Sample records for chromatography

  1. Chromatography. (United States)

    Brantley, L. Reed, Sr.; Demanche, Edna L.; Klemm, E. Barbara; Kyselka, Will; Phillips, Edwin A.; Pottenger, Francis M.; Yamamoto, Karen N.; Young, Donald B.

    This booklet presents some activities on chromatography. Directions for preparing leaf pigment extracts using alcohol are given, and paper chromatography and thin-layer chromatography are described as modifications of the basic principles of chromatography. (KHR)

  2. Applying Chromatography. (United States)

    Klein, Jessie W.; Patev, Paul


    Presents three experiments to introduce students to different kinds of chromatography: (1) paper chromatography; (2) gel filtration chromatography; and (3) reverse-phase liquid chromatography. Written in the form of a laboratory manual, explanations of each of the techniques, materials needed, procedures, and a glossary are included. (PVD)

  3. Neuere Chromatographie (United States)

    Hostettmann, K.


    Besides high-performance liquid chromatography (HPLC) which is now a well-established and currently used technique, several emerging methods for the isolation and separation of natural products are receiving considerable attention. Centrifugal thin-layer chromatography is a very rapid technique, but limited in resolution. Of special interest are the recently developed support-free liquid-liquid chromatography methods such as droplet counter-current chromatography (DCCC) and rotation locular counter-current chromatography (RLCC). This latter method was applied to the separation of the enantiomers of (±)-norephedrine.

  4. Gas chromatography (United States)

    Guiochon, Georges; Guillemin, Claude L.


    Gas chromatography is a powerful separation technique for gas and vapor mixtures. Combining separation and on-line detection permits accurate quantitative analysis of complex mixtures, including traces of compounds down to parts per trillions in some particular cases. The importance of gas chromatography in quality control and process control in the chemical and drug industry, in environmental pollution investigations and in clinical analysis is critical. The principles of the technique are discussed, the main components of a gas chromatograph are described and some idea of the importance of the applications is given.

  5. Green chromatography. (United States)

    Płotka, Justyna; Tobiszewski, Marek; Sulej, Anna Maria; Kupska, Magdalena; Górecki, Tadeusz; Namieśnik, Jacek


    Analysis of organic compounds in samples characterized by different composition of the matrix is very important in many areas. A vast majority of organic compound determinations are performed using gas or liquid chromatographic methods. It is thus very important that these methods have negligible environmental impact. Chromatographic techniques have the potential to be greener at all steps of the analysis, from sample collection and preparation to separation and final determination. The paper summarizes the approaches used to accomplish the goals of green chromatography. While complete elimination of sample preparation would be an ideal approach, it is not always practical. Solventless extraction techniques offer a very good alternative. Where solvents must be used, the focus should be on the minimization of their consumption. The approaches used to make chromatographic separations greener differ depending on the type of chromatography. In gas chromatography it is advisable to move away from using helium as the carrier gas because it is a non-renewable resource. GC separations using low thermal mass technology can be greener because of energy savings offered by this technology. In liquid chromatography the focus should be on the reduction of solvent consumption and replacement of toxic and environmentally hazardous solvents with more benign alternatives. Multidimensional separation techniques have the potential to make the analysis greener in both GC and LC. The environmental impact of the method is often determined by the location of the instrument with respect to the sample collection point.

  6. Ion Chromatography. (United States)

    Mulik, James D.; Sawicki, Eugene


    Accurate for the analysis of ions in solution, this form of analysis enables the analyst to directly assay many compounds that previously were difficult or impossible to analyze. The method is a combination of the methodologies of ion exchange, liquid chromatography, and conductimetric determination with eluant suppression. (Author/RE)

  7. Gas Chromatography (United States)

    Qian, Michael C.

    Gas chromatography (GC) has many applications in the analysis of food products. GC has been used for the determination of fatty acids, triglycerides, cholesterol, gases, water, alcohols, pesticides, flavor compounds, and many more. While GC has been used for other food components such as sugars, oligosaccharides, amino acids, peptides, and vitamins, these substances are more suited to analysis by high performance liquid chromatography. GC is ideally suited to the analysis of volatile substances that are thermally stable. Substances such as pesticides and flavor compounds that meet these criteria can be isolated from a food and directly injected into the GC. For compounds that are thermally unstable, too low in volatility, or yield poor chromatographic separation due to polarity, a derivatization step must be done before GC analysis. The two parts of the experiment described here include the analysis of alcohols that requires no derivatization step, and the analysis of fatty acids which requires derivatization. The experiments specify the use of capillary columns, but the first experiment includes conditions for a packed column.

  8. Instrumentation: Ion Chromatography. (United States)

    Fritz, James S.


    Discusses the importance of ion chromatography in separating and measuring anions. The principles of ion exchange are presented, along with some applications of ion chromatography in industry. Ion chromatography systems are described, as well as ion pair and ion exclusion chromatography, column packings, detectors, and programming. (TW)

  9. Basic Principles of Chromatography (United States)

    Ismail, Baraem; Nielsen, S. Suzanne

    Chromatography has a great impact on all areas of analysis and, therefore, on the progress of science in general. Chromatography differs from other methods of separation in that a wide variety of materials, equipment, and techniques can be used. [Readers are referred to references (1-19) for general and specific information on chromatography.]. This chapter will focus on the principles of chromatography, mainly liquid chromatography (LC). Detailed principles and applications of gas chromatography (GC) will be discussed in Chap. 29. In view of its widespread use and applications, high-performance liquid chromatography (HPLC) will be discussed in a separate chapter (Chap. 28). The general principles of extraction are first described as a basis for understanding chromatography.

  10. Fun with Paper Chromatography. (United States)

    Coleman, Dava; Hounshell, Paul B.


    Discusses paper chromatographic techniques and provides examples of typical classroom activities. Includes description of retardation values obtained during chromatography exercises and suggests using them for math lessons. (JN)

  11. Liquid Chromatography in 1982. (United States)

    Freeman, David H.


    Reviews trends in liquid chromatography including apparatus, factors affecting efficient separation of a mixture (peak sharpness and speed), simplified problem-solving, adsorption, bonded phase chromatography, ion selectivity, and size exclusion. The current trend is to control chemical selectivity by the liquid phase. (Author/JN)

  12. Column Liquid Chromatography. (United States)

    Majors, Ronald E.; And Others


    Reviews literature covering developments of column liquid chromatography during 1982-83. Areas considered include: books and reviews; general theory; columns; instrumentation; detectors; automation and data handling; multidimensional chromatographic and column switching techniques; liquid-solid chromatography; normal bonded-phase, reversed-phase,…

  13. High Performance Liquid Chromatography (United States)

    Talcott, Stephen

    High performance liquid chromatography (HPLC) has many applications in food chemistry. Food components that have been analyzed with HPLC include organic acids, vitamins, amino acids, sugars, nitrosamines, certain pesticides, metabolites, fatty acids, aflatoxins, pigments, and certain food additives. Unlike gas chromatography, it is not necessary for the compound being analyzed to be volatile. It is necessary, however, for the compounds to have some solubility in the mobile phase. It is important that the solubilized samples for injection be free from all particulate matter, so centrifugation and filtration are common procedures. Also, solid-phase extraction is used commonly in sample preparation to remove interfering compounds from the sample matrix prior to HPLC analysis.

  14. Micellar Electrokinetic Chromatography (United States)

    Bald, Edward; Kubalczyk, Paweł

    Since the introduction of micellar electrokinetic chromatography by Terabe, several authors have paid attention to the fundamental characteristics of this separation method. In this chapter the theoretical and practical aspects of resolution optimization, as well as the effect of different separation parameters on the migration behavior are discussed. These among others include fundamentals of separation, retention factor and resolution equation, efficiency, selectivity, and various surfactants and additives. Initial conditions for method development and instrumental approaches such as mass spectrometry detection are also mentioned covering the proposals for overcoming the difficulties arising from the coupling micellar electrokinetic chromatography with mass spectrometry detection.

  15. Micellar liquid chromatography (United States)

    Basova, Elena M.; Ivanov, Vadim M.; Shpigun, Oleg A.


    Background and possibilities of practical applications of micellar liquid chromatography (MLC) are considered. Various retention models in MLC, the effects of the nature and concentration of surfactants and organic modifiers, pH, temperature and ionic strength on the MLC efficiency and selectivity are discussed. The advantages and limitations of MLC are demonstrated. The performance of MLC is critically evaluated in relationship to the reversed-phase HPLC and ion-pair chromatography. The potential of application of MLC for the analysis of pharmaceuticals including that in biological fluids and separation of inorganic anions, transition metal cations, metal chelates and heteropoly compounds is described. The bibliography includes 146 references.

  16. Size fractionation by slalom chromatography and hydrodynamic chromatography


    Dias, Ricardo P.


    Hydrodynamic chromatography, also called separation by flow, is based on the use of the parabolic flow profile occurring in open capillaries or in the pores from a column filled with non-porous particles. The hydrodynamic chromatography separation medium, if any, is much simpler than that from size exclusion chromatography (porous particles), the former technique being used in the size-fractionation of many colloids and macromolecules. The transition between hydrodynamic chromatography (obtai...


    Institute of Scientific and Technical Information of China (English)

    YANGGengliang; TAOZuyi


    The unified theoretical moment expressions for elution chromatography and frontal chromatography when the sorption process is described by a linear model were derived. The moment expressions derived by previous authors can be obtained from these unified theoretical moment expressions. In this paper, a mathematical analysis has been carried out so as to set up a unified theoretical basis for elution and frontal chromatography.

  18. Modern Thin-Layer Chromatography. (United States)

    Poole, Colin F.; Poole, Salwa K.


    Some of the important modern developments of thin-layer chromatography are introduced. Discussed are the theory and instrumentation of thin-layer chromatography including multidimensional and multimodal techniques. Lists 53 references. (CW)

  19. Extending Paper Chromatography Inquiry (United States)

    Finson, Kevin


    One of the "good old" standard activities middle school students seem to enjoy is paper chromatography. The procedures and materials needed are relatively simple and the results can be colorful. All too often, the activity ends just after these colorful results are obtained, cutting short the potential it holds for some further inquiry. With some…

  20. Chromatography in industry

    NARCIS (Netherlands)

    Schoenmakers, P.


    This review focuses on the chromatography research that has been carried out within industry or in close cooperation with industry and that has been reported in the scientific literature between 2006 and mid-2008. Companies in the health care sector, such as pharmaceutical and biotechnology companie

  1. Peak capacity in unidimensional chromatography. (United States)

    Neue, Uwe Dieter


    The currently existing knowledge about peak capacity in unidimensional separations is reviewed. The majority of the paper is dedicated to reversed-phase gradient chromatography, covering specific techniques as well as the subject of peak compression. Other sections deal with peak capacity in isocratic chromatography, size-exclusion chromatography and ion-exchange chromatography. An important topic is the limitation of the separation power and the meaning of the concept of peak capacity for real applications.

  2. High Performance Thin Layer Chromatography. (United States)

    Costanzo, Samuel J.


    Clarifies where in the scheme of modern chromatography high performance thin layer chromatography (TLC) fits and why in some situations it is a viable alternative to gas and high performance liquid chromatography. New TLC plates, sample applications, plate development, and instrumental techniques are considered. (JN)

  3. Separation techniques: Chromatography


    Coskun, Ozlem


    Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis. Proteins can be purified based on characteristics such as size and shape, total charge, hydrophobic groups present on the surface, and binding capacity with the stationary phase. Four separation techniques based on molecular characteristics and interaction type use mechanisms of ion exchange, surface adsorp...

  4. Principles of Liquid Chromatography (United States)

    Bakalyar, Stephen R.

    This article reviews the basic principles of high performance liquid chromatography (HPLC). The introductory section provides an overview of the HPLC technique, placing it in historical context and discussing the elementary facts of the separation mechanism. The next section discusses the nature of resolution, describing the two principal aspects, zone center separation and zone spreading. The third section takes a detailed look at how HPLC is used in practice to achieve a separation. It discusses the three key variables that need to be adjusted: retention, efficiency, and selectivity. A fourth section is concerned with various relationships of practical importance: flow rate, temperature, and pressure. A final section discusses future trends in HPLC.

  5. Flow Rates in Liquid Chromatography, Gas Chromatography and Supercritical Fluid Chromatography: A Tool for Optimization


    Joris Meurs


    This paper aimed to develop a standalone application for optimizing flow rates in liquid chromatography (LC), gas chromatography (GC) and supercritical fluid chromatography (SFC). To do so, Van Deemter’s equation, Knox’ equation and Golay’s equation were implemented in a MATLAB script and subsequently a graphical user interface (GUI) was created. The application will show the optimal flow rate or linear velocity and the corresponding plate height for the set input parameters. Furthermore, a p...

  6. An Undergraduate Column Chromatography Experiment. (United States)

    Danot, M.; And Others


    Background information, list of materials needed, and procedures used are provided for an experiment designed to introduce undergraduate students to the theoretical and technical aspects of column chromatography. The experiment can also be shortened to serve as a demonstration of the column chromatography technique. (JN)

  7. Separation of whey proteins for chromatography liquid


    Abraham D. Giraldo Zuñiga; Edwin E.García Rojas; Jane S. R. Coimbra; Wilmer E. Luera Peña


    This paper describes and compares three chromatographic methods for the analysis and quantification of most abundant proteins in cheese whey, -lactalbumin and -lactoglobulin. The methods were: Reverse-phase high performance liquid chromatography, anion Exchange chromatography and size-exclusion chromatography. The reverse- phase liquid chromatography led to a better separation of whey proteins than size-exclusion chromatography and anion exchange chromatography, this method offered an excel...

  8. Calcium-sensitive immunoaffinity chromatography

    DEFF Research Database (Denmark)

    Henriksen, Maiken L; Lindhardt Madsen, Kirstine; Skjoedt, Karsten


    Immunoaffinity chromatography is a powerful fractionation technique that has become indispensable for protein purification and characterization. However, it is difficult to retrieve bound proteins without using harsh or denaturing elution conditions, and the purification of scarce antigens...... to homogeneity may be impossible due to contamination with abundant antigens. In this study, we purified the scarce, complement-associated plasma protein complex, collectin LK (CL-LK, complex of collectin liver 1 and kidney 1), by immunoaffinity chromatography using a calcium-sensitive anti-collectin-kidney-1 m...... chromatography was superior to the traditional immunoaffinity chromatographies and resulted in a nine-fold improvement of the purification factor. The technique is applicable for the purification of proteins in complex mixtures by single-step fractionation without the denaturation of eluted antigens...

  9. Metal-organic frameworks in chromatography. (United States)

    Yusuf, Kareem; Aqel, Ahmad; ALOthman, Zeid


    Metal-organic frameworks (MOFs) emerged approximately two decades ago and are the youngest class of porous materials. Despite their short existence, MOFs are finding applications in a variety of fields because of their outstanding chemical and physical properties. This review article focuses on the applications of MOFs in chromatography, including high-performance liquid chromatography (HPLC), gas chromatography (GC), and other chromatographic techniques. The use of MOFs in chromatography has already had a significant impact; however, the utilisation of MOFs in chromatography is still less common than other applications, and the number of MOF materials explored in chromatography applications is limited.

  10. Atomic Force Microscope Mediated Chromatography (United States)

    Anderson, Mark S.


    The atomic force microscope (AFM) is used to inject a sample, provide shear-driven liquid flow over a functionalized substrate, and detect separated components. This is demonstrated using lipophilic dyes and normal phase chromatography. A significant reduction in both size and separation time scales is achieved with a 25-micron-length column scale, and one-second separation times. The approach has general applications to trace chemical and microfluidic analysis. The AFM is now a common tool for ultra-microscopy and nanotechnology. It has also been demonstrated to provide a number of microfluidic functions necessary for miniaturized chromatography. These include injection of sub-femtoliter samples, fluidic switching, and sheardriven pumping. The AFM probe tip can be used to selectively remove surface layers for subsequent microchemical analysis using infrared and tip-enhanced Raman spectroscopy. With its ability to image individual atoms, the AFM is a remarkably sensitive detector that can be used to detect separated components. These diverse functional components of microfluidic manipulation have been combined in this work to demonstrate AFM mediated chromatography. AFM mediated chromatography uses channel-less, shear-driven pumping. This is demonstrated with a thin, aluminum oxide substrate and a non-polar solvent system to separate a mixture of lipophilic dyes. In conventional chromatographic terms, this is analogous to thin-layer chromatography using normal phase alumina substrate with sheardriven pumping provided by the AFM tip-cantilever mechanism. The AFM detection of separated components is accomplished by exploiting the variation in the localized friction of the separated components. The AFM tip-cantilever provides the mechanism for producing shear-induced flows and rapid pumping. Shear-driven chromatography (SDC) is a relatively new concept that overcomes the speed and miniaturization limitations of conventional liquid chromatography. SDC is based on a

  11. Extraction chromatography: Progress and opportunities

    Energy Technology Data Exchange (ETDEWEB)

    Dietz, M.L.; Horwitz, E.P.; Bond, A.H. [Argonne National Lab., IL (United States). Chemistry Div.


    Extraction chromatography provides a simple and effective method for the analytical and preparative-scale separation of a variety of metal ions. Recent advances in extractant design, particularly the development of extractants capable of metal ion recognition or of strong complex formation in highly acidic media, have significantly improved the utility of the technique. Advances in support design, most notably the introduction of functionalized supports to enhance metal ion retention, promise to yield further improvements. Column instability remains a significant obstacle, however, to the process-scale application of extraction chromatography. 79 refs.

  12. Thin-Layer and Paper Chromatography. (United States)

    Sherma, Joseph; Fried, Bernard


    Reviews literature on chromatography examining: books, reviews, student experiments; chromatographic systems, techniques, apparatus; detecting and identification of separated zones; preparative chromatography and radiochromatography; and applications related to specific materials (such as acids, alcohols, amino acids, antibiotics, enzymes, dyes,…

  13. Selectivity in microemulsion electrokinetic chromatography

    DEFF Research Database (Denmark)

    Pedersen-Bjergaard, S; Gabel-Jensen, Charlotte; Honoré Hansen, S


    Microemulsion electrokinetic chromatography (MEEKC) is a most promising separation technique providing good selectivity and high separation efficiency of anionic, cationic as well as neutral solutes. In MEEKC lipophilic organic solvents dispersed as tiny droplets in an aqueous buffer by the use...

  14. Liquid phase chromatography on microchips

    DEFF Research Database (Denmark)

    Kutter, Jörg Peter


    explosive development of, in particular, chromatographic separation systems on microchips, has, however, slowed down in recent years. This review takes a closer, critical look at how liquid phase chromatography has been implemented in miniaturized formats over the past several years, what is important...

  15. Bioaffinity chromatography on monolithic supports

    NARCIS (Netherlands)

    Tetala, K.K.R.; Beek, van T.A.


    Affinity chromatography on monolithic supports is a powerful analytical chemical platform because it allows for fast analyses, small sample volumes, strong enrichment of trace biomarkers and applications in microchips. In this review, the recent research using monolithic materials in the field of bi

  16. Illustrating Chromatography with Colorful Proteins (United States)

    Lefebvre, Brian G.; Farrell, Stephanie; Dominiak, Richard S.


    Advances in biology are prompting new discoveries in the biotechnology, pharmaceutical, medical technology, and chemical industries. This paper presents a detailed description of an anion exchange chromatography experiment using a pair of colorful proteins and summarizes the effect of operating parameters on protein separation. This experiment…

  17. Two-dimensional liquid chromatography

    DEFF Research Database (Denmark)

    Græsbøll, Rune

    of this thesis is on online comprehensive two-dimensional liquid chromatography (online LC×LC) with reverse phase in both dimensions (online RP×RP). Since online RP×RP has not been attempted before within this research group, a significant part of this thesis consists of knowledge and experience gained...

  18. Review: Applications of chromatography in forensic sciences

    Directory of Open Access Journals (Sweden)

    Manoj S. Charde


    Full Text Available This article reviews the use of different Chromatography techniques in the forensic science, Chromatographic technique is very sensitive and selective. Different types of chromatography techniques used were Liquid chromatography -mass spectrometry, Gas chromatography–mass spectrometry, Thin layer chromatography, HPTLC in investigating criminal cases of which chemical warfare’s, terrorist attacks, smugglers, drug abuse, alcoholics. This techniques are promising to detect even pictogram or very less, with selectivity and sensitivity.

  19. Lipidomics by Supercritical Fluid Chromatography

    Directory of Open Access Journals (Sweden)

    Laurent Laboureur


    Full Text Available This review enlightens the role of supercritical fluid chromatography (SFC in the field of lipid analysis. SFC has been popular in the late 1980s and 1990s before almost disappearing due to the commercial success of liquid chromatography (LC. It is only 20 years later that a regain of interest appeared when new commercial instruments were introduced. As SFC is fully compatible with the injection of extracts in pure organic solvent, this technique is perfectly suitable for lipid analysis and can be coupled with either highly universal (UV or evaporative light scattering or highly specific (mass spectrometry detection methods. A short history of the use of supercritical fluids as mobile phase for the separation oflipids will be introduced first. Then, the advantages and drawbacks of SFC are discussed for each class of lipids (fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, sterols, prenols, polyketides defined by the LIPID MAPS consortium.

  20. Capillary Separation: Micellar Electrokinetic Chromatography (United States)

    Terabe, Shigeru


    Micellar electrokinetic chromatography (MEKC), a separation mode of capillary electrophoresis (CE), has enabled the separation of electrically neutral analytes. MEKC can be performed by adding an ionic micelle to the running solution of CE without modifying the instrument. Its separation principle is based on the differential migration of the ionic micelles and the bulk running buffer under electrophoresis conditions and on the interaction between the analyte and the micelle. Hence, MEKC's separation principle is similar to that of chromatography. MEKC is a useful technique particularly for the separation of small molecules, both neutral and charged, and yields high-efficiency separation in a short time with minimum amounts of sample and reagents. To improve the concentration sensitivity of detection, several on-line sample preconcentration techniques such as sweeping have been developed.

  1. Affinity chromatography of phosphorylated proteins. (United States)

    Tchaga, Grigoriy S


    This chapter covers the use of immobilized metal ion affinity chromatography (IMAC) for enrichment of phosphorylated proteins. Some requirements for successful enrichment of these types of proteins are discussed. An experimental protocol and a set of application data are included to enable the scientist to obtain high-yield results in a very short time with pre-packed phospho-specific metal ion affinity resin (PMAC).

  2. Lagrangian description of nonlinear chromatography

    Institute of Scientific and Technical Information of China (English)

    LIANG Heng; LIU Xiaolong


    Under the framework of non-equilibrium thermodynamic separation theory (NTST), Local Lagrangian approach (LLA) was proposed to deal with the essential issues of the convection and diffusion (shock waves) phenomena in nonlinear chromatography with recursion equations based on the three basic theorems, Lagrangian description, continuity axiom and local equilibrium assumption (LEA). This approach remarkably distinguished from the system of contemporary chromatographic theories (Eulerian description-partial differential equations), and can felicitously match modern cybernetics.

  3. Lectin affinity chromatography of glycolipids

    Energy Technology Data Exchange (ETDEWEB)

    Torres, B.V.; Smith, D.F.


    Since glycolipids (GLs) are either insoluble or form mixed micelles in water, lectin affinity chromatography in aqueous systems has not been applied to their separation. They have overcome this problem by using tetrahydrofuran (THF) in the mobile phase during chromatography. Affinity columns prepared with the GalNAc-specific Helix pomatia agglutinin (HPA) and equilibrated in THF specifically bind the (/sup 3/H)oligosaccharide derived from Forssman GL indicating that the immobilized HPA retained its carbohydrate-binding specificity in this solvent. Intact Forssman GL was bound by the HPA-column equilibrated in THF and was specifically eluted with 0.1 mg/ml GalNAc in THF. Purification of the Forssman GL was achieved when a crude lipid extract of sheep erythrocyte membranes was applied to the HPA-column in THF. Non-specifically bound GLs were eluted from the column using a step gradient of aqueous buffer in THF, while the addition of GalNAc was required to elute the specifically bound GLs. Using this procedure the A-active GLs were purified from a crude lipid extract of type A human erythrocytes in a single chromatographic step. The use of solvents that maintain carbohydrate-binding specificity and lipid solubility will permit the application of affinity chromatography on immobilized carbohydrate-binding proteins to intact GLs.

  4. Recent progress on countercurrent chromatography modeling


    Wang, Fengkang; Ito, Yoichiro; Wei, Yun


    As countercurrent chromatography is becoming an established method in chromatography for many kinds of products, it is becoming increasingly important to model the process and to be able to predict the peaks for a given process. The CCC industries are looking for rapid methods to analyze the processes of countercurrent chromatography and select suitable solvent system. In this paper, recent progress is reviewed in the development and demonstration of several types of models of countercurrent ...

  5. Ion Exchange and Liquid Column Chromatography. (United States)

    Walton, Harold F.


    Emphasizes recent advances in principles and methodology in ion exchange and chromatography. Two tables list representative examples for inorganic ions and organic compounds. Cites 544 references. (CS)

  6. Thin Layer Chromatography (TLC) of Chlorophyll Pigments. (United States)

    Foote, Jerry


    Background information, list of materials needed, procedures used, and discussion of typical results are provided for an experiment on the thin layer chromatography of chlorophyll pigments. The experiment works well in high school, since the chemicals used are the same as those used in paper chromatography of plant pigments. (JN)

  7. Liquid phase chromatography on microchips. (United States)

    Kutter, Jörg P


    Over the past twenty years, the field of microfluidics has emerged providing one of the main enabling technologies to realize miniaturized chemical analysis systems, often referred to as micro-Total Analysis Systems (uTAS), or, more generally, Lab-on-a-Chip Systems (LOC) [1,2]. While microfluidics was driven forward a lot from the engineering side, especially with respect to ink jet and dispensing technology, the initial push and interest from the analytical chemistry community was through the desire to develop miniaturized sensors, detectors, and, very early on, separation systems. The initial almost explosive development of, in particular, chromatographic separation systems on microchips, has, however, slowed down in recent years. This review takes a closer, critical look at how liquid phase chromatography has been implemented in miniaturized formats over the past several years, what is important to keep in mind when developing or working with separations in a miniaturized format, and what challenges and pitfalls remain.

  8. Polymer nanoparticles in electrokinetic chromatography (United States)

    Hyslop, Jesse Samuel

    This dissertation reports the mobility, methylene selectivity, efficiency, linear solvation relationship (LSER) parameters, and practical chromatographic performance of a large set of NP PSPs and develops the first empirical relationships between NP architecture and chromatographic performance of NP PSPs in EKC. It is found that under typical EKC conditions ionic block chemistry has little effect on performance for 5-10 mer blocks. Solute-PSP interactions appear to be localized on the hydrophobic block of the copolymer with the length of alkyl chains on the hydrophobic block controlling the cohesively and hydrophobicity of the PSP. Small (100 nm) NP PSPs with small hydrophobic NP PSPs providing the best overall performance. This work provides the fundamental understanding of the behavior of RAFT polymerized NP PSPs essential for their further development and application in electrokinetic chromatography. (Abstract shortened by ProQuest.).

  9. Detection in superheated water chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Chienthavorn, O


    Superheated water has been used successfully as an eluent in liquid chromatography and has been coupled to various modes of detection, ultraviolet (UV), fluorescence, and nuclear magnetic resonance spectroscopy (NMR) and mass spectrometry (MS). A number of compounds were examined on poly(styrene-divinylbenzene) (PS-DVB), polybutadiene (PBD), and octadecylsilyl bonded silica (ODS) column with isothermal and temperature programmes. The PS-DVB column was mostly used throughout the project as it was the most stable. Not only pure water could serve as superheated water mobile phase; inorganic buffered water and ion-pairing reagent with a concentration of 1-3 mM of the buffer and reagent were also exploited. It was shown that the pH could be controlled during the separation without salt precipitation and the separations followed a conventional reversed-phase HPLC method. Results from fluorescence detection showed good separation of a series of vitamins, such as pyridoxine, riboflavin, thiamine, and some analgesics. The relationship of riboflavin using the detection was linear and the detection limit was seven times higher than that of a conventional method. Simultaneous separation and identification using superheated water chromatography-NMR was demonstrated. With using a stop flow method, NMR spectra of model drugs, namely barbiturates, paracetamol, caffeine and phenacetin were obtained and the results agreed with reference spectra, confirming a perfect separation. A demonstration to obtain COSY spectrum of salicylamide was also performed. The method was expanded to the coupling of superheated water LC to NMR-MS. Results from the hyphenated detection method showed that deuteration and degradation happened in the superheated water conditions. The methyl group hydrogens of pyrimidine ring of sulfonamide and thiamine were exchanged with deuterium. Thiamine was decomposed to 4-methyl-5-thiazoleethanol and both were deuterated under the conditions. (author)

  10. The Chromatography of Leaves and Inks. (United States)

    Chemecology, 1997


    Describes the use of a simple process known as chromatography to separate and observe the color pigments in leaves, inks, and other materials. Provides some historical background and detailed procedures. (DDR)

  11. Stationary phases for superheated water chromatography

    CERN Document Server

    Saha, S


    This project focused on the comparison of conventional liquid chromatography and superheated water chromatography. It examined the differences in efficiency and retention of a range of different stationary phases. Alkyl aryl ketones and eight aromatic compounds were separated on PBD-zirconia, Xterra RP 18, Luna C sub 1 sub 8 (2) and Oasis HLB columns using conventional LC and superheated water chromatography system. The retention indices were determined in the different eluents. On changing the organic component of the eluent from methanol to acetonitrile to superheated water considerable improvements were found in the peak shapes and column efficiencies on the PBD-zirconia and Oasis HLB columns. PS-DVB, PBD-zirconia and Xterra RP 18 columns have been used in efficiency studies. It was found that simply elevating the column temperature did not increase the efficiency of a separation in superheated water chromatography. The efficiency depended on flow rate, injection volume and also mobile phase preheating sys...

  12. Diagonal chromatography to study plant protein modifications

    NARCIS (Netherlands)

    Walton, Alan; Tsiatsiani, Liana; Jacques, Silke; Stes, Elisabeth; Messens, Joris; Van Breusegem, Frank; Goormachtig, Sofie; Gevaert, Kris


    An interesting asset of diagonal chromatography, which we have introduced for contemporary proteome research, is its high versatility concerning proteomic applications. Indeed, the peptide modification or sorting step that is required between consecutive peptide separations can easily be altered and

  13. High-Performance Liquid Chromatography: An Overview


    Tandia N; Singh SK1; Kumar N; Singh S


    Chromatography is considered extremely powerful separation technique for variety of samples. Thecommon feature of all these techniques is that the different components in a sample mixture aredistributed between two phases, one of which remains stationary while the other the mobile phase, runsthrough the interstices or over the surface of the fixed phase. The movement of the mobile phase resultsin differential migration of the sample components. High performance thin layer chromatography is an...

  14. Low thermal mass liquid chromatography. (United States)

    Gu, Binghe; Cortes, Hernan; Luong, Jim; Pursch, Matthias; Eckerle, Patric; Mustacich, Robert


    A novel technique, low thermal mass liquid chromatography (LTMLC), is introduced in this study. The use of an LTM assembly that utilizes the principle of resistive wire heating and a temperature sensor to accurately deliver unprecedented heating (up to 1800 degrees C/min) or cooling (100 to approximately 200 degrees C/min) rates is reported. With the use of packed microcolumns (heat transfer from the assembly to the mobile phase was obtained. A systematic investigation was conducted to study the performance of the LTMLC technique. Both isocratic and gradient mobile phase conditions were used. For temperature control, isothermal, temperature-increasing, and temperature-decreasing gradients were applied. Three model mixtures, two of which containing neutral and acidic analytes and the other containing neutral, acidic, and basic analytes, were used to study the effect of temperature on elution time, resolution, column efficiency, and selectivity. It was found that the LTMLC experimental setup delivered reliable temperature control, as evidenced by linear van't Hoff plots for neutral and acidic compounds. The effect of temperature on the elution of basic analytes yielded nonlinear van't Hoff plots, explaining the dramatic selectivity changes observed for bases with changes in column temperature. Column efficiency generally increased with the increase in column temperature in the range of 25 to approximately 75 degrees C and decreased in the range of 75 to approximately 150 degrees C at a fixed column flow rate (3 microL/min), when extra column band broadening was taken into account. The increase in efficiency upon the increase in column temperature in the low temperature range was mainly due to the decreased mass transfer term resulting from increased analyte diffusivity. However, under even higher temperatures, the longitudinal diffusion dominated band broadening, explaining the decrease in column efficiency upon a further increase in column temperature. Resolution

  15. Comprehensive two-dimensional liquid chromatography: Ion chromatography × reversed-phase liquid chromatography for separation of low-molar-mass organic acids

    NARCIS (Netherlands)

    S.S. Brudin; R.A. Shellie; P.R. Haddad; P.J. Schoenmakers


    In the work presented here a novel approach to comprehensive two-dimensional liquid chromatography is evaluated. Ion chromatography is chosen for the first-dimension separation and reversed-phase liquid chromatography is chosen for the second-dimension separation mode. The coupling of these modes is

  16. High-performance liquid chromatography of biopolymers

    Energy Technology Data Exchange (ETDEWEB)

    Regnier, F.E.


    The ability to separate biological macromolecules with good resolution on liquid chromatographic columns has depended on the development of suitable packing materials. In size exclusion chromatography, molecules are separated by size on the basis of differential permeation of the packing. Ion exchange, hydrophobic interaction (or reversed-phase), and affinity chromatography are all surface-mediated separation methods, although they depend on different retention mechanisms. High-performance liquid chromatographic columns designed for biopolymers offer major advantages over conventional columns in both speed and resolving power. The exponential growth of literature on the high-performance separation of peptides and proteins in particular indicates that the techniques will become the dominant form of column liquid chromatography. 92 refs., 4 figs.

