of nonmononuclear cells. These contaminating leukocytes could cause artifactual alterations in standard in vitro tests of lymphocyte function. Thus, we compared the in vitro blastogenic response of density gradient purified leukocytes and T-cell purified lymphocytes from 13 burn patients to mitogenic (PHA) and antigenic stimuli. The mitogenic and antigenic response of the patients' density gradient purified leukocytes were impaired compared to healthy volunteers (p less than 0.01). However, when the contaminating nonlymphocytes were removed, the patients' cells responded normally to both stimuli. Thus, density gradient purified mononuclear cells from burn patients are contaminated by leukocytes that are not phenotypically or functionally lymphocytes. Since the lymphocytes from burn patients respond normally to PHA and alloantigens after the contaminating nonlymphocyte cell population has been removed, it appears that in vitro assays of lymphocyte function using density gradient purified leukocytes may give spurious results. PMID:2973771

Simultaneous concentration and purification through gradient deformation chromatography

NASA Technical Reports Server (NTRS)

Velayudhan, A.; Hendrickson, R. L.; Ladisch, M. R.; Mitchell, C. A. (Principal Investigator)

1995-01-01

Mobile-phase additives, commonly used to modulate absorbate retention in gradient elution chromatography, are usually assumed to be either linearly retained or unretained. Previous theoretical work from our laboratory has shown that these modulators, such as salts in ion-exchange and hydrophobic interaction chromatography and organic modifiers in reversed-phase chromatography, can absorb nonlinearly, giving rise to gradient deformation. Consequently, adsorbate peaks that elute in the vicinity of the head of the deformed gradient may exhibit unusual shapes, form shoulders, and/or be concentrated. These effects for a reversed-phase sorbent with aqueous acetonitrile (ACN) as the modulator are verified experimentally. Gradient deformation is demonstrated experimentally and agrees with simulations based on ACN isotherm parameters that are independently determined from batch equilibrium studies using the layer model. Unusual absorbate peak shapes were found experimentally for single-component injections of phenylalanine, similar to those calculated by the simulations. A binary mixture of tryptophan and phenylalanine is used to demonstrate simultaneous concentration and separation, again in agreement with simulations. The possibility of gradient deformation in ion-exchange and hydrophobic interaction chromatography is discussed.

Temperature gradient interaction chromatography of polymers: A molecular statistical model.

PubMed

Radke, Wolfgang; Lee, Sekyung; Chang, Taihyun

2010-11-01

A new model describing the retention in temperature gradient interaction chromatography of polymers is developed. The model predicts that polymers might elute in temperature gradient interaction chromatography in either an increasing or decreasing order or even nearly independent of molar mass, depending on the rate of the temperature increase relative to the flow rate. This is in contrast to solvent gradient elution, where polymers elute either in order of increasing molar mass or molar mass independent. The predictions of the newly developed model were verified with the literature data as well as new experimental data. Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Supercritical fluid processing: opportunities for new resist materials and processes

NASA Astrophysics Data System (ADS)

Gallagher-Wetmore, Paula M.; Ober, Christopher K.; Gabor, Allen H.; Allen, Robert D.

1996-05-01

Over the past two decades supercritical fluids have been utilized as solvents for carrying out separations of materials as diverse as foods, polymers, pharmaceuticals, petrochemicals, natural products, and explosives. More recently they have been used for non-extractive applications such as recrystallization, deposition, impregnation, surface modification, and as a solvent alternative for precision parts cleaning. Today, supercritical fluid extraction is being practiced in the foods and beverage industries; there are commercial plants for decaffeinating coffee and tea, extracting beer flavoring agents from hops, and separating oils and oleoresins from spices. Interest in supercritical fluid processing of polymers has grown over the last ten years, and many new purification, fractionation, and even polymerization techniques have emerged. One of the most significant motivations for applying this technology to polymers has been increased performance demands. More recently, with increasing scrutiny of traditional solvents, supercritical fluids, and in particular carbon dioxide, are receiving widespread attention as 'environmentally conscious' solvents. This paper describes several examples of polymers applications, including a few involving photoresists, which demonstrate that as next- generation advanced polymer systems emerge, supercritical fluids are certain to offer advantages as cutting edge processing tools.

Comparison of Plasma Exosomes by Differential Ultracentrifugation and Solvent Precipitation Methods.

PubMed

Peng, Qiao; Zhang, Jing; Zhou, Gang

2018-06-01

Emerging evidence has identified that exosomes play a pivotal role in intercellular signal transmission. However, the standardized purification techniques to isolate high quality exosomes are still deficient at present. This study was to evaluate reproducibility and efficiency of differential ultracentrifugation and solvent precipitation-based kits by isolating plasma-derived exosomes from oral lichen planus patients. Morphology, exosomal biomarkers, particle size distribution, proteomic components, and protein yield of isolated exosomes were evaluated by transmission electron microscope, western blot, laser diffraction instrument, Coomassie staining, and BCA protein assay kit, respectively. TEM displayed representative cup-shaped morphology of exosomes and western blot identified exosomal biomarkers CD9 and CD63. The size distribution showed that particles by differential ultracentrifugation were mainly from 26.15 nm to 166.5 nm, while some of the particles obtained by solvent precipitation kits were larger than 1,000 nm. In addition, exosomes isolated by solvent precipitation kits showed a significantly higher amount of protein yield due to plasma albumin contamination. Both differential ultracentrifugation and precipitation based kits could successfully isolate plasma exosomes, and exosomes by differential ultracentrifugation were purer and more appropriate for further proteomic analysis.

Modification of the crystal habit of celecoxib for improved processability.

PubMed

Banga, Sheere; Chawla, Garima; Varandani, Deepak; Mehta, B R; Bansal, Arvind K

2007-01-01

Crystallization is often used in the pharmaceutical industry for purification and isolation of drugs, and also as a means of generating polymorphs or isomorphs. The aim of this study was to investigate the role of extrinsic crystallization parameters on the crystallized product, with special emphasis on improving the mechanical properties of acicular celecoxib. Celecoxib isomorphs were prepared using different techniques (solvent crystallization and vapour diffusion) and crystallization conditions (solvents, stirring, degree of supersaturation, crystallization temperature and seeding). Powder X-ray diffractometry, spectroscopic and thermal methods were used to investigate physical characteristics of crystals. Growth kinetics and aggregation dynamics of crystallization in polar and non-polar solvents were simulated using a dynamic light scattering method. The quick appearance of broad peaks over the range of 10-8000 nm in chloroform during crystallization simulation studies indicated faster aggregation in non-polar solvents. Aspect ratio, flow, compressibility and surface area of recrystallized products were also determined. Surface topography was determined by atomic force microscopy and the lath-shaped crystals (aspect ratio of 2-4) exhibited a roughness index of 1.79 in comparison with 2.92 for needles. Overall, the lath-shaped isomorphs exhibited improved flow and better compressibility.

Application of a New Dual Localization-Affinity Purification Tag Reveals Novel Aspects of Protein Kinase Biology in Aspergillus nidulans

PubMed Central

De Souza, Colin P.; Hashmi, Shahr B.; Osmani, Aysha H.; Osmani, Stephen A.

2014-01-01

Filamentous fungi occupy critical environmental niches and have numerous beneficial industrial applications but devastating effects as pathogens and agents of food spoilage. As regulators of essentially all biological processes protein kinases have been intensively studied but how they regulate the often unique biology of filamentous fungi is not completely understood. Significant understanding of filamentous fungal biology has come from the study of the model organism Aspergillus nidulans using a combination of molecular genetics, biochemistry, cell biology and genomic approaches. Here we describe dual localization-affinity purification (DLAP) tags enabling endogenous N or C-terminal protein tagging for localization and biochemical studies in A. nidulans. To establish DLAP tag utility we endogenously tagged 17 protein kinases for analysis by live cell imaging and affinity purification. Proteomic analysis of purifications by mass spectrometry confirmed association of the CotA and NimXCdk1 kinases with known binding partners and verified a predicted interaction of the SldABub1/R1 spindle assembly checkpoint kinase with SldBBub3. We demonstrate that the single TOR kinase of A. nidulans locates to vacuoles and vesicles, suggesting that the function of endomembranes as major TOR cellular hubs is conserved in filamentous fungi. Comparative analysis revealed 7 kinases with mitotic specific locations including An-Cdc7 which unexpectedly located to mitotic spindle pole bodies (SPBs), the first such localization described for this family of DNA replication kinases. We show that the SepH septation kinase locates to SPBs specifically in the basal region of apical cells in a biphasic manner during mitosis and again during septation. This results in gradients of SepH between G1 SPBs which shift along hyphae as each septum forms. We propose that SepH regulates the septation initiation network (SIN) specifically at SPBs in the basal region of G1 cells and that localized gradients of SIN activity promote asymmetric septation. PMID:24599037

Application of a new dual localization-affinity purification tag reveals novel aspects of protein kinase biology in Aspergillus nidulans.

PubMed

De Souza, Colin P; Hashmi, Shahr B; Osmani, Aysha H; Osmani, Stephen A

2014-01-01

Filamentous fungi occupy critical environmental niches and have numerous beneficial industrial applications but devastating effects as pathogens and agents of food spoilage. As regulators of essentially all biological processes protein kinases have been intensively studied but how they regulate the often unique biology of filamentous fungi is not completely understood. Significant understanding of filamentous fungal biology has come from the study of the model organism Aspergillus nidulans using a combination of molecular genetics, biochemistry, cell biology and genomic approaches. Here we describe dual localization-affinity purification (DLAP) tags enabling endogenous N or C-terminal protein tagging for localization and biochemical studies in A. nidulans. To establish DLAP tag utility we endogenously tagged 17 protein kinases for analysis by live cell imaging and affinity purification. Proteomic analysis of purifications by mass spectrometry confirmed association of the CotA and NimXCdk1 kinases with known binding partners and verified a predicted interaction of the SldABub1/R1 spindle assembly checkpoint kinase with SldBBub3. We demonstrate that the single TOR kinase of A. nidulans locates to vacuoles and vesicles, suggesting that the function of endomembranes as major TOR cellular hubs is conserved in filamentous fungi. Comparative analysis revealed 7 kinases with mitotic specific locations including An-Cdc7 which unexpectedly located to mitotic spindle pole bodies (SPBs), the first such localization described for this family of DNA replication kinases. We show that the SepH septation kinase locates to SPBs specifically in the basal region of apical cells in a biphasic manner during mitosis and again during septation. This results in gradients of SepH between G1 SPBs which shift along hyphae as each septum forms. We propose that SepH regulates the septation initiation network (SIN) specifically at SPBs in the basal region of G1 cells and that localized gradients of SIN activity promote asymmetric septation.

WHEAT GERM CELL-FREE TRANSLATION, PURIFICATION, AND ASSEMBLY OF A FUNCTIONAL HUMAN STEAROYL-COA DESATURASE COMPLEX

PubMed Central

Goren, Michael A.; Fox, Brian G.

2008-01-01

A wheat germ cell-free extract was used to perform in vitro translation of human stearoyl-CoA desaturase in the presence of unilamelar liposomes, and near complete transfer of the expressed integral membrane protein into the liposome was observed. Moreover, co-translation of the desaturase along with human cytochrome b5 led to transfer of both membrane proteins into the liposomes. A simple, single step purification via centrifugation in a density gradient yielded proteoliposomes with the desaturase in high purity as judged by capillary electrophoresis. After in vitro reconstitution of the non-heme iron and heme active sites, the function of the reconstituted enzyme complex was demonstrated by conversion of stearoyl-CoA to oleoyl-CoA. This simple translation approach obviates the use of detergents or other lipids to stabilize and isolate a catalytically active integral membrane enzyme. The applicability of cell-free translation to the assembly and purification of other integral membrane protein complexes is discussed. PMID:18765284

Wheat germ cell-free translation, purification, and assembly of a functional human stearoyl-CoA desaturase complex.

PubMed

Goren, Michael A; Fox, Brian G

2008-12-01

A wheat germ cell-free extract was used to perform in vitro translation of human stearoyl-CoA desaturase in the presence of unilamelar liposomes, and near complete transfer of the expressed integral membrane protein into the liposome was observed. Moreover, co-translation of the desaturase along with human cytochrome b(5) led to transfer of both membrane proteins into the liposomes. A simple, single step purification via centrifugation in a density gradient yielded proteoliposomes with the desaturase in high purity as judged by capillary electrophoresis. After in vitro reconstitution of the non-heme iron and heme active sites, the function of the reconstituted enzyme complex was demonstrated by conversion of stearoyl-CoA to oleoyl-CoA. This simple translation approach obviates the use of detergents or other lipids to stabilize and isolate a catalytically active integral membrane enzyme. The applicability of cell-free translation to the assembly and purification of other integral membrane protein complexes is discussed.

Application of hydrophobic interaction displacement chromatography for an industrial protein purification.

PubMed

Sunasara, Khurram M; Xia, Fang; Gronke, Robert S; Cramer, Steven M

2003-05-05

Recently it has been established that low molecular weight displacers can be successfully employed for the purification of proteins in hydrophobic interaction chromatography (HIC) systems. This work investigates the utility of this technique for the purification of an industrial protein mixture. The study involved the separation of a mixture of three protein forms, that differed in the C-terminus, from their aggregate impurities while maintaining the same relative ratio of the three protein forms as in the feed. A batch high-throughput screening (HTS) technique was employed in concert with fluorescence spectroscopy for displacer screening in these HIC systems. This methodology was demonstrated to be an effective tool for identifying lead displacer candidates for a particular protein/stationary-phase system. In addition, these results indicate that surfactants can be employed at concentrations above their CMCs as effective displacers. Displacement of the recombinant proteins with PEG-3400 and the surfactant Big Chap was shown to increase the productivity as compared to the existing step-gradient elution process. Copyright 2003 Wiley Periodicals, Inc.

Purification and protein composition of endogenous rat viruses.

PubMed

Hlubinová, K; Prachar, J; Vrbenská, A; Matoska, J; Simkovic, D

1984-01-01

Endogenous retroviruses are not in the majority of cases the cause of any neoplasia, except for the laboratory conditions. As far as they might serve for the evolution of pathogenic retroviruses more attention should have been paid to them. In this paper we introduce some approaches to the purification of rat endogenous retroviruses to such a degree of purity that enabled satisfactory SDS-PAGE analysis of its structural proteins. Purities of samples obtained by usual purification methods, long-term isopycnic centrifugation at a high gravity force and velocity centrifugation are compared. Protein profile of rat endogenous virus in SDS-PAGE is compared with the ones of other retroviruses. For the first time the evidence was obtained for the striking similarity between electrophoretic protein profile of rat endogenous virus WERC and feline leukemia virus. The major structural proteins of rat endogenous retrovirus and feline leukemia virus cannot be distinguished even when resolution long gradient PAGE had been employed. The accordance of electrophoretic mobilities of major structural proteins in SDS-PAGE can indicate the relatedness of retroviruses.

Suppression of Protonated Organic Solvents in NMR Spectroscopy Using a Perfect Echo Low-Pass Filtration Pulse Sequence.

PubMed

Howe, Peter W A

2018-04-03

Proton NMR spectra are usually acquired using deuterated solvents, but in many cases it is necessary to obtain spectra on samples in protonated solvents. In these cases, the intense resonances of the protonated solvents need to be suppressed to maximize sensitivity and spectral quality. A wide range of highly effective solvent suppression methods have been developed, but additional measures are needed to suppress the 13 C satellites of the solvent. Because the satellites represent 1.1% of the original solvent signal, they remain problematic if unsuppressed. The recently proposed DISPEL pulse sequences suppress 13 C satellites extremely effectively, and this Technical Note demonstrates that combining DISPEL and presaturation results in exceptionally effective solvent suppression. An important element in the effectiveness is volume selection, which is inherent within the DISPEL sequence. Spectra acquired in protonated dimethlysulfoxide and tetrahydrofuran show that optimum results are obtained by modifying the phase cycle, cycling the pulse-field gradients, and using broadband 13 C inversion pulses to reduce the effects of radiofrequency offset and inhomogeneity.

Magnesium Electrorefining in Non-Aqueous Electrolyte at Room Temperature

NASA Astrophysics Data System (ADS)

Kwon, Kyungjung; Park, Jesik; Kusumah, Priyandi; Dilasari, Bonita; Kim, Hansu; Lee, Churl Kyoung

Magnesium, of which application is often limited by its poor corrosion resistance, is more vulnerable to corrosion with existence of metal impurities such as Fe. Therefore, for the refining and recycling of magnesium, high temperature electrolysis using molten salts has been frequently adopted. In this report, the purification of magnesium scrap by electrolysis at room temperature is investigated with non-aqueous electrolytes. An aprotic solvent of tetrahydrofuran (THF) was used as a solvent of the electrolyte. Magnesium scrap was used as anode materials and ethyl magnesium bromide (EtMgBr) was dissolved in THF for magnesium source. The purified magnesium can be uniformly electrodeposited on copper electrode under potentiostatic conditions. The deposits were confirmed by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) analysis.

A Review of Biorefinery Separations for Bioproduct Production via Thermocatalytic Processing.

PubMed

Nguyen, Hannah; DeJaco, Robert F; Mittal, Nitish; Siepmann, J Ilja; Tsapatsis, Michael; Snyder, Mark A; Fan, Wei; Saha, Basudeb; Vlachos, Dionisios G

2017-06-07

With technological advancement of thermocatalytic processes for valorizing renewable biomass carbon, development of effective separation technologies for selective recovery of bioproducts from complex reaction media and their purification becomes essential. The high thermal sensitivity of biomass intermediates and their low volatility and high reactivity, along with the use of dilute solutions, make the bioproducts separations energy intensive and expensive. Novel separation techniques, including solvent extraction in biphasic systems and reactive adsorption using zeolite and carbon sorbents, membranes, and chromatography, have been developed. In parallel with experimental efforts, multiscale simulations have been reported for predicting solvent selection and adsorption separation. We discuss various separations that are potentially valuable to future biorefineries and the factors controlling separation performance. Particular emphasis is given to current gaps and opportunities for future development.

[Simultaneous determination of 6 forbidden colorants in cosmetics by high performance liquid chromatography-tandem mass spectrometry].

PubMed

Lin, Weixuan; Sun, Xingquan; Zhao, Xuerong; Xu, Wei; Guo, Guiyuan

2012-05-01

A method of high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) has been established for the simultaneous determination of six forbidden colorants including Sudan IV, Acid Violet 49, Sudan Blue 2, Solvent Red 49, Basic Violet 1 and Pigment Orange 5 in cream and powdery matrix cosmetics. The samples were extracted with ethanol-acetonitrile (3:2, v/v) solution by ultrasonic technique for 20 min, then centrifuged for purification and enriched by nitrogen blowing sequentially. The analytes were isolated on a Luna C18 column (150 mm x 2.1 mm, 5 microm) by gradient elution with methanol and 10 mmol/L ammonium acetate as the mobile phases, and detected by MS/MS in the multiple reaction monitoring (MRM) mode. The qualitative analysis was based on the retention time and the relative abundance ratio of the characteristic ions, and the quantitative analysis on calibration curve method. The results showed that the limits of quantification (LOQ, S/N= 10) of the six colorants ranged from 0.1 to 10 microg/kg and the average recoveries were from 86.67% to 98.22% with the relative standard deviations (RSDs) from 4.01% to 7.01%. The method is simple and rapid with high sensitivity and good reproducibility, and suitable for the determination of the six forbidden colorants in cosmetics.

Development of a fast and selective UHPLC-DAD-QTOF-MS/MS method for the qualitative and quantitative assessment of destruxin profiles.

PubMed

Taibon, Judith; Sturm, Sonja; Seger, Christoph; Parth, Martin; Strasser, Hermann; Stuppner, Hermann

2014-11-01

A fast and selective ultrahigh-performance liquid chromatography diode array detector (UHPLC-DAD) method combined with an off-line solid phase extraction (SPE) protocol was established to monitor destruxins (dtxs), a secondary metabolite class of highly bioactive cyclic depsipeptides. Sample purification via SPE was tailored to remove both more polar and apolar matrix constituents by applying analyte class-selective washing and elution conditions. To separate and detect destruxin congeners an UHPLC-DAD system hyphenated to a quadrupole-time-of-flight (Q-TOF) hybrid mass spectrometer was utilized. Analyses were performed on a sub-2-μm-particle-size RP-18 column with an acidified (0.02% acetic acid) 12 min water/acetonitrile solvent gradient. In the dtx congener elution zone 22 chromatographic peaks were separated. Four of these were identified by comparison with reference materials as dtx A, dtx B, dtx E, and dtx E-diol; 16 were tentatively assigned as known or novel dtx congeners by the analysis of high resolution UHPLC-DAD-QTOF-MS/MS data recorded in the positive electrospray ionization (ESI) mode. The applicability of the UHPLC-DAD assay to investigate biological materials in a qualitative and quantitative manner was proven by the application of the platform to monitor the dtx production profile of three Metarhizium brunneum strain fungal culture broths.

Quantitation of promethazine and metabolites in urine samples using on-line solid-phase extraction and column-switching

NASA Technical Reports Server (NTRS)

Song, Q.; Putcha, L.; Harm, D. L. (Principal Investigator)

2001-01-01

A chromatographic method for the quantitation of promethazine (PMZ) and its three metabolites in urine employing on-line solid-phase extraction and column-switching has been developed. The column-switching system described here uses an extraction column for the purification of PMZ and its metabolites from a urine matrix. The extraneous matrix interference was removed by flushing the extraction column with a gradient elution. The analytes of interest were then eluted onto an analytical column for further chromatographic separation using a mobile phase of greater solvent strength. This method is specific and sensitive with a range of 3.75-1400 ng/ml for PMZ and 2.5-1400 ng/ml for the metabolites promethazine sulfoxide, monodesmethyl promethazine sulfoxide and monodesmethyl promethazine. The lower limits of quantitation (LLOQ) were 3.75 ng/ml with less than 6.2% C.V. for PMZ and 2.50 ng/ml with less than 11.5% C.V. for metabolites based on a signal-to-noise ratio of 10:1 or greater. The accuracy and precision were within +/- 11.8% in bias and not greater than 5.5% C.V. in intra- and inter-assay precision for PMZ and metabolites. Method robustness was investigated using a Plackett-Burman experimental design. The applicability of the analytical method for pharmacokinetic studies in humans is illustrated.

Chemical isolation of .sup.82 Sr from proton-irradiated Mo targets

DOEpatents

Grant, Patrick M.; Kahn, Milton; O'Brien, Jr., Harold A.

1976-01-01

Spallation reactions are induced in Mo targets with 200-800 MeV protons to produce microcurie to millicurie amounts of a variety of radionuclides. A six-step radiochemical procedure, incorporating precipitation, solvent extractions, and ion exchange techniques, has been developed for the separation and purification of Sr radioactivities from other spallation products and the bulk target material. Radiostrontium can be quantitatively recovered in a sufficiently decontaminated state for use in biomedical generator development.

Production and recovery of Americium-241

DOE Office of Scientific and Technical Information (OSTI.GOV)

Navratil, J.D.

1984-01-01

Americium-241, one of the most useful actinide isotopes, is produced as a by-product of plutonium scrap recovery operations. Rocky Flats (RF) has supplied high-purity americium oxide to the US Department of Energy's Isotope Pool since 1962. Over the years, the evolving separation and purification processes have included such diverse operations as aqueous precipitation, ion exchange, and both molten-salt and organic-solvent extraction. A review is presented of the production and recovery processes of americium-241. 5 references.

Process for purifying geothermal steam

DOEpatents

Li, Charles T.

1980-01-01

Steam containing hydrogen sulfide is purified and sulfur recovered by passing the steam through a reactor packed with activated carbon in the presence of a stoichiometric amount of oxygen which oxidizes the hydrogen sulfide to elemental sulfur which is adsorbed on the bed. The carbon can be recycled after the sulfur has been recovered by vacuum distillation, inert gas entrainment or solvent extraction. The process is suitable for the purification of steam from geothermal sources which may also contain other noncondensable gases.

Process for purifying geothermal steam

DOEpatents

Li, C.T.

Steam containing hydrogen sulfide is purified and sulfur recovered by passing the steam through a reactor packed with activated carbon in the presence of a stoichiometric amount of oxygen which oxidizes the hydrogen sulfide to elemental sulfur which is adsorbed on the bed. The carbon can be recycled after the sulfur has been recovered by vacuum distillation, inert gas entrainment or solvent extraction. The process is suitable for the purification of steam from geothermal sources which may also contain other noncondensable gases.

The application of a monolithic triphenylphosphine reagent for conducting Ramirez gem-dibromoolefination reactions in flow.

PubMed

Roper, Kimberley A; Berry, Malcolm B; Ley, Steven V

2013-01-01

The application of a monolithic form of triphenylphosphine to the Ramirez gem-dibromoolefination reaction using flow chemistry techniques is reported. A variety of gem-dibromides were synthesised in high purity and excellent yield following only removal of solvent and no further off-line purification. It is also possible to perform the Appel reaction using the same monolith and the relationship between the mechanisms of the two reactions is discussed.

The application of a monolithic triphenylphosphine reagent for conducting Ramirez gem-dibromoolefination reactions in flow

PubMed Central

Roper, Kimberley A; Berry, Malcolm B

2013-01-01

Summary The application of a monolithic form of triphenylphosphine to the Ramirez gem-dibromoolefination reaction using flow chemistry techniques is reported. A variety of gem-dibromides were synthesised in high purity and excellent yield following only removal of solvent and no further off-line purification. It is also possible to perform the Appel reaction using the same monolith and the relationship between the mechanisms of the two reactions is discussed. PMID:24062843

Isolation, Purification, and Identification of an Important Pigment, Sepiapterin, from Integument of the lemon Mutant of the Silkworm, Bombyx mori

PubMed Central

Wang, Jing; Wang, Wenjing; Liu, Chaoliang; Meng, Yan

2013-01-01

Sepiapterin is the precursor of tetrahydrobiopterin, an important coenzyme of aromatic amino acid hydroxylases, the lack of which leads to a variety of physiological metabolic diseases or neurological syndromes in humans. Sepiapterin is a main pigment component in the integument of the lemon mutant of the silkworm, Bombyx mori (L.) (Lepidoptera: Bombycidae), and is present there in extremely high content, so lemon is a valuable genetic resource to extract sepiapterin. In this study, an effective experimental system was set up for isolation and purification of sepiapterin from lemon silkworms by optimizing homogenization solvent, elution buffer, and separation chromatographic column. The results showed that ethanol was the most suitable solvent to homogenize the integument, with a concentration of 50% and solid:liquid ratio of 1:20 (g/mL). Sepiapterin was purified successively by column chromatography of cellulose Ecteola, sephadex G-25-150, and cellulose phosphate, and was identified by ultraviolet-visible absorption spectrometry. A stable and accurate high performance liquid chromatography method was constructed to identify sepiapterin and conduct qualitative and quantitative analyses. Sepiapterin of high purity was achieved, and the harvest reached about 40 ug/g of integument in the experiments. This work helps to obtaining natural sepiapterin in large amounts in order to use the lemon B. mori mutant to produce BH4 in vitro. PMID:24773269

Determination of Extraction Process Conditions of Gambier Catechin (Uncaria Gambier Roxb) from Solok Bio Bio Lima Puluh Kota District – West Sumatera

NASA Astrophysics Data System (ADS)

Desni Rahman, Elly; Sari, Ellyta; Burmawi; Frizka; Endah

2018-03-01

Catechin content is the determinant key of quality in gambier trade. The required Catechin content of gambier extracts as a herbal medicinal ingridient is greater than 90%. Mostly, Local gambier that produced by community is not uniform and low quality, thus lowering the price in the export markets. The quality improvement of gambier can be done by extraction and purification processes. This study aims to determine the best extraction process of catechin from Gambier (Uncaria Roxb) which derived from Solok Bio Bio Lima Puluh Kota, West Sumatra. The research methodology includes pre purification: raw materials preparation, washing, filtration, extraction, drying and testing. Washing was done on 100 gr gambier with a variation of water at 500, 600, 700, and 800 ml, heating for an hour at a temperature of 70°C, screened, filtered, and allow to stand until a precipitate is formed, wash repeatedly, filtered, and dried. Further, extract with a solvent variation of : water, etyl acetate, heated at 70°C temperature for 1 hour, then filtered. Filtrate then thickened by using a Rotary evaporator, dried at 50°C temperature for 48 hours and analyzed. The results showed that the best conditions of the extraction process is by using a solvent etyl acetate, at a temperature of 70°C, grading 97.40% catechins.

Preparation, purification, and characterization of aminopropyl-functionalized silica sol.

PubMed

Pálmai, Marcell; Nagy, Lívia Naszályi; Mihály, Judith; Varga, Zoltán; Tárkányi, Gábor; Mizsei, Réka; Szigyártó, Imola Csilla; Kiss, Teréz; Kremmer, Tibor; Bóta, Attila

2013-01-15

A new, simple, and "green" method was developed for the surface modification of 20 nm diameter Stöber silica particles with 3-aminopropyl(diethoxy)methylsilane in ethanol. The bulk polycondensation of the reagent was inhibited and the stability of the sol preserved by adding a small amount of glacial acetic acid after appropriate reaction time. Centrifugation, ultrafiltration, and dialysis were compared in order to choose a convenient purification technique that allows the separation of unreacted silylating agent from the nanoparticles without destabilizing the sol. The exchange of the solvent to acidic water during the purification yielded a stable colloid, as well. Structural and morphological analysis of the obtained aminopropyl silica was performed using transmission electron microscopy (TEM), dynamic light scattering (DLS) and zeta potential measurements, Fourier-transform infrared (FTIR), (13)C and (29)Si MAS nuclear magnetic resonance (NMR) spectroscopies, as well as small angle X-ray scattering (SAXS). Our investigations revealed that the silica nanoparticle surfaces were partially covered with aminopropyl groups, and multilayer adsorption followed by polycondensation of the silylating reagent was successfully avoided. The resulting stable aminopropyl silica sol (ethanolic or aqueous) is suitable for biomedical uses due to its purity. Copyright © 2012 Elsevier Inc. All rights reserved.

A multimodal histamine ligand for chromatographic purification of plasmid DNA.

PubMed

Černigoj, Urh; Vidic, Urška; Barut, Miloš; Podgornik, Aleš; Peterka, Matjaž; Štrancar, Aleš

2013-03-15

To exploit different chromatographic modes for efficient plasmid DNA (pDNA) purification a novel monolithic chromatographic support bearing multimodal histamine (HISA) groups was developed and characterized. Electrostatic charge of HISA groups depends on the pH of the mobile phase, being neutral above pH 7 and becoming positively charged below. As a consequence, HISA groups exhibit predominantly ion-exchange character at low pH values, which decreases with titration of the HISA groups resulting in increased hydrophobicity. This feature enabled separation of supercoiled (sc) pDNA from other plasmid isoforms (and other process related impurities) by adjusting salt or pH gradient. The dynamic binding capacity (DBC) for a 5.1kbp large plasmid at pH 5 was 4.0 mg/ml under low salt binding conditions, remaining relatively high (3.0 mg/ml) even in the presence of 1.0 M NaCl due to the multimodal nature of HISA ligand. Only slightly lower DBC (2.7 mg/ml) was determined under preferentially hydrophobic conditions in 3.0 M (NH(4))(2)SO(4), pH 7.4. Open circular and sc pDNA isoforms were baseline separated in descending (NH(4))(2)SO(4) gradient. Furthermore, an efficient plasmid DNA separation was possible both on analytical as well as on preparative scale by applying the descending pH gradient at a constant concentration (above 3.0 M) of (NH(4))(2)SO(4). Copyright © 2013 Elsevier B.V. All rights reserved.

Hanging drop crystal growth apparatus and method

NASA Technical Reports Server (NTRS)

Carter, Daniel C. (Inventor); Smith, Robbie E. (Inventor)

1989-01-01

An apparatus (10) is constructed having a cylindrical enclosure (16) within which a disc-shaped wicking element (18) is positioned. A well or recess (22) is cut into an upper side (24) of this wicking element, and a glass cover plate or slip (28) having a protein drop disposed thereon is sealably positioned on the wicking element (18), with drop (12) being positioned over well or recess (22). A flow of control fluid is generated by a programmable gradient former (16), with this control fluid having a vapor pressure that is selectively variable. This flow of control fluid is coupled to the wicking element (18) where control fluid vapor diffusing from walls (26) of the recess (22) is exposed to the drop (12), forming a vapor pressure gradient between the drop (12) and the control fluid vapor. Initially, this gradient is adjusted to draw solvent from the drop (12) at a relatively high rate, and as the critical supersaturation point is approached (the point at which crystal nucleation occurs), the gradient is reduced to more slowly draw solvent from the drop (12). This allows discrete protein molecules more time to orient themselves into an ordered crystalline lattice, producing protein crystals which, when processed by X-ray crystallography, possess a high degree of resolution.

PREPARATIVE ISOLATION AND PURIFICATION OF CHEMICAL CONSTITUENTS OF BELAMCANDA BY MPLC, HSCCC AND PREP-HPLC

PubMed Central

Wang, Xiaohong; Liang, Yong; Peng, Cuilin; Xie, Huichun; Pan, Man; Zhang, Tianyou; Ito, Yoichiro

2010-01-01

Combined with medium-pressure liquid chromatography (MPLC) and preparative high-pressure liquid chromatography (Prep-HPLC), high-speed countercurrent chromatography (HSCCC) was successfully applied for separation and purification of isoflavonoids from the extract of belamcanda. HSCCC separation was performed on a two-phase solvent system composed of methyl tert-butyl ether -ethyl acetate - n-butyl alcohol – acetonitrile −0.1% aqueous trifluoroacetic acid at a volume radio of 1:2:1:1:5. Semi-purified peak fractions from HSCCC separation were further purified by Prep-HPLC. Nine well-separated fractions were analyzed by HPLC-UV absorption spectrometry to determine their purities and characterized with ESI-MSn. Except for peaksland VII (unknown) seven compounds were identified as apocynin (peak II), mangiferin (peak III), 7-O-methylmangiferin (peak IV), hispidulin (peak V), 3′-hydroxyltectoridin (peak VI), iristectorin B (peak VII), isoiridin (peak IX). PMID:21552369

High-yield production of pure tagatose from fructose by a three-step enzymatic cascade reaction.

PubMed

Lee, Seon-Hwa; Hong, Seung-Hye; Kim, Kyoung-Rok; Oh, Deok-Kun

2017-08-01

To produce tagatose from fructose with a high conversion rate and to establish a high-yield purification method of tagatose from the reaction mixture. Fructose at 1 M (180 g l -1 ) was converted to 0.8 M (144 g l -1 ) tagatose by a three-step enzymatic cascade reaction, involving hexokinase, plus ATP, fructose-1,6-biphosphate aldolase, phytase, over 16 h with a productivity of 9 g l -1 h -1 . No byproducts were detected. Tagatose was recrystallized from ethanol to a purity of 99.9% and a yield of 96.3%. Overall, tagatose at 99.9% purity was obtained from fructose with a yield of 77%. This is the first biotechnological production of tagatose from fructose and the first application of solvent recrystallization for the purification of rare sugars.

Ethanolic extraction, purification, and partial characterization of a fluorescent toxin from the coral reef crab, Lophozozymus pictor.

PubMed

Lau, C O; Tan, C H; Khoo, H E; Li, Q T; Yuen, R

1995-01-01

A purification procedure for Lophozozymus pictor toxin (LPTX) following ethanolic extraction of whole crab homogenate is described. The ethanol-extracted toxin (LPTX-E) had higher yield and specific activity than the hot aqueous-extracted one (LPTX-H). It was found that LPTX-E was fluorescent and cochromatographed with LPTX-H on two-dimensional thin-layer chromatography. Although LPTX-E, LPTX-H, and palytoxin (P. caribaeorum, PTX) had similar migration and retention times when analysed on high performance capillary electrophoresis and gel permeation-high performance liquid chromatography respectively, LPTX-E and LPTX-H were both fluorescent in contrast to PTX. In addition, LPTX-E had a different retention time compared with PTX when chromatographed on reversed phase high performance liquid chromatography in the solvent system 80% acetonitrile and 0.02 M Tris-HCl, pH 7.2, at a 4:1 ratio, respectively, indicating some differences in their chemical structures.

Quadrupole Magnetic Sorting of Porcine Islets of Langerhans

PubMed Central

Shenkman, Rustin M.; Chalmers, Jeffrey J.; Hering, Bernhard J.; Kirchhof, Nicole

2009-01-01

Islet transplantation is emerging as a treatment option for selected patients with type 1 diabetes. Inconsistent isolation, purification, and recovery of large numbers of high-quality islets remain substantial impediments to progress in the field. Removing islets as soon as they are liberated from the pancreas during digestion and circumventing the need for density gradient purification is likely to result in substantially increased viable islet yields by minimizing exposure to proteolytic enzymes, reactive oxygen intermediates, and mechanical stress associated with centrifugation. This study capitalized on the hypervascularity of islets compared with acinar tissue to explore their preferential enrichment with magnetic beads to enable immediate separation in a magnetic field utilizing a quadrupole magnetic sorting. The results demonstrate that (1) preferential enrichment of porcine islets is achievable, but homogeneous bead distribution within the pancreas is difficult to achieve with current protocols; (2) greater than 70% of islets in the dissociated pancreatic tissue were recovered by quadrupole magnetic sorting, but their purity was low; and (3) infused islets purified by density gradients and subsequently passed through quadrupole magnetic sorting had similar potency as uninfused islets. These results demonstrate proof of concept and define the steps for implementation of this technology in pig and human islet isolation. PMID:19505179

In-pore exchange and diffusion of carbonate solvent mixtures in nanoporous carbon

DOE PAGES

Alam, Todd M.; Osborn Popp, Thomas M.

2016-06-04

High resolution magic angle spinning (HRMAS) 1H NMR spectroscopy has been used to resolve different surface and in-pore solvent environments of ethylene carbonate (EC) and dimethyl carbonate (DMC) mixtures absorbed within nanoporous carbon (NPC). Two dimensional (2D) 1H HRMAS NMR exchange measurements revealed that the inhomogeneous broadened in-pore resonances have pore-to-pore exchange rates on the millisecond timescale. Pulsed-field gradient (PFG) NMR diffusometry revealed the in-pore self-diffusion constants for both EC and DMC were reduced by up to a factor of five with respect to the diffusion in the non-absorbed solvent mixtures.

Determination of Levetiracetam in Human Plasma by Dispersive Liquid-Liquid Microextraction Followed by Gas Chromatography-Mass Spectrometry

PubMed Central

2016-01-01

Levetiracetam (LEV) is an antiepileptic drug that is clinically effective in generalized and partial epilepsy syndromes. The use of this drug has been increasing in clinical practice and intra- or -interindividual variability has been exhibited for special population. For this reason, bioanalytical methods are required for drug monitoring in biological matrices. So this work presents a dispersive liquid-liquid microextraction method followed by gas chromatography-mass spectrometry (DLLME-GC-MS) for LEV quantification in human plasma. However, due to the matrix complexity a previous purification step is required. Unlike other pretreatment techniques presented in the literature, for the first time, a procedure employing ultrafiltration tubes Amicon® (10 kDa porous size) without organic solvent consumption was developed. GC-MS analyses were carried out using a linear temperature program, capillary fused silica column, and helium as the carrier gas. DLLME optimized parameters were type and volume of extraction and dispersing solvents, salt addition, and vortex agitation time. Under chosen parameters (extraction solvent: chloroform, 130 μL; dispersing solvent: isopropyl alcohol, 400 μL; no salt addition and no vortex agitation time), the method was completely validated and all parameters were in agreement with the literature recommendations. LEV was quantified in patient's plasma sample using less than 550 μL of organic solvent. PMID:27830105

Isolation of zymogen granules from rat pancreas.

PubMed

Rindler, Michael J

2006-01-01

This unit describes methods for preparing zymogen granules from rat pancreas. Zymogen granules are storage organelles in pancreatic acinar cells containing digestive enzymes that are released into the pancreatic duct. The protocols in this unit take advantage of the large size (up to 1 microm diameter) and high density (>1.20 g/cm(3) on sucrose gradients) of the granules as compared to other cellular organelles. They use a combination of differential sedimentation and density gradient separation to accomplish the purification. Similar procedures can be used to isolate zymogen granules from mouse pancreas and canine pancreas. A protocol for preparing zymogen granules from dog pancreas is also included.

Purification of lymphocystis disease virus (LDV) grown in tissue culture. Evidences for the presence of two types of viral particles.

PubMed

Robin, J; Berthiaume, L

1981-12-01

Lymphocystis disease virus was highly purified from host cells by precipitation with PEG-6000 and isopycnic centrifugation in a metrizamide gradient. Metrizamide gradient centrifugation produce two distinct bands at equilibrium. As calculated from reconstruction experiments, only 4 and 0.3% respectively of the host DNA and the host proteins were recovered at the position of the bands. The final recovery of infectivity was about 41%. Electron microscopy of the bands showed two types of particles: small and dense particles measuring 100-150 nm and lymphocystis virions that measured about 300-350 nm in diameter.

Two tools for applying chromatographic retention data to the mass-based identification of peptides during hydrogen/deuterium exchange experiments by nano-liquid chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

PubMed

Gershon, P D

2010-12-15

Two tools are described for integrating LC elution position with mass-based data in hydrogen-deuterium exchange (HDX) experiments by nano-liquid chromatography/matrix-assisted laser desorption/ionization mass spectrometry (nanoLC/MALDI-MS, a novel approach to HDX-MS). The first of these, 'TOF2H-Z Comparator', highlights peptides in HDX experiments that are potentially misidentified on the basis of mass alone. The program first calculates normalized values for the organic solvent concentration responsible for the elution of ions in nanoLC/MALDI HDX experiments. It then allows the solvent gradients for the multiple experiments contributing to an MS/MS-confirmed peptic peptide library to be brought into mutual alignment by iteratively re-modeling variables among LC parameters such as gradient shape, solvent species, fraction duration and LC dead time. Finally, using the program, high-probability chromatographic outliers can be flagged within HDX experimental data. The role of the second tool, 'TOF2H-XIC Comparator', is to normalize the LC chromatograms corresponding to all deuteration timepoints of all HDX experiments of a project, to a common reference. Accurate normalization facilitates the verification of chromatographic consistency between all ions whose spectral segments contribute to particular deuterium uptake plots. Gradient normalization in this manner revealed chromatographic inconsistencies between ions whose masses were either indistinguishable or separated by precise isotopic increments. Copyright © 2010 John Wiley & Sons, Ltd.

On the feasibility to conduct gradient liquid chromatography separations in narrow-bore columns at pressures up to 2000bar.

PubMed

De Pauw, Ruben; Swier, Tim; Degreef, Bart; Desmet, Gert; Broeckhoven, Ken

2016-11-18

The limits in operating pressures are extended for narrow-bore columns in gradient elution up to 2000bar. As the required pumps for these pressures are incompatible with common chromatographic solvents and are not suitable to apply a mobile phase composition gradient, a mobile phase delivery and injection system is described and experimentally validated which allows to use any possible chromatographic solvent in isocratic and gradient elution. The mobile phase delivery and injection system also allows to perform multiple separations without the need to depressurize the column. This system consists out of 5 dual on/off valves and two large volume loops in which the gradient and equilibration volume of initial mobile phase are loaded by a commercial liquid chromatography pump. The loops are then flushed toward the column at extreme pressures. The mobile phase delivery and injection system is first evaluated in isocratic elution and shows a comparable performance to a state-of-the-art commercial flow-through-needle injector but with twice the pressure rating. Distortion of the loaded gradient by dispersion in the gradient storage loop is studied. The effect of the most important parameters (such as flow rate, pressure and gradient steepness) is experimentally investigated. Different gradient steepnesses and volumes can be applied at different flow rates and operating pressures with a good repeatability. Due to the isobaric operation of the pumps, the gradient is monitored in real-time by a mass flow meter installed at the detector outlet. The chromatograms are then converted from time to volume-base. A separation of a 19-compound sample is performed on a 300×2.1mm column at 1000bar and on a 600×2.1mm column at 2000bar. The peak capacity was found to increase from 141 to 199 and thus scales with L as is predicted by theory. This allows to conclude that the inlet pressure for narrow-bore columns in gradient elution can be increased up to 2000bar without fundamental pressure-induced limitations. Copyright © 2016 Elsevier B.V. All rights reserved.