  17. Diagonal chromatography to study plant protein modifications. (United States)

    Walton, Alan; Tsiatsiani, Liana; Jacques, Silke; Stes, Elisabeth; Messens, Joris; Van Breusegem, Frank; Goormachtig, Sofie; Gevaert, Kris


    An interesting asset of diagonal chromatography, which we have introduced for contemporary proteome research, is its high versatility concerning proteomic applications. Indeed, the peptide modification or sorting step that is required between consecutive peptide separations can easily be altered and thereby allows for the enrichment of specific, though different types of peptides. Here, we focus on the application of diagonal chromatography for the study of modifications of plant proteins. In particular, we show how diagonal chromatography allows for studying proteins processed by proteases, protein ubiquitination, and the oxidation of protein-bound methionines. We discuss the actual sorting steps needed for each of these applications and the obtained results. This article is part of a Special Issue entitled: Plant Proteomics--a bridge between fundamental processes and crop production, edited by Dr. Hans-Peter Mock.

  18. Liquid chromatography detection unit, system, and method

    Energy Technology Data Exchange (ETDEWEB)

    Derenzo, Stephen E.; Moses, William W.


    An embodiment of a liquid chromatography detection unit includes a fluid channel and a radiation detector. The radiation detector is operable to image a distribution of a radiolabeled compound as the distribution travels along the fluid channel. An embodiment of a liquid chromatography system includes an injector, a separation column, and a radiation detector. The injector is operable to inject a sample that includes a radiolabeled compound into a solvent stream. The position sensitive radiation detector is operable to image a distribution of the radiolabeled compound as the distribution travels along a fluid channel. An embodiment of a method of liquid chromatography includes injecting a sample that comprises radiolabeled compounds into a solvent. The radiolabeled compounds are then separated. A position sensitive radiation detector is employed to image distributions of the radiolabeled compounds as the radiolabeled compounds travel along a fluid channel.

  19. High Performance Liquid Chromatography Experiments to Undergraduate Laboratories (United States)

    Kissinger, Peter T.; And Others


    Reviews the principles of liquid chromatography with electrochemical detection (LCEC), an analytical technique that incorporates the advantages of both liquids chromatography and electrochemistry. Also suggests laboratory experiments using this technique. (MLH)

  20. The History of Ion Chromatography: The Engineering Perspective (United States)

    Evans, Barton


    The development of ion chromatography from an engineering perspective is presented. As ion chromatography became more widely accepted, researchers developed dozens of standard applications that enabled the creation of many low-end instruments.

  1. The Ideal Solvent for Paper Chromatography of Food Dyes. (United States)

    Markow, Peter G.


    Uses paper chromatography with food dyes to provide a simple and inexpensive basis for teaching chromatography. Provides experimental methodology and tabled results. Includes a solvent system comparison (Rf) for seven dyes and twenty-two solvents. (MVL)

  2. Christhin: Quantitative Analysis of Thin Layer Chromatography

    CERN Document Server

    Barchiesi, Maximiliano; Renaudo, Carlos; Rossi, Pablo; Pramparo, María de Carmen; Nepote, Valeria; Grosso, Nelson Ruben; Gayol, María Fernanda


    Manual for Christhin 0.1.36 Christhin (Chromatography Riser Thin) is software developed for the quantitative analysis of data obtained from thin-layer chromatographic techniques (TLC). Once installed on your computer, the program is very easy to use, and provides data quickly and accurately. This manual describes the program, and reading should be enough to use it properly.

  3. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.


    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...

  4. Three dimensional liquid chromatography coupling ion exchange chromatography/hydrophobic interaction chromatography/reverse phase chromatography for effective protein separation in top-down proteomics. (United States)

    Valeja, Santosh G; Xiu, Lichen; Gregorich, Zachery R; Guner, Huseyin; Jin, Song; Ge, Ying


    To address the complexity of the proteome in mass spectrometry (MS)-based top-down proteomics, multidimensional liquid chromatography (MDLC) strategies that can effectively separate proteins with high resolution and automation are highly desirable. Although various MDLC methods that can effectively separate peptides from protein digests exist, very few MDLC strategies, primarily consisting of 2DLC, are available for intact protein separation, which is insufficient to address the complexity of the proteome. We recently demonstrated that hydrophobic interaction chromatography (HIC) utilizing a MS-compatible salt can provide high resolution separation of intact proteins for top-down proteomics. Herein, we have developed a novel 3DLC strategy by coupling HIC with ion exchange chromatography (IEC) and reverse phase chromatography (RPC) for intact protein separation. We demonstrated that a 3D (IEC-HIC-RPC) approach greatly outperformed the conventional 2D IEC-RPC approach. For the same IEC fraction (out of 35 fractions) from a crude HEK 293 cell lysate, a total of 640 proteins were identified in the 3D approach (corresponding to 201 nonredundant proteins) as compared to 47 in the 2D approach, whereas simply prolonging the gradients in RPC in the 2D approach only led to minimal improvement in protein separation and identifications. Therefore, this novel 3DLC method has great potential for effective separation of intact proteins to achieve deep proteome coverage in top-down proteomics.

  5. Instrument platforms for nano liquid chromatography. (United States)

    Šesták, Jozef; Moravcová, Dana; Kahle, Vladislav


    The history of liquid chromatography started more than a century ago and miniaturization and automation are two leading trends in this field. Nanocolumn liquid chromatography (nano LC) and largely synonymous capillary liquid chromatography (capillary LC) are the most recent results of this process where miniaturization of column dimensions and sorbent particle size play crucial role. Very interesting results achieved in the research of extremely miniaturized LC columns at the end of the last century lacked distinctive raison d'être and only advances in mass spectrometry brought a real breakthrough. Configuration of nano LC-electrospray ionization mass spectrometry (LC-ESI-MS) has become a basic tool in bioanalytical chemistry, especially in proteomics. This review discusses and summarizes past and current trends in the realization of nano liquid chromatography (nano LC) platforms. Special attention is given to the mobile phase delivery under nanoflow rates (isocratic, gradient) and sample injection to the nanocolumn. Available detection techniques applied in nano LC separations are also briefly discussed. We followed up the key themes from the original scientific reports over gradual improvements up to the contemporary commercial solutions.

  6. Performance of the same column in supercritical fluid chromatography and in liquid chromatography. (United States)

    Lambert, Nándor; Felinger, Attila


    We have studied the chromatographic behavior of the homologous series of alkylbenzenes (ranging from octylbenzene to octadecylbenzene) on the same C18 reversed-phase column in supercritical fluid chromatography (SFC) and reversed phase liquid chromatography (RPLC) at various experimental conditions, such as different eluent compositions, flow-rates, and mobile phase densities. The first and the second moments of the peaks were used to estimate the overall mass-transfer processes in both chromatographic modes using the stochastic model of chromatography. The results confirm that in SFC - as the density of the mobile phase is influenced by the flow-rate - there is a broader variation of mass-transfer properties than in liquid chromatography. As expected, the optimum mobile phase velocity is higher in SFC, but there is no real difference in the minimum value of plate height, i.e. in the optimum efficiency.

  7. Chromatography purification of canine adenoviral vectors. (United States)

    Segura, María Mercedes; Puig, Meritxell; Monfar, Mercè; Chillón, Miguel


    Canine adenovirus vectors (CAV2) are currently being evaluated for gene therapy, oncolytic virotherapy, and as vectors for recombinant vaccines. Despite the need for increasing volumes of purified CAV2 preparations for preclinical and clinical testing, their purification still relies on the use of conventional, scale-limited CsCl ultracentrifugation techniques. A complete downstream processing strategy for CAV2 vectors based on membrane filtration and chromatography is reported here. Microfiltration and ultra/diafiltration are selected for clarification and concentration of crude viral stocks containing both intracellular and extracellular CAV2 particles. A DNase digestion step is introduced between ultrafiltration and diafiltration operations. At these early stages, concentration of vector stocks with good recovery of viral particles (above 80%) and removal of a substantial amount of protein and nucleic acid contaminants is achieved. The ability of various chromatography techniques to isolate CAV2 particles was evaluated. Hydrophobic interaction chromatography using a Fractogel propyl tentacle resin was selected as a first chromatography step, because it allows removal of the bulk of contaminating proteins with high CAV2 yields (88%). An anion-exchange chromatography step using monolithic supports is further introduced to remove the remaining contaminants with good recovery of CAV2 particles (58-69%). The main CAV2 viral structural components are visualized in purified preparations by electrophoresis analyses. Purified vector stocks contained intact icosahedral viral particles, low contamination with empty viral capsids (10%), and an acceptable total-to-infectious particle ratio (below 30). The downstream processing strategy that was developed allows preparation of large volumes of high-quality CAV2 stocks.

  8. High Performance Liquid Chromatography/Video Fluorometry. Part I. Instrumentation. (United States)


    High Performance Liquid Chromatography /Video...PERIOD COVERED High Performance Liquid Chromatography /Video .. / Fluorometry. Part I. Instrumentation. . Interim/ echnicaliepart,. 6. PERFORMING ORG...34Entered SECURITY CLASSIFICATION OF THIS OlAGE (When Data Entered) II1| III I I I I E I II ... .. High Performance Liquid Chromatography

  9. Separation of proteins by simulated moving bed chromatography

    NARCIS (Netherlands)

    Houwing, J.


    This thesis describes the use of simulated moving bed (SMB) chromatography for separation of proteins. The underlying aim is the increase of the efficiency of process chromatography in terms of a reduction of the sorbent and buffer consumption in comparison to fixed bed chromatography. Both size exc

  10. [Development of metal ions analysis by ion chromatography]. (United States)

    Yu, Hong; Wang, Yuxin


    Analysis of metal ions by ion chromatography, including cation-exchange ion chromatography, anion-exchange ion chromatography and chelation ion chromatography, is reviewed. The cation-exchange ion chromatography is a main method for the determination of metal ions. Stationary phases in cation-exchange ion chromatography are strong acid cation exchanger (sulfonic) and weak acid cation exchanger (carboxylic). Alkali metal ions, alkaline earth metal ions, transition metal ions, rare earth metal ions, ammonium ions and amines can be analyzed by cation-exchange ion chromatography with a suitable detector. The anion-exchange ion chromatography is suitable for the separation and analysis of alkaline earth metal ions, transition metal ions and rare earth metal ions. The selectivity for analysis of metal ions with anion-exchange ion chromatography is good. Simultaneous determination of metal ions and inorganic anions can be achieved using anion-exchange ion chromatography. Chelation ion chromatography is suitable for the determination of trace metal ions in complex matrices. A total of 125 references are cited.

  11. Non-planar microfabricated gas chromatography column (United States)

    Lewis, Patrick R.; Wheeler, David R.


    A non-planar microfabricated gas chromatography column comprises a planar substrate having a plurality of through holes, a top lid and a bottom lid bonded to opposite surfaces of the planar substrate, and inlet and outlet ports for injection of a sample gas and elution of separated analytes. A plurality of such planar substrates can be aligned and stacked to provide a longer column length having a small footprint. Furthermore, two or more separate channels can enable multi-channel or multi-dimensional gas chromatography. The through holes preferably have a circular cross section and can be coated with a stationary phase material or packed with a porous packing material. Importantly, uniform stationary phase coatings can be obtained and band broadening can be minimized with the circular channels. A heating or cooling element can be disposed on at least one of the lids to enable temperature programming of the column.

  12. Retention models for programmed gas chromatography. (United States)

    Castello, G; Moretti, P; Vezzani, S


    The models proposed by many authors for the prediction of retention times and temperatures, peak widths, retention indices and separation numbers in programmed temperature and pressure gas chromatography by starting from preliminary measurements of the retention in isothermal and isobaric conditions are reviewed. Several articles showing the correlation between retention data and thermodynamic parameters and the determination of the optimum programming rate are reported. The columns of different polarity used for the experimental measurement and the main equations, mathematical models and calculation procedures are listed. An empirical approach was used in the early models, followed by the application of thermodynamic considerations, iterative calculation procedures and statistical methods, based on increased computing power now available. Multiple column arrangements, simultaneous temperature and pressure programming, applications of two-dimensional and fast chromatography are summarised.

  13. Protein purification using PDZ affinity chromatography. (United States)

    Walkup, Ward G; Kennedy, Mary B


    PDZ domains function in nature as protein-binding domains within scaffold and membrane-associated proteins. They comprise approximately 90 residues and undergo specific, high-affinity interactions with complementary C-terminal peptide sequences, other PDZ domains, and/or phospholipids. We have previously shown that the specific, strong interactions of PDZ domains with their ligands make them well suited for use in affinity chromatography. This unit provides protocols for the PDZ affinity chromatography procedure that are applicable for the purification of proteins that contain PDZ domains or PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We detail the preparation of affinity resins composed of PDZ domains or PDZ domain peptide ligands coupled to solid supports. These resins can be used to purify proteins containing endogenous or genetically introduced PDZ domains or ligands, eluting the proteins with free PDZ domain peptide ligands.

  14. Basic prinicipal of high performance liquid chromatography


    Ivanova, Violeta


    High-performance liquid chromatography (HPLC) is an analytical technique based on the separation of molecules due to differences in their structure and/or composition. Separation is performed between two phases, mobile and stationary. The molecules in the sample have different affinities and interactions with the stationary support, leading to separation of molecules. Compounds which are longer retained at the stationary phase will elute later, compared to those which are distributed into the...

  15. Recent applications of hydrophilic interaction liquid chromatography in pharmaceutical analysis. (United States)

    Zhang, Qian; Yang, Feng-Qing; Ge, Liya; Hu, Yuan-Jia; Xia, Zhi-Ning


    Hydrophilic interaction liquid chromatography, an alternative liquid chromatography mode, is of particular interest in separating hydrophilic and polar ionic compounds. Compared with traditional liquid chromatography techniques, hydrophilic interaction liquid chromatography offers specific advantages mainly including: (1) relatively green and water-soluble mobile phase composition, which enhances the solubility of hydrophilic and polar ionic compounds; (2) no need for ion-pairing reagents and high content of organic solvent, which benefits mass spectrometry detection; (3) high orthogonality to reverse-phase liquid chromatography, well adapted to two-dimensional liquid chromatography for complicated samples. Therefore, hydrophilic interaction liquid chromatography has been rapidly developed in many areas over the past decades. This review summarizes the recent progress (from 2012 to July 2016) of hydrophilic interaction liquid chromatography in pharmaceutical analysis, with the focus on detecting chemical drugs in various matrices, charactering active compounds of natural products and assessing biotherapeutics through typical structure unit. Moreover, the retention mechanism and behavior of analytes in hydrophilic interaction liquid chromatography as well as some novel hydrophilic interaction liquid chromatography columns used for pharmaceutical analysis are also described.

  16. Isotopic separation by ion chromatography; La separation isotopique par chromatographie ionique

    Energy Technology Data Exchange (ETDEWEB)

    Albert, M.G.; Barre, Y.; Neige, R. [CEA Centre d`Etudes de la Vallee du Rhone, 26 - Pierrelatte (France). Dept. de Technologie de l`Enrichissement


    The isotopic exchange reaction and the isotopic separation factor are first recalled; the principles of ion chromatography applied to lithium isotope separation are then reviewed (displacement chromatography) and the process is modelled in the view of dimensioning and optimizing the industrial process; the various dimensioning parameters are the isotopic separation factor, the isotopic exchange kinetics and the material flow rate. Effects of the resin type and structure are presented. Dimensioning is also affected by physico-chemical and hydraulic parameters. Industrial implementation features are also discussed. 1 fig., 1 tab., 5 refs.

  17. Trends in High Performance Liquid Chromatography for Cultural Heritage. (United States)

    Degano, Ilaria; La Nasa, Jacopo


    The separation, detection and quantitation of specific species contained in a sample in the field of Cultural Heritage requires selective, sensitive and reliable methods. Procedures based on liquid chromatography fulfil these requirements and offer a wide range of applicability in terms of analyte types and concentration range. The main applications of High Performance Liquid Chromatography in this field are related to the separation and detection of dyestuffs in archaeological materials and paint samples by reversed-phase liquid chromatography with suitable detectors. The relevant literature will be revised, with particular attention to sample treatment strategies and future developments. Reversed phase chromatography has also recently gained increasing importance in the analysis of lipid binders and lipid materials in archaeological residues: the main advantages and disadvantages of the new approaches will be discussed. Finally, the main applications of ion chromatography and size exclusion chromatography in the field of Cultural Heritage will be revised in this chapter.

  18. High Performance Liquid Chromatography/Video Fluorometry. Part II. Applications. (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY /VIDEO FLUOROMETRY. PART...REP«T_N&:-ŗ/ High Performance Liquid Chromatography /Video Fluorometry» Part II. Applications« by | Dennis C./Shelly* Michael P./Vogarty and...Data EnlirtdJ REPORT DOCUMENTATION PAGE t. REPORT NUMBER 2 GOVT ACCESSION NO 4. T1TI.F (and Submit) lP-^fffsyva High Performance Liquid Chromatography

  19. Separation of proteins by simulated moving bed chromatography


    Houwing, J.


    This thesis describes the use of simulated moving bed (SMB) chromatography for separation of proteins. The underlying aim is the increase of the efficiency of process chromatography in terms of a reduction of the sorbent and buffer consumption in comparison to fixed bed chromatography. Both size exclusion and ion exchange separations of synthetic mixtures of proteins are considered. The main topics covered are the selection of the liquid to sorbent flow rate ratios, mass transfer effects and ...

  20. Tutorial: simulating chromatography with Microsoft Excel Macros. (United States)

    Kadjo, Akinde; Dasgupta, Purnendu K


    Chromatography is one of the cornerstones of modern analytical chemistry; developing an instinctive feeling for how chromatography works will be invaluable to future generation of chromatographers. Specialized software programs exist that handle and manipulate chromatographic data; there are also some that simulate chromatograms. However, the algorithm details of such software are not transparent to a beginner. In contrast, how spreadsheet tools like Microsoft Excel™ work is well understood and the software is nearly universally available. We show that the simple repetition of an equilibration process at each plate (a spreadsheet row) followed by discrete movement of the mobile phase down by a row, easily automated by a subroutine (a "Macro" in Excel), readily simulates chromatography. The process is readily understood by a novice. Not only does this permit simulation of isocratic and simple single step gradient elution, linear or multistep gradients are also easily simulated. The versatility of a transparent and easily understandable computational platform further enables the simulation of complex but commonly encountered chromatographic scenarios such as the effects of nonlinear isotherms, active sites, column overloading, on-column analyte degradation, etc. These are not as easily simulated by available software. Views of the separation as it develops on the column and as it is seen by an end-column detector are both available in real time. Excel 2010™ also permits a 16-level (4-bit) color gradation of numerical values in a column/row; this permits visualization of a band migrating down the column, much as Tswett may have originally observed, but in a numerical domain. All parameters of relevance (partition constants, elution conditions, etc.) are readily changed so their effects can be examined. Illustrative Excel spreadsheets are given in the Supporting Information; these are easily modified by the user or the user can write his/her own routine.

  1. Extraction and chromatography of carotenoids from pumpkin. (United States)

    Seo, Jung Sook; Burri, Betty Jane C; Quan, Zhejiu; Neidlinger, Terry R


    Vitamin A deficiency is a health problem in Southeast Asia that can be corrected by feeding orange fruits and vegetables such as mango. Pumpkin is a traditional Korean food that is easy to store and is already believed to have health benefits. We extracted carotenoids from pumpkin by liquid-liquid extraction and by supercritical fluid extraction. We measured carotenoids by reversed-phase chromatography with diode array detection. The major carotenoid in pumpkin (> 80%) is beta-carotene, with lesser amounts of lutein, lycopene, alpha-carotene and cis-beta-carotene. Pumpkin is a rich source of beta-carotene and might be useful for preventing Vitamin A deficiency.

  2. Affinity chromatography with an immobilized RNA enzyme.


    Vioque, A; Altman, S


    M1 RNA, the catalytic subunit of Escherichia coli RNase P, has been covalently linked at its 3' terminus to agarose beads. Unlike M1 RNA, which is active in solution in the absence of the protein component (C5) of RNase P, the RNA linked to the beads is active only in the presence of C5 protein. Affinity chromatography of crude extracts of E. coli on a column prepared from the beads to which the RNA has been crosslinked results in the purification of C5 protein in a single step. The protein h...

  3. Separation of Proteins by Electrophoretic Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)

    邺韶骅; 刘铮; 丁富新; 袁乃驹


    A new kind of electrophoretic affinity chromatography (EAC) for bioseparation was proposed,Separation by EAC was conducted in a multicompartment electrolyzer in which the affinity gel media were packed in one of the central compartments.The presence of an electric field accelerated the migration of proteins inside the gel matrix during adsorption and descrption processes,This led to the increase of the overall speed of separation,The present study was focused on the effect of the strength of the electric field on adsorption and desorption processes.

  4. Course on Advanced Analytical Chemistry and Chromatography

    DEFF Research Database (Denmark)

    Andersen, Jens Enevold Thaulov; Fristrup, Peter; Nielsen, Kristian Fog


    Methods of analytical chemistry constitute an integral part of decision making in chemical research, and students must master a high degree of knowledge, in order to perform reliable analysis. At DTU departments of chemistry it was thus decided to develop a course that was attractive to master...... students of different direction of studies, to Ph.D. students and to professionals that need an update of their current state of skills and knowledge. A course of 10 ECTS points was devised with the purpose of introducing students to analytical chemistry and chromatography with the aim of including theory...

  5. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.


    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...... charged metal ions such as Fe3+, Ga3+, Al3+, Zr4+, and Ti4+ has made it possible to enrich phosphorylated peptides from peptide samples. However, the selectivity of most of the metal ions is limited, when working with highly complex samples, e.g., whole-cell extracts, resulting in contamination from...

  6. [Recent advances in capillary scale ion chromatography technology]. (United States)

    Yang, Bingcheng; Diao, Xuefang


    Ion chromatography (IC) has been a well-established technique for the analysis of ionic samples. The aqueous solution used for IC eluent is well suited for bioanalysis in relative to common liquid chromatography. This is especially true for capillary ion chromatography (CIC) due to its advantage of small sample needed. CIC is generally divided into three categories including open tubular, packed and monolithic. In this review, the recent progress of CIC is summarized based on the development of several key components associated with packed column-based system. The development of open tubular ion chromatography is also reviewed.

  7. Extension of the selection of protein chromatography and the rate model to affinity chromatography. (United States)

    Sandoval, G; Shene, C; Andrews, B A; Asenjo, J A


    The rational selection of optimal protein purification sequences, as well as mathematical models that simulate and allow optimization of chromatographic protein purification processes have been developed for purification procedures such as ion-exchange, hydrophobic interaction and gel filtration chromatography. This paper investigates the extension of such analysis to affinity chromatography both in the selection of chromatographic processes and in the use of the rate model for mathematical modelling and simulation. Two affinity systems were used: Blue Sepharose and Protein A. The extension of the theory developed previously for ion-exchange and HIC chromatography to affinity separations is analyzed in this paper. For the selection of operations two algorithms are used. In the first, the value of η, which corresponds to the efficiency (resolution) of the actual chromatography and, Σ, which determines the amount of a particular contaminant eliminated after each separation step, which determines the purity, have to be determined. It was found that the value of both these parameters is not generic for affinity separations but will depend on the type of affinity system used and will have to be determined on a case by case basis. With Blue Sepharose a salt gradient was used and with Protein A, a pH gradient. Parameters were determined with individual proteins and simulations of the protein mixtures were done. This approach allows investigation of chromatographic protein purification in a holistic manner that includes ion-exchange, HIC, gel filtration and affinity separations for the first time.

  8. The weight of flash chromatography: A tool to predict its mass intensity from thin-layer chromatography

    Directory of Open Access Journals (Sweden)

    Freddy Pessel


    Full Text Available Purification by flash chromatography strongly impacts the greenness of a process. Unfortunately, due to the lack of the relevant literature data, very often this impact cannot be assessed thus preventing the comparison of the environmental factors affecting the syntheses. We developed a simple mathematical approach to evaluate the minimum mass intensity of flash chromatography from the retention factor values determined by thin-layer chromatography.

  9. Experimental design in chromatography: a tutorial review. (United States)

    Hibbert, D Brynn


    The ability of a chromatographic method to successful separate, identify and quantitate species is determined by many factors, many of which are in the control of the experimenter. When attempting to discover the important factors and then optimise a response by tuning these factors, experimental design (design of experiments, DoE) gives a powerful suite of statistical methodology. Advantages include modelling by empirical functions, not requiring detailed knowledge of the underlying physico-chemical properties of the system, a defined number of experiments to be performed, and available software to accomplish the task. Two uses of DoE in chromatography are for showing lack of significant effects in robustness studies for method validation, and for identifying significant factors and then optimising a response with respect to them in method development. Plackett-Burman designs are widely used in validation studies, and fractional factorial designs and their extensions such as central composite designs are the most popular optimisers. Box-Behnken and Doehlert designs are becoming more used as efficient alternatives. If it is not possible to practically realise values of the factors required by experimental designs, or if there is a constraint on the total number of experiments that can be done, then D-optimal designs can be very powerful. Examples of the use of DoE in chromatography are reviewed. Recommendations are given on how to report DoE studies in the literature.

  10. Simulating retention in gas-liquid chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Martin, M.G.; Siepmann, J.I.; Schure, M.R.


    Accurate predictions of retention times, retention indices, and partition constants are a long sought-after goal for theoretical studies in chromatography. Configurational-bias Monte Carlo (CBMC) simulations in the Gibbs ensemble using the transferable potentials for phase equilibria-united atom (TraPPE-UA) force field have been carried out to obtain a microscopic picture of the partitioning of 10 alkane isomers between a helium vapor phase and a squalane liquid phase, a protypical gas-liquid chromatography system. The alkane solutes include some topological isomers that differ only in the arrangement of their building blocks (e.g., 2,5-dimethylhexane and 3,4-dimethylhexane), for which the prediction of the retention order is particularly difficult. The Kovats retention indices, a measure of the relative retention times, are calculated directly from the partition constants and are in good agreement with experimental values. The calculated Gibbs free energies of transfer for the normal alkanes conform to Martin's equation which is the basis of linear free energy relationships used in many process modeling packages. Analysis of radial distribution functions and the corresponding energy integrals does not yield evidence for specific retention structures and shows that the internal energy of solvation is not the main driving force for the separation of topological isomers in this system.

  11. Analysis of Tocopherols by High Performance Liquid Chromatography


    Edison, B.


    : Gas chromatography is the key technique for organic components and also for tocopherols analysis. High performance liquid chromatography has an important role to take part in applications such as the handling of less usual samples, prevention of degradation of heat sensitive functional groups and for micro preparative purposes. Many approaches for development of improved methods are suggested, especially for reversed phase applications.

  12. Forensic potential of comprehensive two-dimensional gas chromatography

    NARCIS (Netherlands)

    Sampat, A.; Lopatka, M.; Sjerps, M.; Vivo-Truyols, G.; Schoenmakers, P.; van Asten, A.


    In this study, the application of comprehensive two-dimensional (2D) gas chromatography (GC × GC) in forensic science is reviewed. The peer-reviewed publications on the forensic use of GC × GC and 2D gas chromatography with mass spectrometric detection (GC × GC-MS) have been studied in detail, not o

  13. Thin-Layer Chromatography: Four Simple Activities for Undergraduate Students. (United States)

    Anwar, Jamil; And Others


    Presents activities that can be used to introduce thin-layer chromatography at the undergraduate level in relatively less developed countries and that can be performed with very simple and commonly available apparati in high schools and colleges. Activities include thin-layer chromatography with a test-tube, capillary feeder, burette, and rotating…

  14. Using Ion Exchange Chromatography to Separate and Quantify Complex Ions (United States)

    Johnson, Brian J.


    Ion exchange chromatography is an important technique in the separation of charged species, particularly in biological, inorganic, and environmental samples. In this experiment, students are supplied with a mixture of two substitution-inert complex ions. They separate the complexes by ion exchange chromatography using a "flash"…

  15. Fast centrifugal partition chromatography as a preparative-scale separation technique for citrus flavones (United States)

    Fast centrifugal partition chromatography (FCPC) is a preparative-scale separations methodology based on the principles of counter current chromatography. Separations by FCPC are typically achieved with higher recoveries and with lower solvent use compared to conventional column chromatography. HSCP...

  16. Using ion exchange chromatography to purify a recombinantly expressed protein. (United States)

    Duong-Ly, Krisna C; Gabelli, Sandra B


    Ion exchange chromatography (IEX) separates molecules by their surface charge, a property that can vary vastly between different proteins. There are two types of IEX, cation exhange and anion exchange chromatography. The protocol that follows was designed by the authors for anion exchange chromatography of a recombinantly expressed protein having a pI of 4.9 and containing two cysteine residues and one tryptophan residue, using an FPLC system. Prior to anion exchange, the protein had been salted out using ammonium sulfate precipitation and partially purified via hydrophobic interaction chromatography (see Salting out of proteins using ammonium sulfate precipitation and Use and Application of Hydrophobic Interaction Chromatography for Protein Purification). Slight modifications to this protocol may be made to accommodate both the protein of interest and the availability of equipment.

  17. Chitosan-based membrane chromatography for protein adsorption and separation. (United States)

    Liu, Yezhuo; Feng, Zhicheng; Shao, Zhengzhong; Chen, Xin


    A chitosan-based membrane chromatography was set up by using natural chitosan/carboxymethylchitosan (CS/CMCS) blend membrane as the matrix. The dynamic adsorption property for protein (lysozyme as model protein) was detailed discussed with the change in pore size of the membrane, the flow rate and the initial concentration of the feed solution, and the layer of membrane in membrane stack. The best dynamic adsorption capacity of lysozyme on the CS/CMCS membrane chromatography was found to be 15.3mg/mL under the optimal flow conditions. Moreover, the CS/CMCS membrane chromatography exhibited good repeatability and reusability with the desorption efficiency of ~90%. As an application, lysozyme and ovalbumin were successfully separated from their binary mixture through the CS/CMCS membrane chromatography. This implies that such a natural chitosan-based membrane chromatography may have great potential on the bioseparation field in the future.

  18. Applications of resistive heating in gas chromatography: a review. (United States)

    Jacobs, Matthew R; Hilder, Emily F; Shellie, Robert A


    Gas chromatography is widely applied to separate, identify, and quantify components of samples in a timely manner. Increasing demand for analytical throughput, instrument portability, environmental sustainability, and more economical analysis necessitates the development of new gas chromatography instrumentation. The applications of resistive column heating technologies have been espoused for nearly thirty years and resistively heated gas chromatography has been commercially available for the last ten years. Despite this lengthy period of existence, resistively heated gas chromatography has not been universally adopted. This low rate of adoption may be partially ascribed to the saturation of the market with older convection oven technology, coupled with other analytical challenges such as sampling, injection, detection and data processing occupying research. This article assesses the advantages and applications of resistive heating in gas chromatography and discusses practical considerations associated with adoption of this technology.

  19. Phytochemical analysis of ethanolic extract of Dichrostachys Cinerea W and Arn leaves by a thin layer chromatography, high performance thin layer chromatography and column chromatography


    Vijayalakshmi, M; K Periyanayagam; Kavitha, K; K Akilandeshwari


    Background: The leaves of Dichrostachys cinerea are used as laxative, diuretic, painkiller. It is also used in the treatment of gonorrhoea, boils, oedema, gout, veneral diseases and nasopharyngeal affections, etc. Materials and Methods: The Phytochemical investigation of ethanolic extract of D. cinerea leaves were performed by standard chemical tests, thin layer chromatography (TLC) by using various solvent systems, and by high performance liquid chromatography (HPTLC). Two compounds were...

  20. A Review on Ultra Performance Liquid Chromatography

    Directory of Open Access Journals (Sweden)

    Darasanapalli Meenakshi


    Full Text Available UPLC is a rising chromatographic separation technique whose packing materials have smaller particle size lesser than 2.5μm which improves the speed, resolution and sensitivity of analysis. When many scientists experienced separation barriers with conventional HPLC, UPLC extended and expanded the utility of chromatography. The main advantage is a reduction of analysis time which also reduces solvent consumption. The analysis time, solvent consumption and analysis cost are very important factor in many analytical laboratories. The time spent for optimizing new methods can also be greatly reduced. This results in many analysis in a day and quick results which is of very importance to the industries and research laboratories. UPLC principle is same as that of HPLC that is based on Van Deemter equation but decrease in particle size has increased efficiency at increased flow rates.

  1. A novel amide stationary phase for hydrophilic interaction liquid chromatography and ion chromatography. (United States)

    Shen, Guobin; Zhang, Feifang; Yang, Bingcheng; Chu, Changhu; Liang, Xinmiao


    A novel amide stationary phase (ASP) for hydrophilic interaction liquid chromatography (HILIC) has been prepared via the Click chemistry method. It was based on the strategy that the amino group of Asparagine was easily transferred to the corresponding azido group and then clicked onto terminal alkyne-silica gel in the presence of Cu(I)-based catalyst. For the tested polar compounds including nucleosides and nucleic acid bases, ASP-based column has demonstrated good performance in terms of separation efficiency and column stability, and the retention mechanism was found to match well the typical HILIC retention. In addition, the ASP described here showed much better selectivity in separation of inorganic anions under ion chromatography mode relative to other kinds of commercial ASP.

  2. RSC Chromatography Monographs Quantitative In Silico Chromatography Computational Modelling of Molecular Interactions. (United States)

    Hanai, Toshihiko


    All early chromatographic techniques, starting from the primitive "ancient" chromatography introduced by Tswett in the very early twentieth century, perfected in partition chromatography in the 1940s by Martin and Synge, and extended to a variety of additional separation mechanisms later, were first entirely experimental trial-and-error methods. The early years can also be characterized by searching for theoretical base of various separation techniques that would allow establishing relation between the structure of the analytes and their chromatographic behavior. The advent of computers followed by development of the new software then revolutionized the theoretical approaches and enabled detailed modeling instead of tedious experimentation. This book introduces the readers to the era of computational modeling in which molecular interactions are used to analyze the mechanisms of general molecular interactions with a special focus on biological applications. This article is protected by copyright. All rights reserved.

  3. Peak broadening in paper chromatography and related techniques : IV. The mechanism of the mass transfer term in paper chromatography

    NARCIS (Netherlands)

    Ligny, C.L. de; Remijnse, A.G.


    The mechanism of peak broadening in paper chromatography is investigated by comparing the peak widths obtained in chromatography with those caused only by diffusion, for a set of amino acids of widely differing RF values and with eight kinds of Whatman paper. The results show that longitudinal diff

  4. Peak broadening in paper chromatography and related techniques : III. Peak broadening in thin-layer chromatography on cellulose powder

    NARCIS (Netherlands)

    Ligny, C.L. de; Remijnse, A.G.


    The mechanism of peak broadening in thin-layer chromatography on cellulose powder was investigated by comparing the peak widths obtained in chromatography with those caused only by diffusion in the cellulose powder, for a set of amino acids of widely differing RF values and six kinds of cellulose po

  5. Multifraction separation in countercurrent chromatography (MCSGP). (United States)

    Krättli, Martin; Müller-Späth, Thomas; Morbidelli, Massimo


    The multicolumn countercurrent solvent gradient purification (MCSGP) process is a continuous countercurrent multicolumn chromatography process capable of performing three fraction separations while applying a linear gradient of some modifier. This process can then be used either for the purification of a single species from a multicomponent mixture or to separate a three component mixture in one single operation. In this work, this process is extended to the separation of multifractions, in principle with no limitation. To achieve this goal the MCSGP standard process is extended by introducing one extra separation section per extra fraction to be isolated. Such an extra separation section is realized in this work through a single additional column, so that a n fraction MCSGP process can be realized using a minimum of n columns. Two separation processes were considered to experimentally demonstrate the possibility of realizing a four-fraction MCSGP unit able to purify two intermediate products in a given multicomponent mixture. The first one was a model mixture containing four different proteins. The two proteins eluting in the center of the chromatogram were purified with yields equal to 95% for the early eluting and 92% for the later eluting one. The corresponding purities were 94% and 97%, respectively. Such performance was well superior to that of the batch operation with the same modifier gradient which for the same purity values could not achieve yields larger than 67% and 81%, respectively. Similar performance improvements were found for the second separation where two out of seven charge variants which constitute the mAb Cetuximab currently available on the market have been purified in one single operation using a four-fraction MCSGP unit. In this case, yields of 81% and 65% were obtained with purities of 90% and 89%, respectively. These data compare well with the corresponding data from batch chromatography where with the same gradient and for the same

  6. Simultaneous analysis for water- and fat-soluble vitamins by a novel single chromatography technique unifying supercritical fluid chromatography and liquid chromatography. (United States)

    Taguchi, Kaori; Fukusaki, Eiichiro; Bamba, Takeshi


    Chromatography techniques usually use a single state in the mobile phase, such as liquid, gas, or supercritical fluid. Chromatographers manage one of these techniques for their purpose but are sometimes required to use multiple methods, or even worse, multiple techniques when the target compounds have a wide range of chemical properties. To overcome this challenge, we developed a single method covering a diverse compound range by means of a "unified" chromatography which completely bridges supercritical fluid chromatography and liquid chromatography. In our method, the phase state was continuously changed in the following order; supercritical, subcritical and liquid. Moreover, the gradient of the mobile phase starting at almost 100% CO2 was replaced with 100% methanol at the end completely. As a result, this approach achieved further extension of the polarity range of the mobile phase in a single run, and successfully enabled the simultaneous analysis of fat- and water-soluble vitamins with a wide logP range of -2.11 to 10.12. Furthermore, the 17 vitamins were exceptionally separated in 4min. Our results indicated that the use of dense CO2 and the replacement of CO2 by methanol are practical approaches in unified chromatography covering diverse compounds. Additionally, this is a first report to apply the novel approach to unified chromatography, and can open another door for diverse compound analysis in a single chromatographic technique with single injection, single column and single system.

  7. Studies on Chromatography Fingerprint of Hongqi by High-performance Liquid Chromatography

    Institute of Scientific and Technical Information of China (English)

    Shi Lan FENG; Fang Di HU; Jian Xiong ZHAO; Jing Wen XU; Li Ren CHEN


    Chromatography fingerprint (CFP) of 10 samples of hongqi were studied. 23 common peaks were analyzed, their average similarity was 97.29%. CFP were positioned with main index composition such as formononetin, calycosin and then the contents of index composition were determined. The character and exclusive of CFP of 10 samples of hongqi were clear. CFP and content determination of index composition of hongqi could be used to evaluate the quality of hongqi comprehensively.

  8. Extraction of cocoa proanthocyanidins and their fractionation by sequential centrifugal partition chromatography and gel permeation chromatography. (United States)

    Pedan, Vasilisa; Fischer, Norbert; Rohn, Sascha


    Cocoa beans contain secondary metabolites ranging from simple alkaloids to complex polyphenols with most of them believed to possess significant health benefits. The increasing interest in these health effects has prompted the need to develop techniques for their extraction, fractionation, separation, and analysis. This work provides an update on analytical procedures with a focus on establishing a gentle extraction technique. Cocoa beans were finely ground to an average particle size of centrifugal partition chromatography (SCPC) and gel permeation column chromatography using Sephadex LH-20. For SCPC, a two-phase solvent system consisting of ethyl acetate/n-butanol/water (4:1:5, v/v/v) was successfully applied for the separation of theobromine, caffeine, and representatives of the two main phenolic compound classes flavan-3-ols and flavonols. Gel permeation chromatography on Sephadex LH-20 using a stepwise elution sequence with aqueous acetone has been shown for effectively separating individual flavan-3-ols. Separation was obtained for (-)-epicatechin, proanthocyanidin dimer B2, trimer C1, and tetramer cinnamtannin A2. The purity of alkaloids and phenolic compounds was determined by HPLC analysis and their chemical identity was confirmed by mass spectrometry.

  9. Instrumentation for hand-portable liquid chromatography. (United States)

    Sharma, Sonika; Plistil, Alex; Simpson, Robert S; Liu, Kun; Farnsworth, Paul B; Stearns, Stanley D; Lee, Milton L


    Liquid chromatography (LC) has lagged behind gas chromatography (GC) in developments related to hand-portable instrumentation. In this work, a new battery-operated (24V DC) nano-flow pumping system with a stop-flow injector was developed and integrated with an on-column UV-absorption detector (254nm) that was reduced in size to an acceptable weight and power usage for field operation. The pumping system, which includes nano-flow pump, stepper motor and high-pressure valve weighs only 1.372kg (3lbs) and can generate up to 110.32MPa (16,000psi) pressure. A major advantage of this pump is that it does not employ a splitter, since it was specifically designed for capillary column use. The volume capacity of the pump is 24μL, and a sample volume as low as 10nL can be injected. Flow rate calibration (300nL to 6.12μL per min) was performed, and an accuracy >99.94% was obtained. The percent injection carry-over was found to be low (RSD 0.31%), which makes it practical for quantitative analysis. The detector linear range and limit of detection (LOD) were determined using sodium anthraquinone-2-sulfonate. A linear regression coefficient (R) of 0.9996 was obtained for a plot of log peak area versus log concentration over the range of 3.2μM to 6.5mM, and the LOD (S/N=3) was found to be 7.8fmol (0.13μM). The short term noise of the detector is comparable to commercially available detectors (∼10(-5)AU). In this work, the system was tested in the laboratory using regular line power (120V AC) with an AC to DC adapter. Reversed-phase isocratic separations were performed using a 15.5cm×75μm i.d. fused silica capillary column containing a monolithic stationary phase synthesized from 1,6-hexanediol dimethacrylate. Good retention time repeatability (RSD 0.09-0.74%) was obtained for a mixture containing an unretained marker (i.e., uracil) and a homologous series of alkyl benzenes.

  10. Supercritical fluid chromatography (SFC). Pt. 3. Applications

    Energy Technology Data Exchange (ETDEWEB)

    Leyendecker, D.; Hoefler, F.


    During the last few years, Supercritical Fluid Chromatography (SFC) has been emerged from an academic object of research to an important tool for the analytical chemist. Persuasive SFC-applications have been reported analyzing substrates with low volatilities, either due to high molecular weights or to high polarities, thermally or solvatolytically labile molecules, or analytes without chromophores or electroactive groups. Examples are oligomers and polymers up to a molecular weight of 10,000 g/mol, e.g., polysiloxanes, polyethers (nonionic surfactants), polyesters, polystyrenes and other vinyl polymers, polyenes etc. In addition, SFC is applicable to the analysis of several monomers. In the petroleum and coal industries, SFC was employed for the characterization of high boiling hydrocarbon mixtures as well as for group separations of alkanes, olefins, and aromatics. Due to the high solvent strength of supercritical fluids, several pesticides, herbicides, and fungicides can be determined either separately or in combination with polar metabolites. Physiologically active substances often are thermally labile and polar at the same time. SFC has been successfully applied to the analysis of steroids, prostaglandins, cannabinoids and other drugs, flavours, and medicines. It is also well suited for analyzing natural or synthetic mono-, di-, and triglycerides. (orig.).

  11. Gas chromatography using resistive heating technology. (United States)

    Wang, Anzi; Tolley, H Dennis; Lee, Milton L


    Air bath ovens are standard in conventional gas chromatography (GC) instruments because of their simplicity and reliability for column temperature control. However, their low heating rates, high power consumption and bulky size are in conflict with the increasing demands for fast separation and portable instrumentation. The deficiencies of air bath ovens can be eliminated using resistive heating technology, as the column is conductively heated by compact resistive heaters with low thermal mass. Resistive heating methods were employed in the early years of GC history, and they are emerging again as instrumentation is becoming more compact and sophisticated. Numerous designs have been tested and some have been successfully commercialized. Development of portable GC systems, including lab-on-a-chip devices, greatly benefits from the use of small, low-power resistive heating hardware. High speed GC separations using conventional instruments also can be best achieved with resistive heating modules. Despite some of its own inherent disadvantages, including efficiency loss, complex manufacturing and inconvenient column maintenance, resistive heating is expected to rapidly become a mature technology and even replace oven heating in the not-to-distant future.

  12. Optimizing heterosurface adsorbent synthesis for liquid chromatography (United States)

    Bogoslovskii, S. Yu.; Serdan, A. A.


    The structural and geometric parameters of a silica matrix (SM) for the synthesis of heterosurface adsorbents (HAs) are optimized. Modification is performed by shielding the external surfaces of alkyl-modified silica (AS) using human serum albumin and its subsequent crosslinking. The structural and geometric characteristics of the SM, AS, and HA are measured via low-temperature nitrogen adsorption. It is found that the structural characteristics of AS pores with diameters D 9 nm reduces significantly due to adsorption of albumin. It is concluded that silica gel with a maximum pore size distribution close to 5 nm and a minimal proportion of pores with D > 9 nm is optimal for HA synthesis; this allows us to achieve the greatest similarity between the chromatographic retention parameters for HA and AS. The suitability of the synthesized adsorbents for analyzing drugs in biological fluids through direct sample injection is confirmed by chromatography. It was found that the percentage of the protein fraction detected at the outlet of the chromatographic column is 98%.

  13. Vacuum ultraviolet detector for gas chromatography. (United States)

    Schug, Kevin A; Sawicki, Ian; Carlton, Doug D; Fan, Hui; McNair, Harold M; Nimmo, John P; Kroll, Peter; Smuts, Jonathan; Walsh, Phillip; Harrison, Dale


    Analytical performance characteristics of a new vacuum ultraviolet (VUV) detector for gas chromatography (GC) are reported. GC-VUV was applied to hydrocarbons, fixed gases, polyaromatic hydrocarbons, fatty acids, pesticides, drugs, and estrogens. Applications were chosen to feature the sensitivity and universal detection capabilities of the VUV detector, especially for cases where mass spectrometry performance has been limited. Virtually all chemical species absorb and have unique gas phase absorption cross sections in the approximately 120-240 nm wavelength range monitored. Spectra are presented, along with the ability to use software for deconvolution of overlapping signals. Some comparisons with experimental synchrotron data and computed theoretical spectra show good agreement, although more work is needed on appropriate computational methods to match the simultaneous broadband electronic and vibronic excitation initiated by the deuterium lamp. Quantitative analysis is governed by Beer-Lambert Law relationships. Mass on-column detection limits reported for representatives of different classes of analytes ranged from 15 (benzene) to 246 pg (water). Linear range measured at peak absorption for benzene was 3-4 orders of magnitude. Importantly, where absorption cross sections are known for analytes, the VUV detector is capable of absolute determination (without calibration) of the number of molecules present in the flow cell in the absence of chemical interferences. This study sets the stage for application of GC-VUV technology across a wide breadth of research areas.

  14. High Performance Liquid Chromatography of Vitamin A: A Quantitative Determination. (United States)

    Bohman, Ove; And Others


    Experimental procedures are provided for the quantitative determination of Vitamin A (retinol) in food products by analytical liquid chromatography. Standard addition and calibration curve extraction methods are outlined. (SK)

  15. A generalized theory of chromatography and multistep liquid extraction (United States)

    Chizhkov, V. P.; Boitsov, V. N.


    A generalized theory of chromatography and multistep liquid extraction is developed. The principles of highly efficient processes for fine preparative separation of binary mixture components on a fixed sorbent layer are discussed.

  16. Separation of neutral compounds by microemulsion electrokinetic chromatography

    DEFF Research Database (Denmark)

    Gabel-Jensen, Charlotte; Hansen, Steen Honore'; Pedersen-Bjergaard, S


    The selectivity of microemulsion electrokinetic chromatography (MEEKC) was studied utilizing some uncharged model compounds like aromatic amides, steroids, and esters of nicotinic acid. The cosurfactant of the microemulsion was found to be the most important factor affecting the selectivity...

  17. An Empirical Formula From Ion Exchange Chromatography and Colorimetry. (United States)

    Johnson, Steven D.


    Presents a detailed procedure for finding an empirical formula from ion exchange chromatography and colorimetry. Introduces students to more varied techniques including volumetric manipulation, titration, ion-exchange, preparation of a calibration curve, and the use of colorimetry. (JRH)

  18. Ion Chromatography: An Account of Its Conception and Early Development (United States)

    Small, Hamish


    The conception of ion chromatography and its development into a technique ready for commercialization is described. The pioneering development pointed the way to make ion exclusion an important member of the repertoire of IC methods.

  19. Detection of Two Phosphorus Containing Anacardic Compounds by Paper Chromatography

    Directory of Open Access Journals (Sweden)

    K. Sridharan


    Full Text Available A method based on paper chromatography for detection and determination of tris-tetrahydroanacardoxy phosphite and tris-(2 terat-hydroanacardoxyethyl-phosphite. The Rf values for these compounds in four solvent systems are recorded.

  20. Use of Chromatography Techniques to Separate a Mixture of Substances (United States)

    Donaldson, W.


    Explains the separation of the constituents of mixtures on one piece of chromatography paper. The example presented involves a vitamin C tablet, a disprin tablet, and a glucose tablet. Outlined are two methods for separating the constituents. (GS)

  1. Measurement of Orotic Acid in Urine by Supercritical Fluid Chromatography

    Institute of Scientific and Technical Information of China (English)


    This work presents a simple, rapid and reliable supercritical fluid chromatography (SFC) method for a sensitive measurement of orotic acid in human urine. The samples were diluted with deionized water and analyzed directly without any pretreatment.

  2. Mixed Stationary Liquid Phases for Gas-Liquid Chromatography. (United States)

    Koury, Albert M.; Parcher, Jon F.


    Describes a laboratory technique for use in an undergraduate instrumental analysis course that, using the interpretation of window diagrams, prepares a mixed liquid phase column for gas-liquid chromatography. A detailed procedure is provided. (BT)

  3. Analysis of Tocopherols by High Performance Liquid Chromatography

    Directory of Open Access Journals (Sweden)

    B. Edison


    Full Text Available : Gas chromatography is the key technique for organic components and also for tocopherols analysis. High performance liquid chromatography has an important role to take part in applications such as the handling of less usual samples, prevention of degradation of heat sensitive functional groups and for micro preparative purposes. Many approaches for development of improved methods are suggested, especially for reversed phase applications.

  4. Measurements of Protein-Protein Interactions by Size Exclusion Chromatography


    Bloustine, J.; Berejnov, V.; Fraden, S


    A method is presented for determining second virial coefficients (B2) of protein solutions from retention time measurements in size exclusion chromatography. We determine B2 by analyzing the concentration dependence of the chromatographic partition coefficient. We show the ability of this method to track the evolution of B2 from positive to negative values in lysozyme and bovine serum albumin solutions. Our size exclusion chromatography results agree quantitatively with data obtained by light...

  5. Optimized Separation of Nine Xanthones by Microemulsion Electrokinetic Chromatography

    Institute of Scientific and Technical Information of China (English)


    A microemulsion electrokinetic chromatography method has been firstly used for the separation of the therapeutically important xanthones from Securidaca inappendiculata. The separation of the nine xanthones was systematically optimized with respect to pH, composition of microemulsion, addition of cyclodextrins, applied voltage and column temperature. Baseline separation was successfully achieved for the nine xanthones, which was also compared with that by micellar electrokinetic chromatography.

  6. Discovery and early development of non-suppressed ion chromatography. (United States)

    Fritz, James S; Gjerde, Douglas T


    This year marks the 30th anniversary of the publication of Non-Suppressed Ion Chromatography, which is a method for the rapid separation of anions with on-line conductimetric detection. In this method, the separation column is connected directly to the conductimetric detector. This single-column method is a simpler technique than the original suppressed ion chromatography method, which requires a large suppressor column to reduce the background conductance. In the new method, the background signal is reduced to a manageable level simply by using an ion-exchange separation column of low exchange capacity that lowers the eluent concentration needed for separation. The eluent ion used for separation is chosen based on having large, bulky structure, which lowers the equivalent conductance and facilitates detection of the sample anions. This is a personal account of the initial discovery and early development of non-suppressed ion chromatography. The circumstances for the discovery are recounted by the two authors. Methods are described for determination of anions, cations with indirect detection, and techniques for increasing detection sensitivity. A fundamental equation for the prediction of ion chromatography detector response is given, and the development of several types of detection schemes for ion chromatography is discussed. Finally, the impact of non-suppressed ion chromatography is discussed together with comments on the discovery process.

  7. Towards early detection of the hydrolytic degradation of poly(bisphenol A)carbonate by hyphenated liquid chromatography and comprehensive two-dimensional liquid chromatography

    NARCIS (Netherlands)

    Coulier, L.; Kaal, E.R.; Hankemeier, Th.


    The hydrolytic degradation of poly(bisphenol A)carbonate (PC) has been characterized by various liquid chromatography techniques. Size exclusion chromatography (SEC) showed a significant decrease in molecular mass as a result of hydrolytic degradation, while 'liquid chromatography at critical condit

  8. Characterization of synthetic dyes by comprehensive two-dimensional liquid chromatography combining ion-exchange chromatography and fast ion-pair reversed-phase chromatography. (United States)

    Pirok, Bob W J; Knip, Jitske; van Bommel, Maarten R; Schoenmakers, Peter J


    In the late 19th century, newly invented synthetic dyes rapidly replaced the natural dyes on the market. The characterization of mixtures of these so-called early synthetic dyes is complicated through the occurrence of many impurities and degradation products. Conventional one-dimensional liquid chromatography does not suffice to obtain fingerprints with sufficient resolution and baseline integrity. Comprehensive two-dimensional liquid chromatography (LC×LC) is employed in this study, with ion-exchange chromatography in the first dimension and fast ion-pair liquid chromatography in the second. Retention in the first dimension is largely determined by the number of charges, while the selection of a small ion-pair reagent (tetramethylammonium hydroxide) in the second dimension causes retention to be largely determined by the molecular structure of the dye. As a result, there is a high degree of orthogonality of the two dimensions, similar to the values typically encountered in GC×GC. The proposed LC×LC method shows a theroretical peak capacity of about 2000 in an analysis time of about three hours. Clear, informative fingerprints are obtained that open a way to a more efficient characterization of dyes used in objects of cultural heritage.

  9. Simultaneous determination of cyclodol and diprazin by thin layer chromatography and high performance liquid chromatography. (United States)

    Makharadze, R; Adeishvili, L; Chelidze, T; Imnadze, N; Nizharadze, N


    Ciklodol (trihexyphenidil)--the central and peripheral m-cholinoblocker is currently used with other antipsychotic drugs such as phenotiazines and tricycle antidepressants. For the purpose of simultaneous determination of ciklodol and diprazine, were selected two methods of analysis: Thin Layer Chromatography (TLC) and High Performance Liquid Chromatography (HPLC). During development of TLC method was studied the 10 visualizing system and 24 mobile systems. For individual or simultaneous determination of ciklodol and diprazine were recommended the following solvents' systems: 1. Toluene-acetone-ethanole-25%NH(4)OH (45:45: 7.5:2.5), 2. Hexane-ethyl acetate (15:5), 3. Chloroform-heptene-25%NH(4)OH (16:3:3), 4. Ethylacetate-hexane (10:10), 5. Acetonitrile-metanol (10:10) and 6.Heptene-chloroform-ethanol-25% NH(4)OH (5:10:3:1). As visualizing systems were chosen: Iodine vapors, blacklight (UV254) and reagent of FNP. Reagent of FNP gives colored spot just with diprazine and it is also could be used for separation of both objects in simultaneous analysis. Developed HPLC method of simultaneous determination of ciklodol and diprazine: like mobile phase is recommended: Acetonitril- 0.05M KH(2)PO4 (55:45) (v/v) +H(3)PO(4) (pH3.5), column EC250 x 4.6mm, with solid phase Nucleosil, flow rate 1ml/min, sample volume 40 microl. In given conditions, the retention time of ciklodol is 6.005min and diprazine 7.227min. Developed method of simultaneous determination and separation of ciklodol and diprazine in respective mixtures could be successfully applied as in the pharmaceutical, as well in the chemical-toxicological laboratories.

  10. Hybrid method of solution applied to simulation of pulse chromatography

    Directory of Open Access Journals (Sweden)

    M. A. Cremasco


    Full Text Available In this communication, the method proposed by Cremasco et al. (2003 is applied to predict single and low concentration pulse chromatography. In previous work, a general rate model was presented to describe the breakthrough curve, where a hybrid solution was proposed for the linear adsorption. The liquid phase concentration inside the particle was found analytically and related with the bed liquid phase through Duhamel's Theorem, while the bulk-phase equation was solved by a numerical method. In this paper, this method is applied to describe pulse chromatography of solutes that present linear adsorption isotherms. The simulated results of pulse chromatography are compared with experimental ones for aromatic amino acid experiments from literature.

  11. Application of ion chromatography in clinical studies and pharmaceutical industry. (United States)

    Michalski, Rajmund


    Ion chromatography is a well-established regulatory method for analyzing anions and cations in environmental, food and many other samples. It offers an enormous range of possibilities for selecting stationary and mobile phases. Additionally, it usually helps to solve various separation problems, particularly when it is combined with different detection techniques. Ion chromatography can also be used to determine many ions and substances in clinical and pharmaceutical samples. It provides: availability of high capacity stationary phases and sensitive detectors; simple sample preparation; avoidance of hazardous chemicals; decreased sample volumes; flexible reaction options on a changing sample matrix to be analyzed; or the option to operate a fully-automated system. This paper provides a short review of the ion chromatography applications for determining different inorganic and organic substances in clinical and pharmaceutical samples.

  12. Analysis of protein composition using multidimensional chromatography and mass spectrometry. (United States)

    Link, Andrew J; Washburn, Michael P


    Multidimensional liquid chromatography of peptides produced by protease digestion of complex protein mixtures followed by tandem mass spectrometry can be coupled with automated database searching to identify large numbers of proteins in complex samples. These methods avoid the limitations of gel electrophoresis and in-gel digestions by directly identifying protein mixtures in solution. One method used extensively is named Multidimensional Protein Identification Technology (MudPIT), where reversed-phase chromatography and strong cation-exchange chromatography are coupled directly in a microcapillary column. This column is then placed in line between an HPLC and a mass spectrometer for complex mixture analysis. MudPIT remains a powerful approach for analyzing complex mixtures like whole proteomes and protein complexes. MudPIT is used for quantitative proteomic analysis of complex mixtures to generate novel biological insights.

  13. Ion-Exchange Chromatography: Basic Principles and Application. (United States)

    Cummins, Philip M; Rochfort, Keith D; O'Connor, Brendan F


    Ion-Exchange Chromatography (IEC) allows for the separation of ionizable molecules on the basis of differences in charge properties. Its large sample-handling capacity, broad applicability (particularly to proteins and enzymes), moderate cost, powerful resolving ability, and ease of scale-up and automation have led to it becoming one of the most versatile and widely used of all liquid chromatography (LC) techniques. In this chapter, we review the basic principles of IEC, as well as the broader criteria for selecting IEC conditions. By way of further illustration, we outline basic laboratory protocols to partially purify a soluble serine peptidase from bovine whole brain tissue, covering crude tissue extract preparation through to partial purification of the target enzyme using anion-exchange chromatography. Protocols for assaying total protein and enzyme activity in both pre- and post-IEC fractions are also described.

  14. Resolution of Quadruplex Polymorphism by Size Exclusion Chromatography (United States)

    Miller, M. Clarke; Trent, John O.


    This unit describes a method for the separation of quadruplex species formed from the same sequence via size exclusion chromatography (SEC). Polymorphism is inherent to quadruplex formation, and even relatively simple quadruplex forming sequences, such as the human telomere sequence d(GGG(TTAGGG)3), can form a myriad of possible configurations. High Performance Liquid Chromatography (HPLC), especially reverse phase and anion exchange methods, has been a mainstay of nucleic acids research and purification for many decades. We have applied these methods for the separation of individual quadruplex species formed in a mixture from the same parent sequence. PMID:21638270

  15. Analysis of phenolic type antioxidants; Capillary gas chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Chopra, S.K. (Indian Inst. of Petroleum, Dehradun (India)); Kapoor, V.B. (Indian Inst. of Petroleum, Dehradun (India)); Vishnoi, S.C. (Indian Inst. of Petroleum, Dehradun (India)); Bhagat, S.D. (Indian Inst. of Petroleum, Dehradun (India))


    A simple gas chromatographic (GC) procedure has been developed to estimate the individual alkylated phenols used as antioxidants to improve the shelf life of fuels and lubricants. Preparative gas chromatography was applied for separation and collection in sufficient quantity of the isomers of tertiary butyl, octyl and dodecyl phenols prepared by catalytic alkylation of phenol with isobutylene or its oligomers. The separated fractions were characterised by Infra-red spectrometry (IR) and paper chromatography. Out of several GC columns studies, a high resolution capillary column of 100% Methyl, Silicone gum (SE-30) as stationary phase gave best results. Data generated on various packed and capillary columns are in good agreement. (orig.)

  16. Hypotonic elution, a new desorption principle in immunoadsorbent chromatography

    DEFF Research Database (Denmark)

    Danielsen, Erik Michael; Sjöström, H; Norén, O


    -phlorizin hydrolase (EC and maltase-glucoamylase (EC This elution method proved capable of achieving an acceptable yield (30-70%) while at the same time preserving the purified enzymes in an enzymically active state. It hereby offers a solution to the problem in immunoadsorbent chromatography...... of the enzymes but were considered unlikely on several grounds. Hypotonic elution in immunoadsorbent chromatography, therefore, may have a much broader range of applicability, and the method is recommended to be tried out by workers in other areas of protein chemistry....

  17. Deep eutectic solvents in countercurrent and centrifugal partition chromatography. (United States)

    Roehrer, Simon; Bezold, Franziska; García, Eva Marra; Minceva, Mirjana


    Deep eutectic solvents (DESs) were evaluated as solvents in centrifugal partition chromatography, a liquid-liquid chromatography separation technology. To this end, the partition coefficients of ten natural compounds of different hydrophobicity were determined in non-aqueous biphasic systems containing DES. The influence of the composition of DESs and the presence of water in the biphasic system on the partition coefficient were also examined. In addition, several process relevant physical properties of the biphasic system, such as the density and viscosity of the phases, were measured. A mixture of three to four hydrophobic compounds was successfully separated in a centrifugal partition extractor using a heptane/ethanol/DES biphasic system.

  18. Mathematical model of the dynamics of countercurrent chromatography

    Institute of Scientific and Technical Information of China (English)

    陆向红; 任其龙; 吴平东


    A mathematical model of the dynamic behavior of countercurrent chromatography was proposed, and the model parameters, including the partition coefficient, the axial dispersion coefficient, the intraparticle diffusion coefficient and the external mass ransfer coefficient were calculated by the method of chromatogram moment analysis. Comparison of the experimental chromatograms of caffeine and theophylline determined in this work with the simulated curves computed by the proposed model showed fairly good agreement. Further, the difference between the average identified the partition coefficients by chromatogram moment analysis and the experimental values was small also, and the relationship between the external mass transfer rate and the linear velocity was similar to that obtained with solid-liquid chromatography.

  19. Determination of phytate in high molecular weight, charged organic matrices by two-dimensional size exclusion-ion chromatography (United States)

    A two-dimensional chromatography method for analyzing anionic targets (specifically phytate) in complex matrices is described. Prior to quantification by anion exchange chromatography, the sample matrix was prepared by size exclusion chromatography, which removed the majority of matrix complexities....

  20. Analysis of ganciclovir and its related substances using high performance liquid chromatography and liquid chromatography-mass spectrometry methods

    Institute of Scientific and Technical Information of China (English)


    Objective High performance liquid chromatography(HPLC)and liquid chromatography-mass spectrometry(LC/MS)methods were developed for the determination of ganciclovir and its related substances.Methods A Hypersil ODS2 column(4.6 mm×250 mm,5 μm)was used with a mobile phase of 0.02 M potassium dihydrogen phosphate buffer(pH 6.0)-methanol(92∶8)at a flow rate of 1.0 mL/min,and UV detector set at 254 nm was used for monitoring the eluents.Results The method was simple,rapid,selective and capable of separating all r...

  1. Analysis of Free Fatty Acids on the Fingertips by High Performance Liquid Chromatography. (United States)


    This investigation studied the efficiency of high performance liquid chromatography in the determination of free fatty acids present on the...utilized to eliminate the microbial contamination. The high performance liquid chromatography provided excellent separation of skin fatty acids for

  2. Media selection in ion-exchange chromatography in a single microplate. (United States)

    Cabanne, Charlotte; Santarelli, Xavier


    High-throughput process development is more and more used in chromatography. Limitations are the tools provided by the manufacturers. Here, we describe a method to select chromatographic media for ion-exchange chromatography using a 96-well filter microplate.

  3. MEMS Liquid and Gas Chromatography for Miniaturized Planetary In Situ Instruments (United States)

    Kidd, R. D.; Bae, B.; Willis, P. A.; Noell, A. C.; Scianmarello, N.; Tai, Y.-C.


    Micro-Electro-Mechanical Systems (MEMS) technology to reduce the size, mass and power of the three classical chromatographic technologies: gas chromatography (GC), capillary electrophoresis (CE) and high performance liquid chromatography (HPLC).

  4. Liquid Chromatography Applied to Space System (United States)

    Poinot, Pauline; Chazalnoel, Pascale; Geffroy, Claude; Sternberg, Robert; Carbonnier, Benjamin

    , etc.) which are good examples for one of the two intrinsic features of life (i.e. complexity) would then be searched for. Although these methods are very promising as they have already demonstrated real benefits in terms of sensitivity towards specific compounds of middle/high molecular weight, they cannot be used to detect in one pot a wide range of biopolymer targets with very diverse nature, such as peptides or oligonucleotides. In this context, it would be interesting to develop a “micro-lab” equipped with a miniaturized HPLC-MS as the ones currently developed in the field of biological and medicinal sciences. The objective is to demonstrate unequivocally the presence or absence in space of a wide range of biopolymers thanks to a “one step one pot” instrumentation. We propose to demonstrate the feasibility and the validity of such a concept. For that, we optimize the chromatographic conditions and the mass spectrometer parameters to detect in the range of ppb, proteins and polypeptides biomarkers, while taking into account the space constraints. On a UPLC-HRMS (Q-Exactive and Qq-TOF), different stationary phases (laboratory-made or commercially available), different eluents, gradient flows, temperatures, pressures, and the use of a pre-concentration stage are tested. Dual detection (MS and diode array) is also considered. First experiments have highlighted the ability of such a technique to find ultra-traces level of organic matters under definite space constraints (elution flow, solvents, temperature...). This work is funded by the French Space Agency (CNES) References Glavin DP, Schubert Ml, Botta O, Kminek G, Bada JL (2001) Detecting pyrolysis products from bacteria on Mars. Earth Planet Sc Lett 185:1-2. doi:10.1016/S0012-821X(00)00370-8 Navarro-González R, Navarro KF, de la Rosa J, Iñliguez E, Molina P, Mira LD (2006) The limitations on organic detection in Mars-like soils by thermal volatilization-gas chromatography-MS and their implications for the

  5. Mallow carotenoids determined by high-performance liquid chromatography (United States)

    Mallow (corchorus olitorius) is a green vegetable, which is widely consumed either fresh or dry by Middle East population. This study was carried out to determine the contents of major carotenoids quantitatively in mallow, by using a High Performance Liquid Chromatography (HPLC) equipped with a Bis...

  6. Determination of Caffeine in Beverages by High Performance Liquid Chromatography. (United States)

    DiNunzio, James E.