Synthesis of 2-monoacylglycerols and structured triacylglycerols rich in polyunsaturated fatty acids by enzyme catalyzed reactions.

PubMed

Rodríguez, Alicia; Esteban, Luis; Martín, Lorena; Jiménez, María José; Hita, Estrella; Castillo, Beatriz; González, Pedro A; Robles, Alfonso

2012-08-10

This paper studies the synthesis of structured triacylglycerols (STAGs) by a four-step process: (i) obtaining 2-monoacylglycerols (2-MAGs) by alcoholysis of cod liver oil with several alcohols, catalyzed by lipases Novozym 435, from Candida antartica and DF, from Rhizopus oryzae, (ii) purification of 2-MAGs, (iii) formation of STAGs by esterification of 2-MAGs with caprylic acid catalyzed by lipase DF, from R. oryzae, and (iv) purification of these STAGs. For the alcoholysis of cod liver oil, absolute ethanol, ethanol 96% (v/v) and 1-butanol were compared; the conditions with ethanol 96% were then optimized and 2-MAG yields of around 54-57% were attained using Novozym 435. In these 2-MAGs, DHA accounted for 24-31% of total fatty acids. In the operational conditions this lipase maintained a stable level of activity over at least 11 uses. These results were compared with those obtained with lipase DF, which deactivated after only three uses. The alcoholysis of cod liver oil and ethanol 96% catalyzed by Novozym 435 was scaled up by multiplying the reactant amounts 100-fold and maintaining the intensity of treatment constant (IOT=3g lipase h/g oil). In these conditions, the 2-MAG yield attained was about 67%; these 2-MAGs contained 36.6% DHA. The synthesized 2-MAGs were separated and purified from the alcoholysis reaction products by solvent extraction using solvents of low toxicity (ethanol and hexane); 2-MAG recovery yield and purity of the target product were approximately 96.4% and 83.9%, respectively. These 2-MAGs were transformed to STAGs using the optimal conditions obtained in a previous work. After synthesis and purification, 93% pure STAGs were obtained, containing 38% DHA at sn-2 position and 60% caprylic acid (CA) at sn-1,3 positions (of total fatty acids at these positions), i.e. the major TAG is the STAG with the structure CA-DHA-CA. Copyright © 2012 Elsevier Inc. All rights reserved.

The chromatography-free release, isolation and purification of recombinant peptide for fibril self-assembly.

PubMed

Hartmann, B M; Kaar, W; Yoo, I K; Lua, L H L; Falconer, R J; Middelberg, A P J

2009-12-01

One of the major expenses associated with recombinant peptide production is the use of chromatography in the isolation and purification stages of a bioprocess. Here we report a chromatography-free isolation and purification process for recombinant peptide expressed in Escherichia coli (E. coli). Initial peptide release is by homogenization and then by enzymatic cleavage of the peptide-containing fusion protein, directly in the E. coli homogenate. Release is followed by selective solvent precipitation (SSP) to isolate and purify the peptide away from larger cell contaminants. Specifically, we expressed in E. coli the self-assembling beta-sheet forming peptide P(11)-2 in fusion to thioredoxin. Homogenate was heat treated (55 degrees C, 15 min) and then incubated with tobacco etch virus protease (TEVp) to release P(11)-2 having a native N-terminus. SSP with ethanol at room temperature then removed contaminating proteins in an integrated isolation-purification step; it proved necessary to add 250 mM NaCl to homogenate to prevent P(11)-2 from partitioning to the precipitate. This process structure gave recombinant P(11)-2 peptide at 97% polypeptide purity and 40% overall yield, without a single chromatography step. Following buffer-exchange of the 97% pure product by bind-elute chromatography into defined chemical conditions, the resulting peptide was shown to be functionally active and able to form self-assembled fibrils. To the best of our knowledge, this manuscript reports the first published process for chromatography-free recombinant peptide release, isolation and purification. The process proved able to deliver functional recombinant peptide at high purity and potentially low cost, opening cost-sensitive materials applications for peptide-based materials.

Purification process for .sup.153Gd produced in natural europium targets

DOEpatents

Johnsen, Amanda M; Soderquist, Chuck Z; McNamara, Bruce K; Risher, Darrell R

2013-04-23

An alteration of the traditional zinc/zinc-amalgam reduction procedure which eliminates both the hazardous mercury and dangerous hydrogen gas generation. In order to avoid the presence of water and hydrated protons in the working solution, which can oxidize Eu.sup.2+ and cause hydrogen gas production, a process utilizing methanol as the process solvent is described. While methanol presents some flammability hazard in a radiological hot cell, it can be better managed and is less of a flammability hazard than hydrogen gas generation.

Comparative study of two methods of fractionation bromelain from pineapple core extract (Ananas comosus)

NASA Astrophysics Data System (ADS)

Febriani, K.; Wahyuni, I.; Setiasih, S.; Hudiyono, S.

2017-07-01

The enzyme can be purified by fractional precipitation. This can be done by salt or organic solvent. In this research, purification of bromelain from pineapple core by fractional precipitation was done by 2 compounds, ammonium sulfate, and ethanol. Fractional precipitation by ammonium sulfate proved to be more effective as it yielded a higher specific activity. Specific activity by ethanol and ammonium sulfate is 4.6480 U/mg at 0-60 % saturation and 8.2243 U/mg at 50-80 % saturation.

An eco-compatible strategy for the diversity-oriented synthesis of macrocycles exploiting carbohydrate-derived building blocks.

PubMed

Maurya, Sushil K; Rana, Rohit

2017-01-01

An efficient, eco-compatible diversity-oriented synthesis (DOS) approach for the generation of library of sugar embedded macrocyclic compounds with various ring size containing 1,2,3-triazole has been developed. This concise strategy involves the iterative use of readily available sugar-derived alkyne/azide-alkene building blocks coupled through copper catalyzed azide-alkyne cycloaddition (CuAAC) reaction followed by pairing of the linear cyclo-adduct using greener reaction conditions. The eco-compatibility, mild reaction conditions, greener solvents, easy purification and avoidance of hazards and toxic solvents are advantages of this protocol to access this important structural class. The diversity of the macrocycles synthesized (in total we have synthesized 13 macrocycles) using a set of standard reaction protocols demonstrate the potential of the new eco-compatible approach for the macrocyclic library generation.

Tested Demonstrations.

ERIC Educational Resources Information Center

Gilbert, George L., Ed.

1985-01-01

Background information, procedures, and typical results obtained are provided for two demonstrations. The first involves the colorful complexes of copper(II). The second involves reverse-phase separation of Food, Drug, and Cosmetic (FD & C) dyes using a solvent gradient. (JN)

Proof of concept of a "greener" protein purification/enrichment method based on carboxylate-terminated carbosilane dendrimer-protein interactions.

PubMed

González-García, Estefanía; Maly, Marek; de la Mata, Francisco Javier; Gómez, Rafael; Marina, María Luisa; García, María Concepción

2016-11-01

Protein sample preparation is a critical and an unsustainable step since it involves the use of tedious methods that usually require high amount of solvents. The development of new materials offers additional opportunities in protein sample preparation. This work explores, for the first time, the potential application of carboxylate-terminated carbosilane dendrimers to the purification/enrichment of proteins. Studies on dendrimer binding to proteins, based on protein fluorescence intensity and emission wavelengths measurements, demonstrated the interaction between carboxylate-terminated carbosilane dendrimers and proteins at all tested pH levels. Interactions were greatly affected by the protein itself, pH, and dendrimer concentration and generation. Especially interesting was the interaction at acidic pH since it resulted in a significant protein precipitation. Dendrimer-protein interactions were modeled observing stable complexes for all proteins. Carboxylate-terminated carbosilane dendrimers at acidic pH were successfully used in the purification/enrichment of proteins extracted from a complex sample. Graphical Abstract Images showing the growing turbidity of solutions containing a mixture of proteins (lysozyme, myoglobin, and BSA) at different protein:dendrimer ratios (1:0, 1:1, 1:8, and 1:20) at acidic pH and SDS-PAGE profiles of the corresponsing supernatants. Comparison of SDS-PAGE profiles for the pellets obtained during the purification of proteins present in a complex sample using a conventional "no-clean" method based on acetone precipitation and the proposed "greener" method using carboxylate-terminated carbosilane dendrimer at a 1:20 protein:dendrimer ratio.

Preparative isolation and purification of four flavonoids from the petals of Nelumbo nucifera by high-speed counter-current chromatography.

PubMed

Xingfeng, Guo; Daijie, Wang; Wenjuan, Duan; Jinhua, Du; Xiao, Wang

2010-01-01

Flavonoids, the primary constituents of the petals of Nelumbo nucifera, are known to have antioxidant properties and antibacterial bioactivities. However, efficient methods for the preparative isolation and purification of flavonoids from this plant are not currently available. To develop an efficient method for the preparative isolation and purification of flavonoids from the petals of N. nucifera by high-speed counter-current chromatography (HSCCC). Following an initial clean-up step on a polyamide column, HSCCC was utilised to separate and purify flavonoids. Purities and identities of the isolated compounds were established by HPLC-PAD, ESI-MS, (1)H-NMR and (13)C-NMR. The separation was performed using a two-phase solvent system composed of ethyl acetate-methanol-water-acetic acid (4 : 1 : 5 : 0.1, by volume), in which the upper phase was used as the stationary phase and the lower phase was used as the mobile phase at a flow-rate of 1.0 mL/min in the head-to-tail elution mode. Ultimately, 5.0 mg syringetin-3-O-beta-d-glucoside, 6.5 mg quercetin-3-O-beta-d-glucoside, 12.8 mg isorhamnetin-3-O-beta-d-glucoside and 32.5 mg kaempferol-3-O-beta-d-glucoside were obtained from 125 mg crude sample. The combination of HSCCC with a polyamide column is an efficient method for the preparative separation and purification of flavonoids from the petals of N. nucifera. (c) 2009 John Wiley & Sons, Ltd.

Purification and characterization of haloalkaline thermoactive, solvent stable and SDS-induced protease from Bacillus sp.: a potential additive for laundry detergents.

PubMed

Jain, Deepti; Pancha, Imran; Mishra, Sanjiv K; Shrivastav, Anupama; Mishra, Sandhya

2012-07-01

An extracellular haloalkaline, thermoactive, solvent stable, SDS-induced serine protease was purified and characterized from an alkali-thermo tolerant strain Bacillus sp. SM2014 isolated from reverse osmosis reject. The enzyme was purified to homogeneity with recovery of 54.4% and purity fold of 64. The purified enzyme was composed of single polypeptide of molecular mass about 71 kDa. The enzyme showed optimum activity at alkaline pH 10 and temperature 60°C. The km and Vmax for the enzyme was 0.57 mg/ml and 445.23 U/ml respectively. The enzyme showed novel catalytic ability at high pH (10), temperature (60°C) and salinity (3M). Moreover, the stability of enzyme in organic solvents (50% v/v) of logP ≥ 2 signified the prospective of this enzyme for peptide synthesis. The compatibility of the enzyme with surfactants and various detergent matrices together with wash performance test confirmed its potential applicability in laundry industry. Copyright © 2011 Elsevier Ltd. All rights reserved.

Simulation of elution profiles in liquid chromatography - II: Investigation of injection volume overload under gradient elution conditions applied to second dimension separations in two-dimensional liquid chromatography.

PubMed

Stoll, Dwight R; Sajulga, Ray W; Voigt, Bryan N; Larson, Eli J; Jeong, Lena N; Rutan, Sarah C

2017-11-10

An important research direction in the continued development of two-dimensional liquid chromatography (2D-LC) is to improve the detection sensitivity of the method. This is especially important in applications where injection of large volumes of effluent from the first dimension ( 1 D) column into the second dimension ( 2 D) column leads to severe 2 D peak broadening and peak shape distortion. For example, this is common when coupling two reversed-phase columns and the organic solvent content of the 1 D mobile phase overwhelms the 2 D column with each injection of 1 D effluent, leading to low resolution in the second dimension. In a previous study we validated a simulation approach based on the Craig distribution model and adapted from the work of Czok and Guiochon [1] that enabled accurate simulation of simple isocratic and gradient separations with very small injection volumes, and isocratic separations with mismatched injection and mobile phase solvents [2]. In the present study we have extended this simulation approach to simulate separations relevant to 2D-LC. Specifically, we have focused on simulating 2 D separations where gradient elution conditions are used, there is mismatch between the sample solvent and the starting point in the gradient elution program, injection volumes approach or even exceed the dead volume of the 2 D column, and the extent of sample loop filling is varied. To validate this simulation we have compared results from simulations and experiments for 101 different conditions, including variation in injection volume (0.4-80μL), loop filling level (25-100%), and degree of mismatch between sample organic solvent and the starting point in the gradient elution program (-20 to +20% ACN). We find that that the simulation is accurate enough (median errors in retention time and peak width of -1.0 and -4.9%, without corrections for extra-column dispersion) to be useful in guiding optimization of 2D-LC separations. However, this requires that real injection profiles obtained from 2D-LC interface valves are used to simulate the introduction of samples into the 2 D column. These profiles are highly asymmetric - simulation using simple rectangular pulses leads to peak widths that are far too narrow under many conditions. We believe the simulation approach developed here will be useful for addressing practical questions in the development of 2D-LC methods. Copyright © 2017 Elsevier B.V. All rights reserved.

Application of a chromatography model with linear gradient elution experimental data to the rapid scale-up in ion-exchange process chromatography of proteins.

PubMed

Ishihara, Takashi; Kadoya, Toshihiko; Yamamoto, Shuichi

2007-08-24

We applied the model described in our previous paper to the rapid scale-up in the ion exchange chromatography of proteins, in which linear flow velocity, column length and gradient slope were changed. We carried out linear gradient elution experiments, and obtained data for the peak salt concentration and peak width. From these data, the plate height (HETP) was calculated as a function of the mobile phase velocity and iso-resolution curve (the separation time and elution volume relationship for the same resolution) was calculated. The scale-up chromatography conditions were determined by the iso-resolution curve. The scale-up of the linear gradient elution from 5 to 100mL and 2.5L column sizes was performed both by the separation of beta-lactoglobulin A and beta-lactoglobulin B with anion-exchange chromatography and by the purification of a recombinant protein with cation-exchange chromatography. Resolution, recovery and purity were examined in order to verify the proposed method.

Biodiesel production from ethanolysis of DPO using deep eutectic solvent (DES) based choline chloride - ethylene glycol as co-solvent

NASA Astrophysics Data System (ADS)

Taslim, Indra, Leonardo; Manurung, Renita; Winarta, Agus; Ramadhani, Debbie Aditia

2017-03-01

Biodiesel is usually produced from transesterification using methanol or ethanol as alcohol. However, biodiesel produced using methanol has several disadvantages because methanol is toxic and not entirely bio-based as it is generally produced from petroleum, natural gas and coal. On the other hand, ethanol also has several disadvantages such as lower reactivity in transesterification process and formation of stable emulsion between ester and glycerol. To improve ethanolysis process, deep eutectic solvent (DES) was prepared from choline chloride and ethylene glycol to be used as co-solvent in ethanolysis. Deep eutectic solvent was prepared by mixing choline chloride and ethylene glycol at molar ratio of 1:2, temperature of 80 °C, and stirring speed of 300 rpm for 1 hour. The DES was characterized by its density and viscosity. The ethanolysis of DPO / Degummed Palm Oil was performed at 70 °C, ethanol to oil molar ratio of 9:1, catalyst (potassium hydroxide) concentration of 0.75 wt.% concentration, co-solvent (DES) concentration of 1, 2, 3, 4, 5 and 6 wt.%, stirring speed of 600 rpm, and reaction time of 1 hour. The obtained biodiesel was then characterized by its density, viscosity and ester content. The oil - ethanol phase condition was observed in reaction tube. The oil - ethanol phase with DES tends to form meniscus compared to that without DES. Which implied that oil and ethanol become more slightly miscible, which favours the reaction. Using DES as co-solvent in ethanolysis resulted in an increase in yield and easier purification. The esters properties met the international standards ASTM D6751, with highest yield achieved at 81.72 % with 99.35 % ethyl ester contents at 4% DES concentration.

Purification of Giardia muris cysts by velocity sedimentation.

PubMed Central

Sauch, J F

1984-01-01

Giardia muris cysts were separated from fecal contaminants in primary isolates by unit gravity velocity sedimentation. Crude isolates obtained by centrifugation over 1.0 M sucrose were overlaid onto a Percoll density gradient, 1.01 to 1.03 g/ml. G. muris cysts were well separated from faster-sedimenting fecal debris and slower-sedimenting Spironucleus muris and bacteria in 1.5 h. PMID:6486790

Production of Biodiesel from Candlenut Oil Using a Two-step Co-solvent Method and Evaluation of Its Gaseous Emissions.

PubMed

Pham, Lan Ngoc; Luu, Boi Van; Phuoc, Hung Duong; Le, Hanh Ngoc Thi; Truong, Hoa Thi; Luu, Phuong Duc; Furuta, Masakazu; Imamura, Kiyoshi; Maeda, Yasuaki

2018-05-01

Candlenut oil (CNO) is a potentially new feedstock for biodiesel (BDF) production. In this paper, a two-step co-solvent method for BDF production from CNO was examined. Firstly, esterification of free fatty acids (FFAs) (7 wt%) present in CNO was carried out using a co-solvent of acetonitrile (30 wt%) and H 2 SO 4 as a catalyst. The content of FFAs was reduced to 0.8 wt% in 1 h at 65°C. Subsequent transesterification of the crude oil produced was carried out using a co-solvent of acetone (20 wt%) and 1 wt% potassium hydroxide (KOH). Ester content of 99.3% was obtained at 40°C in 45 min. The water content in BDF was 0.023% upon purification using vacuum distillation at 5 kPa. The components of CNO BDF were characterized using a Fourier-transform infrared spectrometry and gas chromatography-flame ionization detector. The physicochemical properties of BDF satisfied the ASTM D6751-02 standard. The gaseous exhaust emissions from the diesel engine upon combustion of the BDF blends (B0-B100) with petrodiesel were examined. The emissions of carbon monoxide and hydrocarbons were clearly lower, but that of nitrogen oxides was higher in comparison to those from petro-diesel.

Design of protonation constant measurement apparatus for carbon dioxide capturing solvents

NASA Astrophysics Data System (ADS)

Ma'mun, S.; Amelia, E.; Rahmat, V.; Alwani, D. R.; Kurniawan, D.

2016-11-01

Global warming phenomenon has led to world climate change caused by high concentrations of greenhouse gases (GHG), e.g. carbon dioxide (CO2), in the atmosphere. Carbon dioxide is produced in large amount from coal-fired power plants, iron and steel production, cement production, chemical and petrochemical manufacturing, natural gas purification, and transportation. Carbon dioxide emissions seem to rise from year to year; some efforts to reduce the emissions are, therefore, required. Amine-based absorption could be deployed for post-combustion capture. Some parameters, e.g. mass transfer coefficients and chemical equilibrium constants, are required for a vapor-liquid equilibrium modeling. Protonation constant (pKa), as one of those parameters, could then be measured experimentally. Therefore, an experimental setup to measure pKa of CO2 capturing solvents was designed and validated by measuring the pKa of acetic acid at 30 to 70 °C by a potentiometric titration method. The set up was also used to measure the pKa of MEA at 27 °C. Based on the validation results and due to low vapor pressure of CO2 capturing solvents in general, e.g. alkanolamines, the setup could therefore be used for measuring pKa of the CO2 capturing solvents at temperatures up to 70 °C.

Dip coating of sol-gels

NASA Astrophysics Data System (ADS)

Schunk, P. R.; Hurd, A. J.; Brinker, C. J.

Dip coating is the primary means of depositing sol-gel films for precision optical coatings. Sols are typically multicomponent systems consisting of an inorganic phase dispersed in a solvent mixture, with each component differing in volatility and surface tension. This, together with slow coating speeds (less than 1cm/s), makes analysis of the coating process complicated; unlike most high-speed coating methods, solvent evaporation, evolving rheology, and surface tension gradients alter significantly the fluid mechanics of the deposition stage. These phenomena were studied with computer-aided predictions of the flow and species transport fields. The underlying theory involves mass, momentum, and species transport on a domain of unknown shape, with models and constitutive equations for vapor-liquid equilibria and surface tension. Due accounting is made for the unknown position of the free surface, which locates according to the capillary hydrodynamic forces and solvent loss by evaporation. Predictions of the effects of mass transfer, hydrodynamics, and surface tension gradients on final film thickness are compared with ellipsometry measurements of film thickness on a laboratory pilot coater. Although quantitative agreement is still lacking, both experiment and theory reveal that the film profile near the drying line takes on a parabolic shape.

Solvent Flux Method (SFM): A Case Study of Water Access to Candida antarctica Lipase B.

PubMed

Benson, Sven P; Pleiss, Jürgen

2014-11-11

The solvent flux method (SFM) was developed to comprehensively characterize the influx of solvent molecules from the solvent environment into the active site of a protein in the framework of molecular dynamics simulations. This was achieved by introducing a solvent concentration gradient as well as partially reorienting and rescaling the velocity vector of all solvent molecules contained within a spherical volume enclosing the protein, thus inducing an accelerated solvent influx toward the active site. In addition to the detection of solvent access pathway within the protein structure, it is hereby possible to identify potential amino acid positions relevant to solvent-related enzyme engineering with high statistical significance. The method is particularly aimed at improving the reverse hydrolysis reaction rates in nonaqueous media. Candida antarctica lipase B (CALB) binds to a triglyceride-water interface with its substrate entrance channel oriented toward the hydrophobic substrate interface. The lipase-triglyceride-water system served as a model system for SFM to evaluate the influx of water molecules to the active site. As a proof of principle for SFM, a previously known water access pathway in CALB was identified as the primary water channel. In addition, a secondary water channel and two pathways for water access which contribute to water leakage between the protein and the triglyceride-water interface were identified.

Removal of PCB and Other Halogenated Organic Contaminants found in Ex Situ Structures

NASA Technical Reports Server (NTRS)

Quinn, Jacqueline (Inventor); Clausen, Christian (Inventor); Geiger, Cherie L. (Inventor); Coon, Christina (Inventor); Berger, Cristina M. (Inventor); Filipek, Laura B. (Inventor); Milum, Kristen M. (Inventor)

2007-01-01

Emulsified systems or a surfactant-stabilized, biodegradable water-in-solvent emulsion with bimetallic particles contained with the emulsion droplets are useful at removing PCBs from ex situ structures. The hydrophobic emulsion system draws PCBs through the solvent/surfactant membrane. Once inside the membrane, the PCBs diffuse into the bimetallic particles and undergo degradation. The PCBs continue to enter, diffuse, degrade, and biphenyl will exit the particle maintaining a concentration gradient across the membrane and maintaining a driving force of the reaction.

Removal of PCB and other halogenated organic contaminants found in ex situ structures

NASA Technical Reports Server (NTRS)

Quinn, Jacqueline (Inventor); Clausen, Christian (Inventor); Geiger, Cherie L. (Inventor); Coon, Christina (Inventor); Berger, Cristina M. (Inventor); Filipek, Laura B. (Inventor); Milum, Kristen M. (Inventor)

2007-01-01

Emulsified systems of a surfactant-stabilized, biodegradable water-in-solvent emulsion with bimetallic particles contained with the emulsion droplets are useful at removing PCBs from ex situ structures. The hydrophobic emulsion system draws PCBs through the solvent/surfactant membrane. Once inside the membrane, the PCBs diffuse into the bimetallic particles and undergo degradation. The PCBs continue to enter, diffuse, degrade, and biphenyl will exit the particle maintaining a concentration gradient across the membrane and maintaining a driving force of the reaction.

Removal of PCB and other halogenated organic contaminants found in ex situ structures

NASA Technical Reports Server (NTRS)

Clausen, Christian A. (Inventor); Milum, Kristen M. (Inventor); Quinn, Jacqueline W. (Inventor); Berger, Cristina M. (Inventor); Geiger, Cherie L. (Inventor); Filipek, Laura B. (Inventor); Coon, Christina (Inventor)

2009-01-01

Emulsified systems of a surfactant-stabilized, biodegradable water-in-solvent emulsion with bimetallic particles contained with the emulsion droplets are useful at removing PCBs from ex situ structures. The hydrophobic emulsion system draws PCBs through the solvent/surfactant membrane. Once inside the membrane, the PCBs diffuse into the bimetallic particles and undergo degradation. The PCBs continue to enter, diffuse, degrade, and biphenyl will exit the particle maintaining a concentration gradient across the membrane and maintaining a driving force of the reaction.

Semi-Automated Hydrophobic Interaction Chromatography Column Scouting Used in the Two-Step Purification of Recombinant Green Fluorescent Protein

PubMed Central

Murphy, Patrick J. M.

2014-01-01

Background Hydrophobic interaction chromatography (HIC) most commonly requires experimental determination (i.e., scouting) in order to select an optimal chromatographic medium for purifying a given target protein. Neither a two-step purification of untagged green fluorescent protein (GFP) from crude bacterial lysate using sequential HIC and size exclusion chromatography (SEC), nor HIC column scouting elution profiles of GFP, have been previously reported. Methods and Results Bacterial lysate expressing recombinant GFP was sequentially adsorbed to commercially available HIC columns containing butyl, octyl, and phenyl-based HIC ligands coupled to matrices of varying bead size. The lysate was fractionated using a linear ammonium phosphate salt gradient at constant pH. Collected HIC eluate fractions containing retained GFP were then pooled and further purified using high-resolution preparative SEC. Significant differences in presumptive GFP elution profiles were observed using in-line absorption spectrophotometry (A395) and post-run fluorimetry. SDS-PAGE and western blot demonstrated that fluorometric detection was the more accurate indicator of GFP elution in both HIC and SEC purification steps. Comparison of composite HIC column scouting data indicated that a phenyl ligand coupled to a 34 µm matrix produced the highest degree of target protein capture and separation. Conclusions Conducting two-step protein purification using the preferred HIC medium followed by SEC resulted in a final, concentrated product with >98% protein purity. In-line absorbance spectrophotometry was not as precise of an indicator of GFP elution as post-run fluorimetry. These findings demonstrate the importance of utilizing a combination of detection methods when evaluating purification strategies. GFP is a well-characterized model protein, used heavily in educational settings and by researchers with limited protein purification experience, and the data and strategies presented here may aid in development other of HIC-compatible protein purification schemes. PMID:25254496

The Purification and Concentration of Hog Cholera Virus*

PubMed Central

Cunliffe, H. R.; Rebers, P. A.

1968-01-01

Partial purification of hog cholera virus (HCV) using a simple batch-type chromatographic procedure with magnetic ferric oxide (MFO) is described. Infectious HCV was adsorbed from isotonic solutions to MFO and was eluted under conditions of low ionic strength and high pH. Aqueous solutions of 0.01 M sodium cyanide or 0.0003 M ammonium hydroxide effectively dissociated MFO-HCV complexes. The data indicate that 50 to 100% of the original HCV infectivity was recovered concomitant with a 90 to 95% reduction of extraneous organic nitrogen. MFO-purified HCV was concentrated by density gradient type centrifugations in buffered solutions of cesium chloride and sucrose. Prolonged isodensity centrifugations of concentrated MFO-purified HCV indicated a buoyant density of 1.14 to 1.15 gm/ml for the strain of virus used. PMID:15846899

Facile Purification of Milligram to Gram Quantities of Condensed Tannins According to Mean Degree of Polymerization and Flavan-3-ol Subunit Composition.

PubMed

Brown, Ron H; Mueller-Harvey, Irene; Zeller, Wayne E; Reinhardt, Laurie; Stringano, Elisabetta; Gea, An; Drake, Christopher; Ropiak, Honorata M; Fryganas, Christos; Ramsay, Aina; Hardcastle, Emily E

2017-09-13

Unambiguous investigation of condensed tannin (CT) structure-activity relationships in biological systems requires well-characterized, high-purity CTs. Sephadex LH-20 and Toyopearl HW-50F resins were compared for separating CTs from acetone/water extracts, and column fractions analyzed for flavan-3-ol subunits, mean degree of polymerization (mDP), and purity. Toyopearl HW-50F generated fractions with higher mDP values and better separation of procyanidins (PC) and prodelphinidins (PD) but required a prepurification step, needed more time for large scale purifications, and gave poorer recoveries. Therefore, two gradient elution schemes were developed for CT purification on Sephadex LH-20 providing 146-2000 mg/fraction. Fractions were analyzed by thiolysis and NMR spectroscopy. In general, PC/PD ratios decreased and mDP increased during elution. 1 H NMR spectroscopy served as a rapid screening tool to qualitatively determine CT enrichment and carbohydrate impurities present, guiding fractionation toward repurification or 1 H- 13 C HSQC NMR spectroscopy and thiolysis. These protocols provide options for preparing highly pure CT samples.

Simple and Efficient Purification of Recombinant Proteins Using the Heparin-Binding Affinity Tag.

PubMed

Jayanthi, Srinivas; Gundampati, Ravi Kumar; Kumar, Thallapuranam Krishnaswamy Suresh

2017-11-01

Heparin, a member of the glycosaminoglycan family, is known to interact with more than 400 different types of proteins. For the past few decades, significant progress has been made to understand the molecular details involved in heparin-protein interactions. Based on the structural knowledge available from the FGF1-heparin interaction studies, we have designed a novel heparin-binding peptide (HBP) affinity tag that can be used for the simple, efficient, and cost-effective purification of recombinant proteins of interest. HBP-tagged fusion proteins can be purified by heparin Sepharose affinity chromatography using a simple sodium chloride gradient to elute the bound fusion protein. In addition, owing to the high density of positive charges on the HBP tag, recombinant target proteins are preferably expressed in their soluble forms. The purification of HBP-fusion proteins can also be achieved in the presence of chemical denaturants, including urea. Additionally, polyclonal antibodies raised against the affinity tag can be used to detect HBP-fused target proteins with high sensitivity. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

Preparative isolation and purification of capsaicin and dihydrocapsaicin from Capsici Fructus using supercritical fluid extraction combined with high speed countercurrent chromatography.

PubMed

Yan, Rongwei; Zhao, Leilei; Tao, Junfei; Zou, Yong; Xu, Xinjun

2018-05-01

Supercritical fluid extraction with CO 2 (SFE-CO 2 ) was utilized for extraction of capsaicin (CA) and dihydrocapsaicin (DHCA) from Capsici Fructus, and then a two-step enrichment method for separating capsaicinoids from SFE-CO 2 extracts was developed. The process involved extraction with aqueous methanol and crystallization by alkali extraction and acid precipitation. Finally, a consecutive high-speed countercurrent chromatography (HSCCC) separation method was successfully applied in the purification of CA and DHCA from capsaicinoid crystal. The extraction pressure, extraction temperature and volume of co-solvent were optimized at 33 MPa, 41 °C and 75 mL, respectively, using response surface methodology; the extraction rates of CA and DHCA were about 93.18% and 93.49%, respectively. 407.43 mg capsaicinoid crystal was isolated from the SFE-CO 2 extracts obtained from 100 g capsicum powder by the two-step enrichment method. About 506 mg and 184 mg CA and DHCA with purities up to 98.31% and 96.68%, respectively, were obtained from 1 g capsaicinoid crystal in one HSCCC of three consecutive sample loadings without exchanging any solvent system. This method comprising SFE-CO 2 , a two-step enrichment and HSCCC was efficient, powerful and practical for the large-scale preparation of CA and DHCA from Capsici Fructus with high purity and high yield. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Versatile solvent systems for the separation of betalains from processed Beta vulgaris L. juice using counter-current chromatography.

PubMed

Spórna-Kucab, Aneta; Ignatova, Svetlana; Garrard, Ian; Wybraniec, Sławomir

2013-12-15

Two mixtures of decarboxylated and dehydrogenated betacyanins from processed red beet roots (Beta vulgaris L.) juice were fractionated by high performance counter-current chromatography (HPCCC) producing a range of isolated components. Mixture 1 contained mainly betacyanins, 14,15-dehydro-betanin (neobetanin) and their decarboxylated derivatives while mixture 2 consisted of decarboxy- and dehydro-betacyanins. The products of mixture 1 arose during thermal degradation of betanin/isobetanin in mild conditions while the dehydro-betacyanins of mixture 2 appeared after longer heating of the juice from B. vulgaris L. Two solvent systems were found to be effective for the HPCCC. A highly polar, high salt concentration system of 1-PrOH-ACN-(NH4)2SO4 (satd. soln)-water (v/v/v/v, 1:0.5:1.2:1) (tail-to-head mode) enabled the purification of 2-decarboxy-betanin/-isobetanin, 2,17-bidecarboxy-betanin/-isobetanin and neobetanin (all from mixture 1) plus 17-decarboxy-neobetanin, 2,15,17-tridecarboxy-2,3-dehydro-neobetanin, 2-decarboxy-neobetanin and 2,15,17-tridecarboxy-neobetanin (from mixture 2). The other solvent system included heptafluorobutyric acid (HFBA) as ion-pair reagent and consisted of tert-butyl methyl ether (TBME)-1-BuOH-ACN-water (acidified with 0.7% HFBA) (2:2:1:5, v/v/v/v) (head-to-tail mode). This system enabled the HPCCC purification of 2,17-bidecarboxy-betanin/-isobetanin and neobetanin (from mixture 1) plus 2,15,17-tridecarboxy-2,3-dehydro-neobetanin, 2,17-bidecarboxy-2,3-dehydro-neobetanin and 2,15,17-tridecarboxy-neobetanin (mixture 2). The results of this research are crucial in finding effective isolation methods of betacyanins and their derivatives which are meaningful compounds due their colorant properties and potential health benefits regarding antioxidant and cancer prevention. The pigments were detected by LC-DAD and LC-MS/MS techniques. Copyright © 2013 Elsevier B.V. All rights reserved.

Purification of Peroxisomes and Mitochondria from Spinach Leaf by Percoll Gradient Centrifugation 1

PubMed Central

Schwitzguebel, Jean-Paul; Siegenthaler, Paul-André

1984-01-01

A procedure was developed to purify simultaneously peroxisomes and mitochondria from spinach (Spinacia oleracea L.) leaf under isoosmotic and low viscosity conditions. This method involved differential centrifugation and density gradient centrifugation on four layers of Percoll. Chlorophyll-free preparations of highly intact and active organelles were obtained and cross-contamination was negligible. Both organelles were stable for several hours, even if they remained in Percoll. Purified mitochondria were able to carry out the oxidation of different substrates with excellent respiratory control and ADP:O ratios. The method described in the present work was also suitable to purify mitochondria and peroxisomes from potato (Solanum tuberosum L.) tubers. PMID:16663685

Solid-state diode-like chemiluminescence based on serial, immobilized concentration gradients in mixed-valent poly[Ru(vbpy){sub 3}](PF{sub 6}){sub 2} films

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maness, K.M.; Terrill, R.H.; Meyer, T.J.

The electronic conductivity and electrogenerated chemiluminescence (ECL) of thin, electropolymerized films of the fixed-site redox polymer poly[Ru(vbpy){sub 3}](PF{sub 6}){sub 2} (vbpy = 4-vinyl-4`-methyl-2,2`-bipyridine) on Pt interdigitated array electrodes were examined for both solvent-swollen and dry films. In both cases emission arose from {sup *}Ru{sup 2+} produced via the electron-transfer reaction between Ru{sup 3+} and Ru{sup 1+} states within the film (Ru = Ru-(vbpy){sub 3}). Dry films contained fixed concentration gradients of Ru{sup 3+}, Ru{sup 2+}, and Ru{sup 1+} states which were first introduced in an acetonitrile-swollen film via the constant potential oxidation and reduction of Ru{sup 2+} at opposing IDAmore » fingers. The gradients were then immobilized by drying and cooling the film while retaining the inter-electrode bias (2.6V). The resulting dried and cooled films responded rapidly to changes in voltage bias and exhibited diode-like characteristics, conducting and emitting light at biases >2.6 V and undergoing a reverse bias breakdown current, unassociated with light emission, at ca. -5.5 V. At 0{degree}C the optimum quantum efficiency of solid-state ECL emission ({phi}{sub ECL}) was similar to that in solvent-swollen films: 0.0003 photon/electron. In contrast to the dry films, solvent-swollen films were slow to respond to changes in voltage bias and did not exhibit diode-like behavior. 18 refs., 7 figs.« less

A systematic investigation of sample diluents in modern supercritical fluid chromatography.

PubMed

Desfontaine, Vincent; Tarafder, Abhijit; Hill, Jason; Fairchild, Jacob; Grand-Guillaume Perrenoud, Alexandre; Veuthey, Jean-Luc; Guillarme, Davy

2017-08-18

This paper focuses on the possibility to inject large volumes (up to 10μL) in ultra-high performance supercritical fluid chromatography (UHPSFC) under generic gradient conditions. Several injection and method parameters have been individually evaluated (i.e. analyte concentration, injection volume, initial percentage of co-solvent in the gradient, nature of the weak needle wash solvent, nature of the sample diluent, nature of the column and of the analyte). The most critical parameters were further investigated using in a multivariate approach. The overall results suggested that several aprotic solvents including methyl tert-butyl ether (MTBE), dichloromethane, acetonitrile or cyclopentyl methyl ether (CPME) were well adapted for the injection of large volume in UHPSFC, while MeOH was generally the worst alternative. However, the nature of the stationary phase also had a strong impact and some of these diluents did not perform equally on each column. This was due to the existence of a competition in the adsorption of the analyte and the diluent on the stationary phase. This observation introduced the idea that the sample diluent should not only be chosen according to the analyte but also to the column chemistry to limit the interactions between the diluent and the ligands. Other important characteristics of the "ideal" SFC sample diluent were finally highlighted. Aprotic solvents with low viscosity are preferable to avoid strong solvent effects and viscous fingering, respectively. In the end, the authors suggest that the choice of the sample diluent should be part of the method development, as a function of the analyte and the selected stationary phase. Copyright © 2017 Elsevier B.V. All rights reserved.

Complex investigation of extraction techniques applied for cyclitols and sugars isolation from different species of Solidago genus.

PubMed

Ratiu, Ileana-Andreea; Al-Suod, Hossam; Ligor, Magdalena; Ligor, Tomasz; Railean-Plugaru, Viorica; Buszewski, Bogusław

2018-03-15

Cyclitols are phytochemicals naturally occurring in plant material, which attracted an increasing interest due to multiple medicinal attributes, among which the most important are the antidiabetic, antioxidant, and anticancer properties. Due to their valuable properties, sugars are used in the food industry as sweeteners, preservatives, texture modifiers, fermentation substrates, and flavoring and coloring agents. In this study, we report for the first time the quantitative analysis of sugars and cyclitols isolated from Solidago virgaurea L., which was used for the selection of the optimal solvent and extraction technique that can provide the best possible yield. Moreover, the quantities of sugars and cyclitols extracted from two other species, Solidago canadensis and Solidago gigantea, were investigated using the best extraction method and the most appropriate solvent. Comparative analysis of natural plant extracts obtained using five different techniques-maceration, Soxhlet extraction, pressurized liquid extraction, ultrasound-assisted extraction, and supercritical fluid extraction-was performed in order to decide the most suitable, efficient, and economically convenient extraction method. Three different solvents were used. Analysis of samples has been performed by solid-phase extraction for purification and pre-concentration, followed by derivation and GC-MS analysis. Highest efficiency for the total amount of obtained compounds has been reached by PLE, when water was used as a solvent. d-pinitol amount was almost similar for every solvent and for all the extraction techniques involved. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Porous fiber formation in polymer-solvent system undergoing solvent evaporation

NASA Astrophysics Data System (ADS)

Dayal, Pratyush; Kyu, Thein

2006-08-01

Temporal evolution of the fiber morphology during dry spinning has been investigated in the framework of Cahn-Hilliard equation [J. Chem. Phys. 28, 258 (1958)] pertaining to the concentration order parameter or volume fraction given by the Flory-Huggins free energy of mixing [P. J. Flory, Principles of Polymer Chemistry (Cornell University Press, Ithaca, NY, 1953), p. 672] in conjunction with the solvent evaporation rate. To guide the solvent evaporation induced phase separation, equilibrium phase diagram of the starting polymer solution was established on the basis of the Flory-Huggins free energy of mixing. The quasi-steady-state approximation has been adopted to account for the nonconserved nature of the concentration field caused by the solvent loss. The process of solvent evaporation across the fiber skin-air interface was treated in accordance with the classical Fick's law [R. B. Bird et al., Transport Phenomena (J. Wiley, New York, 1960), p. 780]. The simulated morphologies include gradient type, hollow fiber type, bicontinuous type, and host-guest type. The development of these diverse fiber morphologies is explicable in terms of the phase diagram of the polymer solution in a manner dependent on the competition between the phase separation dynamics and rate of solvent evaporation.

Urea transport through composite polyallylamine membranes

NASA Technical Reports Server (NTRS)

Ballou, E. V.; Kubo, L. Y.; Spitze, L. A.; Wydeven, T.; Clark, J. A.

1977-01-01

Polyallylamine composite reverse osmosis membranes were prepared by plasma polymerization and deposition onto small-pored cellulose acetate/cellulose nitrate films. The polyallylamine coated the porous substrate with a thin uniform polymer film which exhibited water permeability and urea rejection, of interest because of the potential application of reverse osmosis to urine purification in closed environmental systems. The flux of C-14 labeled urea was studied under the influence of osmotic gradients provided by sodium chloride solutions. The urea flux was found to be enhanced by an osmotic pressure gradient in the same direction and diminished, but not prevented, by an opposing osmotic pressure gradient. Consideration is given to the mechanism of the urea transport, as well as to the influence of concentration polarization on the experimental results. The minimization of coupled flow in pores of a critical size range is apparently necessary to improve urea rejection.

[Simultaneous determination of vitamins A, D3 and E in infant formula and adult nutritions by online two-dimensional liquid chromatography].

PubMed

Zhang, Yanhai; Qibule, Hasi; Jin, Yan; Wang, Jia; Ma, Wenli

2015-03-01

A rapid method for the simultaneous determination of vitamins A, D3 and E in infant formula and adult nutritions has been developed using online two-dimensional liquid chromatography (2D-LC). First of all, C8 and polar embedded C18 columns were chosen as the first and second dimensional column respectively according to hydrophobic-subtraction model, which constituted excellent orthogonal separation system. The detection wavelengths were set at 263 nm for vitamin D3, 296 nm for vitamin E and 325 nm for vitamin A. The purification of vitamin D3 and quantifications of vitamins A and E were completed simultaneously in the first dimensional separation using the left pump of Dual Gradient LC (DGLC) with methanol, acetonitrile and water as mobile phases. The heart-cutting time window of vitamin D3 was confirmed according to the retention time of vitamin D3 in the first dimensional separation. The elute from the first dimensional column (1-D column) which contained vitamin D3 was collected by a 500 µL sample loop and then taken into the second dimensional column (2-D column) by the right pump of DGLC with methanol, acetonitrile and water as mobile phases. The quantification of vitamin D3 was performed in the second dimensional separation with vitamin D2 as internal standard. At last, this method was applied for the analysis of the three vitamins in milk powder, cheese and yogurt. The injected sample solution with no further purification was pre-treated by hot-saponification using 1. 25 kg/L KOH solution and extracted by petroleum ether solvent. The recoveries of vitamin D3 spiked in all samples were 75.50%-85.00%. There was no statistically significant difference for the results between this method and standard method through t-test. The results indicate that vitamins A, D3 and E in infant formula and adult fortified dairy can be determined rapidly and accurately with this method.

A sialic acid assay in isolation and purification of bovine k-casein glycomacropeptide: a review.