    Describes the equipment, procedures, and results for the determination of caffeine in beverages by high performance liquid chromatography. The method is simple, fast, accurate, and, because sample preparation is minimal, it is well suited for use in a teaching laboratory. (JN)

  7. Multichannel Detection in High-Performance Liquid Chromatography. (United States)

    Miller, James C.; And Others


    A linear photodiode array is used as the photodetector element in a new ultraviolet-visible detection system for high-performance liquid chromatography (HPLC). Using a computer network, the system processes eight different chromatographic signals simultaneously in real-time and acquires spectra manually/automatically. Applications in fast HPLC…

  8. High-Performance Liquid Chromatography-Mass Spectrometry. (United States)

    Vestal, Marvin L.


    Reviews techniques for online coupling of high-performance liquid chromatography with mass spectrometry, emphasizing those suitable for application to nonvolatile samples. Also summarizes the present status, strengths, and weaknesses of various techniques and discusses potential applications of recently developed techniques for combined liquid…

  9. Ultra high performance liquid chromatography of seized drugs

    NARCIS (Netherlands)

    Lurie, I.S.


    The primary goal of this thesis is to investigate the use of ultra high performance liquid chromatography (UHPLC) for the analysis of seized drugs. This goal was largely achieved and significant progress was made in achieving improved separation and detection of drugs of forensic interest.

  10. High-performance liquid chromatography with packed microchips


    Ehlert, Steffen


    This work is explaining the importance of generating a densely packed chromatographic bed to achieve optimum separation efficiency in miniaturized liquid chromatography. The reduction of peak dispersion and enhancement of separation efficiency is the important problem in micro- and nano-separation that is to be solved to generate highly efficient chromatographic systems that can exploit all advantages of miniaturization. Therefore...

  11. A Size Exclusion Chromatography Laboratory with Unknowns for Introductory Students (United States)

    McIntee, Edward J.; Graham, Kate J.; Colosky, Edward C.; Jakubowski, Henry V.


    Size exclusion chromatography is an important technique in the separation of biological and polymeric samples by molecular weight. While a number of laboratory experiments have been published that use this technique for the purification of large molecules, this is the first report of an experiment that focuses on purifying an unknown small…

  12. Quantification of Tea Flavonoids by High Performance Liquid Chromatography (United States)

    Freeman, Jessica D.; Niemeyer, Emily D.


    We have developed a laboratory experiment that uses high performance liquid chromatography (HPLC) to quantify flavonoid levels in a variety of commercial teas. Specifically, this experiment analyzes a group of flavonoids known as catechins, plant-derived polyphenolic compounds commonly found in many foods and beverages, including green and black…

  13. Protein interactions in hydrophobic charge induction chromatography (HCIC). (United States)

    Ghose, Sanchayita; Hubbard, Brian; Cramer, Steven M


    A quantitative understanding of how proteins interact with hydrophobic charge induction chromatographic resins is provided. Selectivity on this mode of chromatography for monoclonal antibodies as compared to other model proteins is probed by means of a linear retention vs pH plot. The pH-dependent adsorption behavior on this mode of chromatography for a hydrophobic, charged solute is described by taking into account the equilibrium between a hydrophobic, charged solute and an ionizable, heterocyclic ligand. By analogy, an equation that is seen to adequately describe macromolecular retention under linear conditions over a range of pH is developed. A preparative, nonlinear isotherm that can capture both pH and salt concentration dependency for proteins is proposed by using an exponentially modified Langmuir isotherm model. This model is seen to successfully simulate adsorption isotherms for a variety of proteins over a range of pHs and mobile phase salt concentrations. Finally, the widely differing retention characteristics of two monoclonal antibodies are used to derive two different strategies for improving separations on this mode of chromatography. A better understanding of protein binding to this class of resins is seen as an important step to future exploitation of this mode of chromatography for industrial scale purification of proteins.

  14. Bioseparations using surfactant-aided size-exclusion chromatography

    NARCIS (Netherlands)

    Horneman, D.A.


    In most bioprocesses one or more chromatographic steps are used in the purification of the product. Size-exclusion chromatography (SEC) is often one of these chromatographic steps. It is based on the difference in size and shape of the components to be separated and is used for the separation of mol

  15. Advancing liquid chromatography- mass spectrometry based technologies for proteome research

    NARCIS (Netherlands)

    Boersema, P.J.


    In proteomics, high-tech nano-liquid chromatography (LC) and mass spectrometry (MS) instrumentation is used to routinely sequence proteins at a large scale. In this thesis, several technological developments are described to advance proteomics and their applicability is demonstrated in several diffe

  16. Characterisation of cholera toxin by liquid chromatography - Electrospray mass spectrometry

    NARCIS (Netherlands)

    Baar, B.L.M. van; Hulst, A.G.; Wils, E.R.J.


    Cholera toxin, one of the toxins that may be generated by various strains of the bacterium Vibrio cholerae, can be considered as a substance possibly used in biological warfare. The possibilities of characterising the toxin by liquid chromatography electrospray mass spectrometry (LC-ES-MS) were inve

  17. Isolation of Methoxyfuranocoumarins From Ammi majus by Centrifugal Partition Chromatography. (United States)

    Bartnik, Magdalena; Mazurek, Anna Katarzyna


    Pure methoxyfuranocoumarins were isolated from Ammi majus L. by use of low-pressure column chromatography (LPCC) followed by centrifugal partition chromatography (CPC). The concentrated petroleum ether extract from fruits of A. majus was fractionated on a silica gel column using a gradient of ethyl acetate in dichloromethane (0-80%, v/v). Coumarin-rich fractions were analyzed by thin-layer chromatography (TLC) and high-performance liquid chromatography with diode array detection (HPLC/DAD). Xanthotoxin (8-MOP) and isopimpinellin (isoP), structurally similar compounds, were isolated in one fraction (FR6). To avoid multistep and long-lasting TLC preparation, optimization of CPC conditions has been performed. In one run, an effective separation of 8-MOP and isoP was achieved. The two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (10 : 8 : 10 : 9; v/v) in an ascending mode (the aqueous phase was a stationary phase, and the organic phase was a mobile phase), with flow rate 3 mL/min and rotation speed 1,600 r.p.m., was used. The identification and high purities of isolated 8-MOP (98.7%) and isoP (100%) were confirmed by HPLC/DAD assay, when compared with standards. The developed CPC method could be applied to the effective isolation of 8-MOP and isoP from plant extracts. The high purity of obtained compounds makes possible further exploitation of these components in biological studies.

  18. Analysis of moniliformin in maize plants using hydrophilic interaction chromatography

    DEFF Research Database (Denmark)

    Sørensen, Jens Laurids; Nielsen, Kristian Fog; Thrane, Ulf


    A novel HPLC method was developed for detection of the Fusarium mycotoxin, moniliformin in whole maize plants. The method is based on hydrophilic interaction chromatography (HILIC) on a ZIC zwitterion column combined with diode array detection and negative electrospray mass spectrometry (ESI...

  19. Recent advances in polymer monoliths for ion-exchange chromatography. (United States)

    Nordborg, Anna; Hilder, Emily F


    The use of polymeric materials in ion-exchange chromatography applications is advantageous because of their typically high mechanical stability and tolerance of a wide range of pH conditions. The possibility of using polymeric monoliths in ion-exchange chromatography is therefore obvious and many of the same strategies developed for polymeric particles have been adapted for use with polymeric monoliths. In this review different strategies for the synthesis of polymeric monoliths with ion-exchange functionality are discussed. The incorporation of ion-exchange functionality by co-polymerization is included, as also are different post-polymerization alterations to the monolith surface such as grafting. The formulations and strategies presented include materials intended for use in analytical separations in ion-exchange chromatography, sample pre-treatment or enrichment applications, and materials for capillary electrochromatography. Finally, examples of the use of polymeric monoliths in ion-exchange chromatography applications are included with examples published in the years 2003 to 2008.

  20. Ion Exchange Chromatography and Spectrophotometry: An Introductory Undergraduate Laboratory Experiment. (United States)

    Foster, N.; And Others


    Describes an experiment in which students use ion exchange chromatography to separate a mixture of chloro complexes of transition metal ions and then use spectrophotometry to define qualitatively the efficiency of the ion exchange columns. Background information, materials needed, and procedures used are included. (JN)

  1. Analysis of essential oils by gas chromatography and mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Masada, Y.


    The book is in two parts: first part Essential Oil includes compositae; labiatae; verbenaceae; oleaceae; umbelliferae; myrtaceae; euphorbiaceae; rutaceae; geraniaceae; rosaceae; lauraceae; myristicaceae; anonaceae; santalaceae; moraceae; piperaceae; zingiberaceae; araceae; gramineae; and cupressaceae written in English and Japanese. Part two includes essential oil; gas chromatography, and mass spectrometry written in Japanese. (DP)

  2. Supercritical fluid chromatography for lipid analysis in foodstuffs. (United States)

    Donato, Paola; Inferrera, Veronica; Sciarrone, Danilo; Mondello, Luigi


    The task of lipid analysis has always challenged separation scientists, and new techniques in chromatography were often developed for the separation of lipids; however, no single technique or methodology is yet capable of affording a comprehensive screening of all lipid species and classes. This review acquaints the role of supercritical fluid chromatography within the field of lipid analysis, from the early developed capillary separations based on pure CO2 , to the most recent techniques employing packed columns under subcritical conditions, including the niche multidimensional techniques using supercritical fluids in at least one of the separation dimensions. A short history of supercritical fluid chromatography will be introduced first, from its early popularity in the late 1980s, to the sudden fall and oblivion until the last decade, experiencing a regain of interest within the chromatographic community. Afterwards, the subject of lipid nomenclature and classification will be briefly dealt with, before discussing the main applications of supercritical fluid chromatography for food analysis, according to the specific class of lipids.

  3. Environmental applications of ion chromatography in Eastern and Central Europe. (United States)

    Michalski, Rajmund


    Environmental analytics is one of the most important applications of ion chromatography. It includes determination of ions in water and wastewater as well as in gaseous and solid ones. Nowadays, ion chromatography has almost completely displaced the classical methods of ion determination in these areas. In spite of the fact that the ion chromatography has been officially present in the scientific world for 36 years, its role and popularity is highly diversified in various countries and regions of the world. In highly industrialized countries, it has been a reference method of water and wastewater analysis for years. In other parts of the world, it is not used and appreciated sufficiently despite its undeniable advantages. The following paper is a short overview of the most highly cited scientific and research institutions that conduct research in terms of environmental applications of ion chromatography in Eastern and Central Europe. Furthermore, the paper presents a list of a number of scientific papers referring to the discussed area, published in the years 1996-2009 in some of the most highly cited international scientific journals, and concerning publications of scientists from Eastern and Central Europe seen against the background of Europe and the world.

  4. How to Use Chromatography as a Science Teaching Aid. (United States)

    Ganis, Frank M.

    Presented are five procedures which permit the effective teaching of chromatography with equipment which is readily available, economical, and simple in design. The first procedure involves a study of solute partition in two immiscible solvents and of countercurrent distribution. The second illustrates the use of unidimensional ascending paper…

  5. Coupling of column liquid chromatography and Fourier transform infrared spectrometry

    NARCIS (Netherlands)

    Somsen, G.W; Gooijer, C; Velthorst, N.H; Brinkman, U.A Th


    This paper provides an extensive overview of the literature on the coupling of column liquid chromatography (LC) and Fourier transform infrared spectrometry (FT-IR). Flow-cell-based FT-IR detection and early solvent-elimination interfaces for LC-FT-IR are discussed in brief. A comprehensive descript

  6. Direct probing of chromatography columns by laser-induced fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    McGuffin, V.L.


    This report summarizes the progress and accomplishments of this research project from September 1, 1989 to February 28, 1993. During this period, we have accomplished all of the primary scientific objectives of the research proposal: (1) constructed and evaluated a laser-induced fluorescence detection system that allows direct examination of the chromatographic column, (2) examined nonequilibrium processes that occur upon solute injection and elution, (3) examined solute retention in liquid chromatography as a function of temperature and pressure, (4) examined solute zone dispersion in liquid chromatography as a function of temperature and pressure, and (5) developed appropriate theoretical models to describe these phenomena. In each of these studies, substantial knowledge has been gained of the fundamental processes that are responsible for chromatographic separations. In addition to these primary research objectives, we have made significant progress in three related areas: (1) examined pyrene as a fluorescent polarity probe insupercritical fluids and liquids as a function of temperature and pressure, (2) developed methods for the class-selective identification of polynuclear aromatic hydrocarbons in coal-derived fluids by microcolumn liquid chromatography with fluorescence quenching detection, and (3) developed methods for the determination of saturated and unsaturated (including omega-3) fatty acids in fish oil extracts by microcolumn liquid chromatography with laser-induced fluorescence detection. In these studies, the advanced separation and detection techniques developed in our laboratory are applied to practical problems of environmental and biomedical significance.

  7. Development of a thermal desorption modulator for gas chromatography

    NARCIS (Netherlands)

    Geus, H.J.; Boer, de J.; Brinkman, U.A.Th.


    The separation space in gas chromatography can be enhanced dramatically by the comprehensive coupling of two independent separation dimensions. An interface between the two columns must accumulate analytes eluting from the first dimension, focus them and at the appropriate moment transfer them to th

  8. Carbon nanotube based stationary phases for microchip chromatography

    DEFF Research Database (Denmark)

    Mogensen, Klaus Bo; Kutter, Jörg Peter


    The objective of this article is to provide an overview and critical evaluation of the use of carbon nanotubes and related carbon-based nanomaterials for microchip chromatography. The unique properties of carbon nanotubes, such as a very high surface area and intriguing adsorptive behaviour, have...

  9. Specialized Gas Chromatography--Mass Spectrometry Systems for Clinical Chemistry. (United States)

    Gochman, Nathan; And Others


    A discussion of the basic design and characteristics of gas chromatography-mass spectrometry systems used in clinical chemistry. A comparison of three specific systems: the Vitek Olfax IIA, Hewlett-Packard HP5992, and Du Pont DP-102 are included. (BB)

  10. Denatured Thermodynamics of Proteins in Weak Cation-exchange Chromatography

    Institute of Scientific and Technical Information of China (English)

    LI Rong; CHEN Guo-Liang


    The thermostability of some proteins in weak cation-exchange chromatography was investigated at 20-80 ℃. The results show that there is a fixed thermal denaturation transition temperature for each protein. The appearance of the thermal transition temperature indicates that the conformations of the proteins are destroyed seriously. The thermal behavior of the proteins in weak cation-exchange and hydrophobic interaction chromatographies were compared in a wide temperature range. It was found that the proteins have a higher thermostability in a weak cation-exchange chromatography system. The thermodynamic parameters(ΔH0, ΔS0) of those proteins were determined by means of Vant Hoff relationship(lnk-1/T). According to standard entropy change(ΔS0), the conformational change of the proteins was judged in the chromatographic process. The linear relationships between ΔH0 and ΔS0 can be used to evaluate "compensation temperature"(β) at the protein denaturation and identify the identity of the protein retention mechanism in weak cation-exchange chromatography.

  11. Use and application of hydrophobic interaction chromatography for protein purification. (United States)

    McCue, Justin T


    The objective of this section is to provide the reader with guidelines and background on the use and experimental application of Hydrophobic Interaction chromatography (HIC) for the purification of proteins. The section will give step by step instructions on how to use HIC in the laboratory to purify proteins. General guidelines and relevant background information is also provided.

  12. Characteristics of elution profile in radial chromatography under linear conditions

    Institute of Scientific and Technical Information of China (English)

    ZHANG; Weibing; SHAN; Yichu; Andreas; Seidel-Morgenster


    Based on the mass balance equations of solute transfer in the radial chromatographic column, the theoretical expression to describe the column efficiency and shape of elution profile is obtained under linear isotherm case.Moreover, the tendency for the variation of column efficiency and symmetry of peak profile is systematically discussed.The results showed that in radial chromatography the relationship between the column efficiency and volumetric flow rate is similar with that relationship in axial chromatography; relatively high column efficiency still can be obtained under high flow rate in radial chromatography.Accompanying the increase of retention factor of solutes and injection time, the column efficiency decreases monotonously.The effect of column diameter and column length on the column efficiency interfere with each other.It is more advantageous to increase the column efficiency by applying columns with larger column diameter and shorter column length.According to the discussion of the effect of diffusion on the column efficiency, radial chromatography is proved to be suitable for the separation of samples with relatively high diffusion coefficient, which predicts its obvious advantage in the preparative separation of samples such as proteins and DNA.

  13. An automatic system for multidimensional integrated protein chromatography. (United States)

    Kong, Yingjun; Li, Xiunan; Bai, Gaoying; Ma, Guanghui; Su, Zhiguo


    An automatic system for multidimensional integrated protein chromatography was designed for simultaneous separation of multiple proteins from complex mixtures, such as human plasma and tissue lysates. This computer-controlled system integrates several chromatographic columns that work independently or cooperatively with one another to achieve efficient high throughputs. The pipelines can be automatically switched either to another column or to a collection container for each UV-detected elution fraction. Environmental contamination is avoided due to the closed fluid paths and elimination of manual column change. This novel system was successfully used for simultaneous preparation of five proteins from the precipitate of human plasma fraction IV (fraction IV). The system involved gel filtration, ion exchange, hydrophobic interaction, and heparin affinity chromatography. Human serum albumin (HSA), transferrin (Tf), antithrombin-III (AT-III), alpha 1-antitrypsin (α1-AT), and haptoglobin (Hp) were purified within 3 h. The following recovery and purity were achieved: 95% (RSD, 2.8%) and 95% for HSA, 80% (RSD, 2.0%) and 99% for Tf, 70% (RSD, 2.1%) and 99% for AT-III, 65% (RSD, 2.0%) and 94% for α1-AT, and 50% (RSD, 1.0%) and 90% for Hp. The results demonstrate that this novel multidimensional integrated chromatography system is capable of simultaneously separating multiple protein products from the same raw material with high yield and purity and it has the potential for a wide range of multi-step chromatography separation processes.

  14. Isolation of bovine serum albumin from whey using affinity chromatography

    NARCIS (Netherlands)

    Besselink, T.; Janssen, A.E.M.; Boom, R.M.


    The adsorption of bovine serum albumin (BSA) to a chromatography resin with immobilised llama antibody fragments as affinity ligands was investigated. The maximum adsorption capacity of the affinity resin was 21.6 mg mL-1 with a Langmuir equilibrium constant of 20.4 mg mg-1. Using packed bed chromat

  15. Modeling of biopharmaceutical processes. Part 2: Process chromatography unit operation

    DEFF Research Database (Denmark)

    Kaltenbrunner, Oliver; McCue, Justin; Engel, Philip;


    Process modeling can be a useful tool to aid in process development, process optimization, and process scale-up. When modeling a chromatography process, one must first select the appropriate models that describe the mass transfer and adsorption that occurs within the porous adsorbent...

  16. Use of a fluorosurfactant in micellar electrokinetic capillary chromatography. (United States)

    de Ridder, R; Damin, F; Reijenga, J; Chiari, M


    A fluorosurfactant, the anionic N-ethyl-N-[(heptadecafluorooctyl)sulfonyl]glycine potassium salt, trade name FC-129 [CAS 2991-51-7] was investigated for possible application in micellar electrokinetic capillary chromatography (MEKC). The surfactant was characterized with conductometric titration and test sample mixtures were investigated in MEKC systems, and compared with sodium dodecylsulphate. An increased efficiency and interesting selectivity differences were observed.

  17. Fungal mycelium--the source of chitosan for chromatography. (United States)

    Kucera, Jiri


    Mycelium of the mold Aspergillus niger was used as a raw material for the preparation of microbial chitosan. Aspergillus niger, the mold used for the production of citric acid, contains approx. 15% of chitin, which can be separated, transformed into chitosan, and used as a sorbent for chromatography. The main advantage of this material in comparison with krill chitosan is the uniformity of particle size leading to the low back-pressure in the column. The other advantage is the fact, that original fibrous structure of mycelial pellets could be stabilized before chitosan preparation by cross-linking with glutaraldehyde. The product prepared by this way -- crosslinked chitosan of uniform particle size, is highly porous, with high water regain and, as a result, low sedimentation velocity. Low sedimentation velocity is not disadvantage in chromatographic application, but may form some problems in batchwise operation. Chitosan as a polymer of glucosamine is anion exchanger in nature and the chromatographic properties of this anion exchanger was demonstrated by the chromatography of bovine blood plasma, glucose oxidase, and chicken pepsinogen. In all cases, the course of chromatography on crosslinked chitosan was compared with the chromatography on MONO Q (bovine blood plasma) or DEAE-cellulose (glucose oxidase, chicken pepsinogen) under the same protocol.

  18. Purification of bacteriocins using size-exclusion chromatography

    Directory of Open Access Journals (Sweden)

    Vivek K. Bajpai


    Full Text Available The bacteriocin purification involves following main steps. a. Extraction of cell-free-supernatant of bacteria. b. Ammonium sulfate precipitation. c. Dialysis. d. Diafiltration using PVP and e. Size-exclusion chromatography. However, depending on the nature of work, the compound could be further analyzed by reverse-phase HPLC, NMR, mass spectrometry and sequencing.

  19. A Thin Layer Chromatography Laboratory Experiment of Medical Importance (United States)

    Sharma, Loretta; Desai, Ankur; Sharma, Ajit


    A thin layer chromatography experiment of medical importance is described. The experiment involves extraction of lipids from simulated amniotic fluid samples followed by separation, detection, and scanning of the lecithin and sphingomyelin bands on TLC plates. The lecithin-to-sphingomyelin ratio is calculated. The clinical significance of this…

  20. Supercritical fluid chromatography of fish, shark and seal oils

    DEFF Research Database (Denmark)

    Borch-Jensen, Christina; Mollerup, Jørgen


    Various natural and treated fish, shark liver and seal oils have been analyzed by supercritical fluid chromatography (SFC) using a non-polar capillary column. The lipids are separated according to molecular mass. The lipid groups found included free fatty acids, cholesterol, squalene, vitamins, wax...


    The combination of micellar electrokinetic chromatography (MEKC) with mass spectrometry (MS) is very attractive for the direct identification of analyte molecules, for the possibility of selectivity enhancement, and for the structure confirmation and analysis in a MS-MS mode. The...

  2. A Better Method for Filling Pasteur Pipet Chromatography Columns (United States)

    Ruekberg, Ben


    An alternative method for the preparation of Pasteur pipet chromatography columns is presented that allows the column to be filled with solvent without bubbles and allows greater control of fluid flow while the materials to be separated are added. Students are required to wear gloves and goggles and caution should be used while handling glass…

  3. Developing Inquiry-Based Labs Using Micro-Column Chromatography (United States)

    Barden-Gabbei, Laura M.; Moffitt, Deborah L.


    Chromatography is a process by which mixtures can be separated or substances can be purified. Biological and chemical laboratories use many different types of chromatographic processes. For example, the pharmaceutical industry uses chromatographic techniques to purify drugs, medical labs use them to identify blood components such as cholesterol,…


    The principal appeal of ion chromatography (IC) as analytical technique lies in the ability to rapidly analyze a mixture of ions of widely varying concentrations and properties in a single elution. It is therefore not surprising that IC has been hampered by the similar ion exchan...

  5. Thin-Layer Chromatography: The "Eyes" of the Organic Chemist (United States)

    Dickson, Hamilton; Kittredge, Kevin W.; Sarquis, Arlyne


    Thin-layer chromatography (TLC) methods are successfully used in many areas of research and development such as clinical medicine, forensic chemistry, biochemistry, and pharmaceutical analysis as TLC is relatively inexpensive and has found widespread application as an easy to use, reliable, and quick analytic tool. The usefulness of TLC in organic…

  6. Alternative solvents can make preparative liquid chromatography greener

    NARCIS (Netherlands)

    Shen, Y.; Chen, B.; Beek, van T.A.


    To make preparative Reversed-Phase High Performance Liquid Chromatography (RP-pHPLC) greener, alternative solvents were considered among others in terms of toxicity, cost, safety, workability, chromatographic selectivity and elution strength. The less toxic solvents ethanol, acetone and ethyl acetat

  7. Liquid chromatography and liquid chromatography-mass spectrometry analysis of donepezil degradation products

    Directory of Open Access Journals (Sweden)

    Mladenović Aleksandar R.


    Full Text Available This study describes the investigation of degradation products of donepezil (DP using stability indicating RP-HPLC method for determination of donepezil, which is a centrally acting reversible acetylcholinesterase inhibitor. In order to investigate the stability of drug and formed degradation products, a forced degradation study of drug sample and finished product under different forced degradation conditions has been conducted. Donepezil hydrochloride and donepezil tablets were subjected to stress degradation conditions recommended by International Conference on Harmonization (ICH. Donepezil hydrochloride solutions were subjected to acid and alkali hydrolysis, chemical oxidation and thermal degradation. Significant degradation was observed under alkali hydrolysis and oxidative degradation conditions. Additional degradation products were observed under the conditions of oxidative degradation. The degradation products observed during forced degradation studies were monitored using the high performance liquid chromatography (HPLC method developed. The parent method was modified in order to obtain LC-MS compatible method which was used to identify the degradation products from forced degradation samples using high resolution mass spectrometry. The mass spectrum provided the precise mass from which derived molecular formula of drug substance and degradation products formed and proved the specificity of the method unambiguously. [Projekat Ministarstva nauke Republike Srbije, br. 172013

  8. Rhamnolipid biosurfactant analysis using online turbulent flow chromatography-liquid chromatography-tandem mass spectrometry. (United States)

    Behrens, Beate; Helmer, Patrick O; Tiso, Till; Blank, Lars M; Hayen, Heiko


    Rhamnolipids are biosurfactants produced by a variety of bacterial species that present a promising alternative to surfactants from petrochemical or oleochemical origin. The success of the fermentation is evaluated by subsequent qualitative and quantitative analysis. However, the sample preparation for high numbers of samples is often laborious and inefficient. In this study an online sample preparation is developed for the qualitative and quantitative analysis of rhamnolipids by LC-MS/MS. Online sample preparation is carried out on a TurboFlow Cyclone MAX column using turbulent flow chromatography. Sample preparation prior the analysis is minimized to a dilution and syringe filtration step leading to an instrumental analysis time of 33min. The limit of detection and the limit of quantification were 0.4ng and 0.6ng on column, respectively. Recovery of the main mono- and di-rhamnolipids from a fermentation sample was 102-104%. Additionally, the rhamnolipid biosynthetic precursors 3-hydroxy(alkanoyloxy)alkanoic acids (HAAs) are covered, albeit extraction is not quantitative (85-90%). The analysis of rhamnolipids from four different microbial species was in good agreement with previous reports. The presented method allows rapid and comprehensive analysis of rhamnolipids with minimal sample preparation directly from the fermentation broth. The application of complementary data-dependent MS/MS acquisition enables non-target screening of rhamnolipids.

  9. Carbon monoxide measurement by gas chromatography; Mesure du monoxyde de carbone par chromatographie en phase gazeuse

    Energy Technology Data Exchange (ETDEWEB)

    Gros, V.; Sarda-Esteve, R.; Bonsang, B.; Ramonet, M.


    Although carbon monoxide (CO) is present in trace quantities in the atmosphere (0.1 ppm -or parts per million in volume- on average), the study of this gas is important. Indeed, its impact on human can be dangerous at high level of concentration on the hand and it constitutes one of the main precursor of ozone in presence of concentration on the one hand and it constitutes one of the main precursor of ozone in presence of other pollutants on the other hand. Finally, CO affects the levels of several important greenhouse gases, through its reaction with hydroxyl radicals (OH). CO is measured in the atmosphere since the mid 60's by various methods. Among them, gas chromatography has the advantage to combine a low detection limit with a high precision. This report details the improvements made on the measurement analyser which allowed to perform automatic CO measurements in remote areas with low mixing ratios of carbon monoxide. This report describes some quality tests and the results of various applications. (authors)

  10. Comparison of Membrane Chromatography and Monolith Chromatography for Lactoferrin and Bovine Serum Albumin Separation

    Directory of Open Access Journals (Sweden)

    Chalore Teepakorn


    Full Text Available These last few decades, membranes and monoliths have been increasingly used as stationary phases for chromatography. Their fast mass transfer is mainly based on convection, which leads to reduced diffusion, which is usually observed in resins. Nevertheless, poor flow distribution, which causes inefficient binding, remains a major challenge for the development of both membrane and monolith devices. Moreover, the comparison of membranes and monoliths for biomolecule separation has been very poorly investigated. In this paper, the separation of two proteins, bovine serum albumin (BSA and lactoferrin (LF, with similar sizes, but different isoelectric points, was investigated at a pH of 6.0 with a BSA-LF concentration ratio of 2/1 (2.00 mg·mL−1 BSA and 1.00 mg·mL−1 LF solution using strong cation exchange membranes and monoliths packed in the same housing, as well as commercialized devices. The feeding flow rate was operated at 12.0 bed volume (BV/min for all devices. Afterward, bound LF was eluted using a phosphate-buffered saline solution with 2.00 M NaCl. Using membranes in a CIM housing from BIA Separations (Slovenia with porous frits before and after the membrane bed, higher binding capacities, sharper breakthrough curves, as well as sharper and more symmetric elution peaks were obtained. The monolith and commercialized membrane devices showed lower LF binding capacity and broadened and non-symmetric elution peaks.

  11. Monitoring stevioside in soju by high-performance liquid chromatography and liquid chromatography/mass spectrometry. (United States)

    Ni, Fan; Ammann, Jeffrey; Mabud, Abdul


    A method using high-performance liquid chromatography (HPLC) with UV absorption detection was developed to monitor stevioside in soju, a distilled spirits product that is commercially available. The method uses a single-step dilution for sample preparation. It completely eliminates the time-consuming process of solid-phase extraction. A method using HPLC/mass spectrometry was optimized to confirm the identities of stevioside and other related impurities, including rebaudioside A, rebaudioside C, and dulcoside. The method was validated. The validation parameters included range (10.1-1007.3 ppm), precision, linearity, accuracy, robustness, system suitability, and intermediate precision. Stevioside standard solutions at 6 concentration levels were prepared for the validation work, including the tests for precision, linearity, and accuracy. The solutions were prepared in triplicate for each concentration. The relative standard deviation for the precision test was 0.999. The average recovery ranged from 95.7 to 101.1% for the soju samples spiked with stevioside standard. The detection limit for stevioside was estimated at 75 ppb. The method was used to screen several soju samples; no detectable stevioside was found in the samples.

  12. Standard flow liquid chromatography for shotgun proteomics in bioenergy research

    Directory of Open Access Journals (Sweden)

    Susana M. González Fernández-Niño


    Full Text Available Over the past ten years the bioenergy and biofuels field has realized significant achievements that have encouraged many follow on efforts centered on biosynthetic production of fuel-like compounds. Key to the success of these efforts has been transformational developments in feedstock characterization and metabolic engineering of biofuel-producing microbes. Lagging far behind these advancements are analytical methods to characterize and quantify systems of interest to the bioenergy field. In particular the utilization of proteomics, while valuable for identifying novel enzymes and diagnosing problems associated with biofuel-producing microbes, is limited by a lack of robustness and limited throughput. Nano-flow liquid chromatography coupled to high-mass accuracy, high-resolution mass spectrometers has become the dominant approach for the analysis of complex proteomic samples, yet such assays still require dedicated experts for data acquisition, analysis, and instrument upkeep. The recent adoption of standard flow chromatography (ca. 0.5 mL/min for targeted proteomics has highlighted the robust nature and increased throughput of this approach for sample analysis. Consequently, we assessed the applicability of standard flow liquid chromatography for shotgun proteomics using samples from Escherichia coli and Arabidopsis thaliana, organisms commonly used as model systems for lignocellulosic biofuels research. Employing 120 minute gradients with standard flow chromatography we were able to routinely identify nearly 800 proteins from E. coli samples, while for samples from Arabidopsis over 1,000 proteins could be reliably identified. An examination of identified peptides indicated that the method was suitable for reproducible applications in shotgun proteomics. Standard flow liquid chromatography for shotgun proteomics provides a robust approach for the analysis of complex samples. To the best of our knowledge this study represents the first attempt

  13. Standard flow liquid chromatography for shotgun proteomics in bioenergy research. (United States)

    González Fernández-Niño, Susana M; Smith-Moritz, A Michelle; Chan, Leanne Jade G; Adams, Paul D; Heazlewood, Joshua L; Petzold, Christopher J


    Over the past 10 years, the bioenergy field has realized significant achievements that have encouraged many follow on efforts centered on biosynthetic production of fuel-like compounds. Key to the success of these efforts has been transformational developments in feedstock characterization and metabolic engineering of biofuel-producing microbes. Lagging far behind these advancements are analytical methods to characterize and quantify systems of interest to the bioenergy field. In particular, the utilization of proteomics, while valuable for identifying novel enzymes and diagnosing problems associated with biofuel-producing microbes, is limited by a lack of robustness and limited throughput. Nano-flow liquid chromatography coupled to high-mass accuracy, high-resolution mass spectrometers has become the dominant approach for the analysis of complex proteomic samples, yet such assays still require dedicated experts for data acquisition, analysis, and instrument upkeep. The recent adoption of standard flow chromatography (ca. 0.5 mL/min) for targeted proteomics has highlighted the robust nature and increased throughput of this approach for sample analysis. Consequently, we assessed the applicability of standard flow liquid chromatography for shotgun proteomics using samples from Escherichia coli and Arabidopsis thaliana, organisms commonly used as model systems for lignocellulosic biofuels research. Employing 120 min gradients with standard flow chromatography, we were able to routinely identify nearly 800 proteins from E. coli samples; while for samples from Arabidopsis, over 1,000 proteins could be reliably identified. An examination of identified peptides indicated that the method was suitable for reproducible applications in shotgun proteomics. Standard flow liquid chromatography for shotgun proteomics provides a robust approach for the analysis of complex samples. To the best of our knowledge, this study represents the first attempt to validate the standard

  14. Determination of a Jet Fuel Metal Deactivator by High Performance Liquid Chromatography (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY Paul C. Hayes, Jr. Fuels Branch...SUPPLEMENTARY NOTES 19. KEY WORDS (Continue on reverse side if necessary and identify by block number) High Performance Liquid Chromatography absorbance...SYMBOL HPLC High Performance Liquid Chromatography P-4 jet propulsion fuel, wide-boiling range, conforming to MIL-T-5624L MDA metal deactivator,

  15. Determination of Stabiliser Contents in Advanced Gun Propellants by Reverse Phase High Performance Liquid Chromatography (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY N"m A.R. TURNER AND A. WHITE...TO biEPROOU.; AND SELL THIS REPORT Determination of Stabiliser Contents in Advanced Gun Propellants by Reverse Phase High Performance Liquid Chromatography A.R...8217/......... .. Availability Cooes Dist Avaiardlo A-i Determination of Stabiliser Contents in Advanced Gun Propellants by Reverse Phase High Performance Liquid Chromatography

  16. Gas chromatography-olfactometry analysis of the volatile compounds of two commercial Irish beef meats

    NARCIS (Netherlands)

    Machiels, D.; Ruth, van S.M.; Posthumus, M.A.; Istasse, L.