PubMed

Nakano, Takuo; Ozimek, Lech

2014-01-01

Sialic acid is a carbohydrate moiety of k-casein glycomacropeptide (GMP), which is a 64 amino acid residue C-terminal sialylated phosphorylated glycopeptide released from k-casein by the action of chymosin during cheese making. GMP lacks aromatic amino acids including phenylalanine, tyrosine, and tryptophan. Because of its unique amino acid composition and various biological activities, GMP is thought to be a potential ingredient for dietetic foods (e.g., a food for PKU patients) and pharmaceuticals. Thus, increased attention has been given to the development of techniques to purify GMP. In this review, techniques of GMP purification described in patents and scientific research papers were introduced. A sialic acid assay is the important method to track GMP isolation and purification processes, for which the thiobarbituric acid reaction with 1-propanol as a chromophore extracting solvent is an inexpensive, practical and specific technique. Sephacryl S-200 gel filtration chromatography, cellulose acetate electrophoresis, and sodium dodecyl sulfate polyacrylamide gel electrophoresis are the major techniques to identify sialic acid specific to GMP. Sephacryl S-200 chromatography and cellulose acetate electrophoresis are also used to detect GMP sialic acid in whey pearmeate and whey added commercial margarine samples. Future research includes development of an economical industrial scale method to produce high purity GMP.

Fermentation and purification strategies for the production of betulinic acid and its lupane-type precursors in Saccharomyces cerevisiae.

PubMed

Czarnotta, Eik; Dianat, Mariam; Korf, Marcel; Granica, Fabian; Merz, Juliane; Maury, Jérôme; Baallal Jacobsen, Simo A; Förster, Jochen; Ebert, Birgitta E; Blank, Lars M

2017-11-01

Microbial production of plant derived, biologically active compounds has the potential to provide economic and ecologic alternatives to existing low productive, plant-based processes. Current production of the pharmacologically active cyclic triterpenoid betulinic acid is realized by extraction from the bark of plane tree or birch. Here, we reengineered the reported betulinic acid pathway into Saccharomyces cerevisiae and used this novel strain to develop efficient fermentation and product purification methods. Fed-batch cultivations with ethanol excess, using either an ethanol-pulse feed or controlling a constant ethanol concentration in the fermentation medium, significantly enhanced production of betulinic acid and its triterpenoid precursors. The beneficial effect of excess ethanol was further exploited in nitrogen-limited resting cell fermentations, yielding betulinic acid concentrations of 182 mg/L, and total triterpenoid concentrations of 854 mg/L, the highest concentrations reported so far. Purification of lupane-type triterpenoids with high selectivity and yield was achieved by solid-liquid extraction without prior cell disruption using polar aprotic solvents such as acetone or ethyl acetate and subsequent precipitation with strong acids. This study highlights the potential of microbial production of plant derived triterpenoids in S. cerevisiae by combining metabolic and process engineering. © 2017 Wiley Periodicals, Inc.

Extraction and purification of high added value compounds from by-products of the winemaking chain using alternative/nonconventional processes/technologies.

PubMed

Yammine, Sami; Brianceau, Sylène; Manteau, Sébastien; Turk, Mohammad; Ghidossi, Rémy; Vorobiev, Eugène; Mietton-Peuchot, Martine

2018-05-24

Grape byproducts are today considered as a cheap source of valuable compounds since existent technologies allow the recovery of target compounds and their recycling. The goal of the current article is to explore the different recovery stages used by both conventional and alternative techniques and processes. Alternative pre-treatments techniques reviewed are: ultrasounds, pulsed electric fields and high voltage discharges. In addition, nonconventional solvent extraction under high pressure, specifically, supercritical fluid extraction and subcritical water extraction are discussed. Finally alternative purification technologies, for example membrane processing were also examined. The intent is to describe the mechanisms involved by these alternative technologies and to summarize the work done on the improvement of the extraction process of phenolic compounds from winery by-products. With a focus on the developmental stage of each technology, highlighting the research need and challenges to be overcome for an industrial implementation of these unitary operations in the overall extraction process. A critical comparison of conventional and alternative techniques will be reviewed for ethe pre-treatment of raw material, the diffusion of polyphenols and the purification of these high added value compounds. This review intends to give the reader some key answers (costs, advantages, drawbacks) to help in the choice of alternative technologies for extraction purposes.

Mechanical properties of ultrahigh molecular weight PHEMA hydrogels synthesized using initiated chemical vapor deposition.

PubMed

Bose, Ranjita K; Lau, Kenneth K S

2010-08-09

In this work, poly(2-hydroxyethyl methacrylate) (PHEMA), a widely used hydrogel, is synthesized using initiated chemical vapor deposition (iCVD), a one-step surface polymerization that does not use any solvents. iCVD synthesis is capable of producing linear stoichiometric polymers that are free from entrained unreacted monomer or solvent and, thus, do not require additional purification steps. The resulting films, therefore, are found to be noncytotoxic and also have low nonspecific protein adsorption. The kinetics of iCVD polymerization are tuned so as to achieve rapid deposition rates ( approximately 1.5 microm/min), which in turn yield ultrahigh molecular weight polymer films that are mechanically robust with good water transport and swellability. The films have an extremely high degree of physical chain entanglement giving rise to high tensile modulus and storage modulus without the need for chemical cross-linking that compromises hydrophilicity.

HPLC-ELSD Quantification and Centrifugal Partition Chromatography Isolation of 8-O-Acetylharpagide from Oxera coronata (Lamiaceae).

PubMed

Remeur, Camille; Le Borgne, Erell; Gauthier, Léa; Grougnet, Raphaël; Deguin, Brigitte; Poullain, Cyril; Litaudon, Marc

2017-05-01

Iridoid glycosides possess highly functionalised monoterpenoid aglycon with several contiguous stereocentres. For the most common, they are often present in quantities reaching several percentage of the fresh plant weight, and thus they may be regarded as starting material for the synthesis of a number of new chiral and bioactive molecules. To quantify and to isolate 8-O-acetylharpagide (AH) from several extracts of Oxera coronata R.P.J. de Kok, a Lamiaceae species endemic to New Caledonia, using HPLC-ELSD (evaporative light scattering detector) and centrifugal partition chromatography (CPC). Oxera coronata produces high amounts of AH in leaves, twigs and fruits. Water and methanol extracts of these plant parts were prepared. The content of AH in each extract was quantified by HPLC-ELSD, using acetonitrile-water (+0.1% formic acid) gradient elution. The HPLC method was validated for precision, linearity, limit of detection (LOD), limit of quantification (LOQ) and accuracy. A ternary solvent system ethyl acetate/n-propanol/water (3:2:5, v/v/v) was selected and applied to recover the target compound using Spot CPC from the leaves aqueous extract. HPLC-ELSD analysis followed by CPC purification led to the efficient isolation of AH from O. coronata leaves aqueous extract. HPLC-ELSD has proven to be a well-adapted detection and quantification method for iridoid glycosides, while CPC confirmed to be an efficient technique for the isolation of polar compounds. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Green and scalable production of colloidal perovskite nanocrystals and transparent sols by a controlled self-collection process

NASA Astrophysics Data System (ADS)

Liu, Shuangyi; Huang, Limin; Li, Wanlu; Liu, Xiaohua; Jing, Shui; Li, Jackie; O'Brien, Stephen

2015-07-01

Colloidal perovskite oxide nanocrystals have attracted a great deal of interest owing to the ability to tune physical properties by virtue of the nanoscale, and generate thin film structures under mild chemical conditions, relying on self-assembly or heterogeneous mixing. This is particularly true for ferroelectric/dielectric perovskite oxide materials, for which device applications cover piezoelectrics, MEMs, memory, gate dielectrics and energy storage. The synthesis of complex oxide nanocrystals, however, continues to present issues pertaining to quality, yield, % crystallinity, purity and may also suffer from tedious separation and purification processes, which are disadvantageous to scaling production. We report a simple, green and scalable ``self-collection'' growth method that produces uniform and aggregate-free colloidal perovskite oxide nanocrystals including BaTiO3 (BT), BaxSr1-xTiO3 (BST) and quaternary oxide BaSrTiHfO3 (BSTH) in high crystallinity and high purity. The synthesis approach is solution processed, based on the sol-gel transformation of metal alkoxides in alcohol solvents with controlled or stoichiometric amounts of water and in the stark absence of surfactants and stabilizers, providing pure colloidal nanocrystals in a remarkably low temperature range (15 °C-55 °C). Under a static condition, the nanoscale hydrolysis of the metal alkoxides accomplishes a complete transformation to fully crystallized single domain perovskite nanocrystals with a passivated surface layer of hydroxyl/alkyl groups, such that the as-synthesized nanocrystals can exist in the form of super-stable and transparent sol, or self-accumulate to form a highly crystalline solid gel monolith of nearly 100% yield for easy separation/purification. The process produces high purity ligand-free nanocrystals excellent dispersibility in polar solvents, with no impurity remaining in the mother solution other than trace alcohol byproducts (such as isopropanol). The afforded stable and transparent suspension/solution can be treated as inks, suitable for printing or spin/spray coating, demonstrating great capabilities of this process for fabrication of high performance dielectric thin films. The simple ``self-collection'' strategy can be described as green and scalable due to the simplified procedure from synthesis to separation/purification, minimum waste generation, and near room temperature crystallization of nanocrystal products with tunable sizes in extremely high yield and high purity.Colloidal perovskite oxide nanocrystals have attracted a great deal of interest owing to the ability to tune physical properties by virtue of the nanoscale, and generate thin film structures under mild chemical conditions, relying on self-assembly or heterogeneous mixing. This is particularly true for ferroelectric/dielectric perovskite oxide materials, for which device applications cover piezoelectrics, MEMs, memory, gate dielectrics and energy storage. The synthesis of complex oxide nanocrystals, however, continues to present issues pertaining to quality, yield, % crystallinity, purity and may also suffer from tedious separation and purification processes, which are disadvantageous to scaling production. We report a simple, green and scalable ``self-collection'' growth method that produces uniform and aggregate-free colloidal perovskite oxide nanocrystals including BaTiO3 (BT), BaxSr1-xTiO3 (BST) and quaternary oxide BaSrTiHfO3 (BSTH) in high crystallinity and high purity. The synthesis approach is solution processed, based on the sol-gel transformation of metal alkoxides in alcohol solvents with controlled or stoichiometric amounts of water and in the stark absence of surfactants and stabilizers, providing pure colloidal nanocrystals in a remarkably low temperature range (15 °C-55 °C). Under a static condition, the nanoscale hydrolysis of the metal alkoxides accomplishes a complete transformation to fully crystallized single domain perovskite nanocrystals with a passivated surface layer of hydroxyl/alkyl groups, such that the as-synthesized nanocrystals can exist in the form of super-stable and transparent sol, or self-accumulate to form a highly crystalline solid gel monolith of nearly 100% yield for easy separation/purification. The process produces high purity ligand-free nanocrystals excellent dispersibility in polar solvents, with no impurity remaining in the mother solution other than trace alcohol byproducts (such as isopropanol). The afforded stable and transparent suspension/solution can be treated as inks, suitable for printing or spin/spray coating, demonstrating great capabilities of this process for fabrication of high performance dielectric thin films. The simple ``self-collection'' strategy can be described as green and scalable due to the simplified procedure from synthesis to separation/purification, minimum waste generation, and near room temperature crystallization of nanocrystal products with tunable sizes in extremely high yield and high purity. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr02351c

Controlling microtube permeability via grafted polymers and solvent quality.

PubMed

Suo, Tongchuan; Whitmore, Mark D

2014-03-21

We examine pressure-driven flow through a microtube with grafted polymers using a "doubly self-consistent field" steady-state theory. Our focus is on the structure of the polymer layer, the tube permeability, and the effects of solvent quality, for different regimes of open and closed tubes. We find that, within experimentally attainable pressure gradients, the flow has very little effect on the grafted layer. However, the polymers, and in particular variations in the solvent quality and cylinder radii, can have large effects on the flow. We find that the permeability can either increase or decrease with either the radius or solvent quality, and we identify the regimes for different behaviors in terms of general parameters that can be used to generalize to other systems. This allows us to identify regimes where the systems are most sensitive to these "tuning" parameters, and we find that they correspond to the boundaries between open and closed tubes identified earlier.

Removing micron size particles from coal liquids

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodgers, B.R.; Westmoreland, P.R.

This paper reports results of an investigation which was undertaken in order to improve purification of liquid fuels obtained by coal liquefaction processes. It is shown that settling and filtration rates increased substantially after agglomeration. And, ground coal was found to be an economical substitute for diatomaceous earth in filtration. The effects of certain solvents on the agglomerating tendencies of solids in the unfiltered oil (UFO) from the SRC and COED processes were determined by oil immersion microscopy. The significant results obtained by these experiments are listed. Economic advantages are presented. 13 references.

Coal liquefaction process

DOEpatents

Wright, Charles H.

1986-01-01

A process for the liquefaction of coal wherein raw feed coal is dissolved in recycle solvent with a slurry containing recycle coal minerals in the presence of added hydrogen at elevated temperature and pressure. The highest boiling distillable dissolved liquid fraction is obtained from a vacuum distillation zone and is entirely recycled to extinction. Lower boiling distillable dissolved liquid is removed in vapor phase from the dissolver zone and passed without purification and essentially without reduction in pressure to a catalytic hydrogenation zone where it is converted to an essentially colorless liquid product boiling in the transportation fuel range.

PLUTONIUM COMPOUNDS AND PROCESS FOR THEIR PREPARATION

DOEpatents

Wolter, F.J.; Diehl, H.C. Jr.

1958-01-01

This patent relates to certain new compounds of plutonium, and to the utilization of these compounds to effect purification or separation of the plutonium. The compounds are organic chelate compounds consisting of tetravalent plutonium together with a di(salicylal) alkylenediimine. These chelates are soluble in various organic solvents, but not in water. Use is made of this property in extracting the plutonium by contacting an aqueous solution thereof with an organic solution of the diimine. The plutonium is chelated, extracted and effectively separated from any impurities accompaying it in the aqueous phase.

Expression, purification, crystallization and preliminary X-ray studies of Lactobacillus jensenii enolase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harris, Paul T.; Raghunathan, Kannan; Spurbeck, Rachel R.

2010-09-02

Recombinant Lactobacillus jensenii enolase fused to a C-terminal noncleavable His tag was expressed in Escherichia coli, purified and crystallized by sitting-drop vapor diffusion. A complete data set was collected to 3.25 {angstrom} resolution. The crystals belonged to space group I4, with unit-cell parameters a = b = 145.31, c = 99.79 {angstrom}. There were two protein subunits in the asymmetric unit, which gave a Matthews coefficient V{sub M} of 2.8 {angstrom}{sup 3} Da{sup -1}, corresponding to 55.2% solvent content.

Comparison of a novel TiO₂/diatomite composite and pure TiO₂ for the purification of phosvitin phosphopeptides.

PubMed

Zhang, Yang; Li, Junhua; Niu, Fuge; Sun, Jun; Dou, Yuan; Liu, Yuntao; Su, Yujie; Zhou, Bei; Xu, Qinqin; Yang, Yanjun

2014-06-01

A novel TiO2/diatomite composite (TD) was prepared and then characterized by scanning electron microscope (SEM) and Fourier Transform Infrared (FTIR). The results of SEM showed that after modification, the porous surface of diatomite was covered with TiO2. Both diatomite and TD had clear disc-shaped structures with average grain diameters of around 25 μm. Then TD and pure TiO2 were applied in the purification of phosvitin phosphopeptides (PPPs) from the digest of egg yolk protein, and a comparative study of adsorption properties of PPPs on TD and TiO2 was performed. In the study of adsorption kinetics, the adsorption equilibrium of PPPs on TD and TiO2 fitted well with the Langmuir model, and the time needed to reach adsorption equilibrium were both around 10 min. The maximum dynamic adsorption capacity of TD (8.15 mg/g) was higher than that of TiO2 (4.96 mg/g). The results of repeated use showed that TD and TiO2 were very stable after being subjected to ten repeated adsorption-elution cycles, and TD could easily be separated from aqueous solution by filtration. On the other hand, the present synthetic technology of TD was very simple, cost-effective, organic solvent-free and available for large-scale preparation. Thus, this separation method not only brings great advantages in the purification of PPPs from egg yolk protein but also provides a promising purification material for the enrichment of phosphopeptides in proteomic researches. Copyright © 2014 Elsevier B.V. All rights reserved.

Preparative isolation and purification of indigo and indirubin from Folium isatidis by high-speed counter-current chromatography.

PubMed

Lü, Hai-tao; Liu, Jing; Deng, Rui; Song, Ji-ying

2012-01-01

Indigo and indirubin are the main active ingredients found in traditional Chinese herbal medicine Folium isatidis. An effective method for the isolation and purification of indigo and indirubin from Folium isatidis is needed. Compared with the conventional column chromatographic techniques, high-speed counter-current chromatography (HSCCC) is a suitable alternative for the enrichment and purification of these target compounds, and eliminates the complications resulting from a solid support matrix. To develop a reliable HSCCC method for isolation and identification of indigo and indirubin in a one-step separation from Folium isatidis. The optimum extracting conditions of indigo and indirubin from Folium isatidis were investigated by orthogonal test L(16) (4(5)). The target compounds were isolated and purified with a solvent system of n-hexane:ethyl acetate:ethanol:water (1:1:1:1, v/v) and the lower phase was used as the mobile phase in the head-to-tail elution mode. The purities of target compounds were tested by HPLC and their structures were identified by UV, IR, electrospray ion source (ESI)-MS, (1) H-NMR and (13) C-NMR analyses. From 165 mg of the crude extract, 5.65 mg of indigo and 1.00 mg of indirubin were obtained by HPLC analysis with purities of 98.4% and 99.0% respectively, and their mean recoveries were 91.0% and 90.7%, respectively. The HSCCC method is effective for the preparative separation and purification of indigo and indirubin in a one-step separation from Folium isatidis. Copyright © 2012 John Wiley & Sons, Ltd.

Purification of Bacteriophages Using Anion-Exchange Chromatography.

PubMed

Vandenheuvel, Dieter; Rombouts, Sofie; Adriaenssens, Evelien M

2018-01-01

In bacteriophage research and therapy, most applications ask for highly purified phage suspensions. The standard technique for this is ultracentrifugation using cesium chloride gradients. This technique is cumbersome, elaborate and expensive. Moreover, it is unsuitable for the purification of large quantities of phage suspensions.The protocol described here, uses anion-exchange chromatography to bind phages to a stationary phase. This is done using an FLPC system, combined with Convective Interaction Media (CIM ® ) monoliths. Afterward, the column is washed to remove impurities from the CIM ® disk. By using a buffer solution with a high ionic strength, the phages are subsequently eluted from the column and collected. In this way phages can be efficiently purified and concentrated.This protocol can be used to determine the optimal buffers, stationary phase chemistry and elution conditions, as well as the maximal capacity and recovery of the columns.

CONTINUOUS CHELATION-EXTRACTION PROCESS FOR THE SEPARATION AND PURIFICATION OF METALS

DOEpatents

Thomas, J.R.; Hicks, T.E.; Rubin, B.; Crandall, H.W.

1959-12-01

A continuous process is presented for separating metal values and groups of metal values from each other. A complex mixture. e.g., neutron-irradiated uranium, can be resolved into component parts. In the present process the values are dissolved in an acidic solution and adjusted to the proper oxidation state. Thenceforth the solution is contacted with an extractant phase comprising a fluorinated beta -diketone in an organic solvent under centain pH conditions whereupon plutonium and zirconium are extracted. Plutonium is extracted from the foregoing extract with reducing aqueous solutions or under specified acidic conditions and can be recovered from the aqueous solution. Zirconium is then removed with an oxalic acid aqueous phase. The uranium is recovered from the residual original solution using hexone and hexone-diketone extractants leaving residual fission products in the original solution. The uranium is extracted from the hexone solution with dilute nitric acid. Improved separations and purifications are achieved using recycled scrub solutions and the "self-salting" effect of uranyl ions.

Preparative isolation and purification of macrolactin antibiotics from marine bacterium Bacillus amyloliquefaciens using high-speed counter-current chromatography in stepwise elution mode.

PubMed

He, Shan; Wang, Hongqiang; Yan, Xiaojun; Zhu, Peng; Chen, Juanjuan; Yang, Rui

2013-01-11

Preparative high-speed counter-current chromatography (HSCCC) was successfully applied to the isolation and purification of two macrolactin antibiotics from marine bacterium Bacillus amyloliquefaciens for the first time using stepwise elution with a pair of two-phase solvent systems composed of n-hexane-ethyl acetate-methanol-water at (1:4:1:4, v/v) and (3:4:3:4, v/v). The preparative HSCCC separation was performed on 300 mg of crude sample yielding macrolactin B (22.7 mg) and macrolactin A (40.4 mg) in a one-step separation, with purities over 95% as determined by HPLC. The structures of these compounds were identified by MS, (1)H NMR and (13)C NMR. Our results demonstrated that HSCCC was an efficient technique to separate marine antibiotics, which provide an approach to solve the problem of their sample availability for drug development. Copyright © 2012 Elsevier B.V. All rights reserved.

Separation and purification of triterpene saponins from roots of Radix phytolaccae by high-speed countercurrent chromatography coupled with evaporative light scattering detection

PubMed Central

Ma, Jie; Chen, Qianliang; Lai, Daowan; Sun, Wenji; Zhang, Tianyou; Ito, Yoichiro

2009-01-01

Coupled with evaporative light scattering detection, high-speed countercurrent chromatography was successfully applied for the first time to separation and purification of four triterpene saponins including esculentoside A, B, C and D from roots of Radix Phytolaccae. The separation was performed with an optimized two-phase solvent system composed of chloroform-methanol-water (4:4:2, v/v) using the lower phase as the mobile phase at a flow rate of 1.5 ml/min,. From 150 mg of crude extract 46.3 mg of esculentoside A, 21.8 mg of esculentoside B, 7.3 mg of esculentoside C, and 13.6 mg of esculentoside D were obtained at purities of 96.7%, 99.2%, 96.5% and 97.8%, respectively, as determined by HPLC analysis. The structures of the four triterpene saponins were identified by ESI-MS,1H NMR and 13C NMR. PMID:20454595

Purification Procedures for Single-Wall Carbon Nanotubes

NASA Technical Reports Server (NTRS)

Gorelik, Olga P.; Nikolaev, Pavel; Arepalli, Sivaram

2001-01-01

This report summarizes the comparison of a variety of procedures used to purify carbon nanotubes. Carbon nanotube material is produced by the arc process and laser oven process. Most of the procedures are tested using laser-grown, single-wall nanotube (SWNT) material. The material is characterized at each step of the purification procedures by using different techniques including scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), transmission electron microscopy (TEM), Raman, X-ray diffractometry (XRD), thermogravimetric analysis (TGA), nuclear magnetic resonance (NMR), and high-performance liquid chromatography (HPLC). The identified impurities are amorphous and graphitic carbon, catalyst particle aggregates, fullerenes, and hydrocarbons. Solvent extraction and low-temperature annealing are used to reduce the amount of volatile hydrocarbons and dissolve fullerenes. Metal catalysts and amorphous as well as graphitic carbon are oxidized by reflux in acids including HCl, HNO3 and HF and other oxidizers such as H2O2. High-temperature annealing in vacuum and in inert atmosphere helps to improve the quality of SWNTs by increasing crystallinity and reducing intercalation.

Silybum marianum pericarp yields enhanced silymarin products.

PubMed

AbouZid, Sameh F; Chen, Shao-Nong; McAlpine, James B; Friesen, J Brent; Pauli, Guido F

2016-07-01

An improved method for the purification of silymarin, the flavonolignan complex from the fruits of milk thistle, Silybum marianum, is reported. The method enables a more efficient extraction of silymarin from the pericarp after it has been separated mechanically from the rest of the fruits. Accelerated solvent extraction (ASE) was employed for each extraction procedure. Quantitation of the eight major silymarin components in the pericarp extract was compared to that of the whole fruit extract using two orthogonal analytical methods. The pericarp extract showed higher silymarin content (2.24-fold by HPLC and 2.12-fold by qHNMR) than whole fruit extract using acetone as an extraction solvent following defatting with hexane. Furthermore, the mg/g recovery of silymarin major components was not diminished by eliminating the hexane defatting step from the pericarp extraction procedure. The efficiencies of acetone, ethanol, and methanol as extraction solvents were compared. Methanol pericarp extract showed the highest content of the silymarin major components, 2.72-fold higher than an extract prepared from the whole fruits using acetone. Finally, all of the major silymarin components showed a higher w/w content in the pericarp extract than in a commercial extract. Copyright © 2016 Elsevier B.V. All rights reserved.

Preparative isolation of huperzines A and B from Huperzia serrata by displacement centrifugal partition chromatography.

PubMed

Toribio, Alix; Delannay, Eldra; Richard, Bernard; Plé, Karen; Zèches-Hanrot, Monique; Nuzillard, Jean-Marc; Renault, Jean-Hugues

2007-01-26

The pH-zone refining centrifugal partition chromatography technique was used to separate the two acetylcholinesterase inhibitors huperzines A and B from a crude alkaloid extract of the club moss Huperzia serrata. Complete co-elution of huperzines A and B was initially observed with the well-known methyl tert-butyl ether-acetonitrile-water (4:1:5, v/v/v) solvent system with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer. An efficient biphasic system was designed on the basis of solvent association that provided selectivity in the elution mode: n-heptane/ethyl acetate/n-propanol/water (5:15:35:45, v/v/v/v). Lowering the bridge solvent content (n-propanol) of this system increased the polarity difference between the two phases thus adapting it to the pH-zone refining mode. Thus, the purification of these compounds was achieved using the biphasic system n-heptane/ethyl acetate/n-propanol/water (10:30:15:45, v/v/v/v) with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer.

Progress on lipid extraction from wet algal biomass for biodiesel production.

PubMed

Ghasemi Naghdi, Forough; González González, Lina M; Chan, William; Schenk, Peer M

2016-11-01

Lipid recovery and purification from microalgal cells continues to be a significant bottleneck in biodiesel production due to high costs involved and a high energy demand. Therefore, there is a considerable necessity to develop an extraction method which meets the essential requirements of being safe, cost-effective, robust, efficient, selective, environmentally friendly, feasible for large-scale production and free of product contamination. The use of wet concentrated algal biomass as a feedstock for oil extraction is especially desirable as it would avoid the requirement for further concentration and/or drying. This would save considerable costs and circumvent at least two lengthy processes during algae-based oil production. This article provides an overview on recent progress that has been made on the extraction of lipids from wet algal biomass. The biggest contributing factors appear to be the composition of algal cell walls, pre-treatments of biomass and the use of solvents (e.g. a solvent mixture or solvent-free lipid extraction). We compare recently developed wet extraction processes for oleaginous microalgae and make recommendations towards future research to improve lipid extraction from wet algal biomass. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Schizosaccharomyces pombe Polysome Profile Analysis and RNA Purification.

PubMed

Wolf, Dieter A; Bähler, Jürg; Wise, Jo Ann

2017-04-03

Polysome profile analysis is widely used by investigators studying the mechanism and regulation of translation. The method described here uses high-velocity centrifugation of whole cell extracts on linear sucrose gradients to separate 40S and 60S ribosomal subunits from 80S monosomes and polysomes. Cycloheximide is included in the lysis buffer to "freeze" polysomes by blocking translation. After centrifugation, the gradient is fractionated and RNA (and/or protein) is prepared from each fraction for subsequent analysis of individual species using northern or western blots. The entire RNA population in each fraction can be analyzed by hybridization to microarrays or by high-throughput RNA sequencing, and the proteins present can be identified by mass spectrometry analysis. © 2017 Cold Spring Harbor Laboratory Press.

Molecular Level Structure and Dynamics of Electrolytes Using 17O Nuclear Magnetic Resonance Spectroscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murugesan, Vijayakumar; Han, Kee Sung; Hu, Jianzhi

2017-03-19

Electrolytes help harness the energy from electrochemical processes by serving as solvents and transport media for redox-active ions. Molecular-level interactions between ionic solutes and solvent molecules – commonly referred to as solvation phenomena – give rise to many functional properties of electrolytes such as ionic conductivity, viscosity, and stability. It is critical to understand the evolution of solvation phenomena as a function of competing counterions and solvent mixtures to predict and design the optimal electrolyte for a target application. Probing oxygen environments is of great interest as oxygens are located at strategic molecular sites in battery solvents and are directlymore » involved in inter- and intramolecular solvation interactions. NMR signals from 17O nuclei in battery electrolytes offer nondestructive bulk measurements of isotropic shielding, electric field gradient tensors, and transverse and longitudinal relaxation rates, which are excellent means for probing structure, bonding, and dynamics of both solute and solvent molecules. This article describes the use of 17O NMR spectroscopy in probing the solvation structures of various electrolyte systems ranging from transition metal ions in aqueous solution to lithium cations in organic solvent mixtures.« less

Optimization of non-linear gradient in hydrophobic interaction chromatography for the analytical characterization of antibody-drug conjugates.

PubMed

Bobály, Balázs; Randazzo, Giuseppe Marco; Rudaz, Serge; Guillarme, Davy; Fekete, Szabolcs

2017-01-20

The goal of this work was to evaluate the potential of non-linear gradients in hydrophobic interaction chromatography (HIC), to improve the separation between the different homologous species (drug-to-antibody, DAR) of commercial antibody-drug conjugates (ADC). The selectivities between Brentuximab Vedotin species were measured using three different gradient profiles, namely linear, power function based and logarithmic ones. The logarithmic gradient provides the most equidistant retention distribution for the DAR species and offers the best overall separation of cysteine linked ADC in HIC. Another important advantage of the logarithmic gradient, is its peak focusing effect for the DAR0 species, which is particularly useful to improve the quantitation limit of DAR0. Finally, the logarithmic behavior of DAR species of ADC in HIC was modelled using two different approaches, based on i) the linear solvent strength theory (LSS) and two scouting linear gradients and ii) a new derived equation and two logarithmic scouting gradients. In both cases, the retention predictions were excellent and systematically below 3% compared to the experimental values. Copyright © 2016 Elsevier B.V. All rights reserved.

An ammonium sulfate/ethanol aqueous two-phase system combined with ultrasonication for the separation and purification of lithospermic acid B from Salvia miltiorrhiza Bunge.

PubMed

Guo, Y X; Han, J; Zhang, D Y; Wang, L H; Zhou, L L

2012-07-01

We studied the effect of ultrasonication extraction technology combined with ammonium sulfate/ethanol aqueous two-phase system (ATPS) for the separation of lithospermic acid B (LAB) from Salvia miltiorrhiza Bunge. According to the literature and preliminary studies, ammonium sulfate concentration, ethanol concentration, pH, ultrasonication power, ultrasonication time and the ratio of solvent-to-solid were investigated using a single factor design to identify the factors affecting separation. Taking into consideration a simultaneous increase in LAB recovery (R (%)) and partition coefficient (K), the best performance of the ATPS was obtained at 25°C and pH 2 using ammonium sulfate 22% (w/w) and ethanol 30% (w/w). To keep the solvent-to-solid ratio at 10, response surface methodology was used to find the optimal ultrasonication power and ultrasonication time. Quadratic models were predicted for LAB yield in the upper phase. Optimal conditions of 572.1 W ultrasonication power and 42.2 min produced a maximum yield of LAB of 42.16 mg g(-1) sample. There was no obvious degradation of LAB with ultrasound under the applied conditions, and the experimental yield of LAB was 42.49 mg g(-1) sample and the purity was 55.28% (w/w), which was much higher than that obtained using conventional extraction. The present study demonstrated that ultrasound coupled with aqueous two-phase systems is very efficient tool for the extraction and purification of LAB from Salvia miltiorrhiza Bunge. Copyright © 2011 Elsevier B.V. All rights reserved.

Engineered catalytic biofilms: Site-specific enzyme immobilization onto E. coli curli nanofibers.

PubMed

Botyanszki, Zsofia; Tay, Pei Kun R; Nguyen, Peter Q; Nussbaumer, Martin G; Joshi, Neel S

2015-10-01

Biocatalytic transformations generally rely on purified enzymes or whole cells to perform complex transformations that are used on industrial scale for chemical, drug, and biofuel synthesis, pesticide decontamination, and water purification. However, both of these systems have inherent disadvantages related to the costs associated with enzyme purification, the long-term stability of immobilized enzymes, catalyst recovery, and compatibility with harsh reaction conditions. We developed a novel strategy for producing rationally designed biocatalytic surfaces based on Biofilm Integrated Nanofiber Display (BIND), which exploits the curli system of E. coli to create a functional nanofiber network capable of covalent immobilization of enzymes. This approach is attractive because it is scalable, represents a modular strategy for site-specific enzyme immobilization, and has the potential to stabilize enzymes under denaturing environmental conditions. We site-specifically immobilized a recombinant α-amylase, fused to the SpyCatcher attachment domain, onto E. coli curli fibers displaying complementary SpyTag capture domains. We characterized the effectiveness of this immobilization technique on the biofilms and tested the stability of immobilized α-amylase in unfavorable conditions. This enzyme-modified biofilm maintained its activity when exposed to a wide range of pH and organic solvent conditions. In contrast to other biofilm-based catalysts, which rely on high cellular metabolism, the modified curli-based biofilm remained active even after cell death due to organic solvent exposure. This work lays the foundation for a new and versatile method of using the extracellular polymeric matrix of E. coli for creating novel biocatalytic surfaces. © 2015 Wiley Periodicals, Inc.

Simple, high-yield purification of xanthine oxidase from bovine milk.

PubMed

Ozer, N; Müftüoglu, M; Ataman, D; Ercan, A; Ogüs, I H

1999-05-13

Xanthine oxidase, a commercially important enzyme with a wide area of application, was extracted from fresh milk, without added preservatives, using toluene and heat. The short purification procedure, with high yield, consisted of extraction, ammonium sulfate fractionation, and DEAE-Sepharose (fast flow) column chromatography. Xanthine oxidase was eluted as a single activity peak from the column using a buffer gradient. The purification fold, specific activity and yield for the purified xanthine oxidase were 328, 10.161 U/mg and 69%, respectively. The enzyme was concentrated by ultrafiltration, although 31% of the activity was lost during concentration, no change in specific activity was observed. Activity and protein gave coincident staining bands on native polyacrylamide gels. The intensity and the number of bands were dependent on the oxidative state(s) of the enzyme; reduction by 2-mercaptoethanol decreased the intensity of the slow-moving bands and increased the intensity of the fastest-moving band. Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), two major bands (molecular masses of 152 and 131 kDa) were observed, accounting for > or = 95% of xanthine oxidase. Native- and SDS-PAGE showed that the purified xanthine oxidase becomes a heterodimer due to endogenous proteases.

A combined simple bubbling method with high performance liquid chromatography purification strategy, higher radiochemical yield and purity and faster preparation of carbon-11-raclopride.

PubMed

Huang, Huacheng; Ning, Yanli; Zhang, Bucheng; Lou, Cen

2015-01-01

Carbon-11-raclopride (¹¹C-R) is a positron-emitting radiotracer successfully used for the study of cognitive control and widely applied in PET imaging. A simple automated preparation of ¹¹C-R by using the reaction of carbon-(11)-methyl triflate (¹¹C-MeOTF) or ¹¹C-methyl iodide (¹¹C-MeI) with demethylraclopride is described. Specifically we used a simple setup applied an additional "U" reaction vessel for ¹¹C-MeOTf compared with ¹¹C-MeI and assessed the influence of several solvents and of the amount of the percussor for ¹¹C-methylation of demethylraclopride by the bubbling method. The reversal of retention order between product and its precursor has been achieved for ¹¹C-R, enabling collection of the purified ¹¹C-R by using the HPLC column after shorter retention time. By the improved radiosynthesis and purification strategy, ¹¹C-R could be prepared with higher radiochemical yield than that of the previous studies. The yield for ¹¹C-MeOTf was 76% and for ¹¹C-CH3I >26% and with better radiochemical purity (>99% based on both ¹¹C-MeOTf and ¹¹C-MeI) as compared to the previously obtained purity of ¹¹C-R using HPLC method with acetonitrile as a part of mobile phase. Furthermore, by using ethanol as the organic modifier, residual solvent analysis prior to human injection could be avoided and ¹¹C-R could be injected directly following simple dilution and sterile filtration. Improved radiosynthesis and HPLC purification in combination with ethanol containing eluent, extremely shortened the time for preparation of ¹¹C-R, gave a higher radiochemical yield and purity for ¹¹C-R and can be used for multiple and faster synthesis of ¹¹C-R and probably for other ¹¹C-labeled radiopharmaceuticals.

Solvents and Parkinson disease: A systematic review of toxicological and epidemiological evidence

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lock, Edward A., E-mail: e.lock@ljmu.ac.uk; Zhang, Jing; Checkoway, Harvey

2013-02-01

Parkinson disease (PD) is a debilitating neurodegenerative motor disorder, with its motor symptoms largely attributable to loss of dopaminergic neurons in the substantia nigra. The causes of PD remain poorly understood, although environmental toxicants may play etiologic roles. Solvents are widespread neurotoxicants present in the workplace and ambient environment. Case reports of parkinsonism, including PD, have been associated with exposures to various solvents, most notably trichloroethylene (TCE). Animal toxicology studies have been conducted on various organic solvents, with some, including TCE, demonstrating potential for inducing nigral system damage. However, a confirmed animal model of solvent-induced PD has not been developed.more » Numerous epidemiologic studies have investigated potential links between solvents and PD, yielding mostly null or weak associations. An exception is a recent study of twins indicating possible etiologic relations with TCE and other chlorinated solvents, although findings were based on small numbers, and dose–response gradients were not observed. At present, there is no consistent evidence from either the toxicological or epidemiologic perspective that any specific solvent or class of solvents is a cause of PD. Future toxicological research that addresses mechanisms of nigral damage from TCE and its metabolites, with exposure routes and doses relevant to human exposures, is recommended. Improvements in epidemiologic research, especially with regard to quantitative characterization of long-term exposures to specific solvents, are needed to advance scientific knowledge on this topic. -- Highlights: ► The potential for organic solvents to cause Parkinson's disease has been reviewed. ► Twins study suggests etiologic relations with chlorinated solvents and Parkinson's. ► Animal studies with TCE showed potential to cause damage to dopaminergic neurons. ► Need to determine if effects in animals are relevant to human exposure levels.« less

Design and Growth of Novel Compounds for Radiation Sensors: Multinary Chalcogenides

NASA Technical Reports Server (NTRS)

Singh, N. B.; Su, Ching-Hua; Nagaradona, Teja; Arnold, Brad; Choa, Fow-Sen

2016-01-01

Increasing threats of radiological weapons have revitalized the researches for low cost large volume ?-ray and neutron ray sensors In the past few years we have designed and grown ternary and quaternary lead and thallium chalcogenides and lead selenoiodides for detectors to meet these challenges. These materials are congruent, can be tailored to enhance the parameters required for radiation sensors. In addition, this class of compounds can be grown by Bridgman method which promises for large volume productions. We have single crystals of several compounds from the melt including Tl3AsSe3, Tl3AsSe3-xSx, TlGaSe2, AgGaGe3Se8, AgxLi1-xAgGaGe3Se8 and PbTlI5-x Sex compounds. Experimental studies indicate that these have very low absorption coefficient, low defect density and can be fabricated in any shape and sizes. These crystals do not require post growth annealing and do not show any second phase precipitates when processed for electrode bonding and other fabrication steps. In this paper we report purification, growth and fabrication of large Tl3AsSe3 (TAS) crystals. We observed that TAS crystals grown by using further purification of as supplied high purity source materials followed by directionally solidified charge showed higher resistivity than previously reported values. TAS also showed constant value as the function of voltage. A low thermal gradient and high purity source material were used to reduce thermal stresses in large crystals. By improving the purification of the as supplied source materials very high quality thallium, selenium and arsenic was achieved for preparing stoichiometric Tl3AsSe3 compound. Low gradient (<20K/cm) and slow growth rate (1-2 cm/day) produced crystals with reduced stress. Crystals did not show any micro cracking during fabrication of crystals grown with high purity and at low thermal gradient. Since thallium is a major component and very sensitive to surface oxidation, removal of surface and bulk oxides is very important. Intentional increase in the growth rate from 1cm/day to higher speed (>5cm/day) showed very different morphologies on the surface of the crystals. Electrical resistivity was one order of magnitude higher than previously reported value and it was observed to be constant as the function of frequency.

Forward Osmosis in Wastewater Treatment Processes.

PubMed

Korenak, Jasmina; Basu, Subhankar; Balakrishnan, Malini; Hélix-Nielsen, Claus; Petrinic, Irena

2017-01-01

In recent years, membrane technology has been widely used in wastewater treatment and water purification. Membrane technology is simple to operate and produces very high quality water for human consumption and industrial purposes. One of the promising technologies for water and wastewater treatment is the application of forward osmosis. Essentially, forward osmosis is a process in which water is driven through a semipermeable membrane from a feed solution to a draw solution due to the osmotic pressure gradient across the membrane. The immediate advantage over existing pressure driven membrane technologies is that the forward osmosis process per se eliminates the need for operation with high hydraulic pressure and forward osmosis has low fouling tendency. Hence, it provides an opportunity for saving energy and membrane replacement cost. However, there are many limitations that still need to be addressed. Here we briefly review some of the applications within water purification and new developments in forward osmosis membrane fabrication.

Separation and purification of five alkaloids from Aconitum duclouxii by counter-current chromatography.

PubMed

Wang, Yarong; Cai, Shining; Chen, Yang; Deng, Liang; Zhou, Xumei; Liu, Jia; Xu, Xin; Xia, Qiang; Lin, Mao; Zhang, Jili; Huang, Weili; Wang, Wenjun; Xiang, Canhui; Cui, Guozhen; Du, Lianfeng; He, Huan; Qi, Baohui

2015-07-01

C19 -diterpenoid alkaloids are the main components of Aconitum duclouxii Levl. The process of separation and purification of these compounds in previous studies was tedious and time consuming, requiring multiple chromatographic steps, thus resulted in low recovery and high cost. In the present work, five C19 -diterpenoid alkaloids, namely, benzoylaconine (1), N-deethylaconitine (2), aconitine (3), deoxyaconitine (4), and ducloudine A (5), were efficiently prepared from A. duclouxii Levl (Aconitum L.) by ethyl acetate extraction followed with counter-current chromatography. In the process of separation, the critical conditions of counter-current chromatography were optimized. The two-phase solvent system composed of n-hexane/ethyl acetate/methanol/water/NH3 ·H2 O (25%) (1:1:1:1:0.1, v/v) was selected and 148.2 mg of 1, 24.1 mg of 2, 250.6 mg of 3, 73.9 mg of 4, and 31.4 mg of 5 were obtained from 1 g total Aconitum alkaloids extract, respectively, in a single run within 4 h. Their purities were found to be 98.4, 97.2, 98.2, 96.8, and 96.6%, respectively, by ultra-high performance liquid chromatography analysis. The presented separation and purification method was simple, fast, and efficient, and the obtained highly pure alkaloids are suitable for biochemical and toxicological investigation. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Characterization of Molecules Binding to the 70K N-terminal Region of Fibronectin by IFAST Purification Coupled with Mass Spectrometry

PubMed Central

Moussavi-Harami, S. Farshid; Annis, Douglas S.; Ma, Wenjiang; Berry, Scott M.; Coughlin, Emma E.; Strotman, Lindsay N.; Maurer, Lisa M.; Westphall, Michael S.; Coon, Joshua J.; Mosher, Deane F.; Beebe, David J.

2013-01-01

Fibronectin (Fn) is a large glycoprotein present in plasma and extracellular matrix and is important for many processes. Within Fn the 70kDa N-terminal region (70k-Fn) is involved in cell-mediated Fn assembly, a process that contributes to embryogenesis, development, and platelet thrombus formation. In addition, major human pathogens including Staphlycoccus aureus and Streptococcus pyogenes, bind the 70k-Fn region by a novel form of protein-protein interaction called β-zipper formation, facilitating bacterial spread and colonization. Knowledge of blood plasma and platelet proteins that interact with 70k-Fn by β-zipper formation is incomplete. In the current study, we aimed to characterize these proteins through affinity purification. For this affinity purification, we used a novel purification technique termed immiscible filtration assisted by surface tension (IFAST). The foundation of this technology is immiscible phase filtration, using a magnet to draw paramagnetic particle (PMP)-bound analyte through an immiscible barrier (oil or organic solvent) that separates an aqueous sample from an aqueous eluting buffer. The immiscible barrier functions to remove unbound proteins via exclusion rather than dilutive washing used in traditional isolation methods. We identified 31 interactors from plasma, of which only seven were previously known to interact with Fn. Furthermore, five proteins were identified to interact with 70k-Fn from platelet lysate, of which one was previously known. These results demonstrate that IFAST offers advantages for proteomic studies of interacting molecules in that the technique requires small sample volumes, can be done with high enough throughput to sample multiple interaction conditions, and is amenable to exploratory mass spectrometric and confirmatory immuno-blotting read-outs. PMID:23750785

Novel linear and step-gradient counter-current chromatography for bio-guided isolation and purification of cytotoxic podophyllotoxins from Dysosma versipellis (Hance).

PubMed

Yang, Zhi; Liu, Xiaoman; Wang, Kuiwu; Cao, Xiaoji; Wu, Shihua

2013-03-01

Dysosma versipellis (Hance) is a famous traditional Chinese medicine for the treatment of snakebite, weakness, condyloma accuminata, lymphadenopathy, and tumors for thousands of years. In this work, four podophyllotoxin-like lignans including 4'-demethylpodophyllotoxin (1), α-peltatin (2), podophyllotoxin (3), β-peltatin (4) as major cytotoxic principles of D. versipellis were successfully isolated and purified by several novel linear and step gradient counter-current chromatography methods using the systems of hexane/ethyl acetate/methanol/water (4:6:3:7 and 4:6:4:6, v/v/v/v). Compared with isocratic elution, linear and step-gradient elution can provide better resolution and save more time for the separation of photophyllotoxin and its congeners. Their cytotoxicities were further evaluated and their structures were validated by high-resolution electrospray TOF MS and nuclear magnetic resonance spectra. All components showed potent anticancer activity against human hepatoma cells HepG2. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Coal liquefaction process

DOEpatents

Wright, C.H.

1986-02-11

A process is described for the liquefaction of coal wherein raw feed coal is dissolved in recycle solvent with a slurry containing recycle coal minerals in the presence of added hydrogen at elevated temperature and pressure. The highest boiling distillable dissolved liquid fraction is obtained from a vacuum distillation zone and is entirely recycled to extinction. Lower boiling distillable dissolved liquid is removed in vapor phase from the dissolver zone and passed without purification and essentially without reduction in pressure to a catalytic hydrogenation zone where it is converted to an essentially colorless liquid product boiling in the transportation fuel range. 1 fig.

Quinolactacins A, B and C: novel quinolone compounds from Penicillium sp. EPF-6. I. Taxonomy, production, isolation and biological properties.

PubMed

Kakinuma, N; Iwai, H; Takahashi, S; Hamano, K; Yanagisawa, T; Nagai, K; Tanaka, K; Suzuki, K; Kirikae, F; Kirikae, T; Nakagawa, A

2000-11-01

Quinolactacins A (1), B (2) and C (3), novel quinolone antibiotics have been found from the cultured broth of a fungal strain isolated from the larvae of the mulberry pyralid Margaronia pyloalis Welker). The fungal strain, EPF-6 was identified as Penicillium sp. from its morphological characteristics. Quinolactacins were obtained from the culture medium by solvent extraction and chromatographic purification. Compound 1 showed inhibitory activity against tumor necrosis factor (TNF) production induced by murine peritoneal macrophages and macrophage-like J774.1 cells stimulated with lipopolysaccharide (LPS).

Tetraethylene glycol promoted two-step, one-pot rapid synthesis of indole-3-[1- 11C]acetic acid

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Sojeong; Qu, Wenchao; Alexoff, David L.

2014-12-12

An operationally friendly, two-step, one-pot process has been developed for the rapid synthesis of carbon-11 labeled indole-3-acetic acid ([ 11]IAA or [ 11]auxin). By replacing an aprotic polar solvent with tetraethylene glycol, nucleophilic [ 11]cyanation and alkaline hydrolysis reactions were performed consecutively in a single pot without a time-consuming intermediate purification step. The entire production time for this updated procedure is 55 min, which dramatically simplifies the entire synthesis and reduces the starting radioactivity required for a whole plant imaging study.

High Performance Liquid Chromatography of Some Analgesic Compounds: An Instrumental Analysis Experiment.

ERIC Educational Resources Information Center

Haddad, Paul; And Others

1983-01-01

Background information, procedures, and results are provided for an experiment demonstrating techniques of solvent selection, gradient elution, pH control, and ion-pairing in the analysis of an analgesic mixture using reversed-phase liquid chromatography on an octadecylsilane column. Although developed using sophisticated/expensive equipment, less…

Process development for robust removal of aggregates using cation exchange chromatography in monoclonal antibody purification with implementation of quality by design.

PubMed

Xu, Zhihao; Li, Jason; Zhou, Joe X

2012-01-01

Aggregate removal is one of the most important aspects in monoclonal antibody (mAb) purification. Cation-exchange chromatography (CEX), a widely used polishing step in mAb purification, is able to clear both process-related impurities and product-related impurities. In this study, with the implementation of quality by design (QbD), a process development approach for robust removal of aggregates using CEX is described. First, resin screening studies were performed and a suitable CEX resin was chosen because of its relatively better selectivity and higher dynamic binding capacity. Second, a pH-conductivity hybrid gradient elution method for the CEX was established, and the risk assessment for the process was carried out. Third, a process characterization study was used to evaluate the impact of the potentially important process parameters on the process performance with respect to aggregate removal. Accordingly, a process design space was established. Aggregate level in load is the critical parameter. Its operating range is set at 0-3% and the acceptable range is set at 0-5%. Equilibration buffer is the key parameter. Its operating range is set at 40 ± 5 mM acetate, pH 5.0 ± 0.1, and acceptable range is set at 40 ± 10 mM acetate, pH 5.0 ± 0.2. Elution buffer, load mass, and gradient elution volume are non-key parameters; their operating ranges and acceptable ranges are equally set at 250 ± 10 mM acetate, pH 6.0 ± 0.2, 45 ± 10 g/L resin, and 10 ± 20% CV respectively. Finally, the process was scaled up 80 times and the impurities removal profiles were revealed. Three scaled-up runs showed that the size-exclusion chromatography (SEC) purity of the CEX pool was 99.8% or above and the step yield was above 92%, thereby proving that the process is both consistent and robust.

[Application of general retention time formula for gradient liquid chromatography in the studies of ladder-like gradient elution].

PubMed

Sun, Xiaoli; Hao, Weiqiang; Wang, Junde; Di, Bin; Chen, Qiang; Zhuang, Wei; Yu, Qiang; Zhang, Peipei

2013-08-01

By not explicitly specifying the type of solvent strength model, the features of ladder-like gradient elution were studied based on the general retention time formula that was derived in our previous work. For the case where the solute is eluted at like gradient, we derived the expression that connects the mobile phase composition (phiR), at which the solute is eluted from the column, with the gradient slope (B). It was shown that phiR will increase with the increase of B in this case. For the case where the solute is eluted at the last isocratic segment of the ladder-like gradient, it was proven that the retention time (tR) will correlate linearly with the reciprocal of the gradient slope (1/B) when the initial and final mobile phase compositions are set to be constant. In experiments, by taking biphenyl as the sample, the values of retention time in isocratic and gradient elution were measured on a C18 column by using a mixture of methanol and water as the mobile phase. The experimental values were found to be well consistent with the theoretical values that were calculated from the expressions. These expressions will be helpful to understand the features of the ladder-like gradient in practice.

Solubility determination of raloxifene hydrochloride in ten pure solvents at various temperatures: Thermodynamics-based analysis and solute-solvent interactions.

PubMed

Ahad, Abdul; Shakeel, Faiyaz; Alfaifi, Omar Ali; Raish, Mohammad; Ahmad, Ajaz; Al-Jenoobi, Fahad I; Al-Mohizea, Abdullah M

2018-06-10

The purpose of the present study was to determine the solubility of raloxifene hydrochloride (RHCl) in ten solvents: water, ethanol, isopropyl alcohol (IPA), ethylene glycol (EG), propylene glycol (PG), polyethylene glycol-400 (PEG-400), Transcutol, 1-butanol, dimethyl sulfoxide (DMSO), and ethyl acetate (EA) at temperatures of 298.2-323.2 K and a pressure of 0.1 MPa. The solubility data obtained was fitted upon "Apelblat and Van't Hoff" equations. The maximum mole fraction solubility of RHCl was obtained in DMSO (5.02 × 10 -2 at 323.2 K), followed by PEG-400 (5.92 × 10 -3 at 323.2 K), EA (3.11 × 10 -3 at 323.2 K), Transcutol (1.22 × 10 -3 at 323.2 K), PG (2.19 × 10 -4 at 323.2 K), 1-butanol (1.96 × 10 -4 at 323.2 K), IPA (1.47 × 10 -4 at 323.2 K), ethanol (7.90 × 10 -5 at 323.2 K), EG (6.65 × 10 -5 at 323.2 K), and water (3.60 × 10 -5 at 323.2 K). Similar fashions were noticed at each studied temperature. The higher solubility of RHCl in DMSO, PEG-400, EA, and Transcutol was possibly referable to their lower polarity in comparison with water. The molecular interactions between the solute and solvent molecules were estimated by calculating parameters like activity coefficients, and more prominent solute-solvent molecular interactions were noted for RHCl-DMSO, RHCl-EA, and RHCl-PEG-400 in comparison with the other solute-solvent combinations. The outcomes of the "apparent thermodynamic analysis" showed that the dissolution of RHCl was "endothermic, spontaneous and entropy-driven" in all investigated solvents. The obtained solubility data of RHCl in commonly used solvents could be useful in the purification, recrystallization, and dosage form design of the drug. Copyright © 2018 Elsevier B.V. All rights reserved.

Determination of 24 personal care products in fish bile using hybrid solvent precipitation and dispersive solid phase extraction cleanup with ultrahigh performance liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry.

PubMed

Yao, Li; Lv, Yin-Zhi; Zhang, Li-Juan; Liu, Wang-Rong; Zhao, Jian-Liang; Liu, You-Sheng; Zhang, Qian-Qian; Ying, Guang-Guo

2018-05-25

Personal care products (PCPs) are ubiquitous in aquatic environments owing to the continuous discharge of domestic wastewater from highly urbanized regions. These PCPs can be adsorbed by fish and thereafter usually enter the bile of the fish through biliary excretion. In this study, a sensitive method based on a combination of hybrid solvent precipitation and dispersive solid phase extraction (d-SPE) purification was developed to simultaneously extract and detect 24 PCPs, namely, 16 biocides, 4 synthetic musks, and 4 benzotriazoles, from fish bile. Hybrid precipitation on solid phase extraction (SPE) tubes was applied to remove phospholipids and proteins, and a d-SPE procedure was used for further purification. The extraction solvents for the hybrid precipitation/SPE tubes and d-SPE materials were optimized. The method performance for bile samples both with and without enzyme hydrolysis using β-glucuronidase/aryl-sulfatase were validated. The 24 PCPs in fish bile were spiked with standard concentrations of 10 ng/mL, 20 ng/mL, 100 ng/mL, and 200 ng/mL to evaluate recoveries, which ranged from 70 to 120% for 16, 16, 22, and 21 analytes with hydrolysis, respectively, and 70-120% for 14, 15, 23, and 23 analytes without hydrolysis, respectively. The quantification limits for target PCPs were in the range 0.26-7.38 ng/mL [excluding musk xylene (MX) and musk ketone (MK)] and 0.20-9.48 ng/mL (excluding MX and MK) for bile samples with and without enzyme hydrolysis, respectively. After enzyme hydrolysis, 12 PCPs were detected in bile from fish collected from the Yangtze River, with a maximum detected concentration of 460 ng/mL, for triclosan (TCS). The hydrolysis reaction indicated that high percentages of glucuronide and sulfate metabolites for some PCPs, i.e. four parabens and TCS, existed in the bile. Copyright © 2018 Elsevier B.V. All rights reserved.

Preconcentration of DNA using magnetic ionic liquids that are compatible with real-time PCR for rapid nucleic acid quantification.

PubMed

Emaus, Miranda N; Clark, Kevin D; Hinners, Paige; Anderson, Jared L

2018-04-28

Nucleic acid extraction and purification represents a major bottleneck in DNA analysis. Traditional methods for DNA purification often require reagents that may inhibit quantitative polymerase chain reaction (qPCR) if not sufficiently removed from the sample. Approaches that employ magnetic beads may exhibit lower extraction efficiencies due to sedimentation and aggregation. In this study, four hydrophobic magnetic ionic liquids (MILs) were investigated as DNA extraction solvents with the goal of improving DNA enrichment factors and compatibility with downstream bioanalytical techniques. By designing custom qPCR buffers, we directly incorporated DNA-enriched MILs including trihexyl(tetradecyl)phosphonium tris(hexafluoroacetylaceto)nickelate(II) ([P 6,6,6,14 + ][Ni(hfacac) 3 - ]), [P 6,6,6,14 + ] tris(hexafluoroacetylaceto)colbaltate(II) ([Co(hfacac) 3 - ]), [P 6,6,6,14 + ] tris(hexafluoroacetylaceto)manganate(II) ([Mn(hfacac) 3 - ]), or [P 6,6,6,14 + ] tetrakis(hexafluoroacetylaceto)dysprosate(III) ([Dy(hfacac) 4 - ]) into reaction systems, thereby circumventing the need for time-consuming DNA recovery steps. Incorporating MILs into the reaction buffer did not significantly impact the amplification efficiency of the reaction (91.1%). High enrichment factors were achieved using the [P 6,6,6,14 + ][Ni(hfacac) 3 - ] MIL for the extraction of single-stranded and double-stranded DNA with extraction times as short as 2 min. When compared to a commercial magnetic bead-based platform, the [P 6,6,6,14 + ][Ni(hfacac) 3 - ] MIL was capable of producing higher enrichment factors for single-stranded DNA and similar enrichment factors for double-stranded DNA. The MIL-based method was applied for the extraction and direct qPCR amplification of mutation prone-KRAS oncogene fragment in plasma samples. Graphical abstract Magnetic ionic liquid solvents are shown to preconcentrate sufficient KRAS DNA template from an aqueous solution in as short as 2 min without using chaotropic salts or toxic organic solvents. By using custom-designed qPCR buffers, DNA can be directly amplified and quantified from four MILs examined in this study.

Ultrasound-assisted extraction of ginseng saponins from ginseng roots and cultured ginseng cells.

PubMed

Wu, J; Lin, L; Chau, F T

2001-10-01

Ultrasound-assisted extraction was evaluated as a simpler and more effective alternative to conventional extraction methods for the isolation of ginsenosides (saponins) from various types of ginseng. The ginseng samples were extracted with different solvents, under either direct sonication by an ultrasound probe horn or indirect sonication in an ultrasound cleaning bath. The ultrasonic extraction was compared with the conventional method of refluxing boiling solvents in a soxhlet extractor, on the yields of both the total saponin isolated by thin-layer chromatography and the individual ginsenosides by high performance liquid chromatography. It was found that the sonication-assisted extraction of ginseng saponins was about three times faster than the traditional extraction method. The ultrasonic extraction was not only more efficient but also convenient for the recovery and purification of the active ingredients of plant materials. In addition, the sonication-assisted extraction can be carried out at lower temperatures which are favorable for the thermally unstable compounds.

Ultrahigh pressure extraction of lignan compounds from Dysosma versipellis and purification by high-speed counter-current chromatography.

PubMed

Zhu, Qing; Liu, Feng; Xu, Meixia; Lin, Xiaojing; Wang, Xiao

2012-09-15

Ultrahigh pressure extraction (UPE) was employed to extract podophyllotoxin and 4'-demethylpodophyllotoxin from Dysosma versipellis. The effects of extraction parameters including extraction solvents, pressure, time and solid/liquid ratio were investigated using a High Hydrostatic Pressure Processor. The optimal condition for UPE of the target compounds was 80% methanol, 200 MPa of pressure, 1 min of extraction time and 1:12 (g/mL) of solid/liquid ratio. Podophyllotoxin and 4'-demethylpodophyllotoxin in the crude extract were purified by high-speed counter-current chromatography (HSCCC) with a two-phase solvent system composed of petroleum ether-ethyl acetate-methanol-water (10:10:8:12, v/v), and the fractions were analyzed by HPLC, ESI-MS and (1)H NMR. As a result, 73.7 mg podophyllotoxin and 16.5mg 4'-demethylpodophyllotoxin with purities over 96% were obtained from 260 mg crude sample in one-step separation. Copyright © 2012 Elsevier B.V. All rights reserved.

The Baker's Yeast Reduction of Keto-Esters in Organic Solvents: A One Week Research Project for Undergraduate Students

NASA Astrophysics Data System (ADS)

North, Michael

1998-05-01

An experiment has been designed which allows final year undergraduate students to carry out a mini-research project in one week and thus get a flavour of the joys and tribulations of conducting chemical research before they undertake a major research project. The experiment is an investigation into the reduction of alpha- or beta-keto esters using non-fermenting Baker's yeast in petroleum ether. There are a number of advantages to this method of using Baker's yeast, including a reduction in the amount of organic solvent used, and a much simplified purification procedure. During the course of the mini-project, the substrate specificity of the yeast is investigated, and the conditions for the optimisation of a particular keto ester are determined. Each product is analysed by a variety of analytical techniques including polarimetry, IR, NMR, and GC. In addition, the use of correct stereochemical nomenclature to describe prochiral, and chiral compounds as well as chemical reactions are discussed.

[Extraction of 10-Deacetyl Baccatin by Supercritical CO2 from Taxus yunnanensis Branches and Leaves].

PubMed

Tang, Yang-qin; Li, Hai-chi; Huang, Wen-jie; Xiong, Yan; Ge, Fa-huan

2015-04-01

To study the supercritical CO2 fluids extraction (SFE) method to extract the components from Taxus yunnanensis. Medicinal meterials were extracted by supercritical CO2, and then purified by industrial chromatography. Using the extraction yield of 10-DAB as the index,single factor test was carried out to investigate the effect of co-solvent, extraction time, extraction pressure, extraction temperature, pressure and temperature of separation kettle I. Then orthogonal experiment was used to optimize the best extraction condition. The suitable extraction condition was as follows: the ratio of co-solvent (80% ethanol) amount and the madicinal materials was 3: 1, Separation kettle I pressure was 14 MPa, separation kettle I temperature was 40 °C, extraction pressure was 25 MPa, extraction temperature was 60 T and extraction time was 90 min. The extract was separated by industrial chromatographic and then crystallized. The supercritical CO2 extraction and purification process of 10-DAB were simple and feasible.

Produced Water Treatment Using the Switchable Polarity Solvent Forward Osmosis (SPS FO) Desalination Process: Preliminary Engineering Design Basis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wendt, Daniel; Adhikari, Birendra; Orme, Christopher

Switchable Polarity Solvent Forward Osmosis (SPS FO) is a semi-permeable membrane-based water treatment technology. INL is currently advancing SPS FO technology such that a prototype unit can be designed and demonstrated for the purification of produced water from oil and gas production operations. The SPS FO prototype unit will used the thermal energy in the produced water as a source of process heat, thereby reducing the external process energy demands. Treatment of the produced water stream will reduce the volume of saline wastewater requiring disposal via injection, an activity that is correlated with undesirable seismic events, as well as generatemore » a purified product water stream with potential beneficial uses. This paper summarizes experimental data that has been collected in support of the SPS FO scale-up effort, and describes how this data will be used in the sizing of SPS FO process equipment. An estimate of produced water treatment costs using the SPS FO process is also provided.« less

Preparative isolation and purification of theaflavins and catechins by high-speed countercurrent chromatography.

PubMed

Wang, Kunbo; Liu, Zhonghua; Huang, Jian-an; Dong, Xinrong; Song, Lubing; Pan, Yu; liu, Fang

2008-05-15

High-speed countercurrent chromatography (HSCCC) has been applied for the separation of theaflavins and catechins. The HSCCC run was carried out with a two-phase solvent system composed of hexane-ethyl acetate-methanol-water-acetic acid (1:5:1:5:0.25, v/v) by eluting the lower aqueous phase at 2 ml/min at 700 rpm. The results indicated that pure theaflavin, theaflavins-3-gallate, theaflavins-3'-gallate and theaflavin-3,3'-digallate could be obtained from crude theaflavins sample and black tea. The structures of the isolated compounds were positively confirmed by (1)H NMR and (13)C NMR, MS analysis, HPLC data and TLC data. Meanwhile, catechins including epigallocatechin gallate, gallocatechin gallate, epicatechin gallate and epigallocatechin were isolated from the aqueous extract of green tea by using the same solvent system. This study developed a modified method combined with enrichment theaflavins method by using HSCCC for separation of four individual theaflavins, especially for better separation of theaflavins monogallates.

Fabrication of cellulose-based aerogels from waste newspaper without any pretreatment and their use for absorbents.

PubMed

Jin, Chunde; Han, Shenjie; Li, Jingpeng; Sun, Qingfeng

2015-06-05

Cellulose-based aerogel (CBA) was prepared from waste newspaper (WNP) without any pretreatment using 1-allyl-3-methyimidazolium chloride (AmImCl) as a solvent via regeneration and an environmentally friendly freeze-drying method. After being treated with trimethylchlorosilane (TMCS) via a simple thermal chemical vapor deposition process, the resulting CBAs were rendered both hydrophobic and oleophilic. Successful silanization on the surface of the porous CBA was verified by a variety of techniques including scanning electron microscopy (SEM), energy-dispersive X-ray analysis (EDX), and water contact angle (WCA) measurements. As a result, the silane-coated, interconnected CBAs not only exhibited good absorption performance for oils (e.g., waste engine oil), but also showed absorption capacity for organic solvents such as chloroform (with a representative weight gain ranging from 11 to 22 times of their own dry weight), making them diversified absorbents for potential applications including sewage purification. Copyright © 2015 Elsevier Ltd. All rights reserved.

Superhydrophobic silica wool—a facile route to separating oil and hydrophobic solvents from water

NASA Astrophysics Data System (ADS)

Crick, Colin R.; Bhachu, Davinder S.; Parkin, Ivan P.

2014-12-01

Silica microfiber wool was systematically functionalized in order to provide an extremely water repellent and oleophilic material. This was carried out using a two-step functionalization that was shown to be a highly effective method for generating an intense water repulsion and attraction for oil. A demonstration of the silica wools application is shown through the highly efficient separation of oils and hydrophobic solvents from water. Water is confined to the extremities of the material, while oil is absorbed into the voids within the wool. The effect of surface functionalization is monitored though observing the interaction of the material with both oils and water, in addition to scanning electron microscope images, x-ray photoelectron spectroscopy and energy dispersive x-ray analysis. The material can be readily utilized in many applications, including the cleaning of oil spills and filtering during industrial processes, as well as further water purification tasks—while not suffering the losses of efficiency observed in current leading polymeric materials.

Estimation of the nucleation kinetics for the anti-solvent crystallisation of paracetamol in methanol/water solutions

NASA Astrophysics Data System (ADS)

Ó'Ciardhá, Clifford T.; Frawley, Patrick J.; Mitchell, Niall A.

2011-08-01

In this work the primary nucleation kinetics have been estimated for the anti-solvent crystallisation of paracetamol in methanol-water solutions from metastable zone widths (MSZW) and induction times at 25 °C. Laser back-scattering via a focused beam reflectance Measurement (FBRM ®) is utilised to detect the onset of nucleation. The theoretical approach of Kubota was employed to estimate the nucleation kinetics, which accounts for the sensitivity of the nucleation detection technique. This approach is expanded in this work to analyse the induction time for an anti-solvent crystallisation process. Solvent composition is known to have a significant impact on the measured induction times and MSZW. The induction time in this paper was measured from 40% to 70% mass water and the MSZW is measured from 40% to 60% mass water. The primary focus of the paper was to gauge the extent of how solvent composition affects nucleation kinetics so that this effect may be incorporated into a population balance model. Furthermore, the effects of solvent composition on the estimated nucleation rates are investigated. The primary nucleation rates were found to decrease with dynamic solvent composition, with the extent of their reduction linked to the gradient of the solubility curve. Finally, both MSZW and induction time methods have been found to produce similar estimates for the nucleation parameters.

Phenolic compounds containing/neutral fractions extract and products derived therefrom from fractionated fast-pyrolysis oils

DOEpatents

Chum, Helena L.; Black, Stuart K.; Diebold, James P.; Kreibich, Roland E.

1993-01-01

A process for preparing phenol-formaldehyde novolak resins and molding compositions in which portions of the phenol normally contained in said resins are replaced by a phenol/neutral fractions extract obtained from fractionating fast-pyrolysis oils. The fractionation consists of a neutralization stage which can be carried out with aqueous solutions of bases or appropriate bases in the dry state, followed by solvent extraction with an organic solvent having at least a moderate solubility parameter and good hydrogen bonding capacity. Phenolic compounds-containing/neutral fractions extracts obtained by fractionating fast-pyrolysis oils from a lignocellulosic material, is such that the oil is initially in the pH range of 2-4, being neutralized with an aqueous bicarbonate base, and extracted into a solvent having a solubility parameter of approximately 8.4-9.11 [cal/cm.sup.3 ].sup.1/2 with polar components in the 1.8-3.0 range and hydrogen bonding components in the 2-4.8 range and the recovery of the product extract from the solvent with no further purification being needed for use in adhesives and molding compounds. The product extract is characterized as being a mixture of very different compounds having a wide variety of chemical functionalities, including phenolic, carbonyl, aldehyde, methoxyl, vinyl and hydroxyl. The use of the product extract on phenol-formaldehyde thermosetting resins is shown to have advantages over the conventional phenol-formaldehyde resins.

Permeability and electrical properties of planar lipid membranes from thylakoid lipids.

PubMed Central

Fuks, B; Homblé, F

1994-01-01

Electrical measurements were carried out on planar lipid membranes from thylakoid lipids. The specific capacitance of membranes formed from decane-containing monogalactosyldiacylglycerol (MGDG), which accounts for 57% of the total lipid content of thylakoids, showed that it adopted a bilayer structure. Solvent-free bilayers of MGDG were not formed, with very rare exceptions, indicating that decane is required to stabilize the planar conformation. However, this cone-shaped lipid produces bilayer structures in combination with other cylindrical thylakoid lipids even in the absence of organic solvent. We compared the properties of solvent-free and decane-containing bilayers from MGDG, soybean lecithin, and the quaternary mixture of lipids similar to that found in vivo. The conductance of decane-MGDG was 26 times higher than that of decane-lecithin. The flux through the decane-lecithin bilayer was found to be slightly dependent on pH, whereas the decane-MGDG membrane was not. The specific conductance of bilayers formed from the quaternary mixture of lipids was 5 to 10 times larger than lecithin (with alkane or not). Further experiments with bilayers made in the presence of a KCl gradient showed that decane-MGDG, decane-MGDG/DGDG/SQDG/PG, and solvent-free MGDG/DGDG/SQDG/PG were cation-selective. The permeability coefficient for potassium ranged from 4.9 to 8.3 x 10(-11) cm s-1. The permeability coefficient for protons in galactolipids, however, was determined to be about six orders of magnitude higher than the value for potassium ions. The HCl permeation mechanism through the lipid membranes was determined from diffusion potentials measured in HCl gradients. Our results suggest that HCl was not transported as neutral molecules. The data is discussed with regard to the function of galactolipids in the ion transport through thylakoid membranes. PMID:8061192

Monitoring gradient profile on-line in micro- and nano-high performance liquid chromatography using conductivity detection.

PubMed

Zhang, Min; Chen, Apeng; Lu, Joann J; Cao, Chengxi; Liu, Shaorong

2016-08-19

In micro- or nano-flow high performance liquid chromatography (HPLC), flow-splitters and gradient elutions are commonly used for reverse phase HPLC separations. When a flow splitter was used at a high split-ratio (e.g., 1000:1 or higher), the actual gradient may deviate away from the programmed gradient. Sometimes, mobile phase concentrations can deviate by as much as 5%. In this work, we noticed that the conductivity (σ) of a gradient decreased with the increasing organic-solvent fraction (φ). Based on the relationship between σ and φ, a method was developed for monitoring gradient profile on-line to record any deviations in these HPLC systems. The conductivity could be measured by a traditional conductivity detector or a capacitively coupled contactless conductivity detector (C(4)D). The method was applied for assessing the performance of an electroosmotic pump (EOP) based nano-HPLC. We also observed that σ value of the gradient changed with system pressure; a=0.0175ΔP (R(2)=0.964), where a is the percentage of the conductivity increase and ΔP is the system pressure in bar. This effect was also investigated. Copyright © 2016. Published by Elsevier B.V.

Spiral counter-current chromatography of small molecules, peptides and proteins using the spiral tubing support rotor.

PubMed

Knight, Martha; Finn, Thomas M; Zehmer, John; Clayton, Adam; Pilon, Aprile

2011-09-09

An important advance in countercurrent chromatography (CCC) carried out in open flow-tubing coils, rotated in planetary centrifuges, is the new design to spread out the tubing in spirals. More spacing between the tubing was found to significantly increase the stationary phase retention, such that now all types of two-phase solvent systems can be used for liquid-liquid partition chromatography in the J-type planetary centrifuges. A spiral tubing support (STS) frame with circular channels was constructed by laser sintering technology into which FEP tubing was placed in 4 spiral loops per layer from the bottom to the top and a cover affixed allowing the tubing to connect to flow-tubing of the planetary centrifuge. The rotor was mounted and run in a P.C. Inc. type instrument. Examples of compounds of molecular weights ranging from <300 to approximately 15,000 were chromatographed in appropriate two-phase solvent systems to assess the capability for separation and purification. A mixture of small molecules including aspirin was completely separated in hexane-ethyl acetate-methanol-water. Synthetic peptides including a very hydrophobic peptide were each purified to a very high purity level in a sec-butanol solvent system. In the STS rotor high stationary phase retention was possible with the aqueous sec-butanol solvent system at a normal flow rate. Finally, the two-phase aqueous polyethylene glycol-potassium phosphate solvent system was applied to separate a protein from a lysate of an Escherichia coli expression system. These experiments demonstrate the versatility of spiral CCC using the STS rotor. Copyright © 2011 Elsevier B.V. All rights reserved.

Isolation and purification of prenylated phenolics from Amorpha fruticosa by high-speed counter-current chromatography.

PubMed

Chen, Chu; Wu, Yan; Chen, Yang; Du, Leilei

2015-08-01

Prenylated phenolics such as amorfrutins are recently identified potent anti-inflammatory and antidiabetic natural products. In this work, high-speed counter-current chromatography was investigated for the isolation and purification of prenylated phenolics from the fruits of Amorpha fruticosa by using a two-phase solvent system composed of n-hexane/ethanol/water (5:4:1, v/v). As a result, 14.2 mg of 5,7-dihydroxy-8-geranylflavanone, 10.7 mg of amorfrutin A and 17.4 mg of amorfrutin B were obtained from 200 mg of n-hexane-soluble crude extract in one step within 250 min. The purities of 5,7-dihydroxy-8-geranylflavanone, amorfrutins A and B were 95.2, 96.7 and 97.1%, respectively, as determined by ultra high performance liquid chromatography. The structural identification was performed by mass spectrometry and (1) H and (13) C NMR spectroscopy. The results indicated that the established method is an efficient and convenient way to purified prenylated phenolics from A. fruticosa extract. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Isolation and purification of diastereoisomeric flavonolignans from silymarin by binary-column recycling preparative high-performance liquid chromatography.

PubMed

Zhao, Weiquan; Yang, Guang; Zhong, Fanyi; Yang, Nan; Zhao, Xin; Qi, Yunpeng; Fan, Guorong

2014-09-01

Silymarin extracted from Silybum marianum (L.) Gaertn consists of a large number of flavonolignans, of which diastereoisomeric flavonolignans including silybin A and silybin B, and isosilybin A and isosilybin B are the main bioactive components, whose preparation from the crude extracts is still a difficult task. In this work, binary-column recycling preparative high-performance liquid chromatography systems without sample loop trapping, where two columns were switched alternately via one or two six-port switching valves, were established and successfully applied to the isolation and purification of the four diastereoisomeric flavonolignans from silymarin. The proposed system showed significant advantages over conventional preparative high-performance liquid chromatography with a single column in increasing efficiency and reducing the cost. To obtain the same amounts of products, the proposed system spends only one tenth of the time that the conventional system spends, and needs only one eleventh of the solvent that the conventional system consumes. Using the proposed system, the four diastereoisomers were successfully isolated from silymarin with purities over 98%. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[Simultaneous isolation and purification of gallic acid and brevifolincarboxylic acid from Polygonum capitatum by high-speed counter-current chromatography].

PubMed

Chen, Xinxia; Zhang, Liyan; Wan, Jinzhi; Liang, Bin; Xie, Yu

2010-08-01

To isolate and purify gallic acid and brevifolincarboxylic acid simultaneously by high-speed counter-current chromatography (HSCCC) from a crude extract of Polygonum capitatum. The biphasic solvent system composed of ethyl acetate-n-butanol-0.44% acetic acid (3:1:5) was used at a flow rate of 2.0 mL x min(-1), while the aqueous phase was selected as the mobile phase and the apparatus was rotated at 860 r x min(-1). The effluent was detected at 272 nm. 51.5 mg of gallic acid and 5.9 mg of brevifolincarboxylic acid were separated from 1.07 g of the crude extract with the purities of 99.7% and 97.5%, respectively, while brevifolincarboxylic acid was obtained firstly from the genus Polygonum. The structures of the compounds were identified by ultraviolet spectrometry (UV), infra-red spectrometry (IR), liquid chromatography/mass spectrometry (LC/MS), time-of-flight mass spectrometry( TOF-MS), 1H-nuclear magnetic resonance (NMR) and 13C-NMR. This method is feasible and rapid for isolation and purification of gallice acid and brevifolincarboxylil acid.

Salting-out extraction of allicin from garlic (Allium sativum L.) based on ethanol/ammonium sulfate in laboratory and pilot scale.

PubMed

Li, Fenfang; Li, Qiao; Wu, Shuanggen; Tan, Zhijian

2017-02-15

Salting-out extraction (SOE) based on lower molecular organic solvent and inorganic salt was considered as a good substitute for conventional polymers aqueous two-phase extraction (ATPE) used for the extraction of some bioactive compounds from natural plants resources. In this study, the ethanol/ammonium sulfate was screened as the optimal SOE system for the extraction and preliminary purification of allicin from garlic. Response surface methodology (RSM) was developed to optimize the major conditions. The maximum extraction efficiency of 94.17% was obtained at the optimized conditions for routine use: 23% (w/w) ethanol concentration and 24% (w/w) salt concentration, 31g/L loaded sample at 25°C with pH being not adjusted. The extraction efficiency had no obvious decrease after amplification of the extraction. This ethanol/ammonium sulfate SOE is much simpler, cheaper, and effective, which has the potentiality of scale-up production for the extraction and purification of other compounds from plant resources. Copyright © 2016 Elsevier Ltd. All rights reserved.

Preparative Isolation and Purification of Flavone C-Glycosides from the Leaves of Ficus microcarpa L. f by Medium-Pressure Liquid Chromatography, High-Speed Countercurrent Chromatography, and Preparative Liquid Chromatography

PubMed Central

Wang, Xiaohong; Liang, Yong; Zhu, Licai; Xie, Huichun; Li, Hang; He, Junting; Pan, Man; Zhang, Tianyou; Ito, Yoichiro

2009-01-01

Combined with medium-pressure liquid chromatography (MPLC) and preparative high-performance liquid chromatography (perp-HPLC), high-speed countercurrent chromatography (HSCCC) was applied for separation and purification of flavone C-glycosides from the crude extract of leaves of Ficus microcarpae L. f. HSCCC separation was performed on a two-phase solvent system composed of methyl tert- butyl ether - ethyl acetate – 1-butanol – acetonitrile – 0.1% aqueous trifluoroacetic acid at a volume ratio of 1:3:1:1:5. Partially resolved peak fractions from HSCCC separation were further purified by preparative HPLC. Four well-separated compounds were obtained and their purities were determined by HPLC. The purities of these peaks were 97.28%, 97.20%, 92.23%, and 98.40%.. These peaks were characterized by ESI-MSn. According to the reference, they were identified as orientin (peak I), isovitexin-3″-O-glucopyranoside (peak II), isovitexin (peak III), and vitexin (peak IV), yielded 1.2 mg, 4.5 mg, 3.3 mg, and 1.8 mg, respectively. PMID:20190866

Controlled synthesis of flexible graphene aerogels macroscopic monolith as versatile agents for wastewater treatment

NASA Astrophysics Data System (ADS)

Dong, Shuying; Xia, Longji; Guo, Teng; Zhang, Fangyuan; Cui, Lingfang; Su, Xianfa; Wang, Dong; Guo, Wei; Sun, Jianhui

2018-07-01

Emerging applications for environmental purification require agents that not only possess high decontamination efficiency, but also are capable of withstanding mechanical deformation without secondary pollution and degradation of performance. To this end, we have controlled synthesis of mechanically flexible graphene aerogel (GA) by vacuum freeze-drying of their hydrogel precursors obtained from heating the aqueous mixtures of graphene oxide (GO) and Vitamin C (VC) without stirring. Through the adaptable process conditions, such as the particle size of carbon, GO concentration, dosage of reducing agent and solution pH, the highly porous, ultralight and mechanically flexible GA are synthesized. Owing to the porous, robust and stable structure, the resulting GA show very promising performance in water purification including enrichment of organic liquid solvents (alcohols, oil and alkanes), removal of hexavalent chromium Cr(VI) and purified industrial wastewater, as well as flexible conductors. The successful creation of the GA may provide new insights into the design of carbon-based aerogels for various applications, as the GA can be prepared via a very simple procedure and available in macroscopic diverse morphologies with tunable porosity.

Isolation, one-step affinity purification, and characterization of a polyextremotolerant laccase from the halophilic bacterium Aquisalibacillus elongatus and its application in the delignification of sugar beet pulp.

PubMed

Rezaei, Shahla; Shahverdi, Ahmad Reza; Faramarzi, Mohammad Ali

2017-04-01

The aim of the present work was to study the ability of a halophilic bacterial laccase to efficient delignification in extreme conditions. Here, a highly stable extracellular laccase showing ligninolytic activity from halophilic Aquisalibacillus elongatus is described. The laccase production was strongly influenced by NaCl and CuSO 4 and under optimal conditions reached 4.8UmL -1 . The monomeric enzyme of 75kDa was purified by a synthetic affinity column with 68.2% yield and 99.8-fold purification. The enzyme showed some valuable features viz. stability against a wide range of organic solvents, salts, metals, inhibitors, and surfactants and specificity to a wide spectrum of substrates diverse in structure and redox potential. It retained more than 50% of the original activity at 25-75°C and pH 5.0-10.0. Furthermore, the enzyme was found to be effective in the delignification of sugar beet pulp in an ionic liquid that makes it useful for industrial applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

One-step fabrication of multifunctional composite polyurethane spider-web-like nanofibrous membrane for water purification.

PubMed

Pant, Hem Raj; Kim, Han Joo; Joshi, Mahesh Kumar; Pant, Bishweshwar; Park, Chan Hee; Kim, Jeong In; Hui, K S; Kim, Cheol Sang

2014-01-15

A stable silver-doped fly ash/polyurathene (Ag-FA/PU) nanocomposite multifunctional membrane is prepared by a facile one-step electrospinning process using fly ash particles (FAPs). Colloidal solution of PU with FAPs and Ag metal precursor was subjected to fabricate nanocomposite spider-web-like membrane using electrospinning process. Presence of N,N-dimethylformamide (solvent of PU) led to reduce silver nitrate into Ag NPs. Incorporation of Ag NPs and FAPs through electrospun PU fibers is proven through electron microscopy and spectroscopic techniques. Presence of these NPs on PU nanofibers introduces several potential physicochemical properties such as spider-web-like nano-neeting for NPs separation, enhanced absorption capacity to remove carcinogenic arsenic (As) and toxic organic dyes, and antibacterial properties with reduce bio-fouling for membrane filter application. Preliminary observations used for above-mentioned applications for water treatment showed that it will be an economically and environmentally friendly nonwoven matrix for water purification. This simple approach highlights new avenues about the utilization of one pollutant material to control other pollutants in scalable and inexpensive ways. Copyright © 2013 Elsevier B.V. All rights reserved.

Ultrastable, highly luminescent quantum dot composites based on advanced surface manipulation strategy for flexible lighting-emitting.

PubMed

Kong, Lingqing; Zhang, Lin; Meng, Zhaohui; Xu, Chuan; Lin, Naibo; Liu, Xiang-Yang

2018-08-03

Although quantum dots (QDs) have remarkable potential application in flexible light emitting diodes (LED), the loss of solvent-protected QDs leads to low quantum yield (QY) and poor stability, severely restricting the development. Flexible QD LEDs (Q-LEDs) with three primary colors were fabricated by mixing CdS/ZnS, CdSe@ZnS/ZnS, and CdSe/CdS QDs with polydimethylsiloxane (PDMS) by in situ hydrosilylation based surface manipulation strategy, which endows the device with highly ultrastable and luminescent performance. The surface manipulation strategy mainly includes the control of solvent dosage, purification times of QDs, concentration of QDs in PDMS, and oxidation on the preparation process of the QDs and PDMS composites. The highest QY of CdSe@ZnS/ZnS-PDMS composite is 82.03%, higher than the QY (80%) of the QD solution. After UV bleaching, organic solvents (acetone, ethanol and water), and heating treatment, the QYs of the QDs and PDMS maintain a high value, manifesting their good stability. Q-LED hybrid light-emitting devices were further fabricated by a molding technique demonstrating satisfied current and thermal stability. Flexible Q-LEDs can be expended to other shapes, such as fibers and blocks, indicating the huge potential of QD-polymer composites for light sources and displays etc.

Preparation of factor VII concentrate using CNBr-activated Sepharose 4B immunoaffinity chromatography

PubMed Central

Mousavi Hosseini, Kamran; Nasiri, Saleh

2015-01-01

Background: Factor VII concentrates are used in patients with congenital or acquired factor VII deficiency or treatment of hemophilia patients with inhibitors. In this research, immunoaffinity chromatography was used to purify factor VII from prothrombin complex (Prothrombin- Proconvertin-Stuart Factor-Antihemophilic Factor B or PPSB) which contains coagulation factors II, VII, IX and X. The aim of this study was to improve purity, safety and tolerability as a highly purified factor VII concentrate. Methods: PPSB was prepared using DEAE-Sephadex and was used as the starting material for purification of coagulation factor VII. Prothrombin complex was treated by solvent/detergent at 24°C for 6 h with constant stirring. The mixture of PPSB in the PBS buffer was filtered and then chromatographed using CNBr-activated Sepharose 4B coupled with specific antibody. Factors II, IX, VII, X and VIIa were assayed on the fractions. Fractions of 48-50 were pooled and lyophilized as a factor VII concentrate. Agarose gel electrophoresis was performed and Tween 80 was measured in the factor VII concentrate. Results: Specific activity of factor VII concentrate increased from 0.16 to 55.6 with a purificationfold of 347.5 and the amount of activated factor VII (FVIIa) was found higher than PPSB (4.4-fold). Results of electrophoresis on agarose gel indicated higher purity of Factor VII compared to PPSB; these finding revealed that factor VII migrated as alpha-2 proteins. In order to improve viral safety, solvent-detergent treatment was applied prior to further purification and nearly complete elimination of tween 80 (2 μg/ml). Conclusion: It was concluded that immuonoaffinity chromatography using CNBr-activated Sepharose 4B can be a suitable choice for large-scale production of factor VII concentrate with higher purity, safety and activated factor VII. PMID:26034723

Preparation of factor VII concentrate using CNBr-activated Sepharose 4B immunoaffinity chromatography.