    The volatile flavour compounds of two commercial Irish beef meats (labelled as conventional and organic) were evaluated by gas chromatography-olfactometry and were identified by gas chromatography-mass spectrometry. The volatile compounds were isolated in a model mouth system. Gas chromatography-olf

  17. Normal and Reversed-Phase Thin Layer Chromatography of Green Leaf Extracts (United States)

    Sjursnes, Birte Johanne; Kvittingen, Lise; Schmid, Rudolf


    Introductory experiments of chromatography are often conducted by separating colored samples, such as inks, dyes, and plant extracts, using filter paper, chalk, or thin layer chromatography (TLC) plates with various solvent systems. Many simple experiments have been reported. The relationship between normal chromatography and reversed-phase…

  18. Theories to support method development in comprehensive two-dimensional liquid chromatography - A review

    NARCIS (Netherlands)

    Bedani, F.; Schoenmakers, P.J.; Janssen, H.-G.


    On-line comprehensive two-dimensional liquid chromatography techniques promise to resolve samples that current one-dimensional liquid chromatography methods cannot adequately deal with. To make full use of the potential of two-dimensional liquid chromatography, optimization is required. Optimization

  19. Chiral Separation of G-type Chemical Warfare Nerve Agents via Analytical Seupercritical Fluid Chromatography (United States)


    by HPLC have not been well characterized . Supercritical fluid chromatography (SFC) is a well- established chiral chromatography technology that offers...smoothed chromatograms. For presentation purposes only smoothed chromatograms are shown. Chromatographic Characterization Chromatography parameters were...under these conditions (data not shown). Various other polysaccharide -type chiral columns (Chiralpak AD-H, Chiralcel OJ-RH, Chiralpak IB) were tested

  20. 21 CFR 862.2260 - High pressure liquid chromatography system for clinical use. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false High pressure liquid chromatography system for... Clinical Laboratory Instruments § 862.2260 High pressure liquid chromatography system for clinical use. (a) Identification. A high pressure liquid chromatography system for clinical use is a device intended to...

  1. 21 CFR 862.2250 - Gas liquid chromatography system for clinical use. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Gas liquid chromatography system for clinical use... Instruments § 862.2250 Gas liquid chromatography system for clinical use. (a) Identification. A gas liquid chromatography system for clinical use is a device intended to separate one or more drugs or compounds from...

  2. Identification of Explosives from Porous Materials: Applications Using Reverse Phase High Performance Liquid Chromatography and Gas Chromatography (United States)

    Miller, C. J.; Elias, G.; Schmitt, N. C.; Rae, C.


    High performance liquid chromatography and gas chromatography techniques are well documented and widely used for the detection of trace explosives from organic solvents. These techniques were modified to identify and quantify explosives extracted from various materials taken from people who had recently handled explosives. Documented techniques were modified to specifically detect and quantify trace levels of the military explosives, RDX, TNT, and PETN from denim, colored flannel, vinyl, and canvas extracted in methanol and filtered using no additional sample cleanup of the sample extract prior to analysis. The filtered methanol extracts were injected directly into several different column types and analyzed by high performance liquid chromatography using ultraviolet detection and/or gas chromatography using electron capture detection. This paper describes general screening methods that were used to determine the presence of explosives (RDX, TNT, and PETN) in unknown samples of denim, colored flannel, vinyl and canvas in addition to techniques that have been optimized for quantification of each explosive from the substrate extracts.

  3. 5-O-caffeoylshikimic acid from Solanum somalense leaves: advantage of centrifugal partition chromatography over conventional column chromatography. (United States)

    Chideh, Saïda; Pilard, Serge; Attoumbré, Jacques; Saguez, Robert; Hassan-Abdallah, Alshaimaa; Cailleu, Dominique; Wadouachi, Anne; Baltora-Rosset, Sylvie


    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.

  4. Analysis of pesticide residues in tobacco with online size exclusion chromatography with gas chromatography and tandem mass spectrometry. (United States)

    Guo, Weiyun; Bian, Zhaoyang; Tang, Gangling; Wang, Deguo; Li, Guanghui; Wang, Jianlong


    An ultrasensitive method for the simultaneous analysis of pesticides residues in tobacco was developed with online size exclusion chromatography with gas chromatography and tandem mass spectrometry. Tobacco samples were extracted with the solvent mixture of cyclohexane and acetone (7:3, v/v) and centrifuged. Then, the supernatant liquors were injected directly into the online size exclusion chromatography with gas chromatography and tandem mass spectrometry without any other purification procedures after being filtered with a 0.22 μm organic phase filter. The matrix interferences were effectively removed and recoveries of most pesticides were in the range of 72-121%. Especially, for chlorothalonil, the analysis efficiency of this method was much more favorable than that of the general method, in which dispersive solid-phase extraction was used as an additional purified procedure. In addition, the limits of quantitation of this method were from 1 to 50 μg/kg. Therefore, a rapid, cost-effective, labor-saving method was proposed in the present work, which was suitable for the analysis of 41 pesticide residues in tobacco.

  5. Preparative separation of polyphenols from artichoke by polyamide column chromatography and high-speed counter-current chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Shu, Xikai; Wang, Mei; Liu, Daicheng [College of Life Science, Shandong Normal University, Jinan, Shandong (China); Wang, Daijie; Lin, Xiaojing; Liu, Jianhua; Wang, Xiao; Huang, Luqi, E-mail: [Shandong Analysis and Test Center, Shandong Academy of Sciences, Jinan, Shandong (China)


    An efficient method for the rapid separation and purification of polyphenols from artichoke by polyamide column chromatography in combination with high-speed counter-current chromatography (HSCCC) was successfully built. The crude ethanol extracts from dry artichoke were first pre-separated by polyamide column chromatography and divided in two parts as sample 1 and sample 2. Then, the samples were further separated by HSCCC and yielded 7.8 mg of chlorogenic acid (compound I), 24.5 mg of luteolin-7-O-{beta}-D-rutinoside (compound II), 18.4 mg of luteolin-7-O-{beta}-D-glucoside (compound III), and 33.4 mg of cynarin (compound IV) with purity levels of 92.0%, 98.2%, 98.5%, and 98.0%, respectively, as determined by high-performance liquid chromatography (HPLC) method. The chemical structures of these compounds were identified by electrospray ionization-mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR). (author)

  6. High pH reversed-phase chromatography with fraction concatenation as an alternative to strong-cation exchange chromatography for two-dimensional proteomic analysis


    Yang, Feng; Shen, Yufeng; Camp, David G.; Smith, Richard D.


    Orthogonal high-resolution separations are critical for attaining improved analytical dynamic range and protein coverage in proteomic measurements. High pH reversed-phase liquid chromatography (RPLC) followed by fraction concatenation affords better peptide analysis than conventional strong-cation exchange (SCX) chromatography applied for the two-dimensional proteomic analysis. For example, concatenated high pH reversed-phase liquid chromatography increased identification for peptides (1.8-fo...

  7. HPLC separation and GC-MS identification of the polar components of coal liquids. [High pressure liquid chromatography, gas chromatography-mass spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Satoh, M.; Tanabe, K.; Uchino, H.; Yokoyama, S.; Sanada, Y.


    The polar components in the 200-400 C fraction of Akabira coal liquids were separated by high pressure liquid chromatography using an amine-based column. By varying the CHCl/sub 3/ concentration in the solvent, basic, neutral and acid fractions were obtained. These were then investigated separately using infrared spectroscopy, flame ionization detection-gas chromatography and gas chromatography-mass spectroscopy. The basic fraction contained alkyltetrahydroquinolines, the neutral fraction, alkylphenols and alkylcarbazoles; and the acid fraction, alkylphenols. 15 references.

  8. Electrochemically modulated liquid chromatography: Theoretical investigations and applications from the perspectives of chromatography and interfacial electrochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Keller, David W.


    Electrochemically modulated liquid chromatography (EMLC) employs a conductive material as both a stationary phase for chromatographic separations and as a working electrode for performing electrochemistry experiments. This dual functionality gives EMLC the capacity to manipulate chromatographic separations by changing the potential applied (E{sub app}) to the stationary phase with respect to an external reference. The ability to monitor retention as a function of E{sub app} provides a means to chromatographically monitor electrosorption processes at solid-liquid interfaces. In this dissertation, the retention mechanism for EMLC is examined from the perspective of electrical double layer theory and interfacial thermodynamics. From the chromatographic data, it is possible to determine the interfacial excess ({Lambda}) of a solute and changes in interfacial tension (d{gamma}) as a function of both E{sub app} and the supporting electrolyte concentration. Taken together, these two experimentally manipulated parameters can be examined within the context of the Gibbs adsorption equation to delineate the contribution of a variety of interfacial properties, including the charge of solute on the stationary phase and the potential of zero charge (PZC), to the mechanism behind EMLC-based retention. The chromatographic probing of interfacial phenomena is complemented by electroanalytical experiments that exploit the ability to monitor the electronic current flowing through an EMLC column. Cyclic voltammetry and chronoamperometry of an EMLC column are used to determine the electronic performance characteristics of an EMLC column. An electrochemical flow injection analysis of a column is provided in which the current required to maintain a constant E{sub app} is monitored and provides a way to examine the influence that acetonitrile and supporting electrolyte composition, flow rate, column backpressure, and ionic strength have on the structure of electrified interfaces.

  9. Electrochemically modulated liquid chromatography: Theoretical investigations and applications from the perspectives of chromatography and interfacial electrochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Keller, David W. [Iowa State Univ., Ames, IA (United States)


    Electrochemically modulated liquid chromatography (EMLC) employs a conductive material as both a stationary phase for chromatographic separations and as a working electrode for performing electrochemistry experiments. This dual functionality gives EMLC the capacity to manipulate chromatographic separations by changing the potential applied (Eapp) to the stationary phase with respect to an external reference. The ability to monitor retention as a function of Eapp provides a means to chromatographically monitor electrosorption processes at solid-liquid interfaces. In this dissertation, the retention mechanism for EMLC is examined from the perspective of electrical double layer theory and interfacial thermodynamics. From the chromatographic data, it is possible to determine the interfacial excess (Λ) of a solute and changes in interfacial tension (dγ) as a function of both Eapp and the supporting electrolyte concentration. Taken together, these two experimentally manipulated parameters can be examined within the context of the Gibbs adsorption equation to delineate the contribution of a variety of interfacial properties, including the charge of solute on the stationary phase and the potential of zero charge (PZC), to the mechanism behind EMLC-based retention. The chromatographic probing of interfacial phenomena is complemented by electroanalytical experiments that exploit the ability to monitor the electronic current flowing through an EMLC column. Cyclic voltammetry and chronoamperometry of an EMLC column are used to determine the electronic performance characteristics of an EMLC column. An electrochemical flow injection analysis of a column is provided in which the current required to maintain a constant Eapp is monitored and provides a way to examine the influence that acetonitrile and supporting electrolyte composition, flow rate, column backpressure, and ionic strength have on the structure of electrified interfaces.

  10. Liquid chromatography and supercritical fluid chromatography as alternative techniques to gas chromatography for the rapid screening of anabolic agents in urine. (United States)

    Desfontaine, Vincent; Nováková, Lucie; Ponzetto, Federico; Nicoli, Raul; Saugy, Martial; Veuthey, Jean-Luc; Guillarme, Davy


    This work describes the development of two methods involving supported liquid extraction (SLE) sample treatment followed by ultra-high performance liquid chromatography or ultra-high performance supercritical fluid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS and UHPSFC-MS/MS) for the screening of 43 anabolic agents in human urine. After evaluating different stationary phases, a polar-embedded C18 and a diol columns were selected for UHPLC-MS/MS and UHPSFC-MS/MS, respectively. Sample preparation, mobile phases and MS conditions were also finely tuned to achieve highest selectivity, chromatographic resolution and sensitivity. Then, the performance of these two methods was compared to the reference routine procedure for steroid analyses in anti-doping laboratories, which combines liquid-liquid extraction (LLE) followed by gas chromatography coupled to tandem mass spectrometry (GC-MS/MS). For this purpose, urine samples spiked with the compounds of interest at five different concentrations were analyzed using the three analytical platforms. The retention and selectivity of the three techniques were very different, ensuring a good complementarity. However, the two new methods displayed numerous advantages. The overall procedure was much faster thanks to high throughput SLE sample treatment using 48-well plates and faster chromatographic analysis. Moreover, the highest sensitivity was attained using UHPLC-MS/MS with 98% of the doping agents detected at the lowest concentration level (0.1ng/mL), against 76% for UHPSFC-MS/MS and only 14% for GC-MS/MS. Finally, the weakest matrix effects were obtained with UHPSFC-MS/MS with 76% of the analytes displaying relative matrix effect between -20 and 20%, while the GC-MS/MS reference method displayed very strong matrix effects (over 100%) for all of the anabolic agents.

  11. Nano-liquid chromatography applied to enantiomers separation. (United States)

    Fanali, Salvatore


    This paper presents the state of the art concerning the separation of chiral compounds by means of nano-liquid chromatography (nano-LC). The enantiomers' separation and determination are a subject of fundamental importance in various application fields such as pharmaceutical industry, biomedicine, food, agrochemical etc. Nano-LC is a miniaturized chromatographic technique offering some advantages over conventional ones such as low consumption of mobile phase, sample volume and amount of chiral stationary phase, reduced costs etc. This is reported in the first part of the paper illustrating the features of the nano-LC. In addition, chiral resolution methods are briefly illustrated. Some chiral selectors, used in high-performance liquid chromatography have also been applied in nano-LC including cyclodextrins, glycopeptide antibiotics, modified polysaccharides etc. This is discussed in the second part of the review. Finally some examples of the applications available in literature are reported.

  12. Analysis of polycyclic aromatic hydrocarbons by supercritical fluid chromatography (SFC)

    Energy Technology Data Exchange (ETDEWEB)

    Leselier, E. [Institut Universitaire de Technologie, 91 - Orsay (France). Letiam


    Usually, elution gradient high performance liquid chromatography using polymeric octadecyl bonded phases is chosen for this kind of separation. Due to the properties of carbon dioxide, low viscosity, high eluting power, separations obtained by supercritical fluid chromatography (SFC) are generally faster than by liquid solvents. This paper reports the study of the PAH separation by SFC. The effects of the nature and percentage of modifiers, pressure, temperature and the choice of the stationary phase (mono or poly-functional, high or low bonded density) are discussed. Results show that coupling two different columns is needed to reach the complete separation of the 16 PAHs requires by the standard of the Environmental Protection Agency (EPA 610). The separation is achieved in 25 minutes with a linear acetonitrile/CO{sub 2} mobile phase gradient. Applied to the analysis of soil extracts, this analytical technique is enable to separate numerous other compounds than standard PAHs. (author) 17 refs.

  13. HD gas analysis with Gas Chromatography and Quadrupole Mass Spectrometer

    CERN Document Server

    Ohta, T; Didelez, J -P; Fujiwara, M; Fukuda, K; Kohri, H; Kunimatsu, T; Morisaki, C; Ono, S; Rouille, G; Tanaka, M; Ueda, K; Uraki, M; Utsuro, M; Wang, S Y; Yosoi, M


    A gas analyzer system has been developed to analyze Hydrogen-Deuteride (HD) gas for producing frozen-spin polarized HD targets, which are used for hadron photoproduction experiments at SPring-8. Small amounts of ortho-H$_{2}$ and para-D$_{2}$ gas mixtures ($\\sim$0.01%) in the purified HD gas are a key to realize a frozen-spin polarized target. In order to obtain reliable concentrations of these gas mixtures in the HD gas, we produced a new gas analyzer system combining two independent measurements with the gas chromatography and the QMS. The para-H$_{2}$, ortho-H$_{2}$, HD, and D$_{2}$ are separated using the retention time of the gas chromatography and the mass/charge. It is found that the new gas analyzer system can measure small concentrations of $\\sim$0.01% for the otho-H$_2$ and D$_2$ with good S/N ratios.

  14. Mass transfer mechanism in chiral reversed phase liquid chromatography. (United States)

    Gritti, Fabrice; Guiochon, Georges


    The mechanism of mass transfer in chiral chromatography was investigated using an experimental protocol already applied in RPLC and HILIC chromatography. The different contributions to the reduced height equivalent to a theoretical plate (HETP) include the longitudinal diffusion HETP term, the solid-liquid mass transfer resistance HETP term, the short-range eddy dispersion HETP term, and the long-range eddy dispersion HETP term. Their accurate measurement permits the determination of the adsorption rate constant kads of trans-stilbene enantiomers on a column packed with Lux 5 μm Cellulose-1 particles. The experimental results demonstrate that the number of adsorption-desorption steps per unit time of chiral compounds on polysaccharide-based chiral stationary phases is four orders of magnitude smaller than that of achiral compounds.

  15. Sol-gel multicapillary columns for gas-solid chromatography. (United States)

    Sidelnikov, Vladimir N; Patrushev, Yuri V; Belov, Yuri P


    In this work, we report the method for the preparation of multicapillary columns (MCCs) for gas-solid chromatography. The porous layer adsorbent is formed on capillary walls by the hydrolysis of aluminum alkoxide in the presence of polypropylene glycol (PPG) and HCl. Porosity and selectivity of the adsorbent depend on reaction conditions and the concentration of PPG. Sol-gel MCCs are well suited for high-speed chromatographic analysis of light hydrocarbons by gas-solid chromatography. Nine-component mixtures of C1-C4 hydrocarbons are separated within 8-12 s. The efficiency of 25-30 cm long alumina sol-gel MCCs consisting of approximately 1400 capillaries of 40 microm diameter is up to 2500-3000 theoretical plates.

  16. Speciation analysis by gas chromatography with plasma source spectrometric detection (United States)

    Łobiński, Ryszard; Adams, Freddy C.

    State-of-the-art species-selective analysis by gas chromatography (GC) with plasma source spectrometric detection is discussed for organometal and organometalloid compounds. Various plasmas, inductively coupled plasma, microwave induced plasma, capacitatively coupled plasma, direct current plasma and alternating current plasma, are characterized and critically compared as sources of radiation for atomic emission spectrometry and sources of ions for mass spectrometry. Interfaces between gas chromatography (packed, wide-bore, capillary and multicapillary) and plasma source spectrometry are characterized. Particular emphasis is given to applications of GC with plasma source detection to real-world analytical problems, which are comprehensively reviewed. The use of plasmas for the acquisition of auxiliary molecular information such as empirical formulae and structural information is discussed. Recent developments relating to sample preparation and presentation to the hyphenated system are addressed. The most significant trends in speciation analysis are highlighted.

  17. Numerical Simulation of Asynchronous Simulated Moving Bed Chromatography

    Institute of Scientific and Technical Information of China (English)



    Asynchronous simulated moving bed chromatography (ASMBC), known also as the "VARICOL" process, is more efficient and flexible than the well-known and traditional simulated moving bed chromatography (SMBC). A detailed model of ASMBC, taking account of non-linear competitive isotherms, mass transfer parameters, and complex port switching schedule parameters, was developed to simulate the complex dynamics of ASMBC.The simulated performance is in close agreement with the experimental data of chiral separation reported in the literature. The simulation results show that ASMBC can achieve the performance similar to SMBC with fewer columns and can achieve better performance than SMBC with the same total column number. All design and operation parameters can be chosen correctly by numerical simulation. This detailed ASMBC model and the numerical technique are useful for design, operation, optimization and scale-up of ASMBC.

  18. Ion chromatography in the manufacture of multilayer circuit boards (United States)

    Smith, Robert E.


    Ion chromatography (IC) has proven useful in analyzing chemical solutions used in the manufacture of multilayer circuit boards. The manufacturing process is described briefly and previously published IC methods are reviewed. Then, methods are described for determining chlorate and chlorite in a brown oxide solution; salicylic acid in an epoxy cure agent; formate, sulfate, and tartrate in an electroless copper bath; anionic detergents in a tin-lead brightener and in a cleaning solution; and aqueous photoresist and nonionic brightener in a tin-lead bath. Anion exchange, reverse phase HPLC on a poly(styrene/divinylbenzene), PS/DVB, column and 2-D liquid chromatography also are described. Chemically suppressed conductivity and photometric detection are used.

  19. Moment equations for chromatography based on Langmuir type reaction kinetics. (United States)

    Miyabe, Kanji


    Moment equations were derived for chromatography, in which the reaction kinetics between solute molecules and functional ligands on the stationary phase was represented by the Langmuir type rate equation. A set of basic equations of the general rate model of chromatography representing the mass balance, mass transfer rate, and reaction kinetics in the column were analytically solved in the Laplace domain. The moment equations for the first absolute moment and the second central moment in the real time domain were derived from the analytical solution in the Laplace domain. The moment equations were used for predicting the chromatographic behavior under hypothetical HPLC conditions. The influence of the parameters relating to the adsorption equilibrium and to the reaction kinetics on the chromatographic behavior was quantitatively evaluated. It is expected that the moment equations are effective for a detailed analysis of the influence of the mass transfer rates and of the Langmuir type reaction kinetics on the column efficiency.

  20. Gas chromatography-vacuum ultraviolet spectroscopy for multiclass pesticide identification. (United States)

    Fan, Hui; Smuts, Jonathan; Walsh, Phillip; Harrison, Dale; Schug, Kevin A


    A new vacuum ultraviolet detector for gas chromatography was recently developed and applied to multiclass pesticide identification. VUV detection features full spectral acquisition in a wavelength range of 115-240nm, where virtually all chemical species absorb. VUV absorption spectra of 37 pesticides across different classes were recorded. These pesticides display rich gas phase absorption features across various classes. Even for isomeric compounds, such as hexachlorocyclohexane (HCH) isomers, the VUV absorption spectra are unique and can be easily differentiated. Also demonstrated is the ability to use VUV data analysis software for deconvolution of co-eluting signals. As a universal detector, VUV provides both qualitative and quantitative information. It offers high specificity, sensitivity (pg on-column detection limits), and a fast data acquisition rate, making it a powerful tool for multiclass pesticide screening when combined with gas chromatography.

  1. Application of fast liquid chromatography for antioxidants analysis

    Directory of Open Access Journals (Sweden)

    Agnieszka Drożdżyńska


    Full Text Available Background. An intensive development of the Fast Liquid Chromatography (FLC has been recently observed. It makes possible to reduce time analysis and improve resolution as well as sensitivity. The aim of this study was to separate the chosen antioxidants optimization using the FLC method. Material  and methods. The three various procedures for antioxidants analysis were compared. Mobile phases containing aqueous solution of formic acid, acetic acid, acetonitrile, and methanol were tested. Limit of detection (LOD, limit of quantification (LOQ, linearity and repeatability of each procedures were determined. Results. Developed procedure enabled to separate all analytes and allowed to get low LOD levels and good repeatability. This procedure was used for antioxidants analysis in buckwheat and buckwheat products. Conclusion. Fast Liquid Chromatography allows to  reduce time analysis and  obtain good  validation parameters.

  2. Affinity Purification of Insulin by Peptide-Ligand Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)


    The affinity heptapeptide (HWWWPAS) for insulin, selected from phage display library,was coupled to EAH Sepharose 4B gel and packed to a 1-mL column. The column was used for the affinity purification of insulin from protein mixture and commercial insulin preparation. It was observed that the minor impurity in the commercial insulin was removed by the affinity chromatography. Nearly 40 mg of insulin could be purified with the 1-mL affinity column. The results revealed the high specificity and capacity of the affinity column for insulin purification. Moreover, based on the analysis of the amino acids in the peptide sequence, shorter peptides were designed and synthesized for insulin chromatography. As a result, HWWPS was found to be a good alternative to HWWWPAS, while the other two peptides with three or four amino acids showed weak affinity for insulin. The results indicated that the peptide sequence of HWWWPAS was quite conservative for specific binding of insulin.

  3. Temperature-Modulated Array High-Performance Liquid Chromatography


    Premstaller, Andreas; Xiao, Wenzhong; Oberacher, Herbert; O'Keefe, Matthew; Stern, David; Willis, Thomas; Huber, Christian G.; Peter J. Oefner


    Using novel monolithic poly(styrene-divinylbenzene) capillary columns with an internal diameter of 0.2 mm, we demonstrate for the first time the feasibility of constructing high-performance liquid chromatography arrays for the detection of mutations by heteroduplex analysis under partially denaturing conditions. In one embodiment, such an array can be used to analyze one sample simultaneously at different temperatures to maximize the detection of mutations in DNA fragments containing multiple...

  4. Stationary phase optimized selectivity supercritical fluid chromatography (SOS-SFC)


    Delahaye, Sander; Lynen, Frederic


    In stationary phase optimized selectivity liquid chromatography (SOS-LC) the stationary phase becomes a tunable parameter by connecting column segments with variable lengths of different stationary phases. An optimization procedure and algorithm based on the PRISMA model for optimization of the mobile phase in LC was developed to apply this strategy for isocratic and gradient separations. An optimized column segment combination, giving the highest separation selectivity for all compounds in a...

  5. Determination of serotonin released from coffee wax by liquid chromatography. (United States)

    Kele, M; Ohmacht, R


    A simple hydrolysis and extraction method was developed for the release of serotonin (5-hydroxytryptamine) from a coffee wax sample obtained from decaffeination of coffee beans. The recoverable amount of serotonin was determined by reversed-phase high-performance liquid chromatography with gradient elution and UV detection, using the standard addition method. Different type of basic deactivated chromatographic columns were used for the separation.

  6. Determination of aminocresol isomers by high-speed liquid chromatography. (United States)

    Sakurai, H; Kito, M

    Aminocresol isomers (4-hydroxy-m-toluidine [II], 3-hydroxy-p-toluidine [II], 2-hydroxy-p-toluidine [III]) and p-aminophenol have been separated and determined by a high-speed liquid Chromatographie method. Since this method is applicable in aqueous media, it was used to investigate the suitability of a haemin-cysteine system as a model for the cytochrome P-450 mono-oxygenase system, by determination of the [I], [II], [III] and p-aminophenol formed.

  7. Mineral Separation in a CELSS by Ion-exchange Chromatography (United States)

    Ballou, E. V.; Spitze, L. A.; Wong, F. W.; Wydeven, T.; Johnson, C. C.


    Operational parameters pertinent to ion exchange chromatography separation were identified. The experiments were performed with 9 mm diameter ion exchange columns and conventional column accessories. The cation separation beds were packed with AG 50W-X2 strong acid cation exchange resin in H(+) form and 200-400 dry mesh particle size. The stripper beds used in some experiments were packed with AG 1-XB strong base cation exchange resin in OH(-) form and 200-400 dry mesh particle size.

  8. Affinity chromatography purification of cytochrome c binding enzymes.


    Azzi, A; Bill, K; Broger, C


    An efficient affinity chromatography procedure for the isolation of mitochondrial cytochrome c oxidase and reductase is described. Saccharomyces cerevisiae cytochrome c was used as a ligand, bound to a thiol-Sepharose 4B gel through cysteine-107. In this way, the site of interaction of cytochrome c with cytochrome oxidase and reductase remained unmodified and available for binding to a number of partner enzymes. The procedure is adequate for the purification of all those proteins having in co...

  9. Determination of chloride in geological samples by ion chromatography (United States)

    Wilson, S.A.; Gent, C.A.


    Samples of silicate rocks are prepared by sodium carbonate fusion and then treated by ion chromatography. The method was tested for geological standards with chloride concentration between 0.003 and 3%. Observed chloride concentrations comparedd favorably with literature values. The relative standard deviation and detection limit for the method were 8% and 7 ppm, respectively. Up to 30 determination per 24-hour period were possible. ?? 1983.

  10. Enantiomeric Separation of Amino Alcohols by Ion-pair Chromatography

    Institute of Scientific and Technical Information of China (English)


    Enantiomers of four amino alcohols were resolved by ion-pair chromatography with (+)-10-camphorsulphonic acid as chiral counter ion. Studies of the influence of the mobile phase composition, the solute structure and the mobile phase flow-rate on separation are presented. Under the optimized conditions, enantiomeric propranolol, norephedrine, metropolol and salbutamol were separated using dichloromethane -1-pentanol (97:3, v/v) as mobile phase on Lichrospher-100-DIOL column.

  11. Host cell protein adsorption characteristics during protein A chromatography. (United States)

    Tarrant, Richard D R; Velez-Suberbie, M Lourdes; Tait, Andrew S; Smales, C Mark; Bracewell, Daniel G


    Protein A chromatography is a critical and 'gold-standard' step in the purification of monoclonal antibody (mAb) products. Its ability to remove >98% of impurities in a single step alleviates the burden on subsequent process steps and facilitates the implementation of platform processes, with a minimal number of chromatographic steps. Here, we have evaluated four commercially available protein A chromatography matrices in terms of their ability to remove host cell proteins (HCPs), a complex group of process related impurities that must be removed to minimal levels. SELDI-TOF MS was used as a screening tool to generate an impurity profile fingerprint for each resin and indicated a number of residual impurities present following protein A chromatography, agreeing with HCP ELISA. Although many of these were observed for all matrices there was a significantly elevated level of impurity binding associated with the resin based on controlled pore glass under standard conditions. Use of null cell line supernatant with and without spiked purified mAb demonstrated the interaction of HCPs to be not only with the resin back-bone but also with the bound mAb. A null cell line column overload and sample enrichment method before 2D-PAGE was then used to determine individual components associated with resin back-bone adsorption. The methods shown allow for a critical analysis of HCP removal during protein A chromatography. Taken together they provide the necessary process understanding to allow process engineers to identify rational approaches for the removal of prominent HCPs.

  12. The Analysis of Metal Finishing Solutions by Ion Chromatography (United States)


    but some Cr(III) is produced during the electrolysis and can negatively effect plate properties and plating efficiency (67, 68). In an operating system... Brines by Indirect Atomic Absorption Spectroscopy", Chem. Prum., 33, 8, 429- 435 (1983). 17. D. Jones, S. Manahan, "Atomic Absorption Detector For... Brine ", J. Chromatogr., 250, 134-7 (1982). 24. T. Nishina, "Analysis of Sulfate Ion by Ion Chromatography", Yogyo Kyokaishi, 92, 7, 410-2 (1984). 25. S


    Institute of Scientific and Technical Information of China (English)

    AanTianwei; XuWeijiang; 等


    A new and an inexpensive adsorbent of chitosan coated silica for immobilized metal affinity chromatography(IMAC) was studied.After a double coating,the chitosan coated on silica beads could be up to 53.4mg/g silica beads.When pH>3.8,the metal ligand Cu2+ was chelated on the coated chitosan with a bound capacity of 14.6mg/g chitosan without introducing iminodiacetic acid(IDA).

  14. Chromatography, solid-phase extraction, and capillary electrochromatography with MIPs. (United States)

    Tóth, Blanka; Horvai, George


    Most analytical applications of molecularly imprinted polymers are based on their selective adsorption properties towards the template or its analogs. In chromatography, solid phase extraction and electrochromatography this adsorption is a dynamic process. The dynamic process combined with the nonlinear adsorption isotherm of the polymers and other factors results in complications which have limited the success of imprinted polymers. This chapter explains these problems and shows many examples of successful applications overcoming or avoiding the problems.

  15. Fiber Bragg grating photoacoustic detector for liquid chromatography. (United States)

    Yang, Qingxin; Loock, Hans-Peter; Kozin, Igor; Pedersen, David


    Fiber Bragg Gratings (FBGs) are known to be sensitive acoustic transducers and have previously been used for the photoacoustic detection of small solid samples. Here, we demonstrate the use of an FBG as an on-line detector for liquid chromatography. The FBG was inserted into a silica capillary and the photoacoustic response from the effluent was generated by a 10 ns pulsed laser. The acoustic pulse was quantified by the FBG through a characteristic change in the reflection spectrum. Good repeatability and linear response were obtained over three orders of magnitude (R(2) > 0.99), and the limit of detection of Coumarin 440 was determined to be 5 microM. The technique was successfully coupled to high performance liquid chromatography and applied to on-line analysis of a three-compound solution. Photoacoustic detection in liquid chromatography using FBGs is a label-free method, which can be applied to the detection of any chromogenic compound irrespective of its fluorogenic properties. It is a simple, inexpensive, and inherently micron-sized technique, insensitive to electromagnetic interference.

  16. Advantages of ion-exchange chromatography for oligonucleotide analysis. (United States)

    Cook, Ken; Thayer, Jim


    The rapid development of therapeutic oligonucleotides (ONs) has created a need for in-depth characterization of ONs, beyond previous requirements. The natural migration to LC-MS requires the use of chromatography with MS-compatible eluents to introduce the large, highly charged biopolymers into the mass spectrometer. Most frequently this employs ion-pair reversed-phase liquid chromatography, which may leave gaps in the characterization, but these can be filled with the use of high-resolution ion-exchange chromatography. Several classes of isobaric isomers are among the impurities that will require further separation prior to MS analysis. This review shows how the use of ion exchange as an additional orthogonal analytical method can be used as standalone or interfaced with MS to achieve the highest possible analytical coverage in the characterization and quantification of impurities present in single- and double-stranded ON formulations. Some of these techniques have been in use for some time and the importance of others is just being recognized.