PubMed

Mousavi Hosseini, Kamran; Nasiri, Saleh

2015-01-01

Factor VII concentrates are used in patients with congenital or acquired factor VII deficiency or treatment of hemophilia patients with inhibitors. In this research, immunoaffinity chromatography was used to purify factor VII from prothrombin complex (Prothrombin- Proconvertin-Stuart Factor-Antihemophilic Factor B or PPSB) which contains coagulation factors II, VII, IX and X. The aim of this study was to improve purity, safety and tolerability as a highly purified factor VII concentrate. PPSB was prepared using DEAE-Sephadex and was used as the starting material for purification of coagulation factor VII. Prothrombin complex was treated by solvent/detergent at 24°C for 6 h with constant stirring. The mixture of PPSB in the PBS buffer was filtered and then chromatographed using CNBr-activated Sepharose 4B coupled with specific antibody. Factors II, IX, VII, X and VIIa were assayed on the fractions. Fractions of 48-50 were pooled and lyophilized as a factor VII concentrate. Agarose gel electrophoresis was performed and Tween 80 was measured in the factor VII concentrate. Specific activity of factor VII concentrate increased from 0.16 to 55.6 with a purificationfold of 347.5 and the amount of activated factor VII (FVIIa) was found higher than PPSB (4.4-fold). RESULTS of electrophoresis on agarose gel indicated higher purity of Factor VII compared to PPSB; these finding revealed that factor VII migrated as alpha-2 proteins. In order to improve viral safety, solvent-detergent treatment was applied prior to further purification and nearly complete elimination of tween 80 (2 μg/ml). It was concluded that immuonoaffinity chromatography using CNBr-activated Sepharose 4B can be a suitable choice for large-scale production of factor VII concentrate with higher purity, safety and activated factor VII.

HPLC-UV Method for the Identification and Screening of Hydroquinone, Ethers of Hydroquinone and Corticosteroids Possibly Used as Skin-Whitening Agents in Illicit Cosmetic Products.

PubMed

Gimeno, Pascal; Maggio, Annie-Françoise; Bancilhon, Marjorie; Lassu, Nelly; Gornes, Hervé; Brenier, Charlotte; Lempereur, Laurent

2016-03-01

Corticosteroids, hydroquinone and its ethers are regulated in cosmetics by the Regulation 1223/2009. As corticosteroids are forbidden to be used in cosmetics and cannot be present as contaminants or impurities, an identification of one of these illicit compounds deliberately introduced in these types of cosmetics is enough for market survey control. In order to quickly identify skin-whitening agents present in illegal cosmetics, this article proposes an HPLC-UV method for the identification and screening of hydroquinone, 3 ethers of hydroquinone and 39 corticosteroids that may be found in skin-whitening products. Two elution gradients were developed to separate all compounds. The main solvent gradient (A) allows the separation of 39 compounds among the 43 compounds considered in 50 min. Limits of detection on skin-whitening cosmetics are given. For compounds not separated, a complementary gradient elution (B) using the same solvents is proposed. Between 2004 and 2009, a market survey on "skin-whitening cosmetic" was performed on 150 samples and highlights that more than half of the products tested do not comply with the Cosmetic Regulation 1223/2009 (amending the Council Directive 76/768/EEC). © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Refolding and simultaneous purification of recombinant human proinsulin from inclusion bodies on protein-folding liquid-chromatography columns.

PubMed

Yuan, Jie; Zhou, Huifang; Yang, Yicong; Li, Weimin; Wan, Yi; Wang, Lili

2015-05-01

Protein-folding liquid chromatography (PFLC) is an effective and scalable method for protein renaturation with simultaneous purification. However, it has been a challenge to fully refold inclusion bodies in a PFLC column. In this work, refolding with simultaneous purification of recombinant human proinsulin (rhPI) from inclusion bodies from Escherichia coli were investigated using the surface of stationary phases in immobilized metal ion affinity chromatography (IMAC) and high-performance size-exclusion chromatography (HPSEC). The results indicated that both the ligand structure on the surface of the stationary phase and the composition of the mobile phase (elution buffer) influenced refolding of rhPI. Under optimized chromatographic conditions, the mass recoveries of IMAC column and HPSEC column were 77.8 and 56.8% with purifies of 97.6 and 93.7%, respectively. These results also indicated that the IMAC column fails to refold rhPI, and the HPSEC column enables efficient refolding of rhPI with a low-urea gradient-elution method. The refolded rhPI was characterized by circular dichroism spectroscopy. The molecular weight of the converted human insulin was further confirmed with SDS-18% PAGE, Matrix-Assisted Laser Desorption/ Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS) and the biological activity assay by HP-RPLC. Copyright © 2014 John Wiley & Sons, Ltd.

Purification and characterization of chromatin-bound DNA-dependent RNA polymerase I from parsley (Petroselinum crispum). Influence of nucleoside triphosphates.

PubMed Central

Grossmann, K; Friedrich, H; Seitz, U

1980-01-01

The isolation and purification of DNA-dependent RNA polymerase I (EC 2.7.7.6) from parsley (Petroselinum crispum) callus cells grown in suspension culture is described. The enzyme was solubilized from isolated chromatin. Purification was achieved by using DEAE- and phospho-cellulose in batches, followed by column chromatography on DEAE- and phospho-cellulose (two columns) and density-gradient centrifugation. The highly purified enzyme was stable over several months. The properties of purified parsley RNA polymerase I were investigated. Optimum concentration for Mn2+ was 1 mM, and for Mg2+ 4-6 mM, Mn2+ was slightly more stimulatory than Mg2+. The enzyme was most active at low ionic strengths [10-20 mM-(NH4)SO4]. The influence of various phosphates was tested: pyrophosphate inhibited RNA polymerase at low concentrations, whereas orthophosphate had no effect on the enzyme activity. ADP was slightly inhibitory, and AMP had no effect on the enzyme reaction. Nucleoside triphosphates and bivalent cations in equimolar concentrations in the range 4-11 mM did not influence the RNA synthesis in vitro. Free nucleoside triphosphates in excess of this 1:1 ratio inhibited the enzyme activity, unlike free bivalent cations, which stimulated RNA polymerase I. PMID:7470092

Efficient purification of Apolipoprotein A1 (ApoA1) from plasma by HEA HyperCel™: An alternative approach.

PubMed

G, Arun Govind; Kamalanathan, Agamudi Shivasankaran; Vijayalakshmi, Mookambeswaran Arunachalam; Venkataraman, Krishnan

2018-01-15

HDL-ApoA1 plays a pivotal role in the prevention of atherosclerosis and cardiovascular diseases. ApoA1 purification from blood plasma has always remained tedious, involving multiple steps, large volumes of plasma and substantial loss in the final yield of pure ApoA1. In this study, a two-step method has been developed and optimized for the purification of ApoA1 from plasma. Plasma was first subjected to 60% ammonium sulphate (NH 4 ) 2 SO 4 precipitation and subsequently, ApoA1 was recovered using mixed mode chromatographic sorbent, HEA HyperCel™. ApoA1 was found to be enriched in 60% (NH 4 ) 2 SO 4 supernatant that was dialyzed and injected onto HEA sorbent with 50 mM phosphate buffer pH 7.4. The bound proteins were eluted by decreasing the pH in step-gradient from pH 7.4 to pH 4.0 and subsequently to pH 3.5 using 50 mM sodium acetate buffer. Gel electrophoresis showed elution of homogeneous apoA1 at pH 3.5, with purity and yield of 63%. An interesting feature of this approach is that the purified ApoA1 was monomeric with a mass of 28,079.30 Da as confirmed by MS analysis. This simple and efficient method of purification of apoA1 serves as an alternative method which can be combined with traditional approaches and has a great potential for biochemical and clinical studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Purification, crystallization and preliminary X-ray diffraction analysis of the kinase domain of human tousled-like kinase 2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garrote, Ana M.; Redondo, Pilar; Montoya, Guillermo, E-mail: gmontoya@cnio.es

2014-02-19

The C-terminal kinase domain of TLK2 (a human tousled-like kinase) has been cloned and overexpressed in Escherichia coli followed by purification to homogeneity. Crystallization experiments in the presence of ATP-γ-S yielded crystals suitable for X-ray diffraction analysis belonging to two different space groups: tetragonal I4{sub 1}22 and cubic P2{sub 1}3. Tousled-like kinases (TLKs) are an evolutionarily conserved family of serine/threonine protein kinases involved in chromatin dynamics, including DNA replication and repair, transcription and chromosome segregation. The two members of the family reported in humans, namely TLK1 and TLK2, localize to the cell nucleus and are capable of forming homo- ormore » hetero-oligomers by themselves. To characterize the role of TLK2, its C-terminal kinase domain was cloned and overexpressed in Escherichia coli followed by purification to homogeneity. Crystallization experiments in the presence of ATP-γ-S yielded crystals suitable for X-ray diffraction analysis belonging to two different space groups: tetragonal I4{sub 1}22 and cubic P2{sub 1}3. The latter produced the best diffracting crystal (3.4 Å resolution using synchrotron radiation), with unit-cell parameters a = b = c = 126.05 Å, α = β = γ = 90°. The asymmetric unit contained one protein molecule, with a Matthews coefficient of 4.59 Å{sup 3} Da{sup −1} and a solvent content of 73.23%.« less

One-step separation and purification of three lignans and one flavonol from Sinopodophyllum emodi by medium-pressure liquid chromatography and high-speed counter-current chromatography.

PubMed

Wang, Ping; Liu, Yongling; Chen, Tao; Xu, Wenhua; You, Jinmao; Liu, Yongjun; Li, Yulin

2013-01-01

Lignans and flavonols are the primary constituents of Sinopodophyllum emodi and have been used as cathartic, anthelmintic, chemotherapeutic and anti-hypertensive compounds. Although these compounds have been isolated, there have been no reports on the separation of 4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin and kaempferol in one step by medium-pressure liquid chromatography (MPLC) and high-speed counter-current chromatography (HSCCC). Development of an efficient method for the preparative separation and purification of three lignans and one flavonol from S. emodi. The precipitate of crude extracts was first separated by MPLC into four parts, numbered GJ-1, GJ-2, GJ-3 and GJ-4. GJ-1 was separated and purified by HSCCC using a solvent system composed of n-hexane:ethyl acetate:methanol:water (1.75:1.5:1:0.75, v/v/v/v). The purities of the target compounds were assessed using high-performance liquid chromatography (HPLC) and chemical structures were identified by (1) H-NMR and (13) C-NMR. The HSCCC and MPLC methods were successfully used for the preparative separation and purification of 4'-demethyl podophyllotoxin (8.5 mg, 92.4%), podophyllotoxin (40.1 mg, 92.1%), deoxypodophyllotoxin (4.6 mg, 98.1%), and kaempferol (1.6 mg, 96.7%) from a 100 mg sample. Three lignans (4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin) and one flavonol (kaempferol) were successfully isolated by HSCCC and MPLC in one step. Copyright © 2013 John Wiley & Sons, Ltd.

At-line hyphenation of high-speed countercurrent chromatography with Sephadex LH-20 column chromatography for bioassay-guided separation of antioxidants from vine tea (Ampelopsis grossedentata).

PubMed

Ma, Ruyi; Zhou, Rongrong; Tong, Runna; Shi, Shuyun; Chen, Xiaoqing

2017-01-01

Vine tea (Ampelopsis grossedentata), a widely used healthy tea, beverage and herbal medicine, exhibited strong antioxidant activity. However, systematic purification of antioxidants, especially for those with similar structures or polarities, is a challenging work. Here, we present a novel at-line hyphenation of high-speed countercurrent chromatography with Sephadex LH-20 column chromatography (HSCCC-Sephadex LH-20 CC) for rapid and efficient separation of antioxidants from vine tea target-guided by 1,1-diphenyl-2-picryl-hydrazyl radical-high performance liquid chromatography (DPPH-HPLC) experiment. A makeup pump, a six-port switching valve and a trapping column were served as interface. The configuration had no operational time and mobile phase limitations between two dimensional chromatography and showed great flexibility without tedious sample-handling procedure. Seven targeted antioxidants were firstly separated by stepwise HSCCC using petroleum ether-ethyl acetate-methanol-water (4:9:4:9, v/v/v/v) and (4:9:5:8, v/v/v/v) as solvent systems, and then co-eluted antioxidants were on-line trapped, concentrated and desorbed to Sephadex LH-20 column for further off-line purification by methanol. It is noted that six elucidated antioxidants with purity over 95% exhibited stronger activity than ascorbic acid (VC). More importantly, this at-line hyphenated strategy could sever as a rapid and efficient pathway for systematic purification of bioactive components from complex matrix. Copyright © 2016 Elsevier B.V. All rights reserved.

Integration of Gas Chromatography Mass Spectrometry Methods for Differentiating Ricin Preparation Methods

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wunschel, David S.; Melville, Angela M.; Ehrhardt, Christopher J.

2012-05-17

The investigation of crimes involving chemical or biological agents is infrequent, but presents unique analytical challenges. The protein toxin ricin is encountered more frequently than other agents and is found in the seeds of the castor plant Ricinus communis. Typically, the toxin is extracted from castor seeds utilizing a variety of different recipes that result in varying purity of the toxin. Moreover, these various purification steps can also leave or differentially remove a variety of exogenous and endogenous residual components with the toxin that may indicate the type and number of purification steps involved. We have applied three gas chromatographicmore » - mass spectrometric (GC-MS) based analytical methods to measure the variation in seed carbohydrates and castor oil ricinoleic acid as well as the presence of solvents used for purification. These methods were applied to the same samples prepared using four previously identified toxin preparation methods starting from four varieties of castor seeds. The individual data sets for seed carbohydrate profiles, ricinoleic acid or acetone amount each provided information capable of differentiating different types of toxin preparations across seed types. However, the integration of the data sets using multivariate factor analysis provided a clear distinction of all samples based on the preparation method and independent of the seed source. In particular the abundance of mannose, arabinose, fucose, ricinoleic acid and acetone were shown to be important differentiating factors. These complementary tools provide a more confident determination of the method of toxin preparation.« less

Automated production of [18 F]FTHA according to GMP.

PubMed

Savisto, Nina; Viljanen, Tapio; Kokkomäki, Esa; Bergman, Jörgen; Solin, Olof

2018-02-01

14-(R,S)-[ 18 F]fluoro-6-thia-heptadecanoic acid is a tracer for fatty acid imaging by positron emission tomography. High demand for this tracer required us to replace semiautomatic synthesis with a fully automated procedure. An automated synthesis device was constructed in-house for multistep nucleophilic 18 F-fluorination and a control system was developed. The synthesis device was combined with a sterile filtration unit and both were qualified. 14-(R,S)-[ 18 F]fluoro-6-thia-heptadecanoic acid was produced according to good manufacturing practice guidelines set by the European Union. The synthesis includes an initial nucleophilic labelling reaction, deprotection, preparative HPLC separation, purification of the final product, and formulation for injection. The duration and temperature of the reaction and hydrolysis were optimized, and the radiochemical stability of the formulated product was determined. The rotary evaporator used to evaporate the solvent after HPLC purification was replaced with solid phase extraction purification. We also replaced the human serum albumin used in the earlier procedure with a phosphate buffer-ascorbic acid mixture in the final formulation solution. From 2011 to 2016, we performed 219 synthesis procedures, 94% of which were successful. The radiochemical yield of 14-(R,S)-[ 18 F]fluoro-6-thia-heptadecanoic acid, decay-corrected to the end of bombardment, was 13% ± 6.3%. The total amount of formulated end product was 1.7 ± 0.8 GBq at end of synthesis. Copyright © 2017 John Wiley & Sons, Ltd.

Tracking control of colloidal particles through non-homogeneous stationary flows

DOE Office of Scientific and Technical Information (OSTI.GOV)

Híjar, Humberto, E-mail: humberto.hijar@lasallistas.org.mx

2013-12-21

We consider the problem of controlling the trajectory of a single colloidal particle in a fluid with steady non-homogeneous flow. We use a Langevin equation to describe the dynamics of this particle, where the friction term is assumed to be given by the Faxén's Theorem for the force on a sphere immersed in a stationary flow. We use this description to propose an explicit control force field to be applied on the particle such that it will follow asymptotically any given desired trajectory, starting from an arbitrary initial condition. We show that the dynamics of the controlled particle can bemore » mapped into a set of stochastic harmonic oscillators and that the velocity gradient of the solvent induces an asymmetric coupling between them. We study the particular case of a Brownian particle controlled through a plane Couette flow and show explicitly that the velocity gradient of the solvent renders the dynamics non-stationary and non-reversible in time. We quantify this effect in terms of the correlation functions for the position of the controlled particle, which turn out to exhibit contributions depending exclusively on the non-equilibrium character of the state of the solvent. In order to test the validity of our model, we perform simulations of the controlled particle moving in a simple shear flow, using a hybrid method combining molecular dynamics and multi-particle collision dynamics. We confirm numerically that the proposed guiding force allows for controlling the trajectory of the micro-sized particle by obligating it to follow diverse specific trajectories in fluids with homogeneous shear rates of different strengths. In addition, we find that the non-equilibrium correlation functions in simulations exhibit the same qualitative behavior predicted by the model, thus revealing the presence of the asymmetric non-equilibrium coupling mechanism induced by the velocity gradient.« less

Streamlined structure elucidation of an unknown compound in a pigment formulation.

PubMed

Yüce, Imanuel; Morlock, Gertrud E

2016-10-21

A fast and reliable quality control is important for ink manufacturers to ensure a constant production grade of mixtures and chemical formulations, and unknown components attract their attention. Structure elucidating techniques seem time-consuming in combination with column-based methods, but especially the low solubility of pigment formulations is challenging the analysis. In contrast, layer chromatography is more tolerant with regard to pigment particles. One PLC plate for NMR and FTIR analyses and one HPTLC plate for recording of high resolution mass spectra, MS/MS spectra and for gathering information on polarity and spectral properties were needed to characterize a structure, exemplarily shown for an unknown component in pigment Red 57:1 to be 3-hydroxy-2-naphtoic acid. A preparative layer chromatography (PLC) workflow was developed that used an Automated Multiple Development 2 (AMD 2) system. The 0.5-mm PLC plate could still be operated in the AMD 2 system and allowed a smooth switch from the analytical to the preparative gradient separation. Through automated gradient development and the resulting focusing of bands, the sharpness of the PLC bands was improved. For NMR, the necessary high load of the target compound on the PLC plate was achieved via a selective solvent extraction that discriminated the polar sample matrix and thus increased the application volume of the extract that could maximally be applied without overloading. By doing so, the yield for NMR analysis was improved by a factor of 9. The effectivity gain through a simple, but thoroughly chosen extraction solvent is often overlooked, and for educational purpose, it was clearly illustrated and demonstrated by an extended solvent screening. Thus, PLC using an automated gradient development after a selective extraction was proven to be a new powerful combination for structural elucidation by NMR. Copyright © 2016 Elsevier B.V. All rights reserved.

Communication: Control of chemical reactions using electric field gradients.

PubMed

Deshmukh, Shivaraj D; Tsori, Yoav

2016-05-21

We examine theoretically a new idea for spatial and temporal control of chemical reactions. When chemical reactions take place in a mixture of solvents, an external electric field can alter the local mixture composition, thereby accelerating or decelerating the rate of reaction. The spatial distribution of electric field strength can be non-trivial and depends on the arrangement of the electrodes producing it. In the absence of electric field, the mixture is homogeneous and the reaction takes place uniformly in the reactor volume. When an electric field is applied, the solvents separate and the reactants are concentrated in the same phase or separate to different phases, depending on their relative miscibility in the solvents, and this can have a large effect on the kinetics of the reaction. This method could provide an alternative way to control runaway reactions and to increase the reaction rate without using catalysts.

Purification, Crystallization, and Preliminary Crystallographic Analysis of Deoxyuridine Triphosphate Nucleotidohydrolase from Arabidopsis Thaliana

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bajaj,M.; Moriyama, H.

2007-01-01

The deoxyuridine triphosphate nucleotidohydrolase gene from Arabidopsis thaliana was expressed and the gene product was purified. Crystallization was performed by the hanging-drop vapour-diffusion method at 298 K using 2 M ammonium sulfate as the precipitant. X-ray diffraction data were collected to 2.2 Angstroms resolution using Cu K{alpha} radiation. The crystal belongs to the orthorhombic space group P212121, with unit-cell parameters a = 69.90, b = 70.86 Angstroms, c = 75.55 Angstroms . Assuming the presence of a trimer in the asymmetric unit, the solvent content was 30%, with a VM of 1.8 Angstroms 3 Da-1.

Assembly of synthetic cellulose I.

PubMed

Lee, J H; Brown, R M; Kuga, S; Shoda, S; Kobayashi, S

1994-08-02

Cellulose microfibrils with an electron diffraction pattern characteristic of crystalline native cellulose I have been assembled abiotically by means of a cellulase-catalyzed polymerization of beta-cellobiosyl fluoride substrate monomer in acetonitrile/acetate buffer. Substantial purification of the Trichoderma viride cellulase enzyme was found to be essential for the formation of the synthetic cellulose I allomorph. Assembly of synthetic cellulose I appears to be a result of a micellar aggregation of the partially purified enzyme and the substrate in an organic/aqueous solvent system favoring the alignment of glucan chains with the same polarity and extended chain conformation, resulting in crystallization to form the metastable cellulose I allomorph.

Quantitative structure-retention relationships applied to development of liquid chromatography gradient-elution method for the separation of sartans.

PubMed

Golubović, Jelena; Protić, Ana; Otašević, Biljana; Zečević, Mira

2016-04-01

QSRR are mathematically derived relationships between the chromatographic parameters determined for a representative series of analytes in given separation systems and the molecular descriptors accounting for the structural differences among the investigated analytes. Artificial neural network is a technique of data analysis, which sets out to emulate the human brain's way of working. The aim of the present work was to optimize separation of six angiotensin receptor antagonists, so-called sartans: losartan, valsartan, irbesartan, telmisartan, candesartan cilexetil and eprosartan in a gradient-elution HPLC method. For this purpose, ANN as a mathematical tool was used for establishing a QSRR model based on molecular descriptors of sartans and varied instrumental conditions. The optimized model can be further used for prediction of an external congener of sartans and analysis of the influence of the analyte structure, represented through molecular descriptors, on retention behaviour. Molecular descriptors included in modelling were electrostatic, geometrical and quantum-chemical descriptors: connolly solvent excluded volume non-1,4 van der Waals energy, octanol/water distribution coefficient, polarizability, number of proton-donor sites and number of proton-acceptor sites. Varied instrumental conditions were gradient time, buffer pH and buffer molarity. High prediction ability of the optimized network enabled complete separation of the analytes within the run time of 15.5 min under following conditions: gradient time of 12.5 min, buffer pH of 3.95 and buffer molarity of 25 mM. Applied methodology showed the potential to predict retention behaviour of an external analyte with the properties within the training space. Connolly solvent excluded volume, polarizability and number of proton-acceptor sites appeared to be most influential paramateres on retention behaviour of the sartans. Copyright © 2015 Elsevier B.V. All rights reserved.

Purification and heterogeneity of human kininogen. Use of DEAE-chromatography, molecular sieving and antibody specific immunosorbents.

PubMed

Hamberg, U; Elg, P; Nissinen, E; Stelwagen, P

1975-01-01

Various methods of preparing human kininogen were investigated with an aim to limit the immunoreactive contaminant proteins to permit purification by immunosorption. A five-step procedure is described giving 7.5% yield of highly purified kininogen (pharmacological purity 14--20) from pooled human plasma, and containing approximately 30% alpha-2HS-glycoprotein and 2.8% albumin. Alpha-2HS could not be removed by polyacrylamide gel electrophoresis or isoelectric focusing in column. Analysis of heterogeneity of kininogen after chromatography on DEAE-Sephadex using various linear gradients and gel filtration on Sephadex G-100 suggested that a minor component may be an aggregate, not included in the yield. It remains uncertain whether this component derives from an occasionally observed high molecular form of active kininogen in the primary purification steps in the 7-12 S sieve fractions from Sephadex G-200, and excluded from further purification by pooling. Purification with immunosorbents was investigated using batch operations with antibody specific polymers prepared with antisera insolubilized with ethylchloroformate. It was found that the adsorption-desorption procedure was favourable for immunization purposes in producing highly specific immunologically pure kininogen. The kininogen obtained by this method or by the removal of contaminant alpha-2HS and albumin with the corresponding antibody specific polymers gave similar heterogenous patterns by polyacrylamide gel electrophoresis, indicating a main band of kininogen and several faintly stained bands which responded only to anti-kininogen. With 200 mug of the kininogen protein purified by immunosorption using monospecific antiserum the kininogen precipitation titre was 1:8 after 6--8 weeks in rabbits. With a polymer prepared with 4 ml anti-kininogen serum (1:8) and incubated with 800 mug highly purified kininogen approximately half the protein was desorbed with 2 M and 3 M sodium iodide in the first adsorption-desorption procedure.

Analysis of translation using polysome profiling

PubMed Central

Chassé, Héloïse; Boulben, Sandrine; Costache, Vlad; Cormier, Patrick

2017-01-01

Abstract During the past decade, there has been growing interest in the role of translational regulation of gene expression in many organisms. Polysome profiling has been developed to infer the translational status of a specific mRNA species or to analyze the translatome, i.e. the subset of mRNAs actively translated in a cell. Polysome profiling is especially suitable for emergent model organisms for which genomic data are limited. In this paper, we describe an optimized protocol for the purification of sea urchin polysomes and highlight the critical steps involved in polysome purification. We applied this protocol to obtain experimental results on translational regulation of mRNAs following fertilization. Our protocol should prove useful for integrating the study of the role of translational regulation in gene regulatory networks in any biological model. In addition, we demonstrate how to carry out high-throughput processing of polysome gradient fractions, for the simultaneous screening of multiple biological conditions and large-scale preparation of samples for next-generation sequencing. PMID:28180329

Separation and purification of γ-aminobutyric acid from fermentation broth by flocculation and chromatographic methodologies.

PubMed

Gao, Qiang; Duan, Qiang; Wang, Depei; Zhang, Yunze; Zheng, Chunyang

2013-02-27

To date, the multifunctional γ-aminobutyric acid (GABA) is mainly produced by microbial fermentation in industry. The purpose of this study was to find an effective method for separation and purification of 31.2 g/L initial GABA from the fermentation broth of Enterococcus raffinosus TCCC11660. To remove the impurities from fermentation broth, flocculation pretreatment using chitosan and sodium alginate was first implemented to facilitate subsequent filtration. Ultrafiltration followed two discontinuous diafiltration steps to effectively remove proteins and macromolecular pigments, and the resulting permeate was further decolored by DA201-CII resin at a high decoloration ratio and GABA recovery. Subsequently, ion exchange chromatography (IEC) with Amberlite 200C resin and gradient elution were applied for GABA separation from glutamate and arginine. Finally, GABA crystals of 99.1% purity were prepared via warm ethanol precipitation twice. Overall, our results reveal that the successive process including flocculation, filtration, ultrafiltration, decoloration, IEC, and crystallization is promising for scale-up GABA extraction from fermentation broth.

A new non-degradative method to purify glycogen.

PubMed

Tan, Xinle; Sullivan, Mitchell A; Gao, Fei; Li, Shihan; Schulz, Benjamin L; Gilbert, Robert G

2016-08-20

Liver glycogen, a complex branched glucose polymer containing a small amount of protein, is important for maintaining glucose homeostasis (blood-sugar control) in humans. It has recently been found that glycogen molecular structure is impaired in diabetes. Isolating the carbohydrate polymer and any intrinsically-attached protein(s) is an essential prerequisite for studying this structural impairment. This requires an effective, non-degradative and efficient purification method to exclude the many other proteins present in liver. Proteins and glycogen have different ranges of molecular sizes. Despite the plethora of proteins that might still be present in significant abundance after other isolation techniques, SEC (size exclusion chromatography, also known as GPC), which separates by molecular size, should separate those extraneous to glycogen from glycogen with any intrinsically associated protein(s). A novel purification method is developed for this, based on preparative SEC following sucrose gradient centrifugation. Proteomics is used to show that the new method compares favourably with current methods in the literature. Copyright © 2016 Elsevier Ltd. All rights reserved.

Purification and stability characterization of a cell regulatory sialoglycopeptide inhibitor

NASA Technical Reports Server (NTRS)

Moos, P. J.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

1995-01-01

Previous attempts to physically separate the cell cycle inhibitory and protease activities in preparations of a purified cell regulatory sialoglycopeptide (CeReS) inhibitor were largely unsuccessful. Gradient elution of the inhibitor preparation from a DEAE HPLC column separated the cell growth inhibitor from the protease, and the two activities have been shown to be distinct and non-overlapping. The additional purification increased the specific biological activity of the CeReS preparation by approximately two-fold. The major inhibitory fraction that eluted from the DEAE column was further analyzed by tricine-SDS-PAGE and microbore reverse phase HPLC and shown to be homogeneous in nature. Two other fractions separated by DEAE HPLC, also devoid of protease activity, were shown to be inhibitory to cell proliferation and most likely represented modified relatives of the CeReS inhibitor. The highly purified CeReS was chemically characterized for amino acid and carbohydrate composition and the role of the carbohydrate in cell proliferation inhibition, stability, and protease resistance was assessed.

Understanding the dissolution of α-zein in aqueous ethanol and acetic acid solutions.

PubMed

Li, Yunqi; Li, Ji; Xia, Qiuyang; Zhang, Boce; Wang, Qin; Huang, Qingrong

2012-10-04

Zein is a corn prolamin that has broad industrial applications because of its unique physical properties. Currently, the high cost of extraction and purification, which is directly related to the dispersion of zein in different solvents, is the major bottleneck of the zein industry. Solution behaviors of zein have been studied for a long time. However, the physical nature of zein in different solvents remains unclear. In this study, small-angle X-ray scattering (SAXS), static light scattering (SLS), and rheology were combined to study the structure and protein-solvent interaction of α-zein in both acetic acid and aqueous ethanol solutions. We found that the like-dissolve-like rule, the partial unfolding, and the protonation of zein are all critical to understanding the solution behaviors. Zein holds an elongated conformation (i.e., prolate ellipsoid) in all solutions, as revealed from SAXS data. There is an "aging effect" for zein in aqueous ethanol solutions, as evidenced by the transition of Newtonian rheological profiles for fresh zein solutions to the non-Newtonian shear thinning behavior for zein solutions after storage at room temperature for 24 h. Such shear thinning behavior becomes more pronounced for zein solutions at higher concentrations. The SLS results clearly show that acetic acid is a better solvent to dissolve zein than aqueous ethanol solution, as supported by a more negative second virial coefficient. This is majorly caused by the protonation of the protein, which was further verified by the dissolution of zein in water (a nonsolvent for zein) with the addition of acids.

Comparison of retention models for polymers 1. Poly(ethylene glycol)s.

PubMed

Bashir, Mubasher A; Radke, Wolfgang

2006-10-27

The suitability of three different retention models to predict the retention times of poly(ethylene glycol)s (PEGs) in gradient and isocratic chromatography was investigated. The models investigated were the linear (LSSM) and the quadratic solvent strength model (QSSM). In addition, a model describing the retention behaviour of polymers was extended to account for gradient elution (PM). It was found that all models are suited to properly predict gradient retention volumes provided the extraction of the analyte specific parameters is performed from gradient experiments as well. The LSSM and QSSM on principle cannot describe retention behaviour under critical or SEC conditions. Since the PM is designed to cover all three modes of polymer chromatography, it is therefore superior to the other models. However, the determination of the analyte specific parameters, which are needed to calibrate the retention behaviour, strongly depend on the suitable selection of initial experiments. A useful strategy for a purposeful selection of these calibration experiments is proposed.

Transition-state optimization by the free energy gradient method: Application to aqueous-phase Menshutkin reaction between ammonia and methyl chloride

NASA Astrophysics Data System (ADS)

Hirao, Hajime; Nagae, Yukihiko; Nagaoka, Masataka

2001-11-01

The transition state (TS) for the Menshutkin reaction H 3N+CH 3Cl→H 3NCH 3++Cl - in aqueous solution was located on the free energy surface (FES) by the free energy gradient (FEG) method. The solute-solvent system was described by a hybrid quantum mechanical and molecular mechanical (QM/MM) method. The reaction path in water was found to deviate largely from that in the gas phase. It was concluded that, in such a reaction including charge separation, TS structure optimization on an FES is inevitable for obtaining valid information about a TS in solution.

Isolation and purification assay of ex vivo photosystem II D1 protein toward integrated biointeraction analysis.

PubMed

Muktiono, B; Schulten, C; Heemken, O; Gandrass, J; Prange, A; Schnabl, H; Cerboncini, C

2008-02-01

Protein extracts of photosystem II were prepared from leaf chloroplasts of different plant species by fast and nondenaturing methods. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and western blot analysis of the proteins obtained showed that the extracts were enriched by D1 proteins, which appeared putatively in association with the 33-kDa oxygen-evolving-complex subunits. In further isolation steps D1 proteins were purified using salt-gradient chromatography (fast protein liquid chromatography) and characterized by western blot and mass spectrometry.

First hyperpolarizability of isomers of pyridinium N-phenoxide betaine dye in solution using the ASEC-FEG method

NASA Astrophysics Data System (ADS)

Torres, E. M.; Georg, H. C.; Fonseca, T. L.; Castro, M. A.

2018-05-01

The linear and nonlinear properties of isomeric forms of pyridinium-N-phenoxide betaine dye were investigated in protic and aprotic solvents using atomistic simulations. We employed the sequential Quantum Mechanics/Molecular Mechanics (S-QM/MM) and the free energy gradient (FEG) methods to optimize the geometry of each isomer in chloroform, acetonitrile, methanol and water. The results show a complex dependence of the first hyperpolarizability with respect to the solvent nature and isomeric form, with a marked effect of conformational changes for para-betaine. Large contrasts of the first hyperpolarizability show a clear distinction between isomeric forms in solution that could be experimentally detected.

Developing procedures for the large-scale purification of human serum butyrylcholinesterase.

PubMed

Saxena, Ashima; Luo, Chunyuan; Doctor, Bhupendra P

2008-10-01

Human serum butyrylcholinesterase (Hu BChE) is the most viable candidate for the prophylactic treatment of organophosphate poisoning. A dose of 200 mg/70 kg is predicted to protect humans against 2x LD(50) of soman. Therefore, the aim of this study was to develop procedures for the purification of gram quantities of this enzyme from outdated human plasma or Cohn Fraction IV-4. The purification of Hu BChE was accomplished by batch adsorption on procainamide-Sepharose-CL-4B affinity gel followed by ion-exchange chromatography on a DEAE-Sepharose column. For the purification of enzyme from Cohn Fraction IV-4, it was resuspended in 25 mM sodium phosphate buffer, pH 8.0, and fat was removed by decantation, prior to batch adsorption on procainamide-Sepharose gel. In both cases, the procainamide gel was thoroughly washed with 25 mM sodium phosphate buffer, pH 8.0, containing 0.05 M NaCl, and the enzyme was eluted with the same buffer containing 0.1 M procainamide. The enzyme was dialyzed and the pH was adjusted to 4.0 before loading on the DEAE column equilibrated in sodium acetate buffer, pH 4.0. The column was thoroughly washed with 25 mM sodium phosphate buffer, pH 8.0 containing 0.05 M NaCl before elution with a gradient of 0.05-0.2M NaCl in the same buffer. The purity of the enzyme following these steps ranged from 20% to 40%. The purity of the enzyme increased to >90% by chromatography on an analytical procainamide affinity column. Results show that Cohn Fraction IV-4 is a much better source than plasma for the large-scale isolation of purified Hu BChE.

Synthesis and Purification of Iodoaziridines Involving Quantitative Selection of the Optimal Stationary Phase for Chromatography

PubMed Central

Boultwood, Tom; Affron, Dominic P.; Bull, James A.

2014-01-01

The highly diastereoselective preparation of cis-N-Ts-iodoaziridines through reaction of diiodomethyllithium with N-Ts aldimines is described. Diiodomethyllithium is prepared by the deprotonation of diiodomethane with LiHMDS, in a THF/diethyl ether mixture, at -78 °Cin the dark. These conditions are essential for the stability of the LiCHI2 reagent generated. The subsequent dropwise addition of N-Ts aldimines to the preformed diiodomethyllithium solution affords an amino-diiodide intermediate, which is not isolated. Rapid warming of the reaction mixture to 0 °C promotes cyclization to afford iodoaziridines with exclusive cis-diastereoselectivity. The addition and cyclization stages of the reaction are mediated in one reaction flask by careful temperature control. Due to the sensitivity of the iodoaziridines to purification, assessment of suitable methods of purification is required. A protocol to assess the stability of sensitive compounds to stationary phases for column chromatography is described. This method is suitable to apply to new iodoaziridines, or other potentially sensitive novel compounds. Consequently this method may find application in range of synthetic projects. The procedure involves firstly the assessment of the reaction yield, prior to purification, by 1H NMR spectroscopy with comparison to an internal standard. Portions of impure product mixture are then exposed to slurries of various stationary phases appropriate for chromatography, in a solvent system suitable as the eluent in flash chromatography. After stirring for 30 min to mimic chromatography, followed by filtering, the samples are analyzed by 1H NMR spectroscopy. Calculated yields for each stationary phase are then compared to that initially obtained from the crude reaction mixture. The results obtained provide a quantitative assessment of the stability of the compound to the different stationary phases; hence the optimal can be selected. The choice of basic alumina, modified to activity IV, as a suitable stationary phase has allowed isolation of certain iodoaziridines in excellent yield and purity. PMID:24893769

Procedure for the determination of retinol and alpha-tocopherol in poultry tissues using capillary gas chromatography with solvent venting injection.

PubMed

Maraschiello, C; García Regueiro, J A

1998-08-28

A procedure designed for the determination of retinol (vitamin A) and alpha-tocopherol (vitamin E) in poultry tissues has been developed. The procedure involves lipid extraction, saponification, solid-phase clean-up and capillary gas chromatography (cGC). Retinol and alpha-tocopherol were determined separately by cGC-flame ionisation detection using a fused-silica open tubular capillary column, 30 m x 0.25 mm I.D. coated with 5% phenylmethylsilicone and with a film thickness of 0.25 micron. Solvent extraction followed by saponification were sufficient to provide a purified extract which was directly analyzed for retinol by cGC in the solvent venting mode. However, in order to accurately determine alpha-tocopherol by cGC, further purification of the extract by solid-phase extraction was necessary. A silica SPE column was used to remove interfering cholesterol from the extract. alpha-Tocopherol was analyzed in its derivatized form. Absolute and relative recoveries for both vitamins from spiked samples were evaluated. Absolute and relative recoveries ranging from 80 to 95% were obtained for both compounds. 5 alpha-Cholestane and alpha-tocopheryl acetate were used as internal standards. Poultry muscle meat and liver tissue were analyzed for their retinol and alpha-tocopherol content and the peaks detected by cGC were confirmed by cGC-mass spectrometry.

MALDI-TOF to compare polysaccharide profiles from commercial health supplements of different mushroom species.

PubMed

López-García, Marta; García, María Sonia Dopico; Vilariño, José Manuel López; Rodríguez, María Victoria González

2016-05-15

In this work MALDI-TOF mass spectroscopy was investigated to characterise the β-glucan profiles of several commercial health supplements, without any derivatisation or purification pre-treatment. The effect of two solvents (water and dimethyl sulfoxide) and two MALDI matrices (2,5-dihydroxybenzoic acid and 2',4',6'-trihydroxyacetophenone) was first evaluated on dextran standards. MALDI-TOF was found as a useful and quick technique to obtain structural information of diverse food supplements based on mushroom extracts. The MALDI polysaccharide profiles of 5 supplements from different mushroom species were qualitatively similar showing [Glucan+Na](+) cations with a peak-to-peak mass difference of 16 Da consistent with the repeating unit of the β-(1→3)-glucan. The profiles strongly depended on the sample solvent used, with m/z values around 5000-8000 for water and 2000 for dimethyl sulfoxide; differences between samples were revealed in the molecular weight of the aqueous preparation, with the highest values for Maitake and Cordyceps species. Copyright © 2015 Elsevier Ltd. All rights reserved.

A solvent-extraction module for cyclotron production of high-purity technetium-99m.

PubMed

Martini, Petra; Boschi, Alessandra; Cicoria, Gianfranco; Uccelli, Licia; Pasquali, Micòl; Duatti, Adriano; Pupillo, Gaia; Marengo, Mario; Loriggiola, Massimo; Esposito, Juan

2016-12-01

The design and fabrication of a fully-automated, remotely controlled module for the extraction and purification of technetium-99m (Tc-99m), produced by proton bombardment of enriched Mo-100 molybdenum metallic targets in a low-energy medical cyclotron, is here described. After dissolution of the irradiated solid target in hydrogen peroxide, Tc-99m was obtained under the chemical form of 99m TcO 4 - , in high radionuclidic and radiochemical purity, by solvent extraction with methyl ethyl ketone (MEK). The extraction process was accomplished inside a glass column-shaped vial especially designed to allow for an easy automation of the whole procedure. Recovery yields were always >90% of the loaded activity. The final pertechnetate saline solution Na 99m TcO 4 , purified using the automated module here described, is within the Pharmacopoeia quality control parameters and is therefore a valid alternative to generator-produced 99m Tc. The resulting automated module is cost-effective and easily replicable for in-house production of high-purity Tc-99m by cyclotrons. Copyright © 2016 Elsevier Ltd. All rights reserved.

Preparative isolation and purification of seven main antioxidants from Eucommia ulmoides Oliv. (Du-zhong) leaves using HSCCC guided by DPPH-HPLC experiment.

PubMed

Dai, Xingping; Huang, Qiong; Zhou, Boting; Gong, Zhicheng; Liu, Zhaoqian; Shi, Shuyun

2013-08-15

Seven antioxidants were purified from Eucommia ulmoides Oliv. leaves using HSCCC guided by DPPH-HPLC experiment. HSCCC was successfully used to separate target antioxidants by three runs with different solvent systems after D101 column chromatography fractionation. Ethyl acetate-n-butanol-water (1:2:3, v/v/v) was selected as the optimum solvent system to purify geniposidic acid. Ethyl acetate-ethanol-water (4:1:5, v/v/v) was used to isolate caffeic acid, chlorogenic acid and ferulic acid. While three flavonoids, quercetin-3-O-sambubioside, rutin and isoquercitrin were purified by petroleum ether-ethyl acetate-methanol-water (1:5:1:5, v/v/v/v). The structures were identified by MS and NMR. Antioxidant activities were assessed, and compounds 2-7 showed strong antioxidant activities. This is the first report about separation of antioxidants from E. ulmoides leaves by HSCCC. The results indicated that the combinative methods using DPPH-HPLC and HSCCC could be widely applied for screening and isolation of antioxidants from complex extracts. Copyright © 2013 Elsevier Ltd. All rights reserved.

Larvicidal Activity of Centaurea bruguierana ssp. belangerana Against Anopheles stephensi Larvae.

PubMed

Khanavi, Mahnaz; Rajabi, Afsaneh; Behzad, Masoud; Hadjiakhoondi, Abbas; Vatandoost, Hassan; Abaee, Mohammad Reza

2011-01-01

In this study, the total 80% of MeOH extract and also petroleum ether, CHCl3, EtOAc, n-BuOH, and the remaining MeOH fractions obtained by solvent-solvent fractionation of the whole flowering samples of Centaurea bruguierana (DC.) Hand.-Mzt. ssp. belangerana (DC.) Bornm. (Asteraceae), namely "Baad-Avard", collected from Borazjan in Bushehr Province (Bushehr, Iran) were investigated for larvicidal activity against malaria vector, Anopheles stephensi Liston, according to WHO methods. The mortality rate of total extract and petroleum ether fraction in concentration of 40 ppm were 28% and 86% respectively and the other fractions were inactive. The probit regression analysis for the dose-response to petroleum ether fraction treatment of larvae exhibited the LC50 and LC90 values of 15.7 ppm and 48.3 ppm, respectively. As results showed, the larvicidal activity of the petroleum ether fraction would be due to the nonpolar compounds in the plant which further isolation and purification would obtain the more active compounds in lower concentrations useful for preparation of biological insecticides.

Larvicidal Activity of Centaurea bruguierana ssp. belangerana Against Anopheles stephensi Larvae

PubMed Central

Khanavi, Mahnaz; Rajabi, Afsaneh; Behzad, Masoud; Hadjiakhoondi, Abbas; Vatandoost, Hassan; Abaee, Mohammad Reza

2011-01-01

In this study, the total 80% of MeOH extract and also petroleum ether, CHCl3, EtOAc, n-BuOH, and the remaining MeOH fractions obtained by solvent-solvent fractionation of the whole flowering samples of Centaurea bruguierana (DC.) Hand.-Mzt. ssp. belangerana (DC.) Bornm. (Asteraceae), namely “Baad-Avard”, collected from Borazjan in Bushehr Province (Bushehr, Iran) were investigated for larvicidal activity against malaria vector, Anopheles stephensi Liston, according to WHO methods. The mortality rate of total extract and petroleum ether fraction in concentration of 40 ppm were 28% and 86% respectively and the other fractions were inactive. The probit regression analysis for the dose-response to petroleum ether fraction treatment of larvae exhibited the LC50 and LC90 values of 15.7 ppm and 48.3 ppm, respectively. As results showed, the larvicidal activity of the petroleum ether fraction would be due to the nonpolar compounds in the plant which further isolation and purification would obtain the more active compounds in lower concentrations useful for preparation of biological insecticides. PMID:24250419

Optimization and development of a SPE-HPLC-UV method to determine astaxanthin in Saccharina japonica.

PubMed

Zhou, Jun; Bi, Wentao; Row, Kyung Ho

2011-04-01

An effective and accurate method including extraction, saponification, and separation was developed to determine astaxanthin (AX) in Saccharina japonica. The optimal extraction conditions with different solvents were investigated. 29.30 μg/g of AX was extracted from dry Saccharina japonica powder by solvent. After subsequent saponification, the extracted amount of AX was increased to 37.26 μg/g. Furthermore, 3 different ionic liquid-based silicas were prepared as sorbents for the solid phase extraction of AX from the extract. By comparing the adsorption isotherms of AX on different ionic liquid-based silicas, suitable sorbent was successfully selected and applied for separation of AX from extract. Astaxanthin, in 3 main forms (free, monoesters, and diesters), can be obtained from marine plants and animals. By extraction with subsequent saponification, the astaxanthin was extracted from Saccharina japonica. And then, ionic liquid-based silicas were used to separate the astaxanthin from the extract solution. This method can be widely applied for determination, or even industrial separation and purification of astaxanthin from many other algae.

Separation and purification of hydrolyzable tannin from Geranium wilfordii Maxim by reversed-phase and normal-phase high-speed counter-current chromatography.

PubMed

Liu, Dan; Su, Zhiguo; Wang, Changhai; Gu, Ming; Xing, Siliang

2010-08-01

Three hydrolyzable tannins, geraniin, corilagin and gallic acid, main active components of Geranium wilfordii Maxim, have been separated and purified in one-step by both reversed-phase and normal-phase high-speed counter-current chromatography. Gallic acid, corilagin and geraniin were purified from 70% aqueous acetone extract of G. wilfordii Maxim with solvent system n-hexane-ethyl acetate-methanol-acetic acid-water (1:10:0.2:0.2:20) by reversed-phase high-speed counter-current chromatography at purities of 94.2, 91.0 and 91.3%, at yields of 89.3, 82.9 and 91.7%, respectively. Gallic acid, corilagin and geraniin were purified with solvent system n-hexane-ethyl acetate-methanol-acetic acid-water (0.2:10:2:1:5) by normal-phase high-speed counter-current chromatography at purities of 85.9, 92.2 and 87.6%, at yields of 87.4, 94.6 and 94.3%, respectively. It was successful for both reversed-phase and normal-phase high-speed counter-current chromatography to separate high-polarity of low-molecular-weight substances.

Prospects for robust biocatalysis: engineering of novel specificity in a halophilic amino acid dehydrogenase.

PubMed

Munawar, Nayla; Engel, Paul C

2013-01-01

Heat- and solvent-tolerant enzymes from halophiles, potentially important industrially, offer a robust framework for protein engineering, but few solved halophilic structures exist to guide this. Homology modelling has guided mutations in glutamate dehydrogenase (GDH) from Halobacterium salinarum to emulate conversion of a mesophilic GDH to a methionine dehydrogenase. Replacement of K89, A163 and S367 by leucine, glycine and alanine converted halophilic GDH into a dehydrogenase accepting L-methionine, L-norleucine and L-norvaline as substrates. Over-expression in the halophilic expression host Haloferax volcanii and three-step purification gave ~98 % pure protein exhibiting maximum activity at pH 10. This enzyme also showed enhanced thermostability and organic solvent tolerance even at 70 °C, offering a biocatalyst resistant to harsh industrial environments. To our knowledge, this is the first reported amino acid specificity change engineered in a halophilic enzyme, encouraging use of mesophilic models to guide engineering of novel halophilic biocatalysts for industrial application. Calibrated gel filtration experiments show that both the mutant and the wild-type enzyme are stable hexamers.

Optimization, Purification, and Starch Stain Wash Application of Two New α-Amylases Extracted from Leaves and Stems of Pergularia tomentosa

PubMed Central

El Abed, Hanen; Belghith, Hafedh; Ben Abdallah, Ferjani; Belghith, Karima

2017-01-01

A continuous research is attempted to fulfil the highest industrial demands of natural amylases presenting special properties. New α-amylases extracted from stems and leaves of Pergularia tomentosa, which is widespread and growing spontaneously in Tunisia, were studied by the means of their activities optimization and purification. Some similarities were recorded for the two identified enzymes: (i) the highest amylase activity showed a promoted thermal stability at 50°C; (ii) the starch substrate at 1% enhanced the enzyme activity; (iii) the two α-amylases seem to be calcium-independent; (iv) Zn2+, Cu2+, and Ag2+ were considered as important inhibitors of the enzyme activity. Following the increased gradient of elution on Mono Q-Sepharose column, an increase in the specific activity of 11.82-fold and 10.92-fold was recorded, respectively, for leaves and stems with the presence of different peaks on the purification profiles. Pergularia amylases activities were stable and compatible with the tested commercial detergents. The combination of plant amylase and detergent allowed us to enhance the wash performance with an increase of 35.24 and 42.56%, respectively, for stems and leaves amylases. Characterized amylases were reported to have a promoted potential for their implication notably in detergent industry as well as biotechnological sector. PMID:29392138

Preparative separation and purification of epigallocatechin gallate from green tea extracts using a silica adsorbent containing β-cyclodextrin.

PubMed

Lai, Shih-Ming; Gu, Jhe-Yu; Huang, Bing-Hao; Chang, Chieh-Ming J; Lee, Wen-Lung

2012-03-01

A silica adsorbent containing β-cyclodextrin (β-CD) has been developed and used for the separation and purification of epigallocatechin gallate (EGCG) from the green tea extracts. The batch adsorption experiments demonstrated that, the β-CD bonded silica adsorbent possessed excellent adsorption equilibrium capacity (> 55 mg/g adsorbent) and adsorption ratio (>95%) for EGCG compared to the other tea catechins and caffeine. The excellent adsorption capacity and selectivity for EGCG are attributed to the specific interactions between β-CD and EGCG. The preparative separation and purification performance of EGCG on the β-CD bonded silica column (220 mm L × 15 mm i.d., 40-63 μm) was then evaluated. The column was operated in the polar organic mode using methanol/acetonitrile/acetic acid as the mobile phase and eluted under a three-step gradient elution program. The sample was dissolved in acetonitrile and loaded on a preparative scale of about 0.8 mg/g adsorbent. Under the optimal chromatographic conditions, the target compound, EGCG, being the most retained species, was obtained at a purity of about 90% with a recovery of about 90%. The productivity of EGCG was about 6 mg per injection, which can be further increased by scaling-up the chromatographic system. Copyright © 2012 Elsevier B.V. All rights reserved.

Simultaneous separation and purification of (-)-epigallocatechin gallate and caffeine from tea extract by size exclusion effect on modified porous adsorption material.

PubMed

Zhang, Xiaofeng; Xu, Yi; Zhang, Qing; Cao, Kun; Mu, Xiuni

2016-09-15

A dual-task method for the simultaneous separation and purification of (-)-epigallocatechin gallate (EGCG) and caffeine (CAF) from crude extract of green tea was established by size exclusion effect onto hydroquinone modified porous adsorbents. The results showed that hydroquinone modified porous adsorbents P4 provided the best separation power due to it has more porous structure and phenolic hydroxyl group. The adsorption-desorption behaviors of EGCG and CAF onto P4 adsorbents were investigated. Adsorption kinetics of EGCG and CAF results showed that the adsorption followed the pseudo-second-order kinetic model. The results also indicated that the equilibrium adsorption data best fit the Langmuir model. Meanwhile, EGCG and CAF were separated successfully onto P4 adsorbents packed columns in a gradient eluent process, and P4 adsorbents exhibited the size exclusion effect for small molecules CAF. Based on the phenolic hydroxyl group and size exclusion effect of P4 adsorbents, the high purity EGCG and CAF were obtained with 40% (v/v) ethanol eluent successively. The process fulfilled the task of simultaneous separation and purification of EGCG and CAF, and proved to be a promising basis for preparations of difficult to obtain active components that have similar polarity and different sizes of molecules and derived from the same natural products. Copyright © 2016 Elsevier B.V. All rights reserved.

One-step separation and purification of hydrolysable tannins from Geranium wilfordii Maxim by adsorption chromatography on cross-linked 12% agarose gel.

PubMed

Liu, Dan; Ma, Yan; Wang, Ye; Su, Zhiguo; Gu, Ming; Janson, Jan-Christer

2011-05-01

The hydrolysable tannins corilagin and geraniin, the major active components of the traditional Chinese medicine Geranium wilfordii Maxim, have been separated and purified from crude extracts in one step by adsorption chromatography on cross-linked 12% agarose gel (Superose 12 10/300 GL). The separation was achieved by gradient elution using mobile phase A composed of 5% ethanol and 5% acetic acid and mobile phase B composed of 30% ethanol and 30% acetic acid. The gradients were composed as follows: 0-240 mL, 0-25% B; 240-480 mL, 25-40% B; after 480 mL, 100% B. The purities of the collected corilagin and geraniin were 92.4 and 87.2%, and the corresponding yields were 88.0 and 76.8%, respectively. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[Development of an automatic vacuum liquid chromatographic device and its application in the separation of the components from Schisandra chinensis (Turz) Baill].

PubMed

Zhu, Jingbo; Liu, Baoyue; Shan, Shibo; Ding, Yanl; Kou, Zinong; Xiao, Wei

2015-08-01

In order to meet the needs of efficient purification of products from natural resources, this paper developed an automatic vacuum liquid chromatographic device (AUTO-VLC) and applied it to the component separation of petroleum ether extracts of Schisandra chinensis (Turcz) Baill. The device was comprised of a solvent system, a 10-position distribution valve, a 3-position changes valve, dynamic axis compress chromatographic columns with three diameters, and a 10-position fraction valve. The programmable logic controller (PLC) S7- 200 was adopted to realize the automatic control and monitoring of the mobile phase changing, column selection, separation time setting and fraction collection. The separation results showed that six fractions (S1-S6) of different chemical components from 100 g Schisandra chinensis (Turcz) Baill. petroleum ether phase were obtained by the AUTO-VLC with 150 mm diameter dynamic axis compress chromatographic column. A new method used for the VLC separation parameters screened by using multiple development TLC was developed and confirmed. The initial mobile phase of AUTO-VLC was selected by taking Rf of all the target compounds ranging from 0 to 0.45 for fist development on the TLC; gradient elution ratio was selected according to k value (the slope of the linear function of Rf value and development times on the TLC) and the resolution of target compounds; elution times (n) were calculated by the formula n ≈ ΔRf/k. A total of four compounds with the purity more than 85% and 13 other components were separated from S5 under the selected conditions for only 17 h. Therefore, the development of the automatic VLC and its method are significant to the automatic and systematic separation of traditional Chinese medicines.

Dramatic nano-fluidic properties of carbon nanotube membranes as a platform for protein channel mimetics

NASA Astrophysics Data System (ADS)

Hinds, Bruce

2013-03-01

Carbon nanotubes have three key attributes that make them of great interest for novel membrane applications: 1) atomically flat graphite surface allows for ideal fluid slip boundary conditions and extremely fast flow rates 2) the cutting process to open CNTs inherently places functional chemistry at CNT core entrance for chemical selectivity and 3) CNT are electrically conductive allowing for electrochemical reactions and application of electric fields gradients at CNT tips. Pressure driven flux of a variety of solvents (H2O, hexane, decane ethanol, methanol) are 4-5 orders of magnitude higher than conventional Newtonian flow [Nature 2005, 438, 44] due to atomically flat graphite planes inducing nearly ideal slip conditions. However this is eliminated with selective chemical functionalization [ACS Nano 2011 5(5) 3867-3877] needed to give chemical selectivity. These unique properties allow us to explore the hypothesis of producing ``Gatekeeper'' membranes that mimic natural protein channels to actively pump through rapid nm-scale channels. With anionic tip functionality strong electroosmotic flow is induced by unimpeded cation flow with similar 10,000 fold enhancements [Nature Nano 2012 7(2) 133-39]. With enhanced power efficiency, carbon nanotube membranes were employed as the active element of a switchable transdermal drug delivery device that can facilitate more effective treatments of drug abuse and addiction. Recently methods to deposit Pt monolayers on CNT surface have been developed making for highly efficient catalytic platforms. Discussed are other applications of CNT protein channel mimetics, for large area robust engineering platforms, including water purification, flow battery energy storage, and biochemical/biomass separations. DOE EPSCoR (DE-FG02-07ER46375) and DARPA, W911NF-09-1-0267

Preparative isolation and purification of lignans from Justicia procumbens using high-speed counter-current chromatography in stepwise elution mode.

PubMed

Zhou, Peijuan; Luo, Qijun; Ding, Lijian; Fang, Fang; Yuan, Ye; Chen, Juanjuan; Zhang, Jinrong; Jin, Haixiao; He, Shan

2015-04-20

Lignans, which are recognized as main constituents in Justicia procumbens, have attracted considerable attention due to their pharmacological activities, including antitumor, anti-hepatitic, cytotoxic, anti-microbial, and anti-virus properties. Preparative high-speed counter-current chromatography (HSCCC) was successfully applied to the isolation and purification of four lignans (justicidin B (1), justicidin A (2), 6'-hydroxyjusticidin C (3) and lignan J1 (4)) from J. procumbens using stepwise elution with a pair of two-phase solvent systems composed of n-hexane-ethyl acetate-methanol-water at (1.3:1:1.3:1, v/v) and (2.5:1:2.5:1, v/v). The preparative HSCCC separation was performed on 300 mg of crude sample yielding compounds 1 (19.7 mg), 2 (9.86 mg), 3 (11.26 mg), and 4 (2.54 mg) in a one-step separation, with purities over 95% as determined by HPLC. The structures of these compounds were identified by MS, 1H-NMR and 13C-NMR. This is the first report on the application of HSCCC to the efficient separation of lignans from J. procumbens.

A polyacrylamide-based silica stationary phase for the separation of carbohydrates using alcohols as the weak eluent in hydrophilic interaction liquid chromatography.

PubMed

Cai, Jianfeng; Cheng, Lingping; Zhao, Jianchao; Fu, Qing; Jin, Yu; Ke, Yanxiong; Liang, Xinmiao

2017-11-17

A hydrophilic interaction liquid chromatography (HILIC) stationary phase was prepared by a two-step synthesis method, immobilizing polyacrylamide on silica sphere particles. The stationary phase (named PA, 5μm dia) was evaluated using a mixture of carbohydrates in HILIC mode and the column efficiency reached 121,000Nm -1 . The retention behavior of carbohydrates on PA stationary phase was investigated with three different organic solvents (acetonitrile, ethanol and methanol) employed as the weak eluent. The strongest hydrophilicity of PA stationary phase was observed in both acetonitrile and methanol as the weak eluent, when compared with another two amide stationary phases. Attributing to its high hydrophilicity, three oligosaccharides (xylooligosaccharide, fructooligosaccharide and chitooligosaccharides) presented good retention on PA stationary phase using alcohols/water as mobile phase. Finally, PA stationary phase was successfully applied for the purification of galactooligosaccharides and saponins of Paris polyphylla. It is feasible to use safer and cheaper alcohols to replace acetonitrile as the weak eluent for green analysis and purification of polar compounds on PA stationary phase. Copyright © 2017. Published by Elsevier B.V.

Mass Spectrometry of Intact Proteins Reveals +98 u Chemical Artifacts Following Precipitation in Acetone.

PubMed

Güray, Melda Z; Zheng, Shi; Doucette, Alan A

2017-02-03

Protein precipitation in acetone is frequently employed ahead of mass spectrometry for sample preconcentration and purification. Unfortunately, acetone is not chemically inert; mass artifacts have previously been observed on glycine-containing peptides when exposed to acetone under acidic conditions. We herein report a distinct chemical modification occurring at the level of intact proteins when incubated in acetone. This artifact manifests as one or more satellite peaks in the MS spectrum of intact protein, spaced 98 u above the mass of the unmodified protein. Other artifacts (+84, +112 u) also appear upon incubation of proteins or peptides in acetone. The reaction is pH-sensitive, being suppressed when proteins are exposed to acetone under acidic conditions. The +98 u artifact is speculated to originate through an intermediate product of aldol condensation of acetone to form diacetone alcohol and mesityl oxide. A +98 u product could originate from nucleophilic attack on mesityl oxide or through condensation with diacetone alcohol. Given the extent of modification possible upon exposure of proteins to acetone, particularly following overnight solvent exposure or incubation at room temperature, an awareness of the variables influencing this novel modification is valued by proteomics researchers who employ acetone precipitation for protein purification.

Separation and Purification of Ombuoside from Gynostemma Pentaphyllum by Microwave-Assisted Extraction Coupled with High-Speed Counter-current Chromatography.

PubMed

Jiang, Wenhui; Shan, Hu; Song, Jiying; Lü, Haitao

2017-01-01

A rapid and efficient method for the separation and purification of ombuoside from Gynostemma pentaphyllum by microwave-assisted extraction coupled with high-speed counter-current chromatography (HSCCC) was successfully developed. Using an orthogonal array design L 9 (3 4 ), the extraction conditions, including microwave power, irradiation time, solid-to-liquid ratio and extraction times, were optimized. Ombuoside was isolated and purified from the crude extraction by HSCCC with two-phase solvent system composed of n-hexane:ethyl acetate:ethanol:water (5:6:5:5, v/v) in a single run. A 210 mg quantity of the crude extract containing 2.16% ombuoside was loaded, yielding 3.9 mg of ombuoside at 96.7% purity. The chemical structure of ombuoside was determined by comparison with the high-performance liquid chromatography retention time of standard substance as well as UV, FT-IR, ESI-MS, 1 H NMR and 13 C NMR spectra. The purified ombuoside had strong 1,1-diphenyl-2-picrylhydrazyl and hydroxyl free radical scavenging activities. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Occupational and environmental hazard assessments for the isolation, purification and toxicity testing of cyanobacterial toxins

PubMed Central

2009-01-01

Cyanobacteria can produce groups of structurally and functionally unrelated but highly potent toxins. Cyanotoxins are used in multiple research endeavours, either for direct investigation of their toxicologic properties, or as functional analogues for various biochemical and physiological processes. This paper presents occupational safety guidelines and recommendations for personnel working in field, laboratory or industrial settings to produce and use purified cyanotoxins and toxic cyanobacteria, from bulk harvesting of bloom material, mass culture of laboratory isolates, through routine extraction, isolation and purification. Oral, inhalational, dermal and parenteral routes are all potential occupational exposure pathways during the various stages of cyanotoxin production and application. Investigation of toxicologic or pharmacologic properties using in vivo models may present specific risks if radiolabelled cyanotoxins are employed, and the potential for occupational exposure via the dermal route is heightened with the use of organic solvents as vehicles. Inter- and intra-national transport of living cyanobacteria for research purposes risks establishing feral microalgal populations, so disinfection of culture equipment and destruction of cells by autoclaving, incineration and/or chlorination is recommended in order to prevent viable cyanobacteria from escaping research or production facilities. PMID:19925679

PREPARATIVE ISOLATION AND PURIFICATION OF THREE GLYCINE-CONJUGATED CHOLIC ACIDS FROM PULVIS FELLIS SUIS BY HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY COUPLED WITH ELSD DETECTION.

PubMed

He, Jiao; Li, Jing; Sun, Wenji; Zhang, Tianyou; Ito, Yoichiro

2012-01-01

Coupled with evaporative light scattering detection, a high-speed counter-current chromatography (HSCCC) method was developed for preparative isolation and purification of three glycine-conjugated cholic acids, glycochenodeoxycholic acid (GCDCA), glycohyodeoxycholic acid (GHDCA) and glycohyocholic acid (GHCA) from Pulvis Fellis Suis (Pig gallbladder bile) for the first time. The separation was performed with a two-phase solvent system consisted of chloroform-methanol-water-acetic acid (65:30:10:1.5, v/v/v/v) by eluting the lower phase in the head-to-tail elution mode. The revolution speed of the separation column, flow rate of the mobile phase and separation temperature were 800 rpm, 2 ml/min and 25 °C, respectively. In a single operation, 33 mg of GCDCA, 38 mg of GHDCA and 23 mg of GHCA were obtained from 200 mg of crude extract with the purity of 95.65%, 96.72% and 96.63%, respectively, in one step separation. The HSCCC fractions were analyzed by high-performance liquid chromatography (HPLC) and the structures of the three glycine-conjugated cholic acids were identified by ESI-MS, (1)H NMR and (13)C NMR.

Purification of proteins from solutions containing residual host cell proteins via preparative crystallization.

PubMed

Hekmat, Dariusch; Breitschwerdt, Peter; Weuster-Botz, Dirk

2015-09-01

To investigate quantitatively and reproducibly a scalable, preparative crystallization method in novel stirred tanks using three different protein solutions containing residual microbial host cell proteins (HCP). Lysozyme from solutions being spiked with up to 15% host cell proteins (HCP) (corresponding to 176,500 ppm) was crystallized within a 2.4-4.6 h at 93.7% yield using NaCl and glycerol. Lipase was crystallized under comparable conditions using NaCl and a mixture of two polyethylene glycols (PEG). Enhanced green fluorescent protein (eGFP) was overexpressed in E. coli yielding a solution containing 23% target protein. Residual HCP content after pre-treatment was 7-16%. eGFP was crystallized from these solutions within 1.75-4 h at 88.7% step yield using ethanol and the same mixture of two PEG as in the case of lipase. HCP contained in the solvent channels of the protein crystals could be removed by diffusive washing yielding final purities at or above 99%. Preparative crystallization can be carried out with fast kinetics and high yields from solutions containing residual impurities and may represent an attractive alternative purification method compared to preparative chromatography, especially at large production scales.

Validation of analytical conditions for determination of polycyclic aromatic hydrocarbons in roasted coffee by gas chromatography-mass spectrometry.

PubMed

Guatemala-Morales, Guadalupe María; Beltrán-Medina, Elisa Alejandra; Murillo-Tovar, Mario Alfonso; Ruiz-Palomino, Priscilla; Corona-González, Rosa Isela; Arriola-Guevara, Enrique

2016-04-15

Polycyclic aromatic hydrocarbons (PAHs) are of significant interest due to their genotoxicity in humans. PAHs quantification in coffee is complex since some of its compounds interfere in the chromatographic analysis, which hinders the reliable determination of the PAHs. Analytical conditions for the ultrasound extraction, purification and quantification of 16 PAHs in roasted coffee were studied. The better extraction efficiency of benzo[a]pyrene (68%) from ground-roasted coffee was achieved with a solvent ratio of Hex:MC (9:1 v/v) and three extraction periods of 20 min, followed by alkaline saponification and purification of the extracts. The detection limits were 0.85-39.32 ng mL(-1), and the quantification limits from 2.84 to 131.05 ng mL(-1), obtained for fluoranthene and chrysene, respectively. The extraction was effective for most of the analytes, with recoveries of 39.8% dibenzo[ah]anthracene and 69.0% benzo[b]fluoranthene. For coffee roasted in a spouted bed reactor, the summation of the 16 PAHs ranged from 3.5 to 16.4 μg kg(-1). Copyright © 2015 Elsevier Ltd. All rights reserved.

PREPARATIVE ISOLATION AND PURIFICATION OF THREE GLYCINE-CONJUGATED CHOLIC ACIDS FROM PULVIS FELLIS SUIS BY HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY COUPLED WITH ELSD DETECTION

PubMed Central

He, Jiao; Li, Jing; Sun, Wenji; Zhang, Tianyou; Ito, Yoichiro

2011-01-01

Coupled with evaporative light scattering detection, a high-speed counter-current chromatography (HSCCC) method was developed for preparative isolation and purification of three glycine-conjugated cholic acids, glycochenodeoxycholic acid (GCDCA), glycohyodeoxycholic acid (GHDCA) and glycohyocholic acid (GHCA) from Pulvis Fellis Suis (Pig gallbladder bile) for the first time. The separation was performed with a two-phase solvent system consisted of chloroform-methanol-water-acetic acid (65:30:10:1.5, v/v/v/v) by eluting the lower phase in the head-to-tail elution mode. The revolution speed of the separation column, flow rate of the mobile phase and separation temperature were 800 rpm, 2 ml/min and 25 °C, respectively. In a single operation, 33 mg of GCDCA, 38 mg of GHDCA and 23 mg of GHCA were obtained from 200 mg of crude extract with the purity of 95.65%, 96.72% and 96.63%, respectively, in one step separation. The HSCCC fractions were analyzed by high-performance liquid chromatography (HPLC) and the structures of the three glycine-conjugated cholic acids were identified by ESI-MS, 1H NMR and 13C NMR. PMID:23008527

Ultramicroelectrode Sensors and Detectors. Considerations of the Stability, Sensitivity, Reproducibility, and Mechanism of Ion Transport in Gas Phase Chromatography and in High Performance Liquid Phase Chromatography

DTIC Science & Technology

1988-07-15

solvents were used. For high performance liquid chromatographic studies, the DNA bases thymine, adenine, cytocine, uracil, and guanine (Aldrich...this experiment. The DNA bases guanine, adenine, cytocine, uracil, and thymine were detected for a gradient elution of a mixture of the bases in a

ANALYSIS OF PROTEIN DIGESTS BY nano- SCX/RP/MSMS WITH pH SALT GRADIENT SCX ELUTION

EPA Science Inventory

The objective of this study was to optimize chromatographic parameters for complex peptide mixture analyses using two dimensional nano-LC/MSMS system. It used a strong cation exchange (SCX) and reversed phase chromatography (RP). The SCX solvent system was designed to promote pep...

New approaches with two cyano columns to the separation of acetaminophen, phenylephrine, chlorpheniramine and related compounds.

PubMed

Olmo, B; García, A; Marín, A; Barbas, C

2005-03-25

The development of new pharmaceutical forms with classical active compounds generates new analytical problems. That is the case of sugar-free sachets of cough-cold products containing acetaminophen, phenylephrine hydrochloride and chlorpheniramine maleate. Two cyanopropyl stationary phases have been employed to tackle the problem. The Discovery cyanopropyl (SUPELCO) column permitted the separation of the three actives, maleate and excipients (mainly saccharine and orange flavour) with a constant proportion of aqueous/ organic solvent (95:5, v/v) and a pH gradient from 7.5 to 2. The run lasted 14 min. This technique avoids many problems related to baseline shifts with classical organic solvent gradients and opens great possibilities to modify selectivity not generally used in reversed phase HPLC. On the other hand, the Agilent Zorbax SB-CN column with a different retention profile permitted us to separate not only the three actives and the excipients but also the three known related compounds: 4-aminophenol, 4-chloracetanilide and 4-nitrophenol in an isocratic method with a run time under 30 min. This method was validated following ICH guidelines and validation parameters showed that it could be employed as stability-indicating method for this pharmaceutical form.

Three-dimensional desirability spaces for quality-by-design-based HPLC development.

PubMed

Mokhtar, Hatem I; Abdel-Salam, Randa A; Hadad, Ghada M

2015-04-01

In this study, three-dimensional desirability spaces were introduced as a graphical representation method of design space. This was illustrated in the context of application of quality-by-design concepts on development of a stability indicating gradient reversed-phase high-performance liquid chromatography method for the determination of vinpocetine and α-tocopheryl acetate in a capsule dosage form. A mechanistic retention model to optimize gradient time, initial organic solvent concentration and ternary solvent ratio was constructed for each compound from six experimental runs. Then, desirability function of each optimized criterion and subsequently the global desirability function were calculated throughout the knowledge space. The three-dimensional desirability spaces were plotted as zones exceeding a threshold value of desirability index in space defined by the three optimized method parameters. Probabilistic mapping of desirability index aided selection of design space within the potential desirability subspaces. Three-dimensional desirability spaces offered better visualization and potential design spaces for the method as a function of three method parameters with ability to assign priorities to this critical quality as compared with the corresponding resolution spaces. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Integration of gas chromatography mass spectrometry methods for differentiating ricin preparation methods.

PubMed

Wunschel, David S; Melville, Angela M; Ehrhardt, Christopher J; Colburn, Heather A; Victry, Kristin D; Antolick, Kathryn C; Wahl, Jon H; Wahl, Karen L

2012-05-07

The investigation of crimes involving chemical or biological agents is infrequent, but presents unique analytical challenges. The protein toxin ricin is encountered more frequently than other agents and is found in the seeds of Ricinus communis, commonly known as the castor plant. Typically, the toxin is extracted from castor seeds utilizing a variety of different recipes that result in varying purity of the toxin. Moreover, these various purification steps can also leave or differentially remove a variety of exogenous and endogenous residual components with the toxin that may indicate the type and number of purification steps involved. We have applied three gas chromatography-mass spectrometry (GC-MS) based analytical methods to measure the variation in seed carbohydrates and castor oil ricinoleic acid, as well as the presence of solvents used for purification. These methods were applied to the same samples prepared using four previously identified toxin preparation methods, starting from four varieties of castor seeds. The individual data sets for seed carbohydrate profiles, ricinoleic acid, or acetone amount each provided information capable of differentiating different types of toxin preparations across seed types. However, the integration of the data sets using multivariate factor analysis provided a clear distinction of all samples based on the preparation method, independent of the seed source. In particular, the abundance of mannose, arabinose, fucose, ricinoleic acid, and acetone were shown to be important differentiating factors. These complementary tools provide a more confident determination of the method of toxin preparation than would be possible using a single analytical method.

Preparative isolation and purification of three stilbene glycosides from the tibetan medicinal plant Rheum tanguticum maxim. Ex Balf. by high-speed counter-current chromatography.

PubMed

Zhao, Xiao-Hui; Han, Fa; Li, Yu-Lin; Yue, Hui-Lan

2013-02-01

Stilbene glycosides are the primary constituents of Rheum tanguticum Maxim. ex Balf., to which different bioactivities has been attributed, including: anti-HIV, anti-oxidant, anti-tumour, anti-malarial, and anti-allergy activity. However, effective methods for the isolation and purification of stilbene glycosides, such as trans-rhapontin, cis-rhapontin and trans-desoxyrhaponticin, from this herb are not currently available. To develop an efficient method for the preparative isolation and purification of three stilbene glycosides from Rheum tanguticum Maxim. ex Balf. via high-speed counter-current chromatography (HSCCC). A solvent system composed of chloroform:n-butanol:methanol:water (4:1:3:2, v/v/v/v) was developed for the separation. The upper phase was used as the stationary phase, and the lower phase was used as the mobile phase. The flow rate was 1.8 mL/min. The apparatus was controlled at 800 rpm and 25 °C, and the effluent was monitored at 280 nm. Chemical constituents were analysed by high-performance liquid chromatography (HPLC), and their structures were identified by ¹H- and ¹³C-NMR. Under the optimised conditions, 25.5 mg trans-rhapontin, 16.0 mg cis-rhapontin and 20.5 mg trans-desoxyrhaponticin were separated from 80 mg crude sample; the isolates had purities of 99.6, 97.2 and 99.2%, respectively. A simple and efficient HSCCC method has been optimised for the preparative separation of stilbene glycosides from Rheum tanguticum Maxim. ex Balf. Copyright © 2012 John Wiley & Sons, Ltd.

Virus elimination during the purification of monoclonal antibodies by column chromatography and additional steps.

PubMed

Roberts, Peter L

2014-01-01

The theoretical potential for virus transmission by monoclonal antibody based therapeutic products has led to the inclusion of appropriate virus reduction steps. In this study, virus elimination by the chromatographic steps used during the purification process for two (IgG-1 & -3) monoclonal antibodies (MAbs) have been investigated. Both the Protein G (>7log) and ion-exchange (5 log) chromatography steps were very effective for eliminating both enveloped and non-enveloped viruses over the life-time of the chromatographic gel. However, the contribution made by the final gel filtration step was more limited, i.e., 3 log. Because these chromatographic columns were recycled between uses, the effectiveness of the column sanitization procedures (guanidinium chloride for protein G or NaOH for ion-exchange) were tested. By evaluating standard column runs immediately after each virus spiked run, it was possible to directly confirm that there was no cross contamination with virus between column runs (guanidinium chloride or NaOH). To further ensure the virus safety of the product, two specific virus elimination steps have also been included in the process. A solvent/detergent step based on 1% triton X-100 rapidly inactivating a range of enveloped viruses by >6 log inactivation within 1 min of a 60 min treatment time. Virus removal by virus filtration step was also confirmed to be effective for those viruses of about 50 nm or greater. In conclusion, the combination of these multiple steps ensures a high margin of virus safety for this purification process. © 2014 American Institute of Chemical Engineers.

Isolation of Exosomes from the Plasma of HIV-1 Positive Individuals

PubMed Central

Konadu, Kateena Addae; Huang, Ming Bo; Roth, William; Armstrong, Wendy; Powell, Michael; Villinger, Francois; Bond, Vincent

2016-01-01

Exosomes are small vesicles ranging in size from 30 nm to 100 nm that are released both constitutively and upon stimulation from a variety of cell types. They are found in a number of biological fluids and are known to carry a variety of proteins, lipids, and nucleic acid molecules. Originally thought to be little more than reservoirs for cellular debris, the roles of exosomes regulating biological processes and in diseases are increasingly appreciated. Several methods have been described for isolating exosomes from cellular culture media and biological fluids. Due to their small size and low density, differential ultracentrifugation and/or ultrafiltration are the most commonly used techniques for exosome isolation. However, plasma of HIV-1 infected individuals contains both exosomes and HIV viral particles, which are similar in size and density. Thus, efficient separation of exosomes from HIV viral particles in human plasma has been a challenge. To address this limitation, we developed a procedure modified from Cantin et. al., 2008 for purification of exosomes from HIV particles in human plasma. Iodixanol velocity gradients were used to separate exosomes from HIV-1 particles in the plasma of HIV-1 positive individuals. Virus particles were identified by p24 ELISA. Exosomes were identified on the basis of exosome markers acetylcholinesterase (AChE), and the CD9, CD63, and CD45 antigens. Our gradient procedure yielded exosome preparations free of virus particles. The efficient purification of exosomes from human plasma enabled us to examine the content of plasma-derived exosomes and to investigate their immune modulatory potential and other biological functions. PMID:26780239

Expression, purification and crystallization of a human protein SH3BGRL at atomic resolution

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yin, Lei; Zhu, De-Yu; Yang, Na

2005-04-01

The protein SH3BGRL, containing both SH3-binding and Homer EVH1-binding motifs, has been crystallized using the hanging-drop vapour-diffusion method. The protein SH3BGRL, containing both SH3-binding and Homer EVH1-binding motifs, has been crystallized using the hanging-drop vapour-diffusion method. The crystals diffract to 0.88 Å resolution and belong to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 28.8886, b = 34.9676, c = 98.0016 Å. Preliminary analysis indicates that the asymmetric unit contains one molecule and has a solvent content of about 34%.

RAPID COMMUNICATION: Large-area uniform ultrahigh-Jc YBa2Cu3O7-x film fabricated by the metalorganic deposition method using trifluoroacetates

NASA Astrophysics Data System (ADS)

Araki, Takeshi; Yamagiwa, Katsuya; Hirabayashi, Izumi; Suzuki, Katsumi; Tanaka, Shoji

2001-07-01

Ultrahigh-Jc YBa2Cu3O7-x (YBCO) films have been successfully fabricated by the metalorganic deposition method using a trifluoroacetate coating solution which is prepared by a newly developed purification technique, the solvent-into-gel (SIG) method. The prepared pure coating solution has less than 0.25% impurities and has a wide flexibility in process conditions to obtain high-Jc YBCO film. Using this feature, we have successfully formed 50 mm diameter YBCO films, which have a critical current density over 10 MA cm-2 (77 K, 0 T) on LaAlO3 single crystalline substrates.

Patulin Production by Penicillium urticae Bainier in Batch Culture1

PubMed Central

Norstadt, Fred A.; McCalla, T. M.

1969-01-01

A still, batch-culture method, with potato dextrose medium and Penicillium urticae Bainier, produced patulin yields of 1.2 to 1.7 g/liter of medium. Incubation was at 25 C for 14 days. Ethyl acetate extraction of condensed culture filtrate and drying with anhydrous MgSO4, followed by solvent change to dry ethyl ether and purification on alumina (pH 4.5), produced pure crystalline patulin. The use of 2-liter, round-bottom flasks and a rotating vacuum evaporator provided versatile equipment and easy manipulation in the operations. Soil from wheat fields provided a convenient natural P. urticae source. Potato dextrose medium was superior to potato sucrose or Raulin-Thom media. Images PMID:5775903

Linearly scaling and almost Hamiltonian dielectric continuum molecular dynamics simulations through fast multipole expansions

NASA Astrophysics Data System (ADS)

Lorenzen, Konstantin; Mathias, Gerald; Tavan, Paul

2015-11-01

Hamiltonian Dielectric Solvent (HADES) is a recent method [S. Bauer et al., J. Chem. Phys. 140, 104103 (2014)] which enables atomistic Hamiltonian molecular dynamics (MD) simulations of peptides and proteins in dielectric solvent continua. Such simulations become rapidly impractical for large proteins, because the computational effort of HADES scales quadratically with the number N of atoms. If one tries to achieve linear scaling by applying a fast multipole method (FMM) to the computation of the HADES electrostatics, the Hamiltonian character (conservation of total energy, linear, and angular momenta) may get lost. Here, we show that the Hamiltonian character of HADES can be almost completely preserved, if the structure-adapted fast multipole method (SAMM) as recently redesigned by Lorenzen et al. [J. Chem. Theory Comput. 10, 3244-3259 (2014)] is suitably extended and is chosen as the FMM module. By this extension, the HADES/SAMM forces become exact gradients of the HADES/SAMM energy. Their translational and rotational invariance then guarantees (within the limits of numerical accuracy) the exact conservation of the linear and angular momenta. Also, the total energy is essentially conserved—up to residual algorithmic noise, which is caused by the periodically repeated SAMM interaction list updates. These updates entail very small temporal discontinuities of the force description, because the employed SAMM approximations represent deliberately balanced compromises between accuracy and efficiency. The energy-gradient corrected version of SAMM can also be applied, of course, to MD simulations of all-atom solvent-solute systems enclosed by periodic boundary conditions. However, as we demonstrate in passing, this choice does not offer any serious advantages.

An integrated process for the recovery of high added-value compounds from olive oil using solid support free liquid-liquid extraction and chromatography techniques.

PubMed

Angelis, Apostolis; Hamzaoui, Mahmoud; Aligiannis, Nektarios; Nikou, Theodora; Michailidis, Dimitris; Gerolimatos, Panagiotis; Termentzi, Aikaterini; Hubert, Jane; Halabalaki, Maria; Renault, Jean-Hugues; Skaltsounis, Alexios-Léandros

2017-03-31

An integrated extraction and purification process for the direct recovery of high added value compounds from extra virgin olive oil (EVOO) is proposed by using solid support free liquid-liquid extraction and chromatography techniques. Two different extraction methods were developed on a laboratory-scale Centrifugal Partition Extractor (CPE): a sequential strategy consisting of several "extraction-recovery" cycles and a continuous strategy based on stationary phase co-current elution. In both cases, EVOO was used as mobile phase diluted in food grade n-hexane (feed mobile phase) and the required biphasic system was obtained by adding ethanol and water as polar solvents. For the sequential process, 17.5L of feed EVOO containing organic phase (i.e. 7L of EVOO treated) were extracted yielding 9.5g of total phenolic fraction corresponding to a productivity of 5.8g/h/L of CPE column. Regarding the second approach, the co-current process, 2L of the feed oil phase (containing to 0.8L of EVOO) were treated at 100mL/min yielding 1.03g of total phenolic fraction corresponding to a productivity of 8.9g/h/L of CPE column. The total phenolic fraction was then fractionated by using stepwise gradient elution Centrifugal Partition Chromatography (CPC). The biphasic solvent systems were composed of n-hexane, ethyl acetate, ethanol and water in different proportions (X/Y/2/3, v/v). In a single run of 4h on a column with a capacity of 1L, 910mg of oleocanthal, 882mg of oleacein, 104mg of hydroxytyrosol were successfully recovered from 5g of phenolic extract with purities of 85%, 92% and 90%, respectively. CPC fractions were then submitted to orthogonal chromatographic steps (adsorption on silica gel or size exclusion chromatography) leading to the isolation of additional eleven compounds belonging to triterpens, phenolic compounds and secoiridoids. Among them, elenolic acid ethylester was found to be new compound. Thin Layer Chromatography (TLC), Nuclear magnetic Resonance (NMR) and High Performance Liquid Chromatography - Diode Array Detector (HPLC-DAD) were used for monitoring and evaluation purposes throughout the entire procedure. Copyright © 2017 Elsevier B.V. All rights reserved.

Directional transport of colloids inside a bath of self-propelling walkers.

PubMed

Merlitz, Holger; Wu, Chenxu; Sommer, Jens-Uwe

2017-05-24

We present a setup in which passive colloids inside a solvent are moved to the boundaries of the container. The directional transport is facilitated by self-propelling microparticles ("walkers") with an activity gradient, which reduces their propulsion in the vicinity of bounding walls. An attractive interaction leads to the adsorption of walkers onto the colloid-surfaces in regions of low walker activity. It is shown that the activity gradient generates a free energy gradient which in turn acts as a driving force on the passive colloids. We carry out molecular dynamics simulations and present approaches to a theoretical description of the involved processes. Although the simulation data are not reproduced on a fully quantitative level, their qualitative features are covered by the model. The effect described here may be applied to facilitate a directional transport of drugs or to eliminate pollutants.

A straightforward method for measuring the range of apparent density of microplastics.

PubMed

Li, Lingyun; Li, Mengmeng; Deng, Hua; Cai, Li; Cai, Huiwen; Yan, Beizhan; Hu, Jun; Shi, Huahong

2018-10-15

Density of microplastics has been regarded as the primary property that affect the distribution and bioavailability of microplastics in the water column. For measuring the density of microplastis, we developed a simple and rapid method based on density gradient solutions. In this study, we tested four solvents to make the density gradient solutions, i.e., ethanol (0.8 g/cm 3 ), ultrapure water (1.0 g/cm 3 ), saturated NaI (1.8 g/cm 3 ) and ZnCl 2 (1.8 g/cm 3 ). Density of microplastics was measured via observing the float or sink status in the density gradient solutions. We found that density gradient solutions made from ZnCl 2 had a larger uncertainty in measuring density than that from NaI, most likely due to a higher surface tension of ZnCl 2 solution. Solutions made from ethanol, ultrapure water, and NaI showed consistent density results with listed densities of commercial products, indicating that these density gradient solutions were suitable for measuring microplastics with a density range of 0.8-1.8 g/cm 3 . Copyright © 2018 Elsevier B.V. All rights reserved.

Method for efficient recovery of high-purity polycarbonates from electronic waste.

PubMed

Weeden, George S; Soepriatna, Nicholas H; Wang, Nien-Hwa Linda

2015-02-17

More than one million tons of polycarbonates from waste electrical and electronic equipment are consigned to landfills at an increasing rate of 3-5% per year. Recycling the polymer waste should have a major environmental impact. Pure solvents cannot be used to selectively extract polycarbonates from mixtures of polymers with similar properties. In this study, selective mixed solvents are found using guidelines from Hansen solubility parameters, gradient polymer elution chromatography, and solubility tests. A room-temperature sequential extraction process using two mixed solvents is developed to recover polycarbonates with high yield (>95%) and a similar purity and molecular weight distribution as virgin polycarbonates. The estimated cost of recovery is less than 30% of the cost of producing virgin polycarbonates from petroleum. This method would potentially reduce raw materials from petroleum, use 84% less energy, reduce emission by 1-6 tons of CO2 per ton of polycarbonates, and reduce polymer accumulation in landfills and associated environmental hazards.