  17. Quantification of in vitro mineralisation using ion chromatography. (United States)

    Souter, Paul; Horner, Alan; Cunningham, Jim C


    Analysis of in vitro mineralisation is an important tool in orthopedic research, allowing assessment of new therapeutic agents and devices; however, access to analytical equipment and accuracy of current methods can be a limiting factor. This current work investigated the use of calcium chelation with citric acid and subsequent analysis by ion chromatography as a method for accurately quantifying the extent of in vitro calcium deposition. Primary human osteoblasts were cultured on tissue culture plastic for 21 days under osteogenic conditions. At 3, 7, 14, and 21 days, alizarin red staining and citric acid calcium chelation of the cultures were performed. The use of alizarin red revealed increased calcium deposition over the culture period but was not sensitive enough to detect mineralisation at early time points after taking in to account background residual staining. The use of ion chromatography gave a limit of detection of 2 μg calcium, sensitive enough to detect mineralisation after 3 days, with no issues relating to background levels. We believe that the use of ion chromatography for quantifying in vitro mineralisation gives researchers an accurate, accessible, and cheap way of assessing novel technologies.

  18. Protein purification by aminosquarylium cyanine dye-affinity chromatography. (United States)

    Silva, M S; Graça, V C; Reis, L V; Santos, P F; Almeida, P; Queiroz, J A; Sousa, F


    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.

  19. Principles of Micellar Electrokinetic Capillary Chromatography Applied in Pharmaceutical Analysis

    Directory of Open Access Journals (Sweden)

    Árpád Gyéresi


    Full Text Available Since its introduction capillary electrophoresis has shown great potential in areas where electrophoretic techniques have rarely been used before, including here the analysis of pharmaceutical substances. The large majority of pharmaceutical substances are neutral from electrophoretic point of view, consequently separations by the classic capillary zone electrophoresis; where separation is based on the differences between the own electrophoretic mobilities of the analytes; are hard to achieve. Micellar electrokinetic capillary chromatography, a hybrid method that combines chromatographic and electrophoretic separation principles, extends the applicability of capillary electrophoretic methods to neutral analytes. In micellar electrokinetic capillary chromatography, surfactants are added to the buffer solution in concentration above their critical micellar concentrations, consequently micelles are formed; micelles that undergo electrophoretic migration like any other charged particle. The separation is based on the differential partitioning of an analyte between the two-phase system: the mobile aqueous phase and micellar pseudostationary phase. The present paper aims to summarize the basic aspects regarding separation principles and practical applications of micellar electrokinetic capillary chromatography, with particular attention to those relevant in pharmaceutical analysis.

  20. Principles of micellar electrokinetic capillary chromatography applied in pharmaceutical analysis. (United States)

    Hancu, Gabriel; Simon, Brigitta; Rusu, Aura; Mircia, Eleonora; Gyéresi, Arpád


    Since its introduction capillary electrophoresis has shown great potential in areas where electrophoretic techniques have rarely been used before, including here the analysis of pharmaceutical substances. The large majority of pharmaceutical substances are neutral from electrophoretic point of view, consequently separations by the classic capillary zone electrophoresis; where separation is based on the differences between the own electrophoretic mobilities of the analytes; are hard to achieve. Micellar electrokinetic capillary chromatography, a hybrid method that combines chromatographic and electrophoretic separation principles, extends the applicability of capillary electrophoretic methods to neutral analytes. In micellar electrokinetic capillary chromatography, surfactants are added to the buffer solution in concentration above their critical micellar concentrations, consequently micelles are formed; micelles that undergo electrophoretic migration like any other charged particle. The separation is based on the differential partitioning of an analyte between the two-phase system: the mobile aqueous phase and micellar pseudostationary phase. The present paper aims to summarize the basic aspects regarding separation principles and practical applications of micellar electrokinetic capillary chromatography, with particular attention to those relevant in pharmaceutical analysis.

  1. Partial characterization of hog renin purified by affinity chromatography. (United States)

    Devaux, C; Ménard, J; Sicard, P; Corvol, P


    A method has been set up to purify renin on a large scale by affinity chromatography using Pepstatin, a potent inhibitor of renin, as a ligand. Pepstatin was covalently coupled to Sepharose via six different spacer 'arms'. The Sepharose-hexamethylenediamino-Pepstatin appeared to be the better derivative for renin purification even at a concentration as low as 160 nmol of Pepstatin/ml of moist gel. Renin was extracted from 100 kg of hog kidneys and semi-purified by ammonium sulfate precipitations and chromatography on DEAE-cellulose. The active fraction (48.5 g of proteins) was applied on a 500-ml affinity column. Renin was eluted in the starting buffer containing 6 M urea. Renin was purified 120-fold by the affinity chromatography step with a 79% recovery. Physico-chemical characterization of highly purified renin was performed. Isoelectrofocusing on a pH gradient from 3 to 6 showed a major peak with an isoelectric point (pI) of 4.95 and a minor peak (pI = 4.70). Polyacrylamide gel electrophoresis, pH 7.8, at different gel concentrations, showed a single peak of renin activity which was found in the major protein band. Molecular size estimated on agarose-acrylamide gel filtration was 40 000. All these physical parameters were similar before and after purification.

  2. Comparison of different transition metal ions for immobilized metal affinity chromatography of selenoprotein P from human plasma

    DEFF Research Database (Denmark)

    Sidenius, U; Farver, O; Jøns, O


    Cu2+, Ni2+, Zn2+, Co2+ and Cd2+ were evaluated in metal ion affinity chromatography for enrichment of selenoprotein P, and immobilized Co2+ affinity chromatography was found to be the most selective chromatographic method. The chromatography was performed by fast protein liquid chromatography and...

  3. Identification of Explosives from Porous Materials: Applications Using Reverse Phase High Performance Liquid Chromatography and Gas Chromatography

    Energy Technology Data Exchange (ETDEWEB)

    C.J. Miller; G. Elias; N.C. Schmitt; C. Rae


    High performance liquid chromatography and gas chromatography techniques are well documented and widely used for the detection of trace explosives from organic solvents. These techniques were modified to specifically identify and quantify explosives extracted from various materials taken from people who had recently handled explosives. Documented techniques were modified to specifically detect and quantify RDX, TNT, and PETN from denim, colored flannel, vinyl, and canvas extracted in methanol using no sample cleanup prior to analysis. The methanol extracts were injected directly into several different column types and analyzed by HPLC-UV and/or GC-ECD. This paper describes general screening methods that were used to determine the presence of explosives in unknown samples and techniques that have been optimized for quantification of each explosive from the substrate extracts.

  4. Analyzing salvia divinorum and its active ingredient salvinorin a utilizing thin layer chromatography and gas chromatography/mass spectrometry. (United States)

    Jermain, John D; Evans, Hiram K


    In recent years, Salvia divinorum has become a major focus by state legislatures throughout the United States looking to prohibit the sale of the psychoactive plant. After researching testing procedures presented in the literature and those employed by crime laboratories throughout the country, it was decided that thin layer chromatography (TLC) and gas chromatography/mass spectrometry (GC/MS) were the methods to use to analyze plant material for salvinorin A. With TLC, salvinorin A was detected from extracted plant material and was easily distinguishable from 13 other Salvia species as well as Cannabis sativa L. (marijuana). When using GC/MS, salvinorin A was best extracted from plant material with chloroform at ambient temperature when using a nonpolar solvent and acetone at ambient temperature when using a polar solvent. By utilizing these techniques, criminalists are now able to confirm the presence of salvinorin A in a submitted plant material suspected to be Salvia divinorum.

  5. Isolation and purification of six iridoid glycosides from gardenia jasminoides fruit by medium-pressure liquid chromatography combined with macroporous resin chromatography. (United States)

    Wang, Yun; Liu, Hui; Shen, Lifeng; Yao, Lan; Ma, Yinlian; Yu, Dingrong; Chen, Jianhong; Li, Puling; Chen, Ying; Zhang, Cun


    Gardeniae fructus is one of the most frequently used herbs in traditional Chinese medicine. In the present study, a process for the enrichment of six iridoid glycosides from Gardeniae fructus was developed using medium-pressure liquid chromatography combined with macroporous resin and reversed-phase chromatography. The purities of different fractions from Gardeniae fructus were assessed using quantitative high-performance liquid chromatography. After fractionation using HPD-100 column chromatography, a 30% ethanol fraction was selected based on high-performance liquid chromatography and liquid chromatography with mass spectrometry qualitative analysis to separate and purify. Based on the orientation analysis results, six compounds-deacetyl asperulosidic acid methyl ester, gardenoside, ixoroside, scandoside methyl ester, genipin-1-O-β-d-gentiobioside, and geniposide-were successfully isolated and purified in three to four combined steps from Gardeniae fructus. The purities of these compounds were found by high-performance liquid chromatography analysis to be 97.9, 98.1, 95.5, 96.3, 97.1, and 98.7%, respectively. Moreover, their structures were elucidated by NMR spectroscopy and liquid chromatography with tandem mass spectrometry. The separation process was highly efficient, rapid, and accurate, making it a potential approach for the large-scale production of iridoids in the laboratory and providing several marker compounds for quality control. This procedure may be meaningful for the purification of other natural products used in traditional Chinese medicine.

  6. Investigation of Pinus mugo essential oil oxygenated fraction by combined use of gas chromatography and dry column chromatography. (United States)

    A, M B; Coran, S A; Giannellini, V; Vincieri, F F; Moneti, G


    The oxygenated compounds of Pinus mugo Turra essential oil were investigated by a combination of GC and dry column chromatography (DCC) coordinated by GC data processing. The collected data resulted in a bar graph ("normalized" gas chromatogram) giving the RRT's and relative amounts of 68 components; 38 of them were identified by MS and IR. The described procedure may be used for essential oil analysis in general.

  7. Zonal rate model for stacked membrane chromatography part II: characterizing ion-exchange membrane chromatography under protein retention conditions. (United States)

    Francis, Patrick; von Lieres, Eric; Haynes, Charles


    The Zonal Rate Model (ZRM) has previously been shown to accurately account for contributions to elution band broadening, including external flow nonidealities and radial concentration gradients, in ion-exchange membrane (IEXM) chromatography systems operated under nonbinding conditions. Here, we extend the ZRM to analyze and model the behavior of retained proteins by introducing terms for intra-column mass transfer resistances and intrinsic binding kinetics. Breakthrough curve (BTC) data from a scaled-down anion-exchange membrane chromatography module using ovalbumin as a model protein were collected at flow rates ranging from 1.5 to 20 mL min(-1). Through its careful accounting of transport nonidealities within and external to the membrane stack, the ZRM is shown to provide a useful framework for characterizing putative protein binding mechanisms and models, for predicting BTCs and complex elution behavior, including the common observation that the dynamic binding capacity can increase with linear velocity in IEXM systems, and for simulating and scaling separations using IEXM chromatography. Global fitting of model parameters is used to evaluate the performance of the Langmuir, bi-Langmuir, steric mass action (SMA), and spreading-type protein binding models in either correlating or fundamentally describing BTC data. When combined with the ZRM, the bi-Langmuir, and SMA models match the chromatography data, but require physically unrealistic regressed model parameters to do so. In contrast, for this system a spreading-type model is shown to accurately predict column performance while also providing a realistic fundamental explanation for observed trends, including an observed increase in dynamic binding capacity with flow rate.

  8. Displacement chromatography of proteins using a retained pH front in a hydrophobic charge induction chromatography column. (United States)

    Pinto, N D S; Frey, Douglas D


    The chromatographic separation of two proteins into a displacement train of two adjoined rectangular bands was accomplished using a novel method for hydrophobic charge induction chromatography (HCIC) which employs a self-sharpening pH front as the displacer. This method exploits the fact that protein elution in HCIC is promoted by a pH change, but is relatively independent of salt effects, so that a retained pH front can be used in place of a traditional displacer in displacement chromatography. The retained pH front was produced using the two adsorbed buffering species tricine and acetic acid. The separation of lysozyme and α-chymotrypsinogen A into adjoined, rectangular bands was accomplished with overall recoveries based on the total mass injected greater than 90 and 70%, respectively. The addition of urea to the buffer system increased the sharpness of the pH front by 36% while the yields of lysozyme and α-chymotrypsinogen A based on the total mass eluted increased from 76% to 99% and from 37% to 85%, respectively, when the purities of both proteins in their product fractions were fixed at 85%. The results demonstrate that the method developed in this study is a useful variant of HCIC and is also a useful alternative to other displacement chromatography methods.

  9. Profiling of drug binding proteins by monolithic affinity chromatography in combination with liquid chromatography-tandem mass spectrometry. (United States)

    Zhang, Xuepei; Wang, Tongdan; Zhang, Hanzhi; Han, Bing; Wang, Lishun; Kang, Jingwu


    A new approach for proteome-wide profiling drug binding proteins by using monolithic capillary affinity chromatography in combination with HPLC-MS/MS is reported. Two immunosuppresive drugs, namely FK506 and cyclosporin A, were utilized as the experimental models for proof-of-concept. The monolithic capillary affinity columns were prepared through a single-step copolymerization of the drug derivatives with glycidyl methacrylate and ethylene dimethacrylate. The capillary chromatography with the affinity monolithic column facilitates the purification of the drug binding proteins from the cell lysate. By combining the capillary affinity column purification and the shot-gun proteomic analysis, totally 33 FK506- and 32 CsA-binding proteins including all the literature reported target proteins of these two drugs were identified. Among them, two proteins, namely voltage-dependent anion-selective channel protein 1 and serine/threonine-protein phosphatase PGAM5 were verified by using the recombinant proteins. The result supports that the monolithic capillary affinity chromatography is likely to become a valuable tool for profiling of binding proteins of small molecular drugs as well as bioactive compounds.

  10. Hyphenated and comprehensive liquid chromatography × gas chromatography-mass spectrometry for the identification of Mycobacterium tuberculosis. (United States)

    Mourão, Marta P B; Denekamp, Ilse; Kuijper, Sjoukje; Kolk, Arend H J; Janssen, Hans-Gerd


    Tuberculosis is one of the world's most emerging public health problems, particularly in developing countries. Chromatography based methods have been used to tackle this epidemic by focusing on biomarker detection. Unfortunately, interferences from lipids in the sputum matrix, particularly cholesterol, adversely affect the identification and detection of the marker compounds. The present contribution describes the serial combination of normal phase liquid chromatography (NPLC) with thermally assisted hydrolysis and methylation followed by gas chromatography-mass spectrometry (THM-GC-MS) to overcome the difficulties of biomarker evaluation. The in-series combination consists of an LC analysis where fractions are collected and then transferred to the THM-GC-MS system. This was either done with comprehensive coupling, transferring all the fractions, or with hyphenated interfacing, i.e. off-line multi heart-cutting, transferring only selected fractions. Owing to the high sensitivity and selectivity of LC as a sample pre-treatment method, and to the high specificity of the MS as a detector, this analytical approach, NPLC × THM-GC-MS, is extremely sensitive. The results obtained indicate that this analytical set-up is able to detect down to 1 × 10(3) mycobacteria/mL of Mycobacterium tuberculosis strain 124, spiked in blank sputum samples. It is a powerful analytical tool and also has great potential for full automation. If further studies demonstrate its usefulness when applied blind in real sputum specimens, this technique could compete with the current smear microscopy in the early diagnosis of tuberculosis.

  11. Cation exchange displacement batch chromatography of proteins guided by screening of protein purification parameters. (United States)

    Kotasińska, Marta; Richter, Verena; Thiemann, Joachim; Schlüter, Hartmut


    Displacement chromatography has been shown to be an effective alternative for protein purification. We investigated in this study sample displacement chromatography, which does not require a displacer molecule. Furthermore, we performed a screening for determination of parameters for an optimal sample displacement chromatography. We screened the affinities of cytochrome C, lysozyme, myoglobin, and ribonuclease A toward a cation exchange material as a function of different pH values and to presence of different concentrations of sodium chloride in the sample application buffer. Sample displacement chromatography in batch chromatography mode for the separation of the protein mixture was studied with a sample application buffer with a pH of 5 and 7. As predicted by the screening experiments, sample displacement chromatography was most effective at pH 7 since this pH guaranteed the largest differences of the affinities of the four proteins toward the stationary phase. In summary, we describe here sample displacement chromatography in the batch chromatography mode for the separation of proteins, which is a simple and fast alternative to conventional displacement chromatography. Systematic screening of chromatographic parameters prior to sample displacement chromatography promises a successful separation of a target protein.

  12. Determination of Selected Colored Smokes on Glass Fiber Discs by High Performance Liquid Chromatography (HPLC) (United States)


    High Performance Liquid Chromatography (HPLC) 12. PERSONAL AUTHOR(S) F F_ n.ipl’prifl. Alan R...GROUP SUB-GROUP High Performance Liquid Chromatography (HPLC), Analytical IMethod, 1,4-diamino-2,3-dihydroanthraquinone, 2-(2 - _ quinolinyl)-1,3...weights, low vapor pressures and low thermal stability. High performance liquid chromatography (HPLC) appears to be the analytical method of choice

  13. [Use of column and thin layer chromatography for detection of vanillyl mandelic acid in urine]. (United States)

    Riabichenko, V V; Barchukov, V G; Chernyĭ, A V; Salenko, Iu A


    The excretion of vanillylmandelic acid was measured by column chromatography of urinary samples on aluminum oxide with subsequent thin-layer chromatography on silica gel. Use of aluminum oxide allowed application of greater urine samples (up to 0.1% of 24-h diuresis) onto chromatographic plates and essentially improved the quality of separation of vanillylmandelic acid from other phenylcarbonic acids by thin-layer chromatography, as well as the specificity and reproducibility of measurements.

  14. Thermal expansion pump for capillary high-performance liquid chromatography. (United States)

    Tao, Qian; Wu, Qian; Zhang, Xiangmin


    A thermal expansion pump (TEP) based on a principle of liquid thermal expansion for capillary high-performance liquid chromatography has been developed. The novel pump is capable of generating a continuous flow at high pressure for constant and stable delivery of binary solvents from nanoliters to microliters per minute without splitting. Theoretical equations for controlling fluidic output of this pump have been established and validated by a series of experiments. Factors affecting flow rate, such as density discrepancy, liquid compressibility, and mass loss in output, were taken into account. An assembly of the pump system employing two groups of thermal expansion pumps (TEPs) working in turns were fabricated, and a controlling strategy for the pump system to maintain a continuous delivery without pressure fluctuation even at switching points was also developed. Both isocratic and gradients of binary solvent delivery by the TEPs were performed. Reproducibility and standard deviation at different flow rates were determined. A capillary high-performance liquid chromatography (micro-HPLC) system consisting of the TEPs, an injection valve, a homemade packed capillary column (20 cm x 100 microm i.d. with 5 microm C18), and a laser-induced fluorescence detector was set up, and sample separations were carried out. Results of RSD = 4% for flow and RSD = 2% for retention times at 500 nL/min were achieved. Such a pump system has almost no moving parts except for the solvent switches. Its overall costs of manufacture and running are very low. It is proven that the TEPs system has great potential and competitive capabilities in capillary liquid chromatography.

  15. Enabling Microliquid Chromatography by Microbead Packing of Microchannels (United States)

    Balvin, Manuel; Zheng, Yun


    The microbead packing is the critical element required in the success of on-chip microfabrication of critical microfluidic components for in-situ analysis and detection of chiral amino acids. In order for microliquid chromatography to occur, there must be a stationary phase medium within the microchannel that interacts with the analytes present within flowing fluid. The stationary phase media are the microbeads packed by the process discussed in this work. The purpose of the microliquid chromatography is to provide a lightweight, low-volume, and low-power element to separate amino acids and their chiral partners efficiently to understand better the origin of life. In order to densely pack microbeads into the microchannels, a liquid slurry of microbeads was created. Microbeads were extracted from a commercially available high-performance liquid chromatography column. The silica beads extracted were 5 microns in diameter, and had surface coating of phenyl-hexyl. These microbeads were mixed with a 200- proof ethanol solution to create a microbead slurry with the right viscosity for packing. A microfilter is placed at the outlet via of the microchannel and the slurry is injected, then withdrawn across a filter using modified syringes. After each injection, the channel is flushed with ethanol to enhance packing. This cycle is repeated numerous times to allow for a tightly packed channel of microbeads. Typical microbead packing occurs in the macroscale into tubes or channels by using highly pressurized systems. Moreover, these channels are typically long and straight without any turns or curves. On the other hand, this method of microbead packing is completed within a microchannel 75 micrometers in diameter. Moreover, the microbead packing is completed into a serpentine type microchannel, such that it maximizes microchannel length within a microchip. Doing so enhances the interactions of the analytes with the microbeads to separate efficiently amino acids and amino acid

  16. Quantification of tryptophan in plasma by high performance liquid chromatography

    Directory of Open Access Journals (Sweden)

    Renata Romanholi Pinhati


    Full Text Available A simple, rapid and selective method using high-performance liquid chromatography with ultraviolet detection (267 nm was applied for the determination of tryptophan in plasma. Separation was carried out on a C18 column (150 x 4.6 mm internal diameter in 6 min. The mobile phase consisted of 5 mM the sodium acetate and acetonitrile (92:8, v/v. The method was shown to be precise and accurate, and good recovery of analyte was achieved, characterizing the method as efficient and reliable for use in laboratory analysis.

  17. Survey of organic acid eluents for anion chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Book, D.E.


    Of all the potential eluents surveyed (including aromatic, sulfonic, phosphonic, among other acids), only the carboxylic acids and the nitrophenols are recommended as eluents for anion chromatography. The concentration of the eluent should be in the range 5 x 10/sup -5/ to 1 x 10/sup -3/ M. The eluent should have the same charge as inorganic anions, a higher charge than organic acid samples. Choice of eluents for separation of halides, chloride and sulfate, multivalent inorganic anions, small alkyl acids, and aromatic acids is discussed. (DLC)

  18. Separation of zirconium by thin-layer chromatography. (United States)

    Oguma, K


    The thin-layer Chromatographie separation of a number of metal ions [Sc, Y, Zr, La, Sm, Th, U(VI), etc.] with solvent mixtures of mesityl oxide, ethanol and 5M nitric acid on silica gel-cellulose (5:1) thin-layer plates is reported. Zirconium remains stationary whilst the other metal ions move with the solvent, thus allowing a selective separation of zirconium from about 20 metal ions in ratios ranging from 100:1 to 1:100. Mixtures of various metal ions can also be separated.

  19. Simultaneous determination of inorganic and organic anions by ion chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Park, Yang Soon; Joe, Ki Soo; Han, Sun Ho; Park, Soon Dal; Choi, Kwang Soon


    Four methods were investigated for the simultaneous determination of several inorganic and organic anions in aqueous solution by ion chromatography. The first is two columns coupled system. The second is the gradient elution system with an anion exchange column. The third is the system with a mixed-mode stationary phase. The fourth is the system with an anion exchange column and the eluant of low conductivity without ion suppressor. The advantages and disadvantages of individual systems were discussed. The suitable methods were proposed for the application to the samples of the nuclear power industry and the environment. (author)

  20. Analysis of uranium isotope separation by redox chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Fujine, S.; Naruse, Y.; Shiba, K.


    Uranium isotope separation by redox chromatography is analytically studied. The periodic withdrawal of products and tails and the introduction of natural feed are simulated on the assumption of a square cascade for a uranium adsorption band. The influences on the separative power and the lead time until product withdrawal are investigated by varying the magnitude of the isotope separation factor, uranium band length, tails concentration, and so on. Simulating calculations indicate that using ion-exchange resins to achieve uranium isotope separation requires a very long lead time for the production of highly enriched uranium.

  1. Separation of sulfoalkylated cyclodextrins with hydrophilic interaction liquid chromatography. (United States)

    Jaramillo, Michael; Kirschner, Daniel L; Dai, Zhipeng; Green, Thomas K


    Determination of the charged state distributions of partially- and fully-substituted sulfoalkylated cyclodextrins was achieved using hydrophilic interaction liquid chromatography (HILIC). HILIC analysis of a spiked sample of the partially sulfopropylated cyclodextrins was achieved using a gradient to baseline resolve the charged states from -1 to -14. The fully-substituted CDs yielded a major peak with some trace impurities and the partially-substituted sulfopropylated cyclodextrins showed a wide range of charge states present in the mixture. Small changes in the structure of the cyclodextrins have a significant impact on the retention times of the various types of cyclodextrins investigated.

  2. Separation of Aniline Derivatives by Micellar Electrokinetic Capillary Chromatography

    Institute of Scientific and Technical Information of China (English)

    Jun LI; Zhuo Bin YUAN


    A micellar electrokinetic capillary chromatography (MECC) was developed for the determination of aniline and 6 substituted anilines.The seven components were separated within 25 min in the buffer solution of 40 mmol/L sodium borate and 100 mmol/L SDS.It was found that the separation was dependent on operating voltage, pH value, borate and SDS concentrations.The analytical performance was examined in terms of linear response and reproducibility.Wastewater was determined by the established method.

  3. A novel ionic liquids grafted polysiloxane for capillary gas chromatography

    Institute of Scientific and Technical Information of China (English)

    Qing Quan Wei; Mei Ling Qi; Ruo Nong Fu


    A new ionic liquids grafted polysiloxane used as stationary phase for capillary gas chromatography(CGC)is described.The stationary phase of 1-vinyl-3-hexylimidazolium hexafluorophosphate anchored to polysiloxane(PMHS-[VHIm][PF6])was synthesized,characterized and coated onto capillary columns by static coating.The results show that the present stationary phase exhibits a very good chromatographic resolution and selectivity for Grob test mixture and alcohols with baseline resolution and symmetry peaks.The present work suggests that novel stationary phase has a great potential for further development and application.

  4. Gas-liquid chromatography in lunar organic analysis. (United States)

    Gehrke, C. W.


    Gas-liquid chromatography (GLC) is a powerful and sensitive method for the separation and detection of organic compounds at nanogram levels. The primary requirement for successful analyses is that the compounds of interest must be volatile under the chromatographic conditions employed. Nonvolatile organic compounds must be converted to volatile derivatives prior to analysis. The derivatives of choice must be both amenable to chromatographic separation and be relatively stable. The condition of volatility necessitates the development of efficient derivatization reactions for important groups of compounds as amino acids, carbohydrates, nucleosides, etc. Trimethylsilylation and trifluoroacetylation represent specific areas of recent prominence. Some relevant practical aspects of GLC are discussed.

  5. Purification of flavonoids from licorice using an off-line preparative two-dimensional normal-phase liquid chromatography/reversed-phase liquid chromatography method. (United States)

    Fan, Yunpeng; Fu, Yanhui; Fu, Qing; Cai, Jianfeng; Xin, Huaxia; Dai, Mei; Jin, Yu


    An orthogonal (71.9%) off-line preparative two-dimensional normal-phase liquid chromatography/reversed-phase liquid chromatography method coupled with effective sample pretreatment was developed for separation and purification of flavonoids from licorice. Most of the nonflavonoids were firstly removed using a self-made Click TE-Cys (60 μm) solid-phase extraction. In the first dimension, an industrial grade preparative chromatography was employed to purify the crude flavonoids. Click TE-Cys (10 μm) was selected as the stationary phase that provided an excellent separation with high reproducibility. Ethyl acetate/ethanol was selected as the mobile phase owing to their excellent solubility for flavonoids. Flavonoids co-eluted in the first dimension were selected for further purification using reversed-phase liquid chromatography. Multiple compounds could be isolated from one normal-phase fraction and some compounds with bad resolution in one-dimensional liquid chromatography could be prepared in this two-dimensional system owing to the orthogonal separation. Moreover, this two-dimensional liquid chromatography method was beneficial for the preparation of relatively trace flavonoid compounds, which were enriched in the first dimension and further purified in the second dimension. Totally, 24 flavonoid compounds with high purity were obtained. The results demonstrated that the off-line two-dimensional liquid chromatography method was effective for the preparative separation and purification of flavonoids from licorice.

  6. Monitoring of cefepime in human serum and plasma by micellar electrokinetic capillary chromatography: Improvement of sample preparation and validation by liquid chromatography coupled to mass spectrometry. (United States)

    Šestáková, Nela; Theurillat, Regula; Sendi, Parham; Thormann, Wolfgang


    Cefepime monitoring in deproteinized human serum and plasma by micellar electrokinetic capillary chromatography and liquid chromatography coupled to mass spectrometry in presence of other drugs is reported. For micellar electrokinetic capillary chromatography, sample preparation comprised dodecylsulfate protein precipitation at pH 4.5 using an increased buffer concentration compared to that of a previous assay and removal of hydrophobic compounds with dichloromethane. This provided robust conditions for cefepime analysis in the presence of sulfamethoxazole and thus enabled its determination in samples of patients that receive co-trimoxazole. The liquid chromatography assay is based upon use of a column with a pentafluorophenyl-propyl modified and multi-endcapped stationary phase and the coupling to electrospray ionization with a single quadrupole detector. The performances of both assays with multi-level internal calibration were assessed with calibration and control samples and both assays were determined to be robust. Cefepime levels monitored by micellar electrokinetic capillary chromatography in samples from patients that were treated with cefepime only and with cefepime and co-trimoxazole were found to compare well with those obtained by liquid chromatography coupled to mass spectrometry. Cefepime drug levels determined by micellar electrokinetic capillary chromatography could thereby be validated. This article is protected by copyright. All rights reserved.

  7. Fundamentals of the Theory of Capillary Gas-Liquid-Solid Chromatography (United States)

    Berezkin, Viktor G.; Zolotarev, P. P.


    It is shown that the known instances of the separation of volatile compounds actually refer to gas-liquid-solid chromatography and not gas-liquid chromatography, the walls of the capillary column fulfilling the function of the solid. The theories of the retention and spreading of the chromatographic bands in elution gas-liquid-solid chromatography are examined. The results of theoretical and experimental studies indicate the need to take into account the role of adsorption in capillary gas-liquid chromatography. The bibliography includes 140 references.

  8. Discussions about high performance liquid chromatography upgrading to ion chromatography%液相色谱升级改造为离子色谱问题探讨

    Institute of Scientific and Technical Information of China (English)

    施超欧; 马浩; 姚宝龙; 黄彩勇


    离子色谱作为色谱的一个分支,在原理和仪器结构上与液相色谱密切相关。本文总结了液相色谱升级为离子改造色谱的经验。%As a part of chromatography ,the functions and structures of instruments between high per-formance liquid chromatography and ion chromatography are closely related .In this paper ,HPLC upgra-ded to IC was discused .

  9. Thermodynamic modeling of protein retention in mixed-mode chromatography: An extended model for isocratic and dual gradient elution chromatography. (United States)

    Lee, Yi Feng; Graalfs, Heiner; Frech, Christian


    An extended model is developed to describe protein retention in mixed-mode chromatography based on thermodynamic principles. Special features are the incorporation of pH dependence of the ionic interaction on a mixed-mode resin and the addition of a water term into the model which enables one to describe the total number of water molecules released at the hydrophobic interfaces upon protein-ligand binding. Examples are presented on how to determine the model parameters using isocratic elution chromatography. Four mixed-mode anion-exchanger prototype resins with different surface chemistries and ligand densities were tested using isocratic elution of two monoclonal antibodies at different pH values (7-10) and encompassed a wide range of NaCl concentrations (0-5M). U-shape mixed-mode retention curves were observed for all four resins. By taking into account of the deprotonation and protonation of the weak cationic functional groups in these mixed-mode anion-exchanger prototype resins, conditions which favor protein-ligand binding via mixed-mode strong cationic ligands as well as conditions which favor protein-ligand binding via both mixed-mode strong cationic ligands and non-hydrophobic weak cationic ligands were identified. The changes in the retention curves with pH, salt, protein, and ligand can be described very well by the extended model using meaningful thermodynamic parameters like Gibbs energy, number of ionic and hydrophobic interactions, total number of released water molecules as well as modulator interaction constant. Furthermore, the fitted model parameters based on isocratic elution data can also be used to predict protein retention in dual salt-pH gradient elution chromatography.

  10. Method transfer from high-pressure liquid chromatography to ultra-high-pressure liquid chromatography. II. Temperature and pressure effects. (United States)

    Åsberg, Dennis; Samuelsson, Jörgen; Leśko, Marek; Cavazzini, Alberto; Kaczmarski, Krzysztof; Fornstedt, Torgny


    The importance of the generated temperature and pressure gradients in ultra-high-pressure liquid chromatography (UHPLC) are investigated and compared to high-pressure liquid chromatography (HPLC). The drug Omeprazole, together with three other model compounds (with different chemical characteristics, namely uncharged, positively and negatively charged) were used. Calculations of the complete temperature profile in the column at UHPLC conditions showed, in our experiments, a temperature difference between the inlet and outlet of 16 °C and a difference of 2 °C between the column center and the wall. Through van't Hoff plots, this information was used to single out the decrease in retention factor (k) solely due to the temperature gradient. The uncharged solute was least affected by temperature with a decrease in k of about 5% while for charged solutes the effect was more pronounced, with k decreases up to 14%. A pressure increase of 500 bar gave roughly 5% increase in k for the uncharged solute, while omeprazole and the other two charged solutes gave about 25, 20 and 15% increases in k, respectively. The stochastic model of chromatography was applied to estimate the dependence of the average number of adsorption/desorption events (n) and the average time spent by a molecule in the stationary phase (τs) on temperature and pressure on peak shape for the tailing, basic solute. Increasing the temperature yielded an increase in n and decrease in τs which resulted in less skew at high temperatures. With increasing pressure, the stochastic modeling gave interesting results for the basic solute showing that the skew of the peak increased with pressure. The conclusion is that pressure effects are more pronounced for both retention and peak shape than the temperature effects for the polar or charged compounds in our study.