Determination of zearalenone and its metabolites in endometrial cancer by coupled separation techniques.

PubMed

Gadzała-Kopciuch, Renata; Cendrowski, Krzysztof; Cesarz, Anna; Kiełbasa, Paweł; Buszewski, Bogusław

2011-10-01

This study presents a selective method of isolation of zearalenone (ZON) and its metabolite, α-zearalenol (α-ZOL), in neoplastically changed human tissue by accelerated solvent and ultrasonic extractions using a mixture of acetonitrile/water (84/16% v/v) as the extraction solvent. Extraction effectiveness was determined through the selection of parameters (composition of the solvent mixture, temperature, pressure, number of cycles) with tissue contamination at the level of nanograms per gram. The produced acetonitrile/water extracts were purified, and analytes were enriched in columns packed with homemade molecularly imprinted polymers. Purified extracts were determined by liquid chromatography (LC) coupled with different detection systems (diode array detection--DAD and mass spectrometry--MS) involving the Ascentis RP-Amide as a stationary phase and gradient elution. The combination of UE-MISPE-LC (ultrasonic extraction--molecularly imprinted solid-phase extraction--liquid chromatography) produced high (R≈95-98%) and repeatable (RSD<3%) recovery values for ZON and α-ZOL. © The Author(s) 2011. This article is published with open access at Springerlink.com

Solubilities of carbon dioxide in aqueous potassium carbonate solutions mixed with physical solvents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, S.B.; Lee, H.; Lee, K.H.

1998-09-01

The removal of acidic gases such as CO{sub 2}, H{sub 2}S, and COS from gas streams is a very important operation for petrochemical, oil refineries, ammonia manufacture, coal gasification, and natural gas purification plants. Here, the solubilities of carbon dioxide in aqueous potassium carbonate (K{sub 2}CO{sub 3}) solutions mixed with physical solvents were measured at 298.2 and 323.2 K with a CO{sub 2} partial-pressure range of 5 kPa to 2 MPa. 1,2-propanediol and propylene carbonate were selected as physical solvents. The aqueous solutions treated in this study were 5 mass% K{sub 2}CO{sub 3}-15 mass% 1,2-propanediol and propylene carbonate were selectedmore » as physical solvents. The aqueous solutions treated in this study were 5 mass% K{sub 2}CO{sub 3}-15 mass% propylene carbonate. The experimental solubility results were presented by the mole ratio of CO{sub 2} and K{sub 2}CO{sub 3} contained in the liquid mixture. The addition of 1,2-propanediol to 5 mass% K{sub 2}CO{sub 3} solution lowered the solubility of CO{sub 2} at constant temperature and pressure conditions when CO{sub 2} partial-pressure range of 5 kPa to 2 MPa. In the case of propylene carbonate the addition of propylene carbonate increased the experimental solubilities in the region of low CO{sub 2} partial pressures and decreased as the CO{sub 2} partial pressure was increased above atmospheric. The solubilities of CO{sub 2} decreased with increasing temperature in the range of 298.2 to 323.2 K.« less

Extraction and identification of flavonoids from parsley extracts by HPLC analysis

NASA Astrophysics Data System (ADS)

Stan, M.; Soran, M. L.; Varodi, C.; Lung, I.

2012-02-01

Flavonoids are phenolic compounds isolated from a wide variety of plants, and are valuable for their multiple properties, including antioxidant and antimicrobial activities. In the present work, parsley (Petroselinum crispum L.) extracts were obtained by three different extraction techniques: maceration, ultrasonic-assisted and microwave-assisted solvent extractions. The extractions were performed with ethanol-water mixtures in various ratios. From these extracts, flavonoids like the flavones apigenin and luteolin, and the flavonols quercetin and kaempferol were identified using an HPLC Shimadzu apparatus equipped with PDA and MS detectors. The separation method involved a gradient step. The mobile phase consisted of two solvents: acetonitrile and distilled water with 0.1% formic acid. The separation was performed on a RP-C18 column.

Highly sensitive detection of naphthalene in solvent vapor using a functionalized PBG refractive index sensor.

PubMed

Girschikofsky, Maiko; Rosenberger, Manuel; Belle, Stefan; Brutschy, Malte; Waldvogel, Siegfried R; Hellmann, Ralf

2012-01-01

We report an optical refractive index sensor system based on a planar Bragg grating which is functionalized by substituted γ-cyclodextrin to determine low concentrations of naphthalene in solvent vapor. The sensor system exhibits a quasi-instantaneous shift of the Bragg wavelength and is therefore capable for online detection. The overall shift of the Bragg wavelength reveals a linear relationship to the analyte concentration with a gradient of 12.5 ± 1.5 pm/ppm. Due to the spectral resolution and repeatability of the interrogation system, this corresponds to acquisition steps of 80 ppb. Taking into account the experimentally detected signal noise a minimum detection limit of 0.48 ± 0.05 ppm is deduced.

Microscale solution manipulation using photopolymerized hydrogel membranes and induced charge electroosmosis micropumps

NASA Astrophysics Data System (ADS)

Paustian, Joel Scott

Microfluidic technology is playing an ever-expanding role in advanced chemical and biological devices, with diverse applications including medical diagnostics, high throughput research tools, chemical or biological detection, separations, and controlled particle fabrication. Even so, local (microscale) modification of solution properties within microchannels, such as pressure, solute concentration, and voltage remains a challenge, and improved spatiotemporal control would greatly enhance the capabilities of microfluidics. This thesis demonstrates and characterizes two microfluidic tools to enhance local solution control. I first describe a microfluidic pump that uses an electrokinetic effect, Induced-Charge Electroosmosis (ICEO), to generate pressure on-chip. In ICEO, steady flows are driven by AC fields along metal-electrolyte interfaces. I design and microfabricate a pump that exploits this effect to generate on-chip pressures. The ICEO pump is used to drive flow along a microchannel, and the pressure is measured as a function of voltage, frequency, and electrolyte composition. This is the first demonstration of chip-scale flows driven by ICEO, which opens the possibility for ICEO pumping in self-contained microfluidic devices. Next, I demonstrate a method to create thin local membranes between microchannels, which enables local diffusive delivery of solute. These ``Hydrogel Membrane Microwindows'' are made by photopolymerizing a hydrogel which serves as a local ``window'' for solute diffusion and electromigration between channels, but remains a barrier to flow. I demonstrate three novel experimental capabilities enabled by the hydrogel membranes: local concentration gradients, local electric currents, and rapid diffusive composition changes. I conclude by applying the hydrogel membranes to study solvophoresis, the migration of particles in solvent gradients. Solvent gradients are present in many chemical processes, but migration of particles within these gradients is not well understood. An improved understanding would allow solvophoresis to be engineered (e.g. for coatings and thin film deposition) or reduced (e.g. in fouling processes during reactions and separations). Toward this end, I perform velocity measurements of colloidal particles at various ethanol-water concentrations and gradient strengths. The velocity was found to depend on the mole fraction via the equation u = DSP▿ln X, where u is the velocity, DSP is the mobility, and X is the ethanol mole fraction.

Separation and purification of polyphenols from red wine extracts using high speed counter current chromatography.

PubMed

Li, Yuanyuan; Li, Lingxi; Cui, Yan; Zhang, Shuting; Sun, Baoshan

2017-06-01

Polyphenols are important compounds of red wine owing to their contribution to sensory properties and antioxidant activities. In this study, high-speed counter-current chromatography (HSCCC) coupled with semi-preparative HPLC was used for large-scale separation and purification of polyphenols from red wine extracts. With the solvent system of hexane-ethyl acetate-water (1-50-50), various oligomeric procyanidins including monomer catechin, epicatechin, dimers B1, B2; phenolic acids including coutaric acid, caftaric acid and other type of polyphenols were largely separated within 370min and most of these compounds presented high yields (0.97mg to 13.79mg) with high purity (90.34% to 98.91%) after the semi-preparative HPLC isolation. Using the solvent system of Methyl tert-Butyl Ether (MTBE) - n-butyl alcohol- acetonitrile-water (1-40-1-50, acidified with 0.01% trifluoroacetic acid (TFA)) by one-step HSCCC of 100mg of the red wine extracts, the major anthocyanins, i.e., malvidin-3-O-glucoside, delphinidin-3-O-glucoside and peonidin-3-O-glucoside, as well as two polymeric proanthocyanidin fractions were successfully separated one another within 320min. The yields of malvidin-3-O-glucoside, delphinidin-3-O-glucoside and peonidin-3-O-glucoside were 12.12mg, 1.78mg and 11.57mg with the purity of 92.74%, 91.03% and 91.21%, respectively. Thiolysis-UPLC analysis indicated that the two polymeric proanthocyanidin fractions presented high purity, with mean degree of polymerization of 7.66±0.12 and 6.20±0.09, respectively. The further experiments on the antioxidant activities by DPPH radical test, FRAP assay and ABTS method showed that all of the isolated procyandins and anthocyanins and the two polymeric proanthocyanidin fractions, with exception of phenolic acids possessed much greater antioxidant activities compared to standard Trolox andl-ascorbic acid (2-14 times). Copyright © 2017 Elsevier B.V. All rights reserved.

Algorithmic cooling in liquid-state nuclear magnetic resonance

NASA Astrophysics Data System (ADS)

Atia, Yosi; Elias, Yuval; Mor, Tal; Weinstein, Yossi

2016-01-01

Algorithmic cooling is a method that employs thermalization to increase qubit purification level; namely, it reduces the qubit system's entropy. We utilized gradient ascent pulse engineering, an optimal control algorithm, to implement algorithmic cooling in liquid-state nuclear magnetic resonance. Various cooling algorithms were applied onto the three qubits of C132-trichloroethylene, cooling the system beyond Shannon's entropy bound in several different ways. In particular, in one experiment a carbon qubit was cooled by a factor of 4.61. This work is a step towards potentially integrating tools of NMR quantum computing into in vivo magnetic-resonance spectroscopy.

Large scale micro-photometry for high resolution pH-characterization during electro-osmotic pumping and modular micro-swimming

NASA Astrophysics Data System (ADS)

Niu, Ran; Khodorov, Stanislav; Weber, Julian; Reinmüller, Alexander; Palberg, Thomas

2017-11-01

Micro-fluidic pumps as well as artificial micro-swimmers are conveniently realized exploiting phoretic solvent flows based on local gradients of temperature, electrolyte concentration or pH. We here present a facile micro-photometric method for monitoring pH gradients and demonstrate its performance and scope on different experimental situations including an electro-osmotic pump and modular micro-swimmers assembled from ion exchange resin beads and polystyrene colloids. In combination with the present microscope and DSLR camera our method offers a 2 μm spatial resolution at video frame rate over a field of view of 3920 × 2602 μm2. Under optimal conditions we achieve a pH-resolution of 0.05 with about equal contributions from statistical and systematical uncertainties. Our quantitative micro-photometric characterization of pH gradients which develop in time and reach out several mm is anticipated to provide valuable input for reliable modeling and simulations of a large variety of complex flow situations involving pH-gradients including artificial micro-swimmers, microfluidic pumping or even electro-convection.

Parapoxvirus papillomatosis in the muskoxen (Ovibos moschatus): genetical differences between the virus causing new outbreak in a vaccinated herd, the vaccine virus and a local orf virus.

PubMed

Moens, U; Wold, I; Mathiesen, S D; Jørgensen, T; Sørensen, D; Traavik, T

1990-01-01

Since 1981 a domesticated muskoxen herd had been successfully vaccinated against papillomatosis with homogenated, glutaraldehyde inactivated papilloma tissue. In the fall of 1985 a new clinical outbreak of disease occurred, affecting previously infected as well as vaccinated animals. The purification of parapox virions directly from papilloma tissue and orf scabs collected in a local sheep farm was followed by restriction endonuclease analysis of viral DNA. The morphological identity of purified virus was controlled by electron microscopy. Comparison of restriction endonuclease digests (10 different enzymes) by gel electrophoresis demonstrated that the muskoxen parapoxvirus from the new outbreak 1985 differed considerably from the 2 other isolates (muskoxen 1981 and local orf). The latter viruses demonstrated a high degree of homology, but differences were evident after digestion with the enzyme EcoRI. During metrizamide gradient purification minor bands containing morphologically intact virions were isolated in addition to the major fractions. The restriction enzyme digests indicated that the virions of the minor bands differed from those in the major bands.

Involvement of the Golgi apparatus in the synthesis and secretion of hydroxyproline-rich cell wall glycoproteins. [carrot

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gardiner, M.; Chrispeels, M.J.

1975-01-01

Pulse labeling of carrot root phloem parenchyma (Daucus carota L. ev. Nantes) tissue with /sup 14/C-proline followed by fractionation of the cytoplasmic organelles on sucrose gradients was used to determine the identiy of the membranous organelles involved in the secretion of the hydroxyproline-rich glycoproteins of the cell wall. Identification of the organelles was done through electron-microscopical observations and through the localization of marker enzymes on the sucrose gradients. Enrichment of the organelles involved in secretion was determined by measuring the percentage of the incorporated radioactivity present as /sup 14/C-hydroxyproline. The Golgi apparatus (dictyosome) was found to be a major sitemore » of glycoprotein transport. This identification was based on the observed enrichment of dictyosomes paralleling the purification of newly synthesized cell-wall glycoproteins. A marker enzyme for the Golgi apparatus, inosinediphosphatase, banded with the newly synthesized cell wall glycoproteins on sequential isopycnic and rate zonal sucrose gradients. Marker enzymes for the endoplasmic reticulum and the plasma memebrane were clearly separated from the dictyosome-rich fraction. UDP-arabinose arabinosyl transferase, an enzyme involved in the glycosylation of the peptide moiety of this glycoprotein, also banded with the dictyosomes on both kinds of gradients. The results suggest an important role of the Golgi apparatus in the biosynthesis and the secretion of the cell wall glycoproteins of higher plants. (auth)« less

Protein purification by aminosquarylium cyanine dye-affinity chromatography.

PubMed

Silva, M S; Graça, V C; Reis, L V; Santos, P F; Almeida, P; Queiroz, J A; Sousa, F

2013-12-01

The most selective purification method for proteins and other biomolecules is affinity chromatography. This method is based on the unique biological-based specificity of the biomolecule-ligand interaction and commonly uses biological ligands. However, these ligands may present some drawbacks, mainly because of their cost and lability. Dye-affinity chromatography overcomes the limitations of biological ligands and is widely used owing to the low cost of synthetic dyes and to their resistance to biological and chemical degradation. In this work, immobilized aminosquarylium cyanine dyes are used in order to exploit affinity interactions with standard proteins such as lysozyme, α-chymotrypsin and trypsin. These studies evaluate the affinity interactions occurring between the immobilized ligand and the different proteins, as a reflection of the sum of several molecular interactions, namely ionic, hydrophobic and van der Waals, spread throughout the structure, in a defined spatial manner. The results show the possibility of using an aminosquarylium cyanine dye bearing a N-hexyl pendant chain, with a ligand density of 1.8 × 10(-2) mmol of dye/g of chromatographic support, to isolate lysozyme, α-chymotrypsin and trypsin from a mixture. The application of a decreasing ammonium sulfate gradient resulted in the recovery of lysozyme in the flowthrough. On the other hand, α-chymotrypsin and trypsin were retained, involving different interactions with the ligand. In conclusion, this study demonstrates the potential applicability of ligands such as aminosquarylium cyanine dyes for the separation and purification of proteins by affinity chromatography. Copyright © 2013 John Wiley & Sons, Ltd.

Solute-Solvent Charge-Transfer Excitations and Optical Absorption of Hydrated Hydroxide from Time-Dependent Density-Functional Theory.

PubMed

Opalka, Daniel; Sprik, Michiel

2014-06-10

The electronic structure of simple hydrated ions represents one of the most challenging problems in electronic-structure theory. Spectroscopic experiments identified the lowest excited state of the solvated hydroxide as a charge-transfer-to-solvent (CTTS) state. In the present work we report computations of the absorption spectrum of the solvated hydroxide ion, treating both solvent and solute strictly at the same level of theory. The average absorption spectrum up to 25 eV has been computed for samples taken from periodic ab initio molecular dynamics simulations. The experimentally observed CTTS state near the onset of the absorption threshold has been analyzed at the generalized-gradient approximation (GGA) and with a hybrid density-functional. Based on results for the lowest excitation energies computed with the HSE hybrid functional and a Davidson diagonalization scheme, the CTTS transition has been found 0.6 eV below the first absorption band of liquid water. The transfer of an electron to the solvent can be assigned to an excitation from the solute 2pπ orbitals, which are subject to a small energetic splitting due to the asymmetric solvent environment, to the significantly delocalized lowest unoccupied orbital of the solvent. The distribution of the centers of the excited state shows that CTTS along the OH(-) axis of the hydroxide ion is avoided. Furthermore, our simulations indicate that the systematic error arising in the calculated spectrum at the GGA originates from a poor description of the valence band energies in the solution.

Purification of antibiotics from the biocontrol agent Streptomyces anulatus S37 by centrifugal partition chromatography.

PubMed

Couillerot, Olivier; Loqman, Souad; Toribio, Alix; Hubert, Jane; Gandner, Léa; Nuzillard, Jean-Marc; Ouhdouch, Yedir; Clément, Christophe; Barka, Essaid Ait; Renault, Jean-Hugues

2014-01-01

A novel actinomycete strain, Streptomyces anulatus S37, has been isolated from the rhizosphere of healthy Moroccan Vitis vinifera on the basis on its ability to promote grapevine growth and to induce natural defences against various phytopathogens. In the present work, the main bioactive metabolites produced by S. anulatus S37 were isolated. A crude n-BuOH extract of the S37 fermentation broth was firstly partitioned in a biphasic solvent system composed of n-heptane, methanol, and water (5:1.5:3.5, v/v). The most active organic fraction (1.1g) as revealed by TLC-bioautography was subsequently separated by a two-step centrifugal partition chromatography procedure. The first separation was performed in the ascending mode at 6mL/min with the biphasic solvent system n-heptane, ethyl acetate, methanol and water (2:1:2:1, v/v), to finally recover 40mg of a pure compound identified as streptochlorin by NMR spectroscopy. In a second separation, the solvent system n-heptane, acetonitrile, and water (5:5:4, v/v) was used in the ascending mode at 3mL/min to purify 135mg of nigericin and 53mg of piericidin A1. Assays performed with the three compounds have confirmed their inhibitory impact on the growth of Botryris cinerea in dual confrontation and also on V. vinifera L. plantlets. Copyright © 2013 Elsevier B.V. All rights reserved.

Synthesis of N-vinylpyrrolidone modified acrylic acid copolymer in supercritical fluids and its application in dental glass-ionomer cements.

PubMed

Moshaverinia, Alireza; Roohpour, Nima; Billington, Richard W; Darr, Jawwad A; Rehman, Ihtesham U

2008-07-01

Compressed fluids such as supercritical CO(2) offer marvellous opportunities for the synthesis of polymers, particularly in applications in medicine and dentistry. It has several advantages in comparison to conventional polymerisation solvents, such as enhanced kinetics and simplified solvent removal process. In this study, poly(acrylic acid-co-itaconic acid-co-N-vinylpyrrolidone) (PAA-IA-NVP), a modified glass-ionomer polymer, was synthesised in supercritical CO(2) (sc-CO(2)) and methanol as a co-solvent. The synthesised polymer was characterized by (1)H-NMR, Raman and FT-IR spectroscopy and viscometry. The molecular weight of the final product was also measured using static light scattering method. The synthesised polymers were subsequently used in several glass ionomer cement formulations (Fuji II commercial GIC) in which mechanical strength (compressive strength (CS), diametral tensile strength (DTS) and biaxial flexural strength (BFS)) and handling properties (working and setting time) of the resulting cements were evaluated. The polymerisation reaction in sc-CO(2)/methanol was significantly faster than the corresponding polymerisation reaction in water and the purification procedures were simpler for the former. Furthermore, glass ionomer cement samples made from the terpolymer prepared in sc-CO(2)/methanol exhibited higher CS and DTS and comparable BFS compared to the same polymer synthesised in water. The working properties of glass ionomer formulations made in sc-CO(2)/methanol were comparable and in selected cases better than the values of those made from polymers synthesised in water.

Isolation of sutherlandins A, B, C and D from Sutherlandia frutescens (L.) R. Br. by counter-current chromatography using spiral tubing support rotors.

PubMed

Chen, Cuiping; Folk, William R; Lazo-Portugal, Rodrigo; Finn, Thomas M; Knight, Martha

2017-07-28

Spiral countercurrent-chromatography has great potential for improving the capacity and efficiency of purification of secondary metabolites, and here we describe applications useful for the isolation of flavonoids from the widely used South African medicinal plant, Sutherlandia frutescens (L.) R. Br. In the spiral tubing support rotor, STS-4 for high-speed counter-current chromatography, several polar butanol aqueous solvent systems were selected using a logK plot, and the novel flavonol glycosides (sutherlandins A-D) were well separated by the optimized solvent system (ethyl acetate:n-butanol:acetic acid:water; 5:1:0.3:6 by vol.). The yield of purified flavonoids from 0.9g extract varied from 8.6mg to 54mg of the sutherlandins for a total of 85.3mg. The same extract was fractionated in the new STS-12 rotor of the same outside dimensions but with more radial channels forming 12 loops of the tubing instead of 4. The rotor holds more layers and increased length of tubing. From 0.9g extract the STS-12 rotor yielded more recovery of 110.4mg total with amounts varying from 11.2mg to 64mg of the sutherlandins and apparent increased separation efficiency as noted by less volume of each fraction peak. Thus from 1-g amounts of extract, good recovery of the flavonoids was achieved in the butanol aqueous solvent system. Copyright © 2017 Elsevier B.V. All rights reserved.

Production of acetone butanol ethanol (ABE) by a hyper-producing mutant strain of Clostridium beijerinckii BA101 and recovery by pervaporation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Qureshi, N.; Blaschek, H.P.

1999-07-01

A silicone membrane was used to study butanol separation from model butanol solutions and fermentation broth. Depending upon the butanol feed concentration in the model solution and pervaporation conditions, butanol selectivities of 20.88--68.32 and flux values of 158.7--215.4 g m{sup {minus}2} h{sup {minus}1} were achieved. Higher flux values were obtained at higher butanol concentrations using air as sweep gas. In an integrated process of butanol fermentation--recovery, solvent productivities were improved to 200% of the control batch fermentation productivities. In a batch reactor the hyper-butanol-producing mutant strain C. beijerinckii BA101 utilized 57.3 g/L glucose and produced 24.2 g/L total solvents, whilemore » in the integrated process it produced 51.5 g/L (culture volume) total solvents. Concentrated glucose medium was also fermented. The C. beijerinckii BA101 mutant strain was not negatively affected by the pervaporative conditions. In the integrated experiment, acids were not produced. With the active fermentation broth, butanol selectivity was reduced by a factor of 2--3. However, the membrane flux was not affected by the active fermentation broth. The butanol permeate concentration ranged from 26.4 to 95.4 g/L, depending upon butanol concentration in the fermentation broth. Since the permeate of most membranes contains acetone, butanol, and ethanol, it is suggested that distillation be used for further purification.« less

A novel nonionic surfactant- and solvent-stable alkaline serine protease from Serratia sp. SYBC H with duckweed as nitrogen source: production, purification, characteristics and application.

PubMed

Li, G Y; Cai, Y J; Liao, X R; Yin, J

2011-07-01

A novel nonionic surfactant- and hydrophilic solvent-stable alkaline serine protease was purified from the culture supernatant of Serratia sp. SYBC H with duckweed as nitrogen source. The molecular mass of the purified protease is about 59 kDa as assayed via SDS-PAGE. The protease is highly active over the pH range between 5.0 and 11.0, with the maximum activity at pH 8.0. It is also fairly active over the temperature range between 30 and 80°C, with the maximum activity at 40°C. The protease activity was substantially stimulated by Mn(2+) and Na(+) (5 mM), up to 837.9 and 134.5% at 40°C, respectively. In addition, Mn(2+) enhanced the thermostability of the protease significantly at 60°C. Over 90% of its initial activity remained even after incubating for 60 min at 40°C in 50% (v/v) hydrophilic organic solvents such as DMF, DMSO, acetone and MeOH. The protease retained 81.7, 83.6 and 76.2% of its initial activity in the presence of nonionic surfactants 20% (v/v) Tween 80, 25% (v/v) glycerol and Triton X-100, respectively. The protease is strongly inhibited by PMSF, suggesting that it is a serine protease. Washing experiments revealed that the protease has an excellent ability to remove blood stains.

Ionic liquid molecularly imprinted polymers for application in pipette-tip solid-phase extraction coupled with gas chromatography for rapid screening of dicofol in celery.

PubMed

Yan, Hongyuan; Yang, Chen; Sun, Yunyun; Row, Kyung Ho

2014-09-26

A new type of ionic liquid molecularly imprinted polymers (IL-MIPs) synthesized by precipitation polymerization using 1-allyl-3-methylimidazolium bromide as an auxiliary solvent and α-chloro-dichlorodiphenyltrichloroethane (α-chloro-DDT) as the template was applied as a selective sorbent of minimized pipette tip-solid-phase extraction (PT-SPE) for rapid isolation and extraction of dicofol (DCF) from celery samples. The pretreatment procedure of celery samples involved only 2.0mg of IL-MIPs, 0.8 mL of acetonitrile-water (ACN-H2O; 1:1, v/v) (washing solvent), and 1.0 mL of acetone-10% acetic acid (HOAc) (elution solvent). Compared with molecularly imprinted polymers (MIPs), ionic liquid-non-imprinted polymers (IL-NIPs) and conventional sorbents such as C18, Si, NH2, and Al2O3-N, IL-MIPs showed higher adsorption and purification capacity to DCF in aqueous solution. Good linearity for DCF was observed in the range from 2.3 to 232.5 ng g(-1) (r(2)=0.9995). The average recoveries at three spiking levels ranged from 86.6% to 101.9% with relative standard deviations (RSDs) of ≤ 6.5% (n=3). The presented IL-MIPs-PT-SPE-GC/ECD method combines the advantages of MIPs, IL, and PT-SPE, and can be used in aqueous conditions with high affinity and selectivity to analytes of complex samples. Copyright © 2014 Elsevier B.V. All rights reserved.

Tar Management and Recycling in Biomass Gasification and Syngas Purification

NASA Astrophysics Data System (ADS)

McCaffrey, Zach

Removal of tars is critical to the design and operation of biomass gasification systems as most syngas utilization processing equipment (e.g. internal combustion engines, gas turbines, fuel cells, and liquid fuel synthesis reactors) have a low tolerance for tar. Capturing and disposal of tar is expensive due to equipment costs, high hazardous waste disposal costs where direct uses cannot be found, and system energy losses incurred. Water scrubbing is an existing technique commonly used in gasification plants to remove contaminants and tar; however using water as the absorbent is non-ideal as tar compounds have low or no water solubility. Hydrophobic solvents can improve scrubber performance and this study evaluated tar solubility in selected solvents using slip-streams of untreated syngas from a laboratory fluidized bed reactor operated on almond composite feedstock using both air and steam gasification. Tar solubility was compared with Hansen's solubility theory to examine the extent to which the tar removal can be predicted. As collection of tar without utilization leads to a hazardous waste problem, the study investigated the effects of recycling tars back into the gasifier for destruction. Prior to experiments conducted on tar capture and recycle, characterizations of the air and steam gasification of the almond composite mix were made. This work aims to provide a better understanding of tar collection and solvent selection for wet scrubbers, and to provide information for designing improved tar management systems for biomass gasification.

Linearly scaling and almost Hamiltonian dielectric continuum molecular dynamics simulations through fast multipole expansions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lorenzen, Konstantin; Mathias, Gerald; Tavan, Paul, E-mail: tavan@physik.uni-muenchen.de

2015-11-14

Hamiltonian Dielectric Solvent (HADES) is a recent method [S. Bauer et al., J. Chem. Phys. 140, 104103 (2014)] which enables atomistic Hamiltonian molecular dynamics (MD) simulations of peptides and proteins in dielectric solvent continua. Such simulations become rapidly impractical for large proteins, because the computational effort of HADES scales quadratically with the number N of atoms. If one tries to achieve linear scaling by applying a fast multipole method (FMM) to the computation of the HADES electrostatics, the Hamiltonian character (conservation of total energy, linear, and angular momenta) may get lost. Here, we show that the Hamiltonian character of HADESmore » can be almost completely preserved, if the structure-adapted fast multipole method (SAMM) as recently redesigned by Lorenzen et al. [J. Chem. Theory Comput. 10, 3244-3259 (2014)] is suitably extended and is chosen as the FMM module. By this extension, the HADES/SAMM forces become exact gradients of the HADES/SAMM energy. Their translational and rotational invariance then guarantees (within the limits of numerical accuracy) the exact conservation of the linear and angular momenta. Also, the total energy is essentially conserved—up to residual algorithmic noise, which is caused by the periodically repeated SAMM interaction list updates. These updates entail very small temporal discontinuities of the force description, because the employed SAMM approximations represent deliberately balanced compromises between accuracy and efficiency. The energy-gradient corrected version of SAMM can also be applied, of course, to MD simulations of all-atom solvent-solute systems enclosed by periodic boundary conditions. However, as we demonstrate in passing, this choice does not offer any serious advantages.« less

Purification and Chemical Control of Molten Li2BeF 4 for a Fluoride Salt Cooled Reactor

NASA Astrophysics Data System (ADS)

Kelleher, Brian Christopher

Out of the many proposed generation IV, high-temperature reactors, the molten salt reactor (MSR) is one of the most promising. The first large scale MSR, the molten salt reactor experiment (MSRE), operated from 1965 to 1969 using Li2BeF4, or flibe, as a coolant and solvent for uranium fluoride fuel, at maximum temperatures of 654°C, for over 15000 hours. The MSRE experienced no concept breaking surprises and was considered a success. Newly proposed designs of molten salt reactors use solid fuels, making them less exotic compared to the MSRE. However, any molten salt reactor will require a great deal of research pertaining to the chemical and mechanical mastery of molten salts in order to prepare it for commercialization. To supplement the development of new molten salt reactors, approximately 100 kg of flibe was purified using the standard hydrofluorination process. Roughly half of the purified salt was lithium-7 enriched salt from the secondary loop of the MSRE. Purification rids the salt of impurities and reduces its capacity for corrosion, also known as the redox potential. The redox potential of flibe was measured at various stages of purification for the first time using a dynamic beryllium reference electrode. These redox measurements have been superimposed with metal impurities measurements found by neutron activation analysis. Lastly, reductions of flibe with beryllium metal have been investigated. Over reductions have been performed, which have shown to decrease redox potential while seemingly creating a beryllium-beryllium halide system. Recommendations of the lowest advisable redox potential for corrosion tests are included along with suggestions for future work.

Optimization of extractants, purifying packings, and eluents for analytical extraction of organochlorine pesticides in Hydragric Acrisols.

PubMed

Zhao, Zhenhua; Xia, Liling; Wang, Fang; Jiang, Xin; Gao, Yanzheng

2012-08-01

In this study, we screened for an economic, rapid, and efficient hypotoxic pretreatment method for organochlorine pesticides in soil samples for gas chromatography (GC) analysis. The analytical extraction efficiencies of 11 different extractants, nine types of solid-phase purification (SPP) cartridge packings, and three types of eluents for 13 organochlorine pesticides (OCPs) in spiked and natural Chinese red soil (Hydragric Acrisols) were evaluated using an ultrasonic extraction and solid-phase purification method. High percent recoveries (85-106%) were obtained for the 13 organochlorine pesticides in soil when petroleum ether/acetone/water (10:5:2, v/v) was used an extractant. They were purified using celite SPP cartridge packing and eluted with 9 mL of dichloromethane/petroleum ether (1:9, v/v). The OCPs purification pretreatment of Hydragric Acrisols, using the above method, meets the GC analysis requirements. Compared with other traditional pretreatment methods for OCPs in soil samples, this method has several advantages, such as a short extraction time, reducing the amount of solvent, having no emulsion phenomenon, and hypotoxicity to the laboratory technicians. The concentrations of 1,1,1,-trichloro-2(p-chlorophenyl)-2-(o-chlorophenyl) ethane (DDTs; 3.42-8.08 ng g(-1)) in field soils were higher than the hexachlorocyclohexane concentration (2.94-6.12 ng g(-1)). The 1,1-dichloro-2,2-bis(p-chlorophenyl) ethylene (p,p'-DDE) + 1,1-dichloro-2,2-bis(p-chlorophenyl)-ethane (p,p'-DDD)/p,p'-DDT ratio in this field soil was approximately 2.7, suggesting that no new DDT pollution source was introduced into the sampling site.

Design and optimization of a chromatographic purification process for Streptococcus pneumoniae serotype 23F capsular polysaccharide by a Design of Experiments approach.

PubMed

Ji, Yu; Tian, Yang; Ahnfelt, Mattias; Sui, Lili

2014-06-27

Multivalent pneumococcal vaccines were used worldwide to protect human beings from pneumococcal diseases. In order to eliminate the toxic organic solutions used in the traditional vaccine purification process, an alternative chromatographic process for Streptococcus pneumoniae serotype 23F capsular polysaccharide (CPS) was proposed in this study. The strategy of Design of Experiments (DoE) was introduced into the process development to solve the complicated design procedure. An initial process analysis was given to review the whole flowchart, identify the critical factors of chromatography through FMEA and chose the flowthrough mode due to the property of the feed. A resin screening study was then followed to select candidate resins. DoE was utilized to generate a resolution IV fractional factorial design to further compare candidates and narrow down the design space. After Capto Adhere was selected, the Box-Behnken DoE was executed to model the process and characterize all effects of factors on the responses. Finally, Monte Carlo simulation was used to optimize the process, test the chosen optimal conditions and define the control limit. The results of three scale-up runs at set points verified the DoE and simulation predictions. The final results were well in accordance with the EU pharmacopeia requirements: Protein/CPS (w/w) 1.08%; DNA/CPS (w/w) 0.61%; the phosphorus content 3.1%; the nitrogen 0.315% and the Methyl-pentose percentage 47.9%. Other tests of final pure CPS also met the pharmacopeia specifications. This alternative chromatographic purification process for pneumococcal vaccine without toxic organic solvents was successfully developed by the DoE approach and proved scalability, robustness and suitability for large scale manufacturing. Copyright © 2014 Elsevier B.V. All rights reserved.

Continuous gradient scaffolds for rapid screening of cell-material interactions and interfacial tissue regeneration

PubMed Central

Bailey, Brennan M.; Nail, Lindsay N.; Grunlan, Melissa A.

2013-01-01

In tissue engineering, the physical and chemical properties of the scaffold mediates cell behavior including regeneration. Thus, a strategy that permits rapid screening of cell-scaffold interactions is critical. Herein, we have prepared eight “hybrid” hydrogel scaffolds in the form of continuous gradients such that a single scaffold contains spatially varied properties. These scaffolds are based on combining an inorganic macromer [methacrylated star polydimethylsiloxane, PDMSstar-MA] and organic macromer [poly(ethylene glycol)diacrylate, PEG-DA] as well both aqueous and organic fabrication solvents. Having previously demonstrated its bioactivity and osteoinductivity, PDMSstar-MA is a particularly powerful component to incorporate into instructive gradient scaffolds based on PEG-DA. The following parameters were varied to produce the different gradients or gradual transitions in: (1) the wt% ratio of PDMSstar-MA to PEG-DA macromers, (2) the total wt% macromer concentration, (3) the number average molecular weight (Mn) of PEG-DA and (4) the Mn of PDMSstar-MA. Upon dividing each scaffold into four “zones” perpendicular to the gradient, we were able to demonstrate the spatial variation in morphology, bioactivity, swelling and modulus. Among these gradient scaffolds are those in which swelling and modulus are conveniently decoupled. In addition to rapid screening of cell-material interactions, these scaffolds are well-suited for regeneration of interfacial tissues (e.g. osteochondral tissues) that transition from one tissue type to another. PMID:23707502

Intensified process for the purification of an enzyme from inclusion bodies using integrated expanded bed adsorption and refolding.

PubMed

Hutchinson, Matthew H; Chase, Howard A

2006-01-01

This work describes the integration of expanded bed adsorption (EBA) and adsorptive protein refolding operations in an intensified process used to recover purified and biologically active proteins from inclusion bodies expressed in E. coli. Delta(5)-3-Ketosteroid isomerase with a C-terminal hexahistidine tag was expressed as inclusion bodies in the cytoplasm of E. coli. Chemical extraction was used to disrupt the host cells and simultaneously solubilize the inclusion bodies, after which EBA utilizing immobilized metal affinity interactions was used to purify the polyhistidine-tagged protein. Adsorptive refolding was then initiated in the column by changing the denaturant concentration in the feed stream from 8 to 0 M urea. Three strategies were tested for performing the refolding step in the EBA column: (i) the denaturant was removed using a step change in feed-buffer composition, (ii) the denaturant was gradually removed using a gradient change in feed-buffer composition, and (iii) the liquid flow direction through the column was reversed and adsorptive refolding performed in the packed bed. Buoyancy-induced mixing disrupted the operation of the expanded bed when adsorptive refolding was performed using either a step change or a rapid gradient change in feed-buffer composition. A shallow gradient reduction in denaturant concentration of the feed stream over 30 min maintained the stability of the expanded bed during adsorptive refolding. In a separate experiment, buoyancy-induced mixing was completely avoided by performing refolding in a settled bed, which achieved comparable yields to refolding in an expanded bed but required a slightly more complex process. A total of 10% of the available KSI-(His(6)) was recovered as biologically active and purified protein using the described purification and refolding process, and the yield was further increased to 19% by performing a second iteration of the on-column refolding operation. This process should be applicable for other polyhistidine tagged proteins and is likely to have the greatest benefit for proteins that tend to aggregate when refolded by dilution.

Preparative isolation and purification of three sesquiterpenoid lactones from Eupatorium lindleyanum DC. by high-speed counter-current chromatography.

PubMed

Yan, Guilong; Ji, Lilian; Luo, Yuming; Hu, Yonghong

2012-07-27

A high-speed counter-current chromatography (HSCCC) method was established for the preparative separation of three sesquiterpenoid lactones from Eupatorium lindleyanum DC. The two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (1:4:2:3, v/v/v/v) was selected. From 540 mg of the n-butanol fraction of Eupatorium lindleyanum DC., 10.8 mg of 3β-hydroxy-8β-[4'-hydroxytigloyloxy]-costunolide, 17.9 mg of eupalinolide A and 19.3 mg of eupalinolide B were obtained in a one-step HSCCC separation, with purities of 91.8%, 97.9% and 97.1%, respectively, as determined by HPLC. Their structures were further identified by ESI-MS and ¹H-NMR.

Isolation and identification of curcumin and bisacurone from rhizome extract of temu glenyeh (Curcuma soloensis. Val)

NASA Astrophysics Data System (ADS)

Vitasari, Rista A.; Wibowo, Fajar R.; Marliyana, Soerya D.; Widyo Wartono, M.

2016-02-01

Temu glenyeh (Curcuma soloensis. Val) is one of the medicinal plants that grow in Surakarta. This plant is similar with C. longa and C. Xanthoriza. Chemical constituents from an extract of the plant have never been studied. In this paper, we report the isolation of a terpenoid and curcumin from the rhizome of C. soloensis. The isolation was employed by soxhlet apparatus using acetone as solvent. The fractionation and purification of the compound from the acetone extracts were undertaken by vacuum liquid chromatography and flash chromatography. Identification of compounds used spectroscopy methods, such as FTIR, NMR (1H NMR, 13C NMR, COSY, HSQC and HMBC) and GC-MS. Isolated compounds were identified as curcumin (1) and bisacurone (2).

Purification of astaxanthin from mutant of Phaffia rhodozyma JH-82 which isolated from forests trees of Iran.

PubMed

Golkhoo, Shokufeh; Barantalab, Fatemeh; Ahmadi, Ali Reza; Hassan, Zuhair Muhammad

2007-03-01

Astaxanthin have been extracted and purified from mutant isolate of Phaffia rhodozyma JH-82. Purified astaxanthin was identified by spectrophotometric, TLC and HPLC analysis and were compared with synthetic astaxanthin. Results of TLC analysis indicated that isolate of P. rhodozyma JH-82 were able to produce nine different carotenoids and high level of carotenoids was belong to astaxanthin. Results of this study for pure astaxanthin production indicated that mutant of JH-82 of P. rhodozyma (230 microg g(-1)(dried yeast)) produced more astaxanthin than natural isolate JH-80 (140 microg g(-1)(dried yeast)). The HPLC spectrum showed retention time 11 min for both purified and synthetic astaxanthin and solvent was CDCl3.

Expression, purification, crystallization and preliminary X-ray studies of Vibrio cholerae pseudopilin EpsH

DOE Office of Scientific and Technical Information (OSTI.GOV)

Raghunathan, Kannan; Vago, Frank S.; Ball, Terry

2010-01-12

EpsH is a minor pseudopilin protein of the Vibrio cholerae type II secretion system. A truncated form of EpsH with a C-terminal noncleavable His tag was constructed and expressed in Escherichia coli, purified and crystallized by sitting-drop vapor diffusion. A complete data set was collected to 1.71 {angstrom} resolution. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 53.39, b = 71.11, c = 84.64 {angstrom}. There were two protein molecules in the asymmetric unit, which gave a Matthews coefficient V{sub M} of 2.1 {angstrom}{sup 3} Da{sup -1}, corresponding to 41.5% solvent content.

Electro-kinetic Separation of Rare Earth Elements Using a Redox-Active Ligand.

PubMed

Fang, Huayi; Cole, Bren E; Qiao, Yusen; Bogart, Justin A; Cheisson, Thibault; Manor, Brian C; Carroll, Patrick J; Schelter, Eric J

2017-10-16

Purification of rare earth elements is challenging due to their chemical similarities. All of the deployed separation methods rely on thermodynamic properties, such as distribution equilibria in solvent extraction. Rare-earth-metal separations based on kinetic differences have not been examined. Herein, we demonstrate a new approach for rare-earth-element separations by exploiting differences in the oxidation rates within a series of rare earth compounds containing the redox-active ligand [{2-(tBuN(O))C 6 H 4 CH 2 } 3 N] 3- . Using this method, a single-step separation factor up to 261 was obtained for the separation of a 50:50 yttrium-lutetium mixture. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Gas stream purifier

NASA Technical Reports Server (NTRS)

Adam, Steven J.

1994-01-01

A gas stream purifier has been developed that is capable of removing corrosive acid, base, solvent, organic, inorganic, and water vapors as well as particulates from an inert mixed gas stream using only solid scrubbing agents. This small, lightweight purifier has demonstrated the ability to remove contaminants from an inert gas stream with a greater than 99 percent removal efficiency. The Gas Stream Purifier has outstanding market and sales potential in manufacturing, laboratory and science industries, medical, automotive, or any commercial industry where pollution, contamination, or gas stream purification is a concern. The purifier was developed under NASA contract NAS9-18200 Schedule A for use in the international Space Station. A patent application for the Gas Stream Purifier is currently on file with the United States Patent and Trademark Office.

European Scientific Notes. Volume 36, Number 11.

DTIC Science & Technology

1982-11-30

and polyisocyanides appear to by S. Piccarolo (Univ. of Palermo , Italy) of the be unique in their behavior in solution, thermal expansion of...of solvent down polyethylenes was investigated by D. Curto an activity gradient coupled with time- (Univ. of Palermo , Italy). When the rheo- dependent...ambitious in Paris. Projects are projected to start in program for a number of reasons, which were 1983 in Pakistan, India, Colombia , and Saudi discussed

Electro-hydrodynamic spray synthesis and low temperature spectroscopic characterization of Perovskite thin films

NASA Astrophysics Data System (ADS)

Sarang, Som; Ishihara, Hidetaka; Tung, Vincent; Ghosh, Sayantani

Utilizing a Marangoni flow inspired electrospraying technique, we synthesize hybrid perovskite (PVSK) thin films with broad absorption spectrum and high crystallinity. The precursor solvents are electrosprayed onto an indium tin oxide (ITO) substrate, resulting in a gradient force developing between the droplet surface and the bulk due to the varying vapor pressure in the bi-solvent system. This gradient force helps the droplets propagate and merge with surrounding ones, forming a uniform thin film with excellent morphological and topological characteristics, as evident from the average power conversion efficiency (PCE) of 16%. In parallel, we use low temperature static and dynamic photoluminescence spectroscopy to probe the grain boundaries and defects in the synthesized PVSK thin films. At 120 K, the emergence of the low temperature orthorhombic phase is accompanied by reduction in lifetimes by an order of magnitude, a result attributed to charge transfer between the orthorhombic and tetragonal domains, as well as due to a crossover from free charge carrier to excitonic recombination. Our fabrication technique and optical studies help in advancement of PVSK based technology by providing unique insights into the fundamental physics of these novel materials. This research was supported by National Aeronautics and Space administration (NASA) Grant No: NNX15AQ01A.