  11. High performance mini-gas chromatography-flame ionization detector system based on micro gas chromatography column (United States)

    Zhu, Xiaofeng; Sun, Jianhai; Ning, Zhanwu; Zhang, Yanni; Liu, Jinhua


    Monitoring Volatile organic compounds (VOCs) was a very important measure for preventing environmental pollution, therefore, a mini gas chromatography (GC) flame ionization detector (FID) system integrated with a mini H2 generator and a micro GC column was developed for environmental VOC monitoring. In addition, the mini H2 generator was able to make the system explode from far away due to the abandoned use of a high pressure H2 source. The experimental result indicates that the fabricated mini GC FID system demonstrated high repeatability and very good linear response, and was able to rapidly monitor complicated environmental VOC samples.

  12. Resolving Isomeric Glycopeptide Glycoforms with Hydrophilic Interaction Chromatography (HILIC). (United States)

    Huang, Yining; Nie, Yongxin; Boyes, Barry; Orlando, Ron


    The ability to resolve glycans while attached to tryptic peptides would greatly facilitate glycoproteomics, as this would enable site-specific glycan characterization. Peptide/glycopeptide separations are typically performed using reversed-phase liquid chromatography (RPLC), where retention is driven by hydrophobic interaction. As the hydrophilic glycans do not interact significantly with the RPLC stationary phase, it is difficult to resolve glycopeptides that differ only in their glycan structure, even when these differences are large. Alternatively, glycans interact extensively with the stationary phases used in hydrophilic interaction chromatography (HILIC), and consequently, differences in glycan structure have profound chromatographic shifts in this chromatographic mode. Here, we evaluate HILIC for the separation of isomeric glycopeptide mixtures that have the same peptide backbone but isomeric glycans. Hydrophilic functional groups on both the peptide and the glycan interact with the HILIC stationary phase, and thus, changes to either of these moieties can alter the chromatographic behavior of a glycopeptide. The interactive processes permit glycopeptides to be resolved from each other based on differences in their amino acid sequences and/or their attached glycans. The separations of glycans in HILIC are sufficient to permit resolution of isomeric N-glycan structures, such as sialylated N-glycan isomers differing in α2-3 and α2-6 linkages, while these glycans remain attached to peptides.

  13. Determination of Free-Energy Relationships Using Gas Chromatography (United States)

    Snow, Nicholas H.


    By performing a few straightforward analyses on a gas chromatograph, it is possible to calculate the free energy, enthalpy, and entropy changes that occur when a compound transfers between the mobile and stationary phases. The partition theory of chromatography allows this transfer to be expressed as a chemical equlibrium. By calculating the equilibrium constant for this reaction from chromatographic retention times, the standard free energy change may be determined, and from this, the standard enthalpy and entropy changes. Also, by calculating these values at several temperatures for structurally related compounds, it is possible to explore the relationship between chromatographic retention, standard free energy, and the structure of a compound. These calculations were performed for groups of homologous alcohols, acetates, and hydrocarbons on packed and capillary column gas chromatographs, using both polar and non-polar columns, and on computer simulation software. It is seen that for homologous compounds, the relationship between standard free energy change in partitioning and hydrocarbon chain length for this reaction is linear. It is also seen that gas chromatography represents a useful tool for the calculation and comparison of thermodynamic properties of compounds and that straightforward exercise of this type allows training of students in chromatographic analysis, basic partition theory, thermodynamic relationships, and linear free energy relationships.

  14. Capillary supercritical fluid chromatography-mass spectrometry (SFC-MS)

    Energy Technology Data Exchange (ETDEWEB)

    Kalinoski, H.T.; Udseth, H.R.; Chess, E.K.; Smith, R.D.


    The physical and chemical characteristics of supercritical fluids have prompted the development of supercritical fluid chromatography (SFC) for the analysis of labile and less volatile compounds. High resolution chromatographic separations with efficiencies approaching those of gas chromatography and high speed analyses are possible in capillary SFC using pressure programming methods and narrow bore columns. Further refinement of the SFC-mass spectrometry interface (SFC-MS) provides the basis for extension to more polar fluid systems with greater solvating power and the selectivity and sensitivity of mass spectrometric detection. The use of polar modified fluids has been facilitated by advances in understanding of supercritical fluid phase behavior. Fluid mixtures have been prepared for analysis of more polar, higher molecular weight analytes, that allow mild chromatographic temperatures and allow full exploitation of selectivity offered through control of fluid pressure (i.e., density). Continuing development of the SFC-MS interface has led to designs which can be near routinely applied with fluids such as CO/sub 2/, and providing enhanced transport of truly nonvolatile compounds to the mass spectrometer ionization regions. These advances also include an SFC interface to a high resolution, dual electric magnetic sector instrument, allowing supercritical fluid solvents to be explited for on-line extraction-mass spectrometry for characterization of complex, often otherwise intractable, materials. 26 refs., 5 figs., 1 tab.

  15. Temperature-responsive chromatography for the separation of biomolecules. (United States)

    Kanazawa, Hideko; Okano, Teruo


    Temperature-responsive chromatography for the separation of biomolecules utilizing poly(N-isopropylacrylamide) (PNIPAAm) and its copolymer-modified stationary phase is performed with an aqueous mobile phase without using organic solvent. The surface properties and function of the stationary phase are controlled by external temperature changes without changing the mobile-phase composition. This analytical system is based on nonspecific adsorption by the reversible transition of a hydrophilic-hydrophobic PNIPAAm-grafted surface. The driving force for retention is hydrophobic interaction between the solute molecules and the hydrophobized polymer chains on the stationary phase surface. The separation of the biomolecules, such as nucleotides and proteins was achieved by a dual temperature- and pH-responsive chromatography system. The electrostatic and hydrophobic interactions could be modulated simultaneously with the temperature in an aqueous mobile phase, thus the separation system would have potential applications in the separation of biomolecules. Additionally, chromatographic matrices prepared by a surface-initiated atom transfer radical polymerization (ATRP) exhibit a strong interaction with analytes, because the polymerization procedure forms a densely packed polymer, called a polymer brush, on the surfaces. The copolymer brush grafted surfaces prepared by ATRP was an effective tool for separating basic biomolecules by modulating the electrostatic and hydrophobic interactions. Applications of thermally responsive columns for the separations of biomolecules are reviewed here.

  16. Structures of multidomain proteins adsorbed on hydrophobic interaction chromatography surfaces. (United States)

    Gospodarek, Adrian M; Sun, Weitong; O'Connell, John P; Fernandez, Erik J


    In hydrophobic interaction chromatography (HIC), interactions between buried hydrophobic residues and HIC surfaces can cause conformational changes that interfere with separations and cause yield losses. This paper extends our previous investigations of protein unfolding in HIC chromatography by identifying protein structures on HIC surfaces under denaturing conditions and relating them to solution behavior. The thermal unfolding of three model multidomain proteins on three HIC surfaces of differing hydrophobicities was investigated with hydrogen exchange mass spectrometry (HXMS). The data were analyzed to obtain unfolding rates and Gibbs free energies for unfolding of adsorbed proteins. The melting temperatures of the proteins were lowered, but by different amounts, on the different surfaces. In addition, the structures of the proteins on the chromatographic surfaces were similar to the partially unfolded structures produced in the absence of a surface by temperature as well as by chemical denaturants. Finally, it was found that patterns of residue exposure to solvent on different surfaces at different temperatures can be largely superimposed. These findings suggest that protein unfolding on various HIC surfaces might be quantitatively related to protein unfolding in solution and that details of surface unfolding behavior might be generalized.

  17. Multimodal chromatography: an efficient tool in downstream processing of proteins. (United States)

    Kallberg, Kristian; Johansson, Hans-Olof; Bulow, Leif


    Chromatography has become an indispensable tool for the purification of proteins. Since the regulatory demands on protein purity are expected to become stricter, the need for generating improved resins for chromatographic separations has increased. More advanced scientific investigations of protein structure/function relationships, in particular, have also been a driving force for generating more sophisticated chromatographic materials for protein separations. As a consequence, the development of alternative chromatographic strategies has been very rapid during the past decade and several new ligands have been designed and explored both in the laboratory and in large-scale industrial settings. This review describes some of these efforts using multimodal chromatography, where two or more physicochemical properties are used to enhance the specificity of the interactions between the protein and the ligand on the chromatographic matrix. In addition to experimental studies, computer modeling of ligand-protein binding has improved the design of ligands for protein recognition. The use of descriptors as well as in silico docking methods have been implemented to design multimodal resins in several instances.

  18. Micellar electrokinetic chromatography of organic and peroxide-based explosives. (United States)

    Johns, Cameron; Hutchinson, Joseph P; Guijt, Rosanne M; Hilder, Emily F; Haddad, Paul R; Macka, Mirek; Nesterenko, Pavel N; Gaudry, Adam J; Dicinoski, Greg W; Breadmore, Michael C


    CE methods have been developed for the analysis of organic and peroxide-based explosives. These methods have been developed for deployment on portable, in-field instrumentation for rapid screening. Both classes of compounds are neutral and were separated using micellar electrokinetic chromatography (MEKC). The effects of sample composition, separation temperature, and background electrolyte composition were investigated. The optimised separation conditions (25 mM sodium tetraborate, 75 mM sodium dodecyl sulfate at 25°C, detection at 200 nm) were applied to the separation of 25 organic explosives in 17 min, with very high efficiency (typically greater than 300,000 plates m(-1)) and high sensitivity (LOD typically less than 0.5 mg L(-1); around 1-1.5 μM). A MEKC method was also developed for peroxide-based explosives (10 mM sodium tetraborate, 100 mM sodium dodecyl sulfate at 25°C, detection at 200 nm). UV detection provided LODs between 5.5 and 45.0 mg L(-1) (or 31.2-304 μM), which is comparable to results achieved using liquid chromatography. Importantly, no sample pre-treatment or post-column reaction was necessary and the peroxide-based explosives were not decomposed to hydrogen peroxide. Both MEKC methods have been applied to pre-blast analysis and for the detection of post-blast residues recovered from controlled, small scale detonations of organic and peroxide-based explosive devices.

  19. Analysis of Some Biogenic Amines by Micellar Liquid Chromatography

    Directory of Open Access Journals (Sweden)

    Irena Malinowska


    Full Text Available Micellar liquid chromatography (MLC with the use of high performance liquid chromatography (HPLC was used to determine some physicochemical parameters of six biogenic amines: adrenaline, dopamine, octopamine, histamine, 2-phenylethylamine, and tyramine. In this paper, an influence of surfactant’s concentration and pH of the micellar mobile phase on the retention of the tested substances was examined. To determine the influence of surfactant’s concentration on the retention of the tested amines, buffered solutions (at pH 7.4 of ionic surfactant—sodium dodecyl sulfate SDS (at different concentrations with acetonitrile as an organic modifier (0.8/0.2 v/v were used as the micellar mobile phases. To determine the influence of pH of the micellar mobile phase on the retention, mobile phases contained buffered solutions (at different pH values of sodium dodecyl sulfate SDS (at 0.1 M with acetonitrile (0.8/0.2 v/v. The inverse of value of retention factor (1/ versus concentration of micelles ( relationships were examined. Other physicochemical parameters of solutes such as an association constant analyte—micelle (ma—and partition coefficient of analyte between stationary phase and water (hydrophobicity descriptor (swΦ were determined by the use of Foley’s equation.

  20. Ion chromatography in the manufacture of multilayer circuit boards (United States)

    Smith, Robert E.


    Ion chromatography (IC) has proven useful in analyzing chemical solutions used in the manufacture of multilayer circuit boards. Unlike other chemical quantification techniques, IC provides results on ions not expected in the production solutions. Thus, solution contamination and break-down products can be monitored in every phase of the circuit board manufacturing. During the first phase, epoxy laminates experience an etchback, first in chromic acid, which can be analyzed for trace chloride and sulfate, then in ammonium bifluoride/HCl, which can be analyzed for fluoride and chloride. Following a wet-blasting to roughen up the surface, 20 microinches of copper are deposited using an electroless bath. Again, IC is applicable for monitoring formate, tartarate, and sulfate levels. Next, an acid copper bath is used to electroplate the through holes with 0.001 inches of ductile copper. This bath is analyzed for trace chloride. Photoimaging is then performed, and the organic solvents used can be assayed for trace ionic chloride. Finally, a fluoroboric acid-based tin-lead bath is used to deposit a solderable alloy. This bath is analyzed for fluoroborate, tin, and lead. In addition, mobile phase ion chromatography (MPIC) is used to monitor the nonionic organic brighteners in the baths.

  1. Supercritical Fluid Chromatography- A Hybrid of GC and LC

    Directory of Open Access Journals (Sweden)

    Kaushal K Chandrul


    Full Text Available

    High performance specifications and unique functionality of chromatographic techniques is a demand of pharmaceutical industry and research. This leads to the origin of Supercritical Fluid Chromatography (SFC. It is a rapidly expanding analytical technique. The main feature that differentiates SFC from other chromatographic techniques is the replacement of either the liquid or gas mobile phase with a supercritical fluid mobile phase. It is considered a hybrid of GC and LC technique. High diffusion coefficient and low viscosity of supercritical fluids is responsible for high speed analysis, high efficiency and high sensitivity. Low mobile-phase flow rate, density programming and compatability with GC and LC detectors make SFC a versatile chromatographic technique in analytical research and development. It has a unique characteristic of analyzing thermo labile or non-volatile substances. This review highlights the role of supercritical fluid chromatography in the separation of polymers, thermally labile pesticides, fatty acids, metal chelates and organometallic compounds, chiral and achiral molecules, identification and analysis of polar samples, explosives, drugs of abuse and application of SFC in forensic science (fingerprinting. 

  2. Supercritical fluid chromatography-A Hybrid of GC and LC

    Directory of Open Access Journals (Sweden)

    Neha Sethi


    Full Text Available High performance specifications and unique functionality of chromatographic techniques is a demand of pharmaceutical industry and research. This leads to the origin of Supercritical Fluid Chromatography (SFC. It is a rapidly expanding analytical technique. The main feature that differentiates SFC from other chromatographic techniques is the replacement of either the liquid or gas mobile phase with a supercritical fluid mobile phase. It is considered a hybrid of GC and LC technique. High diffusion coefficient and low viscosity of supercritical fluids is responsible for high speed analysis, high efficiency and high sensitivity. Low mobile-phase flow rate, density programming and compatability with GC and LC detectors make SFC a versatile chromatographic technique in analytical re-search and development. It has a unique characteristic of analyzing thermo labile or non-volatile substances. This review highlights the role of supercritical fluid chromatography in the separation of polymers, thermally labile pesticides, fatty acids, metal chelates and organometallic compounds, chiral and achiral molecules, identification and analysis of polar samples, explosives, drugs of abuse and application of SFC in forensic science (fingerprint-ing.

  3. Microfluidic liquid chromatography system for proteomic applications and biomarker screening. (United States)

    Lazar, Iulia M; Trisiripisal, Phichet; Sarvaiya, Hetal A


    A microfluidic liquid chromatography (LC) system for proteomic investigations that integrates all the necessary components for stand-alone operation, i.e., pump, valve, separation column, and electrospray interface, is described in this paper. The overall size of the LC device is small enough to enable the integration of two fully functional separation systems on a 3 in. x 1 in. glass microchip. A multichannel architecture that uses electroosmotic pumping principles provides the necessary functionality for eluent propulsion and sample valving. The flow rates generated within these chips are fully consistent with the requirements of nano-LC platforms that are routinely used in proteomic applications. The microfluidic device was evaluated for the analysis of a protein digest obtained from the MCF7 breast cancer cell line. The cytosolic protein extract was processed according to a shotgun protocol, and after tryptic digestion and prefractionation using strong cation exchange chromatography (SCX), selected sample subfractions were analyzed with conventional and microfluidic LC platforms. Using similar experimental conditions, the performance of the microchip LC was comparable to that obtained with benchtop instrumentation, providing an overlap of 75% in proteins that were identified by more than two unique peptides. The microfluidic LC analysis of a protein-rich SCX fraction enabled the confident identification of 77 proteins by using conventional data filtering parameters, of 39 proteins with p screening applications.

  4. Permanent gas analysis using gas chromatography with vacuum ultraviolet detection. (United States)

    Bai, Ling; Smuts, Jonathan; Walsh, Phillip; Fan, Hui; Hildenbrand, Zacariah; Wong, Derek; Wetz, David; Schug, Kevin A


    The analysis of complex mixtures of permanent gases consisting of low molecular weight hydrocarbons, inert gases, and toxic species plays an increasingly important role in today's economy. A new gas chromatography detector based on vacuum ultraviolet (VUV) spectroscopy (GC-VUV), which simultaneously collects full scan (115-240 nm) VUV and UV absorption of eluting analytes, was applied to analyze mixtures of permanent gases. Sample mixtures ranged from off-gassing of decomposing Li-ion and Li-metal batteries to natural gas samples and water samples taken from private wells in close proximity to unconventional natural gas extraction. Gas chromatography separations were performed with a porous layer open tubular column. Components such as C1-C5 linear and branched hydrocarbons, water, oxygen, and nitrogen were separated and detected in natural gas and the headspace of natural gas-contaminated water samples. Of interest for the transport of lithium batteries were the detection of flammable and toxic gases, such as methane, ethylene, chloromethane, dimethyl ether, 1,3-butadiene, CS2, and methylproprionate, among others. Featured is the capability for deconvolution of co-eluting signals from different analytes.

  5. Protein Nitrogen Determination by Kjeldahl Digestion and Ion Chromatography. (United States)

    Wang, Hsiaoling; Pampati, Nagarani; McCormick, William M; Bhattacharyya, Lokesh


    We report development and validation of a simple, rapid, and accurate method for the quantitation of protein nitrogen, which combines Kjeldahl digestion and ion chromatography with suppressed conductivity detection and requires nanomolar amount of nitrogen in samples (≥10 μg protein). The mechanism of suppressed conductivity detection does not permit analysis of samples containing copper (present in Kjeldahl digestion solution) and aluminum (present in many vaccines as adjuvants) due to precipitation of their hydroxides within the suppressor. We overcame this problem by including 10 μM oxalic acid in Kjeldahl digests and in the eluent (30 mM methanesulfonic acid). The chromatography is performed using an IonPac CS-16 cation exchange column by isocratic elution. The method reduces the digestion time to less than 1 h and eliminates the distillation and titration steps of the Kjeldahl method, thereby reducing the analysis time significantly and improving precision and accuracy. To determine protein nitrogen in samples containing non-protein nitrogen, proteins are precipitated by a mixture of deoxycholate and trichloroacetic acid and the precipitates are analyzed after dissolving in KOH. The method is particularly useful for biological samples that are limited and can also be applied to food, environmental, and other materials.

  6. Modelling counter-current chromatography: a chemical engineering perspective. (United States)

    Kostanian, A E


    In conventional chromatography, a solute is usually viewed to be longitudinally transported only in the mobile phase, remaining longitudinally motionless in the stationary phase. In counter-current chromatography, both phases undergo intense mixing in the variable force field of a coil planet centrifuge and longitudinal dispersion of matter in the stationary phase is not to be excluded. To take into account longitudinal mixing in both phases, a cell model of chromatographic process is proposed in which the number of perfectly mixed cells n is determined by the rates of mixing in stationary (Ds) and mobile (Dm) phases by the equation n = LF/(2ADc)/(1 + Sf(lambda - 1)) with A = K(D)D(S)/Dm (F, L, Ac and KD are the mobile phase flow-rate, column length, column cross-section and distribution ratio, respectively). This equation has been derived by comparing the discontinuous cell model with continuous diffusion assuming equilibrium conditions. Parameter determination and their relationships are discussed.

  7. Purification of amide alkaloids from Piper longum L. using preparative two-dimensional normal-phase liquid chromatography × reversed-phase liquid chromatography. (United States)

    Li, Kuiyong; Zhu, Wenya; Fu, Qing; Ke, Yanxiong; Jin, Yu; Liang, Xinmiao


    A comprehensive off-line two-dimensional liquid chromatography (2D-LC) method coupling normal phase liquid chromatography (NPLC) and reversed phase liquid chromatography (RPLC) was developed for separation and purification of amide alkaloids from Piper longum L. In the first dimension, the crude alkaloid fractions were separated in NPLC mode and 20 fractions were collected. Then fractions 5-20 were selected for further purification in RPLC mode in the second dimension. The purities of RPLC fractions with similar structures were all identified accurately by ultra performance liquid chromatography (UPLC). In total, 28 compounds with high purity were obtained and their structures were comprehensively characterized by electrospray ionization-mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR) spectroscopy. The results demonstrate that this 2D NPLC × RPLC method with good orthogonality (58.3%) was effective for the preparative separation and purification of amide alkaloids from Piper longum L.

  8. Novel techniques for petroleum analysis. Vol. 2. Fundamenals of chromatography, instruments and application - an extension of the application according to environmental analysis and forensic analysis according to the hydrocarbons in petroleum. 2. new rev. and enl. ed.; Neue Mineraloelanalyse. Bd. 2. Chromatographie in Grundlagen, Geraeten und Anwendung - Anwendung erweitert um die Umwelt- und forensische Analyse bezueglich der Mineraloelkohlenwasserstoffe

    Energy Technology Data Exchange (ETDEWEB)

    Kaegler, S.H.; Schindlbauer, H. (eds.)


    The authors of the book under consideration report on the state-of-the-art of the chromatographic analysis technique and its applications. In particular the chapter of pure petroleum analysis was extended by the environmental analysis and forensic analysis. Following the introduction to chromatography the authors report on columnchromatography under standard conditions, high-pressure liquid chromatography (HPLC), ion chromatography, gel permeation chromatography, thin-film chromatography, gas chromatography as well as the linkage of gas chromatography with mass spectrometry.

  9. Evaluation between ultrahigh pressure liquid chromatography and high-performance liquid chromatography analytical methods for characterizing natural dyestuffs. (United States)

    Serrano, Ana; van Bommel, Maarten; Hallett, Jessica


    An evaluation was undertaken of ultrahigh pressure liquid chromatography (UHPLC) in comparison to high-performance liquid chromatography (HPLC) for characterizing natural dyes in cultural heritage objects. A new UHPLC method was optimized by testing several analytical parameters adapted from prior UHPLC studies developed in diverse fields of research. Different gradient elution programs were tested on seven UHPLC columns with different dimensions and stationary phase compositions by applying several mobile phases, flow rates, temperatures, and runtimes. The UHPLC method successfully provided more improved data than that achieved by the HPLC method. Indeed, even though carminic acid has shown circa 146% higher resolution with HPLC, UHPLC resulted in an increase of 41-61% resolution and a decrease of 91-422% limit of detection, depending on the dye compound. The optimized method was subsequently assigned to analyse 59 natural reference materials, in which 85 different components were ascribed with different physicochemical properties, in order to create a spectral database for future characterization of dyes in cultural heritage objects. The majority of these reference samples could be successfully distinguished with one single method through the examination of these compounds' retention times and their spectra acquired with a photodiode array detector. These results demonstrate that UHPLC analyses are extremely valuable for the acquisition of more precise chromatographic information concerning natural dyes with complex mixtures of different and/or closely related physicochemical properties, essential for distinguishing similar species of plants and animals used to colour cultural heritage objects.

  10. Differentiation of coloured inks of inkjet printer cartridges by thin layer chromatography and high performance liquid chromatography. (United States)

    Poon, N L; Ho, S S H; Li, C K


    Document examiners are frequently asked to determine whether or not a colour printout has originated from a particular inkjet printer. The printer can rarely be identified unless some unique defects or irregularities of the printer are present on the printout. However, it is possible to decipher the make and/or model of the printer by comparing the ink-profile of the questioned printout with that of a seized inkjet printer cartridge or from one in a database. This paper presents an overview of a systematic approach to characterising and discriminating the inks of different inkjet printer cartridges using thin layer chromatography (TLC) and high pressure liquid chromatography (HPLC) with multi-wavelength ultra-violet/visible (UV/Vis) detection. Ink samples from 23 different inkjet printer cartridges (including originals and substitutes) of different brands and colour printouts, printed by known printers were examined with newly developed chromatographic methods. Subsequently, a database of the ink-profiles was generated. The methods provide a useful tool for discriminating coloured inks in inkjet printer cartridges of different brands.

  11. Phytochemical Profile of Erythrina variegata by Using High-Performance Liquid Chromatography and Gas Chromatography-Mass Spectroscopy Analyses. (United States)

    Muthukrishnan, Suriyavathana; Palanisamy, Subha; Subramanian, Senthilkumar; Selvaraj, Sumathi; Mari, Kavitha Rani; Kuppulingam, Ramalingam


    Natural products derived from plant sources have been utilized to treat patients with numerous diseases. The phytochemical constituents present in ethanolic leaf extract of Erythrina variegata (ELEV) were identified by using high-performance liquid chromatography (HPLC) and gas chromatography-mass spectroscopy (GC-MS) analyses. Shade dried leaves were powdered and extracted with ethanol for analyses through HPLC to identify selected flavonoids and through GC-MS to identify other molecules. The HPLC analysis of ELEV showed the presence of gallic and caffeic acids as the major components at concentrations of 2.0 ppm and 0.1 ppm, respectively, as well as other components. GC-MS analysis revealed the presence of 3-eicosyne; 3,7,11,15-tetramethyl-2-hexadecen-1-ol; butanoic acid, 3-methyl-3,7-dimethyl-6-octenyl ester; phytol; 1,2-benzenedicarboxylic acid, diundecyl ester; 1-octanol, 2-butyl-; squalene; and 2H-pyran, 2-(7-heptadecynyloxy) tetrahydro-derivative. Because pharmacopuncture is a new evolving natural mode that uses herbal extracts for treating patients with various ailments with minimum pain and maximum effect, the results of this study are particularly important and show that ELEV possesses a wide range of phytochemical constituents, as indicated above, as effective active principle molecules that can be used individually or in combination to treat patients with various diseases.

  12. Phytochemical Profile of Erythrina variegata by Using High-Performance Liquid Chromatography and Gas Chromatography-Mass Spectroscopy Analyses

    Directory of Open Access Journals (Sweden)

    Suriyavathana Muthukrishnan


    Full Text Available Natural products derived from plant sources have been utilized to treat patients with numerous diseases. The phytochemical constituents present in ethanolic leaf extract of Erythrina variegata (ELEV were identified by using high-performance liquid chromatography (HPLC and gas chromatography-mass spectroscopy (GC-MS analyses. Shade dried leaves were powdered and extracted with ethanol for analyses through HPLC to identify selected flavonoids and through GC-MS to identify other molecules. The HPLC analysis of ELEV showed the presence of gallic and caffeic acids as the major components at concentrations of 2.0 ppm and 0.1 ppm, respectively, as well as other components. GC-MS analysis revealed the presence of 3-eicosyne; 3,7,11,15-tetramethyl-2-hexadecen-1-ol; butanoic acid, 3-methyl-3,7-dimethyl-6-octenyl ester; phytol; 1,2-benzenedicarboxylic acid, diundecyl ester; 1-octanol, 2-butyl-; squalene; and 2H-pyran, 2-(7-heptadecynyloxy tetrahydro-derivative. Because pharmacopuncture is a new evolving natural mode that uses herbal extracts for treating patients with various ailments with minimum pain and maximum effect, the results of this study are particularly important and show that ELEV possesses a wide range of phytochemical constituents, as indicated above, as effective active principle molecules that can be used individually or in combination to treat patients with various diseases.

  13. Comprehensive characterization of Stevia rebaudiana using two-dimensional reversed-phase liquid chromatography/hydrophilic interaction liquid chromatography. (United States)

    Fu, Qing; Guo, Zhimou; Zhang, Xiuli; Liu, Yanfang; Liang, Xinmiao


    Two-dimensional reversed-phase liquid chromatography/hydrophilic interaction liquid chromatography (2D-RPLC/HILIC) system was successfully applied for comprehensive characterization of steviol glycosides from Stevia rebaudiana. The experiments were performed in offline mode using an XCharge C18 column in first dimension and an XAmide column in second dimension. In first dimension, preliminary separation of Stevia aqueous extract was accomplished and 30 fractions were collected. Then fractions 1-20 were selected for further purification and 13 compounds with high purity were obtained in second dimension. Comprehensive characterization of these compounds was completed by determination of their retention time, accurate molecular weight, diagnostic fragmentation ions, and nuclear magnetic resonance spectroscopy. As a result, all nine known steviol glycosides, as well as other four steviol glycosides were fully purified. The result demonstrated that this procedure is an effective approach for the preparative separation and comprehensive characterization of steviol glycosides in Stevia. This 2D-RPLC/HILIC method will be a promising tool for the purification of low-abundance compounds from natural products.

  14. Comparison of liquid chromatography-microchip/mass spectrometry to conventional liquid chromatography-mass spectrometry for the analysis of steroids. (United States)

    Ahonen, Linda; Keski-Rahkonen, Pekka; Saarelainen, Taija; Paviala, Jenni; Ketola, Raimo A; Auriola, Seppo; Poutanen, Matti; Kostianen, Risto


    The feasibility of a microfluidic-based liquid chromatography-electrospray ionization/mass spectrometric system (HPLC-Chip/ESI/MS) was studied and compared to a conventional narrow-bore liquid chromatography-electrospray ionization/mass spectrometric (LC-ESI/MS) system for the analysis of steroids. The limits of detection (LODs) for oxime derivatized steroids, expressed as concentrations, were slightly higher with the HPLC-Chip/MS system (50-300 pM) using an injection volume of 0.5 μL than with the conventional LC-ESI/MS (10-150 pM) using an injection volume of 40 μL. However, when the LODs are expressed as injected amounts, the sensitivity of the HPLC-Chip/MS system was about 50 times higher than with the conventional LC-ESI/MS system. The results indicate that the use of HPLC-Chip/MS system is clearly advantageous only in the analysis of low-volume samples. Both methods showed good linearity and good quantitative and chromatographic repeatability. In addition to the instrument comparisons with oxime derivatized steroids, the feasibility of the HPLC-Chip/MS system in the analysis of non-derivatized and oxime derivatized steroids was compared. The HPLC-Chip/MS method developed for non-derivatized steroids was also applied to the quantitative analysis of 15 mouse plasma samples.

  15. Simultaneous achiral-chiral analysis of pharmaceutical compounds using two-dimensional reversed phase liquid chromatography-supercritical fluid chromatography. (United States)

    Venkatramani, C J; Al-Sayah, Mohammad; Li, Guannan; Goel, Meenakshi; Girotti, James; Zang, Lisa; Wigman, Larry; Yehl, Peter; Chetwyn, Nik


    A new interface was designed to enable the coupling of reversed phase liquid chromatography (RPLC) and supercritical fluid chromatography (SFC). This online two-dimensional chromatographic system utilizing RPLC in the first dimension and SFC in the second was developed to achieve simultaneous achiral and chiral analysis of pharmaceutical compounds. The interface consists of an eight-port, dual-position switching valve with small volume C-18 trapping columns. The peaks of interest eluting from the first RPLC dimension column were effectively focused as sharp concentration pulses on small volume C-18 trapping column/s and then injected onto the second dimension SFC column. The first dimension RPLC separation provides the achiral purity result, and the second dimension SFC separation provides the chiral purity result (enantiomeric excess). The results are quantitative enabling simultaneous achiral, chiral analysis of compounds. The interface design and proof of concept demonstration are presented. Additionally, comparative studies to conventional SFC and case studies of the applications of 2D LC-SFC in pharmaceutical analysis is presented.

  16. Industrial application of green chromatography - II. Separation and analysis of preservatives in skincare products using subcritical water chromatography. (United States)

    Yang, Y; Kapalavavi, B; Gujjar, L; Hadrous, S; Marple, R; Gamsky, C


    Several high-temperature liquid chromatography (HTLC) and subcritical water chromatography (SBWC) methods have been successfully developed in this study for separation and analysis of preservatives contained in Olay skincare creams. Efficient separation and quantitative analysis of preservatives have been achieved on four commercially available ZirChrom and Waters XBridge columns at temperatures ranging from 100 to 200°C. The quantification results obtained by both HTLC and SBWC methods developed for preservatives analysis are accurate and reproducible. A large number of replicate HTLC and SBWC runs also indicate no significant system building-up or interference for skincare cream analysis. Compared with traditional HPLC separation carried out at ambient temperature, the HTLC methods can save up to 90% methanol required in the HPLC mobile phase. However, the SBWC methods developed in this project completely eliminated the use of toxic organic solvents required in the HPLC mobile phase, thus saving a significant amount of money and making the environment greener. Although both homemade and commercial systems can accomplish SBWC separations, the SBWC methods using the commercial system for preservative analysis are recommended for industrial applications because they can be directly applied in industrial plant settings.

  17. Recent advances in ultra-high performance liquid chromatography for the analysis of traditional chinese medicine (United States)

    Traditional Chinese medicines (TCMs) have been widely used for the prevention and treatment of various diseases for thousands of years in China. Ultra Performance Liquid Chromatography (UHPLC) is a relatively new technique offering new possibilities in liquid chromatography. This paper reviews recen...

  18. Using reaction flow chromatography for the analysis of amino acid: Derivatisation with fluorescamine reagent

    NARCIS (Netherlands)

    Pravadali-Cekic, S.; Jones, A.; Kazarian, A.A.; Paull, B.; Soliven, A.; Ritchie, H.; Camenzuli, M.; Dennis, G.R.; Shalliker, R.A.