Calixarene-Mediated Liquid-Membrane Transport of Choline Conjugates.

PubMed

Adhikari, Birendra Babu; Fujii, Ayu; Schramm, Michael P

2014-05-01

A series of supramolecular calixarenes efficiently transport distinct molecular species through a liquid membrane when attached to a receptor-complementary choline handle. Calix-[6]arene hexacarboxylic acid was highly effective at transporting different target molecules against a pH gradient. Both carboxylic- and phosphonic-acid-functionalized calix[4]arenes effect transport without requiring a pH or ion gradient. NMR binding studies, two-phase solvent extraction, and three-phase transport experiments reveal the necessary and subtle parameters to effect the transport of molecules attached to a choline "handle". On the other hand, rescorin[4]arene cavitands, which have similar guest recognition profiles, did not transport guest molecules. These developments reveal new approaches towards attempting synthetic-receptor-mediated selective small-molecule transport in vesicular and cellular systems.

High-performance liquid chromatography with nuclear magnetic resonance detection applied to organosilicon polymers. Part 2. Comparison with other methods.

PubMed

Blechta, Vratislav; Kurfürst, Milan; Sýkora, Jan; Schraml, Jan

2007-03-23

LC-NMR utilizing (1)H and (29)Si NMR spectroscopy is ideally suited for the analysis of silicones. It is shown that reversed phase gradient LC-NMR surpasses standard gel permeation chromatography (GPC) and diffusion ordered spectroscopy (DOSY) in the analysis of model hydride terminated polydimethylsiloxane. (1)H and (29)Si NMR in the stopped-flow arrangement leads to full identification of the components. Concentration gradient introduces a dependence of the (29)Si shifts on solvent composition, this dependence can be substantially reduced by a proposed method of referencing. It is shown that the ADEQUATE version of powerful but insensitive 2D INADEQUATE experiment can be used for complete line assignment.

Low-density resin impregnated ceramic article and method for making the same

NASA Technical Reports Server (NTRS)

Tran, Huy K. (Inventor); Henline, William D. (Inventor); Hsu, Ming-ta S. (Inventor); Rasky, Daniel J. (Inventor); Riccitiello, Salvatore R. (Inventor)

1997-01-01

A low-density resin impregnated ceramic article advantageously employed as a structural ceramic ablator comprising a matrix of ceramic fibers. The fibers of the ceramic matrix are coated with an organic resin film. The organic resin can be a thermoplastic resin or a cured thermosetting resin. In one embodiment, the resin is uniformly distributed within the ceramic article. In a second embodiment, the resin is distributed so as to provide a density gradient along at least one direction of the ceramic article. The resin impregnated ceramic article is prepared by providing a matrix of ceramic fibers; immersing the matrix of ceramic fibers in a solution of a solvent and an organic resin infiltrant; and removing the solvent to form a resin film on the ceramic fibers.

Isolation and purification of macrocyclic components from Penicillium fermentation broth by high-speed counter-current chromatography.

PubMed

Gao, Xiang; Zhuang, Rongqiang; Guo, Jiannan; Bao, Jian; Fang, Meijuan; Liu, Yan; Xu, Pengxiang; Zhao, Yufen

2010-02-01

In this paper, high-speed counter-current chromatography (HSCCC), assisted with ESI-MS, was first successfully applied to the preparative separation of three macrolide antibiotics, brefeldin A (12.6 mg, 99.0%), 7'-O-formylbrefeldin A (6.5 mg, 95.0%) and 7'-O-acetylbrefeldin A (5.0 mg, 92.3%) from the crude extract of the microbe Penicillium SHZK-15. Considering the chemical nature and partition coefficient (K) values of the three target compounds, a two-step HSCCC isolation protocol was developed in order to obtain products with high purity. In the two-step method, the crude ethyl acetate extract was first fractionated and resulted in two peak fractions by HSCCC using solvent system n-hexane/ethyl acetate/methanol/water (HEMWat) (3:7:5:5 v/v/v/v), then purified using solvent systems HEMWat (3:5:3:5 v/v/v/v) and HEMWat (7:3:5:5 v/v/v/v) for each fraction. The purities and structures of the isolated compounds were determined by HPLC, X-ray crystallography, ESI-MS and NMR. The results demonstrated that HSCCC is a fast and efficient technique for systematic isolation of bioactive compounds from the microbes.

Development of validated high-performance thin layer chromatography for quantification of aristolochic acid in different species of the Aristolochiaceae family.

PubMed

Agrawal, Poonam; Laddha, Kirti

2017-04-01

This study was undertaken to isolate and quantify aristolochic acid in Aristolochia indica stem and Apama siliquosa root. Aristolochic acid is an important biomarker component present in the Aristolochiaceae family. The isolation method involved simple solvent extraction, precipitation and further purification, using recrystallization. The structure of the compound was confirmed using infrared spectroscopy, mass spectrometry and nuclear magnetic resonance. A specific and rapid high-performance thin layer chromatography (HPTLC) method was developed for analysis of aristolochic acid. The method involved separation on the silica gel 60 F 254 plates using the single solvent system of n-hexane: chloroform: methanol. The method showed good linear relationship in the range 0.4-2.0 μg/spot with r 2  = 0.998. The limit of detection and limit of quantification were 62.841 ng/spot and 209.47 ng/spot, respectively. The proposed validated HPTLC method was found to be an easy to use, accurate and convenient method that could be successfully used for standardization and quality assessment of herbal material as well as formulations containing different species of the Aristolochiaceae family. Copyright © 2016. Published by Elsevier B.V.

Nanoarmored Enzymes for Organic Enzymology: Synthesis and Characterization of Poly(2-Alkyloxazoline)-Enzyme Conjugates.

PubMed

Leurs, Melanie; Tiller, Joerg C

2017-01-01

The properties of enzymes can be altered significantly by modification with polymers. Numerous different methods are known to obtain such polymer-enzyme conjugates (PECs). However, there is no universal method to render enzymes into PECs that are fully soluble in organic solvents. Here, we present a method, which achieves such high degree of modification of proteins that the majority of modified enzymes will be soluble in organic solvents. This is achieved by preparing poly(2-alkyloxazoline)s (POx) with an NH 2 end group and coupling this functional polymer via pyromellitic acid dianhydride onto the amino groups of the respective protein. The resulting PECs are capable of serving as surfactants for unmodified proteins, rendering the whole mixture organosoluble. Depending on the nature of the POx and the molecular weight and the nature of the enzyme, the PECs are soluble in chloroform or even toluene. Another advantage of this method is that the poly(2-alkyloxazoline) can be activated with the coupling agent and used for the enzyme conjugation without further purification. The POx-enzyme conjugates generated by this modification strategy show modulated catalytic activity in both, aqueous and organic, systems. © 2017 Elsevier Inc. All rights reserved.

Characterization and structure elucidation of antibacterial compound of Streptomyces sp. ECR77 isolated from east coast of India.

PubMed

Thirumurugan, D; Vijayakumar, R

2015-05-01

Forty marine actinobacteria were isolated from the sediments of east coast (Bay of Bengal) region of Tamilnadu, India. Morphologically distinct colonies were primarily tested against fish pathogenic bacteria such as Vibrio cholerae, V. parahaemolyticus, V. alginolyticus, Pseudomonas fluorescens and Aeromonas hydrophila by cross-streak plate method. The secondary metabolites produced by the highly potential strain cultured on starch casein broth were extracted separately with various solvents such as alcohol, ethyl acetate, methanol, petroleum ether and chloroform. The antibacterial assay of the bioactive compounds was tested against the fish pathogenic bacteria by well diffusion method. Of the various solvents used, the ethyl acetate extract of the isolate had good antibacterial activity. The potential strain was identified as Streptomyces labedae by phenotypic, 16S rRNA gene sequence and phylogenetic analysis. Purification of the biologically active compounds by column chromatography led to isolation of 27 fractions. The biologically active fraction was re-chromatographed on a silica gel column to obtain a single active compound, namely N-isopentyltridecanamide. The structure of the compounds was elucidated on the basis of ultra violet, Fourier transform infrared and nuclear magnetic resonance spectra.

Measurement and correlation of the solubility of gossypol acetic acid and gossypol acetic acid of optical activity in different solvents

NASA Astrophysics Data System (ADS)

Zhang, B.; Tang, H.; Liu, X. Y.; Zhai, X.; Yao, X. C.

2018-01-01

The equilibrium method was used to measure the solubility of gossypol acetic acid and gossypol acetic acid of optical activity in isopropyl alcohol, ethanol, acetic acid and ethyl acetate at temperature from 288.15 to 315.15. The Empirical equation and the Apelblat equation model were adopted to correlate the experimental data. For gossypol acetic acid, the root-mean-square deviations (RMSD) were observed in the range of 0.023-4.979 and 0.0112-0.614 for the Empirical equation and the Apelblat equation, respectively. For gossypol acetic acid of optical activity, the RMSD were observed in the range of 0.021-2.211 and 0.021-2.243 for the Empirical equation and the Apelblat equation, individually. And the maximum relative average deviation was 7.5%. Both equations offered an accurate mathematical expression of the experimental results. The calculated solubility showed a good relationship with the experimental solubility for most of solvents. This study provided valuable datas not only for optimizing the process of purification of gossypol acetic acid of optical activity in industry but also for further theoretical studies.

Liquefaction of corn stover and preparation of polyester from the liquefied polyol.

PubMed

Yu, Fei; Liu, Yuhuan; Pan, Xuejun; Lin, Xiangyang; Liu, Chengmei; Chen, Paul; Ruan, Roger

2006-01-01

This research investigated a novel process to prepare polyester from corn stover through liquefaction and crosslinking processes. First, corn stover was liquefied in organic solvents (90 wt% ethylene glycol and 10 wt% ethylene carbonate) with catalysts at moderate temperature under atmospheric pressure. The effect of liquefaction temperature, biomass content, and type of catalyst, such as H2SO4, HCl, H3PO4, and ZnCl2, was evaluated. Higher liquefaction yield was achieved in 2 wt% sulfuric acid, 1/4 (w/w) stover to liquefying reagent ratio; 160 degrees C temperature, in 2 h. The liquefied corn stover was rich in polyols, which can be directly used as feedstock for making polymers without further separation or purification. Second, polyester was made from the liquefied corn stover by crosslinking with multifunctional carboxylic acids and/or cyclic acid anhydrides. The tensile strength of polyester is about 5 MPa and the elongation is around 35%. The polyester is stable in cold water and organic solvents and readily biodegradable as indicated by 82% weight loss when buried in damp soil for 10 mo. The results indicate that this novel polyester could be used for the biodegradable garden mulch film production.

[Studies on preparative technology and quantitative determination for extracts of total saponin in roof of Panax japonicus].

PubMed

He, Yu-min; Lu, Ke-ming; Yuan, Ding; Zhang, Chang-cheng

2008-11-01

To explore the optimum extraction and purification condition of the total saponins in the root of Panax japonicus (RPJ), and establish its quality control methods. Designed L16 (4(5)) orthogonal test with the extraction rate of total saponins as index, to determine the rational extraction process, and the techniques of water-saturated n-butanol extraction and acetone precipitation were applied to purify the alcohol extract of RPJ. Total saponins were detected by spectrophotometry and its triterpenoidal sapogenin oleanolic acid detected by HPLC. The optimum conditions of total saponins from RPJ was as follows: the material was pulverized, dipped in 60% ethanol aqueous solution as extract solvent at 10 times of volume, and refluxed 3 times for 3 h each time. Extractant of water-saturated n-butanol with extraction times of 3 and precipitant of acetone with precipitation amount of 4-5 times were included in the purification process, which would obtain the quality products. The content of total saponins could reach to 83.48%, and oleanolic acid to 38.30%. The optimized preparative technology is stable, convenient and practical. The extract rate of RPJ was high and steady with this technology, which provided new evidence for industrializing production of the plant and developing new drug.

Environmentally Friendly Procedure Based on Supercritical Fluid Chromatography and Tandem Mass Spectrometry Molecular Networking for the Discovery of Potent Antiviral Compounds from Euphorbia semiperfoliata.

PubMed

Nothias, Louis-Félix; Boutet-Mercey, Stéphanie; Cachet, Xavier; De La Torre, Erick; Laboureur, Laurent; Gallard, Jean-François; Retailleau, Pascal; Brunelle, Alain; Dorrestein, Pieter C; Costa, Jean; Bedoya, Luis M; Roussi, Fanny; Leyssen, Pieter; Alcami, José; Paolini, Julien; Litaudon, Marc; Touboul, David

2017-10-27

A supercritical fluid chromatography-based targeted purification procedure using tandem mass spectrometry and molecular networking was developed to analyze, annotate, and isolate secondary metabolites from complex plant extract mixture. This approach was applied for the targeted isolation of new antiviral diterpene esters from Euphorbia semiperfoliata whole plant extract. The analysis of bioactive fractions revealed that unknown diterpene esters, including jatrophane esters and phorbol esters, were present in the samples. The purification procedure using semipreparative supercritical fluid chromatography led to the isolation and identification of two new jatrophane esters (13 and 14) and one known (15) and three new 4-deoxyphorbol esters (16-18). The structure and absolute configuration of compound 16 were confirmed by X-ray crystallography. This compound was found to display antiviral activity against Chikungunya virus (EC 50 = 0.45 μM), while compound 15 proved to be a potent and selective inhibitor of HIV-1 replication in a recombinant virus assay (EC 50 = 13 nM). This study showed that a supercritical fluid chromatography-based protocol and molecular networking can facilitate and accelerate the discovery of bioactive small molecules by targeting molecules of interest, while minimizing the use of toxic solvents.

Antitubercular constituents from Premna odorata Blanco.

PubMed

Lirio, Stephen B; Macabeo, Allan Patrick G; Paragas, Erickson M; Knorn, Matthias; Kohls, Paul; Franzblau, Scott G; Wang, Yuehong; Aguinaldo, Ma Alicia M

2014-06-11

Premna odorata Blanco (Lamiaceae) is a medicinal plant traditionally used in Albay Province, in southeastern Luzon, Philippines to treat tuberculosis. This study aimed to determine the antitubercular property of the crude extract and sub-extracts of the leaves, and to isolate the bioactive principles from the active fractions. Through extraction, solvent polarity-based fractionation and silica gel chromatography purification of the DCM sub-extract, compound mixtures from the bioactive fractions were isolated and screened for their in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv using the colorimetric Microplate Alamar Blue assay (MABA). The crude methanolic extract and sub-extracts showed poor inhibitory activity against Mycobacterium tuberculosis H37Rv (MIC≥128µg/mL). However, increased inhibitory potency was observed for fractions eluted from the DCM sub-extract (MIC=54 to 120µg/mL). Further purification of the most active fraction (MIC=54µg/mL) led to the isolation of a 1-heneicosyl formate (1), 4:1 mixture of β-sitosterol (2), stigmasterol (3) and diosmetin (4), which were identified through GC-MS analysis (with dereplication) and NMR experiments. The MIC of compound 1 was 8µg/mL. The results of this study provide scientific basis for the traditional use of Premna odorata as treatment for tuberculosis. Copyright © 2014. Published by Elsevier Ireland Ltd.

[18F]FEPPA a TSPO Radioligand: Optimized Radiosynthesis and Evaluation as a PET Radiotracer for Brain Inflammation in a Peripheral LPS-Injected Mouse Model.

PubMed

Vignal, Nicolas; Cisternino, Salvatore; Rizzo-Padoin, Nathalie; San, Carine; Hontonnou, Fortune; Gelé, Thibaut; Declèves, Xavier; Sarda-Mantel, Laure; Hosten, Benoît

2018-06-07

[ 18 F]FEPPA is a specific ligand for the translocator protein of 18 kDa (TSPO) used as a positron emission tomography (PET) biomarker for glial activation and neuroinflammation. [ 18 F]FEPPA radiosynthesis was optimized to assess in a mouse model the cerebral inflammation induced by an intraperitoneal injection of Salmonella enterica serovar Typhimurium lipopolysaccharides (LPS; 5 mg/kg) 24 h before PET imaging. [ 18 F]FEPPA was synthesized by nucleophilic substitution (90 °C, 10 min) with tosylated precursor, followed by improved semi-preparative HPLC purification (retention time 14 min). [ 18 F]FEPPA radiosynthesis were carried out in 55 min (from EOB). The non-decay corrected radiochemical yield were 34 ± 2% ( n = 17), and the radiochemical purity greater than 99%, with a molar activity of 198 ± 125 GBq/µmol at the end of synthesis. Western blot analysis demonstrated a 2.2-fold increase in TSPO brain expression in the LPS treated mice compared to controls. This was consistent with the significant increase of [ 18 F]FEPPA brain total volume of distribution ( V T ) estimated with pharmacokinetic modelling. In conclusion, [ 18 F]FEPPA radiosynthesis was implemented with high yields. The new purification/formulation with only class 3 solvents is more suitable for in vivo studies.

Separation and purification of four flavonol diglucosides from the flower of Meconopsis integrifolia by high-speed counter-current chromatography.

PubMed

Huang, Yanfei; Han, Yatao; Chen, Keli; Huang, Bisheng; Liu, Yuan

2015-12-01

Flavonoids are the main components of Meconopsis integrifolia (Maxim.) Franch, which is a traditional Tibetan medicine. However, traditional chromatography separation requires a large quantity of raw M. integrifolia and is very time consuming. Herein, we applied high-speed counter-current chromatography in the separation and purification of flavonoids from the ethanol extracts of M. integrifolia flower. Ethyl acetate/n-butanol/water (2:3:5, v/v/v) was selected as the optimum solvent system to purify the four components, namely quercetin-3-O-β-d-glucopyrannosy-(1→6)-β-d-glucopyranoside (compound 1, 60 mg), quercetin 3-O-[2'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 2, 40 mg), quercetin 3-O-[3'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 3, 11 mg), and quercetin 3-O-[6'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 4, 16 mg). Among the four compounds, 3 and 4 were new acetylated flavonol diglucosides. After the high-speed counter-current chromatography separation, the purities of the four flavonol diglucosides were 98, 95, 90, and 92%, respectively. The structures of these compounds were identified by mass spectrometry and NMR spectroscopy. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Preparative isolation and purification of phlorotannins from Ecklonia cava using centrifugal partition chromatography by one-step.

PubMed

Lee, Ji-Hyeok; Ko, Ju-Young; Oh, Jae-Young; Kim, Chul-Young; Lee, Hee-Ju; Kim, Jaeil; Jeon, You-Jin

2014-09-01

Various bioactive phlorotannins of Ecklonia cava (e.g., dieckol, eckol, 6,6-bieckol, phloroglucinol, phloroeckol, and phlorofucofuroeckol-A) are reported. However, their isolation and purification are not easy. Centrifugal partition chromatography (CPC) can be used to efficiently purify the various bioactive-compounds efficiently from E. cava. Phlorotannins are successfully isolated from the ethyl acetate (EtOAc) fraction of E. cava by CPC with a two-phase solvent system comprising n-hexane:EtOAc:methanol:water (2:7:3:7, v/v) solution. The dieckol (fraction I, 40.2mg), phlorofucofuroeckol-A (fraction III, 31.1mg), and fraction II (34.1mg) with 2,7-phloroglucinol-6,6-bieckol and pyrogallol-phloroglucinol-6,6-bieckol are isolated from the crude extract (500 mg) by a one-step CPC system. The purities of the isolated dieckol and phlorofucofuroeckol-A are ⩾90% according to high performance liquid chromatography (HPLC) and electrospray ionization multi stage tandem mass spectrometry analyses. The purified 2,7-phloroglucinol-6,6-bieckol and pyrogallol-phloroglucinol-6,6-bieckol are collected from fraction II by recycle-HPLC. Thus, the CPC system is useful for easy and simple isolation of phlorotannins from E. cava. Copyright © 2014 Elsevier Ltd. All rights reserved.

5-O-caffeoylshikimic acid from Solanum somalense leaves: advantage of centrifugal partition chromatography over conventional column chromatography.

PubMed

Chideh, Saïda; Pilard, Serge; Attoumbré, Jacques; Saguez, Robert; Hassan-Abdallah, Alshaimaa; Cailleu, Dominique; Wadouachi, Anne; Baltora-Rosset, Sylvie

2014-09-01

Solanum somalense leaves, used in Djibouti for their medicinal properties, were extracted by MeOH. Because of the high polyphenol and flavonoid contents of the extract, respectively, determined at 80.80 ± 2.13 mg gallic acid equivalent/g dry weight and 24.4 ± 1.01 mg quercetin equivalent/g dry weight, the isolation and purification of the main polyphenols were carried out by silica gel column chromatography and centrifugal partition chromatography. Column chromatography led to 11 enriched fractions requiring further purification, while centrifugal partition chromatography allowed the easy recovery of the main compound of the extract. In a solvent system composed of CHCl3/MeOH/H2O (9.5:10:5), 21.8 mg of this compound at 97% purity was obtained leading to a yield of 2.63%. Its structure was established as 5-O-caffeoylshikimic acid by mass spectrometry and NMR spectroscopy. This work shows that S. somalense leaves contain very high level of 5-O-caffeoylshikimic acid (0.74% dry weight), making it a potential source of production of this secondary metabolite that is not commonly found in nature but could be partly responsible of the medicinal properties of S. somalense leaves. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Semi-Preparative Isolation and Purification of Three Tauro-Conjugated Cholic Acids from Pulvis Fellis Suis by HSCCC Coupled with ELSD Detection.

PubMed

He, Jiao; Zhang, Yongmin; Ito, Yoichiro; Sun, Wenji

2011-01-01

Coupled with evaporative light scattering detection, a high-speed counter-current chromatography (HSCCC) method was applied to the separation and purification of three tauro-conjugated cholic acids of taurochenodeoxycholic acid (TCDCA), taurohyodeoxycholic acid (THDCA) and taurohyocholic acid (THCA) from Pulvis Fellis Suis (Pig gallbladder bile) for the first time. The two-phase solvent system composed of chloroform-methanol-water-acetic acid (4:4:2:0.3, v/v/v/v) was selected for the one-step separation where the lower phase was used as the mobile phase in the head to tail elution mode. The revolution speed of the separation column, flow rate of the mobile phase and separation temperature were 800 rpm, 1.5 ml/min and 25°C respectively. From 100 mg of the crude extract, 10.2 mg of TCDCA, 11.8 mg of THDCA and 5.3 mg of THCA were obtained with the purity of 94.6%, 96.5% and 95.4%, respectively. in one step separation The HSCCC fractions were analyzed by high-performance liquid chromatography (HPLC) and the structures of the three tauro-conjugated cholic acids were identified by ESI-MS, (1)H NMR and (13)C NMR.

Metal- and additive-free photoinduced borylation of haloarenes.

PubMed

Mfuh, Adelphe M; Schneider, Brett D; Cruces, Westley; Larionov, Oleg V

2017-03-01

Boronic acids and esters have critical roles in the areas of synthetic organic chemistry, molecular sensors, materials science, drug discovery, and catalysis. Many of the current applications of boronic acids and esters require materials with very low levels of transition metal contamination. Most of the current methods for the synthesis of boronic acids, however, require transition metal catalysts and ligands that must be removed via additional purification procedures. This protocol describes a simple, metal- and additive-free method of conversion of haloarenes directly to boronic acids and esters. This photoinduced borylation protocol does not require expensive and toxic metal catalysts or ligands, and it produces innocuous and easy-to-remove by-products. Furthermore, the reaction can be carried out on multigram scales in common-grade solvents without the need for reaction mixtures to be deoxygenated. The setup and purification steps are typically accomplished within 1-3 h. The reactions can be run overnight, and the protocol can be completed within 13-16 h. Two representative procedures that are described in this protocol provide details for preparation of a boronic acid (3-cyanopheylboronic acid) and a boronic ester (1,4-benzenediboronic acid bis(pinacol)ester). We also discuss additional details of the method that will be helpful in the application of the protocol to other haloarene substrates.

Negatively charged polysulfone membranes with hydrophilicity and antifouling properties based on in situ cross-linked polymerization.

PubMed

Zhu, Lijing; Song, Haiming; Zhang, Dawei; Wang, Gang; Zeng, Zhixiang; Xue, Qunji

2017-07-15

Polysulfone (PSf) membrane has been widely used in water separation and purification, although, membrane fouling is still a serious problem limiting its potential. We aim to improve the antifouling of PSf membranes via a very simple and efficient method. In this work, antifouling PSf membranes were fabricated via in situ cross-linked polymerization coupled with non-solvent induced phase separation. In brief, acrylic acid (AA) and vinyltriethoxysilane (VTEOS) were copolymerized in PSf solution, then directly casted into membranes without purification. With the increase of monomers concentration, the morphology of the as-cast membranes changed from a finger-like morphology to a fully sponge-like structure due to the increased viscosity and decreased precipitation rate of the polymer solutions. Meanwhile, the hydrophilicity and electronegativity of modified membranes were highly improved leading to inhibited protein adsorption and improved antifouling property. Furthermore, in order to further find out the different roles player by AA and VTESO, the modified membrane without VTEOS was prepared and characterized. The results indicated that AA is more effective in the membrane hydrophilicity improvement, VTEOS is more crucial to improve membrane stability. This work provides valuable guidance for fabricating PSf membranes with hydrophilicity and antifouling property via in situ cross-linked polymerization. Copyright © 2017 Elsevier Inc. All rights reserved.

Chromatographic matrix based on hydrogel-coated reticulated polyurethane foams, prepared by gamma irradiation

NASA Astrophysics Data System (ADS)

Sánchez, Mirna L.; Giménez, Claudia Y.; Delgado, Juan F.; Martínez, Leandro J.; Grasselli, Mariano

2017-12-01

Novel chromatographic materials for protein purification with high adsorption capacity and fouling resistance are highly demanded to improve downstream processes. Here, we describe a novel adsorptive material based on reticulated polyurethane foam (rPUF) coated with a functional hydrogel layer. rPUF provides physical rigidity through its macroscopic structure, whereas the hydrogel layer provides capacity to adsorb proteins by specific interactions. The hydrogel coating process was performed by the dip-coating method, using a polyvinyl alcohol (PVA) solution. The PVA hydrogel was linked to the rPUF material by using a radiation-induced crosslinking process in aqueous ethanol solution. The ethanol in the solvent mixture allowed a balance between PVA swelling and PVA dissolution during the irradiation step. The resulting material showed higher thermal stability than the non-irradiated one. In addition, a simultaneous radiation-induced grafting polymerization (SRIGP) was done by simple addition of glycidyl methacrylate monomer into the irradiation solution. In a further step, sulfonic ligands were included specifically in the hydrogel layer, which contained around 200% of PVA respect to the original rPUF. Materials were characterized by FT-IR, thermogravimetric analysis, SEM microscopy and EDX analysis. The cation-exchange rPUF material was functionally characterized by the Langmuir isotherm and a dynamic adsorption experiment to analyze the chromatographic properties for protein purification processes.

Extraction, purification and characterization of a protease from Micrococcus sp. VKMM 037.

PubMed

Manikandan, Muthu; Kannan, Vijayaraghavan; Pasić, Lejla

2011-10-01

The haloalkaliphilic bacterium Micrococcus sp. VKMM 037, isolated from an effluent of the caustic soda industry, was found to produce a protease. Maximal proteolytic activity was observed in cell culture grown at 40 degrees C using 2% (w/v) glycerol, 2% (w/v) beef extract and 2% (w/v) peptone as nutrients in medium also containing 0.85 M NaCl with a pH of 10.0. An efficient purification procedure combining ammonium sulphate precipitation and Q-Sepharose ion-exchange chromatography was developed. The purified 41 kDa protease was stable in a temperature range between 20 degrees C and 60 degrees C. The protease remained active over a wide range of pH values (4.0-12.0) and NaCl concentrations (0-3.42 M) with an optimum at pH 10.0 and 0.85 M NaCl, respectively. Furthermore, the enzyme remained stable or was only marginally inhibited in the presence of various organic solvents, surfactants and reducing agents. The purified protease of Micrococcus sp. VKMM 037 efficiently removed blood stains within 40 minutes of treatment. Given the biochemical characteristics determined, this novel protease could be exploited as an additive in the detergent industry and also for the synthesis of biomolecules and the degradation of protein.

Microwave-Assisted Extraction and Purification of Arctiin and Arctigenin from Fructus Arctii by High-Speed Countercurrent Chromatography.

PubMed

Lü, Haitao; Sun, Zhaoyun; Shan, Hu; Song, Jiying

2016-03-01

An efficient method for the rapid extraction, separation and purification of bioactive lignans, arctiin and arctigenin, from Fructus arctii by microwave-assisted extraction coupled with high-speed countercurrent chromatography was developed. The optimal extraction conditions of arctiin and arctigenin were evaluated by orthogonal array. Arctigenin could be converted from arctiin by hydrochloric acid hydrolysis. The separations were performed at a preparative scale with two-phase solvents composed of ethyl acetate-ethanol-water (5 : 1 : 5, v/v/v) for arctiin, and n-hexane-ethyl acetate-ethanol-water (4 : 4 : 3 : 4, v/v/v/v) for arctigenin. From 500 mg of crude extract sample, 122.3 mg of arctiin and 45.7 mg of arctigenin were obtained with the purity of 98.46 and 96.57%, and the recovery of 94.3 and 81.6%, respectively. Their structures were determined by comparison with the high-performance liquid chromatography retention time of standard substance as well as UV, FT-IR, electrospray ion source (ESI)-MS, (1)H-NMR and (13)C-NMR spectrum. According to the antioxidant activity assay, arctigenin had stronger 1,1-diphenyl-2-picrylhydrazyl free radicals scavenging activity. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Identification and determination of the saikosaponins in Radix bupleuri by accelerated solvent extraction combined with rapid-resolution LC-MS.

PubMed

Yang, Yun-Yun; Tang, You-Zhi; Fan, Chun-Lin; Luo, Hui-Tai; Guo, Peng-Ran; Chen, Jian-Xin

2010-07-01

A method based on accelerated solvent extraction combined with rapid-resolution LC-MS for efficient extraction, rapid separation, online identification and accurate determination of the saikosaponins (SSs) in Radix bupleuri (RB) was developed. The RB samples were extracted by accelerated solvent extraction using 70% aqueous ethanol v/v as solvent, at a temperature of 120 degrees C and pressure of 100 bar, with 10 min of static extraction time and three extraction cycles. Rapid-resolution LC separation was performed by using a C(18) column at gradient elution of water (containing 0.5% formic acid) and acetonitrile, and the major constituents were well separated within 20 min. A TOF-MS and an IT-MS were used for online identification of the major constituents, and 27 SSs were identified or tentatively identified. Five major bioactive SSs (SSa, SSc, SSd, 6''-O-acetyl-SSa and 6''-O-acetyl-SSd) with obvious peak areas and good resolution were chosen as benchmark substances, and a triple quadrupole MS operating in multiple-reaction monitoring mode was used for their quantitative analysis. A total of 16 RB samples from different regions of China were analyzed. The results indicated that the method was rapid, efficient, accurate and suitable for use in the quality control of RB.

Analysis of flavonoids from lotus (Nelumbo nucifera) leaves using high performance liquid chromatography/photodiode array detector tandem electrospray ionization mass spectrometry and an extraction method optimized by orthogonal design.

PubMed

Chen, Sha; Wu, Ben-Hong; Fang, Jin-Bao; Liu, Yan-Ling; Zhang, Hao-Hao; Fang, Lin-Chuan; Guan, Le; Li, Shao-Hua

2012-03-02

The extraction protocol of flavonoids from lotus (Nelumbo nucifera) leaves was optimized through an orthogonal design. The solvent was the most important factor comparing solvent, solvent:tissue ratio, extraction time, and temperature. The highest yield of flavonoids was achieved with 70% methanol-water and a solvent:tissue ratio of 30:1 at 4 °C for 36 h. The optimized analytical method for HPLC was a multi-step gradient elution using 0.5% formic acid (A) and CH₃CN containing 0.1% formic acid (B), at a flow rate of 0.6 mL/min. Using this optimized method, thirteen flavonoids were simultaneously separated and identified by high performance liquid chromatography coupled with photodiode array detection/electrospray ionization mass spectrometry (HPLC/DAD/ESI-MS(n)). Five of the bioactive compounds are reported in lotus leaves for the first time. The flavonoid content of the leaves of three representative cultivars was assessed under the optimized extraction and HPLC analytical conditions, and the seed-producing cultivar 'Baijianlian' had the highest flavonoid content compared with rhizome-producing 'Zhimahuoulian' and wild floral cultivar 'Honglian'. Copyright © 2012 Elsevier B.V. All rights reserved.

Phase-field approach to implicit solvation of biomolecules with Coulomb-field approximation

NASA Astrophysics Data System (ADS)

Zhao, Yanxiang; Kwan, Yuen-Yick; Che, Jianwei; Li, Bo; McCammon, J. Andrew

2013-07-01

A phase-field variational implicit-solvent approach is developed for the solvation of charged molecules. The starting point of such an approach is the representation of a solute-solvent interface by a phase field that takes one value in the solute region and another in the solvent region, with a smooth transition from one to the other on a small transition layer. The minimization of an effective free-energy functional of all possible phase fields determines the equilibrium conformations and free energies of an underlying molecular system. All the surface energy, the solute-solvent van der Waals interaction, and the electrostatic interaction are coupled together self-consistently through a phase field. The surface energy results from the minimization of a double-well potential and the gradient of a field. The electrostatic interaction is described by the Coulomb-field approximation. Accurate and efficient methods are designed and implemented to numerically relax an underlying charged molecular system. Applications to single ions, a two-plate system, and a two-domain protein reveal that the new theory and methods can capture capillary evaporation in hydrophobic confinement and corresponding multiple equilibrium states as found in molecular dynamics simulations. Comparisons of the phase-field and the original sharp-interface variational approaches are discussed.

Analysis for residual host cell proteins and DNA in process streams of a recombinant protein product expressed in Escherichia coli cells.

PubMed

Rathore, Anurag Singh; Sobacke, S E; Kocot, T J; Morgan, D R; Dufield, R L; Mozier, N M

2003-08-21

Analyses of crude samples from biotechnology processes are often required in order to demonstrate that residual host cell impurities are reduced or eliminated during purification. In later stages of development, as the processes are further developed and finalized, there is a tremendous volume of testing required to confirm the absence of residual host cell proteins (HCP) and DNA. Analytical tests for these components are very challenging since (1). they may be present at levels that span a million-fold range, requiring substantial dilutions; (2). are not a single component, often existing as fragments and a variety of structures; (3). require high sensitivity for final steps in process; and (4). are present in very complex matrices including other impurities, the product, buffers, salts and solvents. Due to the complex matrices and the variety of potential analytes, the methods of analysis are not truly quantitative for all species. Although these limitations are well known, the assays are still very much in demand since they are required for approval of new products. Methods for final products, described elsewhere, focus on approaches to achieve regulatory requirements. The study described herein will describe the technical rationale for measuring the clearance of HCP and DNA in the entire bioprocessing to purification from an Escherichia coli-derived expression system. Three analytical assays, namely, reversed-phase high-performance liquid chromatography (RP-HPLC), enzyme-linked immunosorbent assay (ELISA), and Threshold Total DNA Assay, were utilized to quantify the protein product, HCP and DNA, respectively. Product quantification is often required for yield estimation and is useful since DNA and HCP results are best expressed as a ratio to product for calculation of relative purification factors. The recombinant E. coli were grown to express the protein of interest as insoluble inclusion bodies (IB) within the cells. The IB were isolated by repeated homogenization and centrifugation and the inclusion body slurry (IBS) was solubilized with urea. After refolding the product, the solution was loaded on several commonly used ion exchangers (CM, SP, DEAE, and Q). Product was eluted in a salt gradient mode and fractions were collected and analyzed for product, HCP and DNA. The IBS used for this study contained about 15 mg/ml product, 38 mg/ml HCP and 1.1 mg/ml DNA. Thus, the relative amounts of HCP and DNA in the IBS was excessive, and about 10(3) times greater than typical (because the cells and IB were not processed with the normal number of washing steps during isolation). This was of interest since similar samples may be encountered when working with non-inclusion body systems, such as periplasmic expressions, or in cases where the upstream unit operations under-perform in IB cleaning. The study described herein describes the development of three robust methods that provide the essential process data needed. These findings are of general interest to other projects since applications of similar analytical technology may be used as a tool to develop processes, evaluate clearance of impurities, and produce a suitable product.

Control of Evaporation Behavior of an Inkjet-Printed Dielectric Layer Using a Mixed-Solvent System

NASA Astrophysics Data System (ADS)

Yang, Hak Soon; Kang, Byung Ju; Oh, Je Hoon

2016-01-01

In this study, the evaporation behavior and the resulting morphology of inkjet-printed dielectric layers were controlled using a mixed-solvent system to fabricate uniform poly-4-vinylphenol (PVP) dielectric layers without any pinholes. The mixed-solvent system consisted of two different organic solvents: 1-hexanol and ethanol. The effects of inkjet-printing variables such as overlap condition, substrate temperature, and different printing sequences (continuous and interlacing printing methods) on the inkjet-printed dielectric layer were also investigated. Increasing volume fraction of ethanol (VFE) is likely to reduce the evaporation rate gradient and the drying time of the inkjet-printed dielectric layer; this diminishes the coffee stain effect and thereby improves the uniformity of the inkjet-printed dielectric layer. However, the coffee stain effect becomes more severe with an increase in the substrate temperature due to the enhanced outward convective flow. The overlap condition has little effect on the evaporation behavior of the printed dielectric layer. In addition, the interlacing printing method results in either a stronger coffee stain effect or wavy structures of the dielectric layers depending on the VFE of the PVP solution. All-inkjet-printed capacitors without electrical short circuiting can be successfully fabricated using the optimized PVP solution (VFE = 0.6); this indicates that the mixed-solvent system is expected to play an important role in the fabrication of high-quality inkjet-printed dielectric layers in various printed electronics applications.

Method and system for optical figuring by imagewise heating of a solvent

DOEpatents

Rushford, Michael C.

2005-08-30

A method and system of imagewise etching the surface of a substrate, such as thin glass, in a parallel process. The substrate surface is placed in contact with an etchant solution which increases in etch rate with temperature. A local thermal gradient is then generated in each of a plurality of selected local regions of a boundary layer of the etchant solution to imagewise etch the substrate surface in a parallel process. In one embodiment, the local thermal gradient is a local heating gradient produced at selected addresses chosen from an indexed array of addresses. The activation of each of the selected addresses is independently controlled by a computer processor so as to imagewise etch the substrate surface at region-specific etch rates. Moreover, etching progress is preferably concurrently monitored in real time over the entire surface area by an interferometer so as to deterministically control the computer processor to image-wise figure the substrate surface where needed.

Solvent-Induced Crystallization in Poly(Ethylene Terephthalate) during Mass Transport

NASA Astrophysics Data System (ADS)

Ouyang, Hao

2001-03-01

The solvent transport in poly(ethylene terephthalate) (PET) and related phase transformation were investigated. The data of mass sorption were analyzed according to Harmon¡¦s model for Case I (Fickian), Case II (swelling) and anomalous transport. This transport process in PET is accompanied by the induced crystallization of the original amorphous state. The transformation was studied by wide angle x-ray scattering (WAXS), small angle x-ray scattering (SAXS), Differential Scanning Calorimeter (DSC), density gradient column, and Fourier Transform Infra-Red (FTIR). During this process, the matrix is under a compressive strain that causes different kinetic path of crystallization as compared to that by thermal annealing. This state of strain will assist the development of the solvent-induced crystallization. It also can be explained in terms of the principle of Le Chatelier if the local equilibrium is assumed. The model regarding the crystallization was proposed in terms of the study of long period L, the crystal thickness lc and the thickness of amorphous layer la, obtained from the linear correlation function and interface distribution function.

Comparative studies on performance of CCC and preparative RP-HPLC in separation and purification of steroid saponins from Dioscorea zingiberensis C.H.Wright

PubMed Central

Zhang, Xinxin; Liang, Jinru; Zhang, Yongmin; Liu, Jianli; Sun, Wenji; Ito, Yoichiro

2015-01-01

Steroid saponins from Dioscorea zingiberensis C.H.Wright were separated for the first time using two chromatographic methods for comparison: counter-current chromatography (CCC) coupled with evaporative light scattering detector (ELSD) and preparative reversed phase high-performance liquid chromatography (RP-HPLC) with an ultraviolet detector. Ethyl acetate-n-butanol-methanol-water (4:1:2:4, v/v) was chosen as the two-phase solvent system for CCC, while the acetonitrile-water (25:75 for the first step and15:85 for the second step, v/v) was used as the mobile phase in the preparative RP-HPLC. The following five steroid saponins were purified by theses two chromatographic methods, in one-step operation by CCC and by two-step operation in preparative RP-HPLC: 1) 26-O-β-D- glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside (compound A), 2) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 4) 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside (compound B), 3) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside (compound C), 4) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-{α-L-rhamnopyranosyl-(1→4)-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→2)]}-β-D-glucopyranoside (compound D) and 5) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-[β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosy-(1→2)]-β-D-glucopyranoside (compound E). The purities of these five steroid saponins separated by both methods were over 95%, and structural identification of these compounds was performed by ESI-MS, and 13C NMR. Comparison of these two established approaches revealed that CCC required a longer separation time but with less solvent consumption, whereas preparative RP-HPLC gave a shorter separation time but with higher solvent consumption. These results demonstrated that either of these two methods of different separation mechanism is feasible, economical and efficient for rapid preparative isolation and purification of steroid saponins from Dioscorea zingiberensis C.H.Wright. PMID:26726306

Comparative studies on performance of CCC and preparative RP-HPLC in separation and purification of steroid saponins from Dioscorea zingiberensis C.H.Wright.

PubMed

Zhang, Xinxin; Liang, Jinru; Zhang, Yongmin; Liu, Jianli; Sun, Wenji; Ito, Yoichiro

2015-03-01

Steroid saponins from Dioscorea zingiberensis C.H.Wright were separated for the first time using two chromatographic methods for comparison: counter-current chromatography (CCC) coupled with evaporative light scattering detector (ELSD) and preparative reversed phase high-performance liquid chromatography (RP-HPLC) with an ultraviolet detector. Ethyl acetate-n-butanol-methanol-water (4:1:2:4, v/v) was chosen as the two-phase solvent system for CCC, while the acetonitrile-water (25:75 for the first step and15:85 for the second step, v/v) was used as the mobile phase in the preparative RP-HPLC. The following five steroid saponins were purified by theses two chromatographic methods, in one-step operation by CCC and by two-step operation in preparative RP-HPLC: 1) 26-O-β-D- glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside ( compound A ), 2) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 4) 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside ( compound B ), 3) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside ( compound C ), 4) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-{α-L-rhamnopyranosyl-(1→4)-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→2)]}-β-D-glucopyranoside ( compound D ) and 5) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-[β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosy-(1→2)]-β-D-glucopyranoside ( compound E ). The purities of these five steroid saponins separated by both methods were over 95%, and structural identification of these compounds was performed by ESI-MS, and 13 C NMR. Comparison of these two established approaches revealed that CCC required a longer separation time but with less solvent consumption, whereas preparative RP-HPLC gave a shorter separation time but with higher solvent consumption. These results demonstrated that either of these two methods of different separation mechanism is feasible, economical and efficient for rapid preparative isolation and purification of steroid saponins from Dioscorea zingiberensis C.H.Wright.