    Reaction flow (RE) chromatography with fluorescamine reagent and UV-vis detection was used for the analysis of amino acids. The performance advantage of RF chromatography was tested against the conventional post-column derivatisation (PCD) methods. The results of the study verified that greater sens

  19. Identification of Cyclopentenyl Fatty Acids by Gas Liquid Chromatography and Mass Spectrometry

    DEFF Research Database (Denmark)

    Shukla, V. K. S.; Abdel-Moety, E. M.; Larsen, Elfinn;


    The straight chain fatty acids and the cyclopentenyl fatty acids present in the lipids of Hydnocarpus wightiana seeds were separated as their pyrrolidides by means of gas chromatography. A gas chromatography-mass spectrometry system confirmed the complete separation and permitted the identification...

  20. Determination of Vinyl Chloride at ug/l. Level in Water by Gas Chromatography (United States)

    Bellar, Thomas A.; And Others


    A quantitative method for the determination of vinyl chloride in water is presented. Vinyl chloride is transfered to the gas phase by bubbling inert gas through the water. After concentration on silica gel or Carbosieve-B, determination is by gas chromatography. Confirmation of vinyl chloride is by gas chromatography-mass spectrometry. (Author/BT)

  1. Determination of Roxarsone in feeds using solid phase extraction and liquid chromatography with ultraviolet detection. (United States)

    Sapp, R E; Davidson, S


    A method is presented for detection and quantitation of Roxarsone in poultry feed by liquid chromatography. The drug is extracted by phosphate buffer and determined by solid phase extraction and reversed-phase liquid chromatography. Recoveries of the sample spikes and fortified field samples agree closely with those obtained by the standard spectrophotometric method.

  2. Normal- and reverse-phase paper chromatography of leaf extracts of dandelions


    Du Toit, Maria H.; Eggen, Per-Odd; Kvittigen, Lise; Partali, Vassilia; Schmid, Rudolf


    This demonstration describes how normal and reverse phase chromatography can be illustrated using only chromatography paper for the separation of extracts of dandelions.

  3. Quasi-normal phase chromatography of nitrogen-containing adamantane derivatives (United States)

    Prokopov, S. V.; Tyrina, E. V.; Davankov, V. A.; Il'in, M. M.; Kurbatova, S. V.


    The chromatographic retention of adamantyl-containing amidrazones and triazoles is studied under the conditions of quasi-normal phase (QNP) and reversed phase high-performance liquid chromatography using partially sulfonated hypercrosslinked polystyrene as a stationary phase. The considerable effect of the sorbent-sorbate π interactions on the retention factor of the analytes, particularly under conditions of QNP chromatography, is revealed.

  4. An efficient hydrophilic interaction liquid chromatography separation of 7 phospholipid classes based on a diol column

    NARCIS (Netherlands)

    Zhu, C.; Dane, A.; Spijksma, G.; Wang, M.; Greef, J. van der; Luo, G.; Hankemeier, T.; Vreeken, R.J.


    A hydrophilic interaction liquid chromatography (HILIC) - ion trap mass spectrometry method was developed for separation of a wide range of phospholipids. A diol column which is often used with normal phase chromatography was adapted to separate different phospholipid classes in HILIC mode using a m

  5. 21 CFR 862.2270 - Thin-layer chromatography system for clinical use. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Thin-layer chromatography system for clinical use... Instruments § 862.2270 Thin-layer chromatography system for clinical use. (a) Identification. A thin-layer... a mixture. The mixture of compounds is absorbed onto a stationary phase or thin layer of...

  6. Isolation of Three Components from Spearmint Oil: An Exercise in Column and Thin-Layer Chromatography (United States)

    Davies, Don R.; Johnson, Todd M.


    A simple experiment for undergraduate organic chemistry students to separate a colorless mixture using column chromatography and then monitor the outcome of the separation using thin-layer chromatography (TLC) and infrared spectroscopy(IR) is described. The experiment teaches students the principle and techniques of column and thin-layer…

  7. New methods and materials for solid phase extraction and high performance liquid chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Dumont, Philip John [Iowa State Univ., Ames, IA (United States)


    This paper describes methods for solid phase extraction and high performance liquid chromatography (HPLC). The following are described: Effects of Resin Sulfonation on the Retention of Polar Organic Compounds in Solid Phase Extraction; Ion-Chromatographic Separation of Alkali Metals In Non-Aqueous Solvents; Cation-Exchange Chromatography in Non-Aqueous Solvents; and Silicalite As a Stationary Phase For HPLC.

  8. Using Aspen to Teach Chromatographic Bioprocessing: A Case Study in Weak Partitioning Chromatography for Biotechnology Applications (United States)

    Evans, Steven T.; Huang, Xinqun; Cramer, Steven M.


    The commercial simulator Aspen Chromatography was employed to study and optimize an important new industrial separation process, weak partitioning chromatography. This case study on antibody purification was implemented in a chromatographic separations course. Parametric simulations were performed to investigate the effect of operating parameters…

  9. Investigating the Retention Mechanisms of Liquid Chromatography Using Solid-Phase Extraction Cartridges (United States)

    O'Donnell, Mary E.; Musial, Beata A.; Bretz, Stacey Lowery; Danielson, Neil D.; Ca, Diep


    Liquid chromatography (LC) experiments for the undergraduate analytical laboratory course often illustrate the application of reversed-phase LC to solve a separation problem, but rarely compare LC retention mechanisms. In addition, a high-performance liquid chromatography instrument may be beyond what some small colleges can purchase. Solid-phase…

  10. Applications of nanomaterials in liquid chromatography: opportunities for separation with high efficiency and selectivity. (United States)

    Zhang, Zhengxiang; Wang, Zhiyong; Liao, Yiping; Liu, Huwei


    During recent decades, great efforts have been made to improve the chemical stability, selectivity, and separation efficiency of stationary phases in liquid chromatography. Significant progress has been achieved, especially after the introduction of nanomaterials into separation science. This review covers the applications of nanomaterials playing various roles in liquid chromatography. Future possibilities for developing nanomaterial-based stationary phases are also discussed.

  11. Preparative isolation and analysis of alcohol dehydrogenase inhibitors from Glycyrrhiza uralensis root using ultrafiltration combined with high-performance liquid chromatography and high-speed countercurrent chromatography. (United States)

    Chen, Miao; Liu, Liangliang; Chen, Xiaoqing


    A simple, rapid, and effective assay based on ultrafiltration combined with high-performance liquid chromatography and high-speed countercurrent chromatography was developed for screening and purifying alcohol dehydrogenase inhibitors from Glycyrrhiza uralensis root extract. Experiments were carried out to optimize binding conditions including alcohol dehydrogenase concentration, incubation time, temperature, and pH. By comparing the chromatograms, three compounds were found possessing alcohol dehydrogenase binding activity in Glycyrrhiza uralensis root. Under the target-guidance of ultrafiltration combined with the high-performance liquid chromatography experiment, liquiritin (1), isoliquiritin (2), and liquiritigenin (3) were separated by high-speed countercurrent chromatography using ethyl acetate/methanol/water (5:1:4) as the solvent system. The alcohol dehydrogenase inhibitory activities of these three isolated compounds were assessed; compound 2 showed strongest inhibitory activity with an IC50 of 8.95 μM. The results of the present study indicated that the combinative method using ultrafiltration, high-performance liquid chromatography and high-speed countercurrent chromatography could be widely applied for the rapid screening and isolation of enzyme inhibitors from complex mixtures.

  12. Analysis of allergens in tubeimu saponin extracts by using rat basophilic leukemia 2H3 cell-based affinity chromatography coupled to liquid chromatography and mass spectrometry. (United States)

    Zhang, Tao; Han, Shengli; Liu, Qi; Guo, Ying; He, Langchong


    An affinity two-dimensional chromatography method was developed for the recognition, separation, and identification of allergic components from tubeimu saponin extracts, a preparation often injected to treat various conditions as indicated by traditional Chinese medicine. Rat basophilic leukemia-2H3 cell membranes were used as the stationary phase of a membrane affinity chromatography column to capture components with affinity for mast cells that could be involved in a degranulation reaction. The retained components were enriched and analyzed by membrane affinity chromatography with liquid chromatography and mass spectrometry via a port switch valve. Suitability and reliability of the method was investigated using appropriate standards, and then, the method was applied to identify components retained from tubeimu saponin extracts. Tubeimoside A was identified in this way as a potential allergen, and degranulation assays confirmed that tubeimoside A induces RBL-2H3 cell degranulation in a dose-dependent manner. An increase in Ca(2+) influx indicated that degranulation induced by tubeimoside A is likely Ca(2+) dependent. Coupled with the degranulation assay, RBL-2H3 cell-based affinity chromatography coupled with liquid chromatography and mass spectrometry is an effective method for screening and identifying allergic components from tubeimu saponin extracts.

  13. Isolation and purification of uremic middle molecules by multi-step liquid chromatography

    Institute of Scientific and Technical Information of China (English)

    储结根; 何炳林; 刘晓航; 袁直


    Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and re-versed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular c

  14. Advances in Capillary Chromatography%毛细管色谱的进展

    Institute of Scientific and Technical Information of China (English)


    Capillary columns are used in both capillary liquid chromatography and capillary electrochromatography. The design for capillary liquid chromatography is discussed in comparison with capillary gas chromatography. The difference of diffusion coefficient in gas and liquid phase is a key role. The study for obtaining a high performance capillary liquid chromatography is discussed. Capillary electrochromatography is recently interesting for its instinct ability to realize a high performance chromatography. Capillary electrochromatography with and without pressurized flow is reviewed briefly. Instrumentation for capillary electrochromatography with pressurized flow is discussed. The port of splitting, and gradient elution of both solution and potential are described. The new findings of both the variation of column resistance and capacity factor according to the value of applied electric voltage are also discussed.

  15. Discrimination of eight chloramphenicol isomers by liquid chromatography tandem mass spectrometry in order to investigate the natural occurence of chloramphenicol

    NARCIS (Netherlands)

    Berendsen, B.J.A.; Zuidema, T.; Jong, de J.; Stolker, A.A.M.; Nielen, M.W.F.


    This paper describes the discrimination of eight different isomers of chloramphenicol (CAP), an antibiotic banned for use in food producing animals, by reversed phase and chiral liquid chromatography in combination with tandem mass spectrometric detection. Previously, by liquid chromatography couple

  16. Surface Characterization of Glass Fiber by Inverse Gas Chromatography

    Institute of Scientific and Technical Information of China (English)

    HUANG Xiaohua; LI Bin; SHI Baoli; GUO Xuefei; JIA Lina


    The surface properties of glass fiber were quantificationally analyzed by inverse gas chromatography (IGC). Five n-alkanes (C6, C7, C8, C9, and C10) were chosen as apolar probes to characterize the dispersive component of surface free energy. Trichloromethane (CHCl3), acetone,and tetrahydrofuran (THF) were chosen as polar probes to detect the Lewis acid-base parameters. It is found that the dispersive components of free energy are 32.3, 30.5, 27.5, and 26.9 Mj/m2 at 70,80, 90, and 100 ℃, respectively. The Lewis acidic number Ka of the glass fiber is 0.512 4, and the basic number Kb is 2.862. The results mean the glass fiber is a Lewis basic material.

  17. Separation of carbohydrates using hydrophilic interaction liquid chromatography. (United States)

    Fu, Qing; Liang, Tu; Li, Zhenyu; Xu, Xiaoyong; Ke, Yanxiong; Jin, Yu; Liang, Xinmiao


    A strategy was developed to rapidly evaluate chromatographic properties of hydrophilic interaction chromatography (HILIC) columns for separating carbohydrates. Seven HILIC columns (Silica, Diol, TSK Amide-80, XAmide, Click Maltose, Click β-CD, and Click TE-Cys columns) were evaluated by using three monosaccharide and seven disaccharides as probes. The influence of column temperature on the peak shape and tautomerization of carbohydrates, as well as column selectivity were investigated. The influence of surface charge property on the retention was also studied by using glucose, glucuronic acid, and glucosamine, which indicated that buffer salt concentration and pH value in mobile phase was necessary to control the ionic interactions between ionic carbohydrates and HILIC columns. According to evaluation results, the XAmide column was selected as an example to establish experimental schemes for separation of complex mixtures of oligosaccharide.

  18. Protein identification using nano liquid chromatography-tandem mass spectrometry. (United States)

    Negroni, Luc


    Tandem mass spectrometry is an efficient technique for the identification of peptides on the basis of their fragmentation pattern (MS/MS scan). It can generate individual spectra for each peptide, thereby creating a powerful tool for protein identification on the basis of peptide characterization. This important advance in automatic data acquisition has allowed an efficient association between liquid chromatography and tandem mass spectrometry, and the use of nanocolumns and nanoelectrospray ionization has dramatically increased the efficiency of this method. Now large sets of peptides can be identified at a femtomole level. At the end of the process, batch processing of the MS/MS spectra produces peptide lists that identify purified proteins or protein mixtures with high confidence.

  19. Hansen solubility parameters for polyethylene glycols by inverse gas chromatography. (United States)

    Adamska, Katarzyna; Voelkel, Adam


    Inverse gas chromatography (IGC) has been applied to determine solubility parameter and its components for nonionic surfactants--polyethylene glycols (PEG) of different molecular weight. Flory-Huggins interaction parameter (chi) and solubility parameter (delta(2)) were calculated according to DiPaola-Baranyi and Guillet method from experimentally collected retention data for the series of carefully selected test solutes. The Hansen's three-dimensional solubility parameters concept was applied to determine components (delta(d), delta(p), delta(h)) of corrected solubility parameter (delta(T)). The molecular weight and temperature of measurement influence the solubility parameter data, estimated from the slope, intercept and total solubility parameter. The solubility parameters calculated from the intercept are lower than those calculated from the slope. Temperature and structural dependences of the entopic factor (chi(S)) are presented and discussed.

  20. Micellar electrokinetic chromatography (MEKC) separation of furanonaphthoquinones from Tabebuia impetiginosa. (United States)

    Koyama, J; Morita, I; Kino, A; Tagahara, K


    The separation of nine furanonaphthoquinones by micellar electrokinetic chromatography (MEKC) is described. The running electrolytes used in this method were 0.03 M sodium dodecyl sulphate (SDS) in 0.09 M borate buffer (pH 9) containing 10% methanol, with an applied voltage of 20 kV. Application of this technique in the determination of the main furanonaphthoquinones, 5-hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione, 8-hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione, and 2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione, of Tabebuia impetiginosa is demonstrated in this paper.

  1. Liquid chromatography mass spectrometry for analysis of microbial metabolites

    DEFF Research Database (Denmark)

    Klitgaard, Andreas

    are still to be discovered. The main analytical technique used to investigate production of products from these diverse organisms is liquid-chromatography coupled to mass spectrometry (LC-MS). With the development of new and improved analytical instrumentation for chemical analysis, the time needed...... to human health. Because of this, methods for detection and analysis of these compounds are vital. Estimates suggest that there are around 1.5 million different fungal species on Earth, dwarfing the number of plants estimated to 300,000, meaning that there potentially are many more interesting compounds...... to perform a single analytical run has decreased, while the amount of information obtained from each of these analytical runs has increased drastically. Consequently, the limiting step in chemical analysis of a microorganism is no longer the analytical run itself, but rather analysis of the resulting data...

  2. Determination of sulphite in wines using suppressed ion chromatography. (United States)

    Yoshikawa, Kenji; Uekusa, Yuki; Sakuragawa, Akio


    Suppressed ion chromatography with the use of a conductivity detector was developed for the determination of sulphite ions in wine samples. When a mixed solution of sodium carbonate, sodium bicarbonate, and acetone was used as the mobile phase, simultaneous determination of eight inorganic anions (i.e., fluoride, chloride, nitrite, nitrate, sulphite, phosphate, sulphate, and thiosulphate) was completed in approximately 25 min. Linearity, reproducibility, and detection limits were determined for the proposed method. In the case of sulphite detection, a linear calibration curve with a good correlation coefficient of 0.9992 was obtained from the peak height of sulphite with a relative standard deviation (n = 6) 1.48%. In addition, the detection limit of sulphite was 0.27 mg/L at a signal-to-noise ratio of 3. Further, the developed method was applied for the determination of sulphite contained in several wine samples.

  3. Feedlot lamb meat fatty acids profile characterization employing gas chromatography

    Directory of Open Access Journals (Sweden)

    M.I. Cruz-Gonzalez


    Full Text Available Fat is an important constituent in diet, not only as an energy source, but for its essential fatty acids associated to fats in foods, considering that some polyunsaturated fatty acids like linoleic, linolenic and arachidonic cannot be synthesized by superior animals like humans. Scientific evidence show that the fatty acids ingest can affect the thrombotic tendency, cardiac rhythm, endothelial function systematic inflammation, insulin sensibility and oxidative stress. Samples from 21 ovine crossbreds from Pelibuey, Blackbelly, Dorper and Katahadin (40 kg average weight feed with corn based balanced diets were taken from loin area 18 h after refrigeration. Saturated and polyunsaturated fatty acids levels were analyzed by gas chromatography. Results in this work showed that the healthy fatty acids levels are higher as compared to saturated fatty acids levels, indicating that this meat can influence consumer’s buying choice decision regarded to their health.

  4. Purification of Chironex fleckeri venom components using Chironex immunoaffinity chromatography. (United States)

    Naguib, A M; Bansal, J; Calton, G J; Burnett, J W


    A comparison of the purification of the nematocyst venom of Chironex fleckeri by affinity immunochromatography using 13 different monoclonal antibodies was made. Varying degrees of purification of mouse lethal factor, hemolysin and dermonecrotic factors, as well as antigen positive proteins were achieved with each of the monoclonal antibodies. Although the protein curves of the chromatography were similar, each of the monoclonal antibody columns had a distinctive pharmacological and SDS-PAGE profile. At least two hemolysins (120,000 and 70,000 molecular weight), two dermonecrotic principles (120,000, less than 120,000) and three lethal factors (120,000, 70,000 and 14,500 molecular weight) were detected. The degree to which aggregation and fragmentation affects the molecular weights of these proteins is not known. It appears that multiple pharmacological activities are present within the same molecule since it is only with great difficulty that a pharmacological activity can be assigned to a specific molecular weight.

  5. Virus inactivation by protein denaturants used in affinity chromatography. (United States)

    Roberts, Peter L; Lloyd, David


    Virus inactivation by a number of protein denaturants commonly used in gel affinity chromatography for protein elution and gel recycling has been investigated. The enveloped viruses Sindbis, herpes simplex-1 and vaccinia, and the non-enveloped virus polio-1 were effectively inactivated by 0.5 M sodium hydroxide, 6 M guanidinium thiocyanate, 8 M urea and 70% ethanol. However, pH 2.6, 3 M sodium thiocyanate, 6 M guanidinium chloride and 20% ethanol, while effectively inactivating the enveloped viruses, did not inactivate polio-1. These studies demonstrate that protein denaturants are generally effective for virus inactivation but with the limitation that only some may inactivate non-enveloped viruses. The use of protein denaturants, together with virus reduction steps in the manufacturing process should ensure that viral cross contamination between manufacturing batches of therapeutic biological products is prevented and the safety of the product ensured.

  6. Superheated water chromatography--a green technology for the future. (United States)

    Smith, Roger M


    Reversed phase liquid chromatography using superheated water as the mobile phase, at temperatures between 100 and 250 degrees C, offers a number of advantages for the analyst. It is an environmentally clean solvent, reducing solvent usage and disposal costs. It has advantages in detection, allowing UV spectra to be monitored down to short wavelengths, as well as a compatibility with universal flame ionisation detection and mass spectroscopy. By employing deuterium oxide as the eluent, solvent free NMR spectra can be measured. The development of newer more thermally stable stationary phases, including hybrid phases, have expanded the analytes that can be examined and these now range from alkylbenzenes, phenols, alkyl aryl ketones and a number of pharmaceuticals to carboxylic acids, amino acids, and carbohydrates. Very few compounds have been found to be unstable during the analysis. The separation methods can be directly coupled to superheated water extraction providing a totally solvent free system for sample extraction and analysis.

  7. Estimation of brassylic acid by gas chromatography-mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Mohammed J. Nasrullah, Erica N. Pfarr, Pooja Thapliyal, Nicholas S. Dusek, Kristofer L. Schiele, Christy Gallagher-Lein, and James A. Bahr


    The main focus of this work is to estimate Brassylic Acid (BA) using gas chromatography-mass spectrometry (GC-MS). BA is a product obtained from the oxidative cleavage of Erucic Acid (EA). BA has various applications for making nylons and high performance polymers. BA is a 13 carbon compound with two carboxylic acid functional groups at the terminal end. BA has a long hydrocarbon chain that makes the molecule less sensitive to some of the characterization techniques. Although BA can be characterized by NMR, both the starting material (EA) and products BA and nonanoic acid (NA) have peaks at similar {delta}, ppm values. Hence it becomes difficult for the quick estimation of BA during its synthesis.

  8. Large scale protein separations: engineering aspects of chromatography. (United States)

    Chisti, Y; Moo-Young, M


    The engineering considerations common to large scale chromatographic purification of proteins are reviewed. A discussion of the industrial chromatography fundamentals is followed by aspects which affect the scale of separation. The separation column geometry, the effect of the main operational parameters on separation performance, and the physical characteristics of column packing are treated. Throughout, the emphasis is on ion exchange and size exclusion techniques which together constitute the major portion of commercial chromatographic protein purifications. In all cases, the state of current technology is examined and areas in need of further development are noted. The physico-chemical advances now underway in chromatographic separation of biopolymers would ensure a substantially enhanced role for these techniques in industrial production of products of new biotechnology.

  9. Nano-liquid chromatography in pharmaceutical and biomedical research. (United States)

    Gama, Mariana Roberto; Collins, Carol H; Bottoli, Carla B G


    Miniaturized separation techniques have emerged as environmentally friendly alternatives to available separation methods. Nano-liquid chromatography (nano-LC), microchip devices and nano-capillary electrophoresis are miniaturized methods that minimize reagent consumption and waste generation. Furthermore, the low levels of analytes, especially in biological samples, promote the search for more highly sensitive techniques; coupled to mass spectrometry, nano-LC has great potential to become an indispensable tool for routine analysis of biomolecules. This short review presents the fundamental aspects of nano-LC analytical instrumentation, discussing practical considerations and the primary differences between miniaturized and conventional instrumentation. Some theoretical aspects are discussed to better explain both the potential and the principal limitations of nano-LC. Recent pharmaceutical and biomedical applications of this separation technique are also presented to indicate the satisfactory performance for complex matrices, especially for proteomic analysis, that is obtained with nano-LC.

  10. Separation of capsaicin from capsaicinoids by macroporous resin adsorption chromatography. (United States)

    Liu, Changxia; Liu, Ruican; Zhang, Peng; Chen, Yiming; Xu, Tao; Wang, Fang; Tan, Tianwei; Liu, Chunqiao


    The aim of present study is to develop an efficient and low-cost method for capsaicin production isolated from capsaicinoids by macroporous resin adsorption chromatography. HZ816 resin has shown the best adsorption and desorption capacities for capsaicin among other resins. To optimize the operating parameters for separation, initial concentration, diameter-to-height ratio, mobile phase ratio, and crystallization method were investigated. When capsaicinoids solution (5 g/L) was loaded onto the column (diameter-to-height ratio = 1:12) with ethanol/1% w/w NaOH (4:6, v/v) as the mobile phase, capsaicin was purified most effectively. By using acid neutralization as the crystallization method, the purity of capsaicin improved from 90.3 to 99.5% with 82.3% yield. In conclusion, this study provides a simple and low-cost method for the industrial-scale production of high-purity capsaicin.

  11. Determination of Stereoisomers of Epalrestat by Liquid Chromatography

    Institute of Scientific and Technical Information of China (English)

    齐美玲; 王鹏; 杨建军


    An isocratic reversed-phase liquid chromatographic (LC) method is described for the determination of epalrestat and its three stereoisomers (degradation impurities) in drug substance. The LC separation system consisted of a Hypersil C18 column (250 mm×4.6 mm, 5 μm) with a mobile phase comprising methanol, acetonitrile and water (volume ratio 60∶1∶50, pH 4.5) delivered at a flow rate of 1.6 mL/min and UV detection at 280 nm. The proposed LC method is simple and selective for the determination of the stereoisomers of epalrestat in the drug substance with a limit of detection and quantification of 3.9 μg/mL and 4.9 μg/mL, respectively. The stereoisomers were identified by liquid chromatography-mass spectrometry (LC-MS).

  12. High performance thin layer chromatography profile of Cassytha filiformis

    Institute of Scientific and Technical Information of China (English)

    Mythili Sathiavelu; Sathiavelu Arunachalam


    Objective: To study the phenols, flavonoids, saponin profile of the medicinal plant Cassytha filiformis (C. filiformis) using high performance thin layer chromatography (HPTLC). Methods:The extracts were tested to determine the presence of various phytochmeicals like alkaloids, phenolic compounds, flavonoids, carbohydrates, glycosides, saponins, terpenoids, tannins, fixed oils, fats and protein and aminoacids (Harborne and Harborne, 1998). HPTLC studies were carried out by Harborne and Wagner et al method. Different compositions of the mobile phase for HPTLC analysis were tested in order to obtain high resolution and reproducible peaks. Results: The results of the preliminary phytochemical studies confirm the presence of phenols, alkaloids, carbohydrates, saponins, flavanoids, terpenoids and tannins in the methanolic extracts of C. filiformis. The methanolic extracts of C. filiformis displayed the presence of 13 types of phenolic substances with 13 different Rf values ranging from 0.01 to 0.96. The results illustrated the presence of 9 different types of flavonoides with 9 different Rf values ranging from 0.01 to 0.97. The results of HPTLC analysis of saponins demonstrated the presence of 11 different types of saponins with 11 different Rf values ranging from 0.04 to 0.92. Conclusions: In the present study we observed the phenols, flavonoids, saponin profile of the medicinal plant C. filiformis using high performance thin layer chromatography (HPTLC). Hence it was concluded that the phenolic compounds present in the methonolic extract could be responsible for antioxidant activities. Plant derived antioxidants, especially phenols and flavonoids, have been described to have various properties like anticancer, antiaging and prevention of cardiovascular diseases. Furthur, separation and characterization of the bioactive compound from the plant is to be evaluated and reported in near future.

  13. Towards Chip Scale Liquid Chromatography and High Throughput Immunosensing

    Energy Technology Data Exchange (ETDEWEB)

    Ni, Jing [Iowa State Univ., Ames, IA (United States)


    This work describes several research projects aimed towards developing new instruments and novel methods for high throughput chemical and biological analysis. Approaches are taken in two directions. The first direction takes advantage of well-established semiconductor fabrication techniques and applies them to miniaturize instruments that are workhorses in analytical laboratories. Specifically, the first part of this work focused on the development of micropumps and microvalves for controlled fluid delivery. The mechanism of these micropumps and microvalves relies on the electrochemically-induced surface tension change at a mercury/electrolyte interface. A miniaturized flow injection analysis device was integrated and flow injection analyses were demonstrated. In the second part of this work, microfluidic chips were also designed, fabricated, and tested. Separations of two fluorescent dyes were demonstrated in microfabricated channels, based on an open-tubular liquid chromatography (OT LC) or an electrochemically-modulated liquid chromatography (EMLC) format. A reduction in instrument size can potentially increase analysis speed, and allow exceedingly small amounts of sample to be analyzed under diverse separation conditions. The second direction explores the surface enhanced Raman spectroscopy (SERS) as a signal transduction method for immunoassay analysis. It takes advantage of the improved detection sensitivity as a result of surface enhancement on colloidal gold, the narrow width of Raman band, and the stability of Raman scattering signals to distinguish several different species simultaneously without exploiting spatially-separated addresses on a biochip. By labeling gold nanoparticles with different Raman reporters in conjunction with different detection antibodies, a simultaneous detection of a dual-analyte immunoassay was demonstrated. Using this scheme for quantitative analysis was also studied and preliminary dose-response curves from an immunoassay of a

  14. A Method for the Quantitation of Trace Levels of Dimethyl Sulfoxide in Urine by High Performance Liquid Chromatography (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY by...for the sample cleanup and concentration, followed by separation by reversed phase high performance liquid chromatography . EXPERIMENTAL Materials...DIMETHYL SULFOXIDE IN URINE BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY 4. AUTHORS (Last name, first name, middle initial. If military, show rank, e.g.

  15. Determination of Dimethyl Sulfoxide (DMSO), Ethanol (ETOH), Formamide (F) and Glycerol/Formal (GF) by High Performance Liquid Chromatography (HPLC) (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY (HPLC...Classification) (U) Determination of Dimethyl Sulfoxide (DMSO), Ethanol, (ETOH), Formamide (F), and Glycerol/ Formal (GF) by High Performance Liquid Chromatography (HPLC...and 5). High performance liquid chromatography (HPLC) was the analytical method of choice for analyzing DMSO, ethanol, formamide and

  16. Determination of 5-Bromo-2’-Deoxyuridine (BrdU) in Well Water by High Performance Liquid Chromatography (HPLC) (United States)


    HIGH PERFORMANCE LIQUID CHROMATOGRAPHY (HPLC...Securrty Classification) Determination of 5-Bromo-2’-Deoxyuridine (BrdU) in Well Water by High Performance Liquid Chromatography (hPLC) 12. PERSONAL...PLOT OF BrdU STABILITY VERSUS TIME ....................... 10 ii DETERMINATION OF 5-BROMO-2’-DEOXY-URIDINE (BrdU) IN WELL WATER BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

  17. Comparison of preparative reversed phase liquid chromatography and countercurrent chromatography for the kilogram scale purification of crude spinetoram insecticide. (United States)

    DeAmicis, Carl; Edwards, Neil A; Giles, Michael B; Harris, Guy H; Hewitson, Peter; Janaway, Lee; Ignatova, Svetlana


    Reversed phase HPLC (RP-HPLC) and high performance countercurrent chromatography (HPCCC) were compared for the pilot scale purification of two semi-synthetic spinosyns, spinetoram-J and spinetoram-L, the major components of the commercial insecticide spinetoram. Two, independently performed, 1 kg, purification campaigns were compared. Each method resulted in the isolation of both components at a purity of >97% and yields for spinetoram-J and spinetoram-L of >93% and ≥ 63% of theoretical, respectively. The HPCCC process produced a 2-fold higher throughput and consumed approximately 70% less solvent than preparative scale RP-HPLC, the volume of product containing fractions from HPCCC amounted to 7% of that produced by HPLC and so required much less post-run processing.

  18. Screening anti-tumor compounds from Ligusticum wallichii using cell membrane chromatography combined with high-performance liquid chromatography and mass spectrometry. (United States)

    Zhang, Tao; Ding, Yuanyuan; An, Hongli; Feng, Liuxin; Wang, Sicen


    Tyrosine 367 Cysteine-fibroblast growth factor receptor 4 cell membrane chromatography combined with high-performance liquid chromatography and mass spectrometry was developed. Tyrosine 367 Cysteine-HEK293 cells were used as cell membrane stationary phase. Specificity and reproducibility of the cell membrane chromatography was evaluated using 1-tert-butyl-3-{2-[4-(diethylamino)butylamino]-6-(3,5-dimethoxyphenyl)pyrido[2,3-d]pyrimidin-7-yl}urea, Nimodipine and dexamethasone acetate. Then, anti-tumor components acting on Tyrosine 367 Cysteine-fibroblast growth factor receptor 4 were screened and identified from extracts of Ligusticum wallichii. Components from the extract were retained on the cell membrane chromatographic column. The retained fraction was directly eluted into high-performance liquid chromatography with mass spectrometry system for separation and identification. Finally, Levistolide A was identified as an active component from Ligusticum wallichii extracts. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide-formazan colorimetric assay revealed that Levistolide A inhibits proliferation of overexpressing the mutated receptor cells with dose-dependent manner. Phosphorylation of fibroblast growth factor receptor 4 was also decrease under Levistolide A treatment. Flex dock simulation verified that Levistolide A could bind with the tyrosine kinase domain of fibroblast growth factor receptor 4. Therefore, Levistolide A screened by the cell membrane chromatography combined with high-performance liquid chromatography and mass spectrometry can arrest cell growth. In conclusion, the two-dimensional high-performance liquid chromatography method can screen and identify potential anti-tumor ingredients which specifically act on the tyrosine kinase domain of the mutated fibroblast growth factor receptor 4. This article is protected by copyright. All rights reserved.

  19. Measurement of choline and choline metabolite concentrations using high-pressure liquid chromatography and gas chromatography-mass spectrometry. (United States)

    Pomfret, E A; daCosta, K A; Schurman, L L; Zeisel, S H


    We have developed a reproducible and sensitive procedure for the isolation and measurement of choline, phosphocholine, glycerophosphocholine, phosphatidylcholine, lysophosphatidylcholine and acetylcholine in a single 100-mg sample of biological tissue. Tissues were spiked with 14C-methyl- and 2H-methyl- or 15N-choline labeled internal standards for each compound. They were extracted with chloroform/methanol/water and the aqueous and organic phases were dried. The organic phase was resuspended in chloroform/methanol (1/1, v/v) and an aliquot was applied to a silica-gel thin-layer chromatography plate. The plate was developed in chloroform/methanol/water (65/30/4, v/v). Segments which cochromatographed with external standards of phosphatidylcholine and lysophosphatidylcholine were stained, scraped, and hydrolyzed in 6 M methanolic-HCl at 80 degrees C for 60 min, liberating free choline. The aqueous phase was resuspended in methanol/water and injected onto a silica HPLC column. Choline and its metabolites were eluted using a binary nonlinear gradient of acetonitrile/ethanol/acetic acid/1 M ammonium acetate/water/0.1 M sodium phosphate (800/68/2/3/127/10, v/v changing to 400/68/44/88/400/10, v/v). Peaks were detected with an on-line radiometric detector, collected, and dried under vacuum. Each choline ester was digested in 6 M HCl at 80 degrees C to form choline. Choline was then converted to the propionyl ester and demethylated with sodium benzenethiolate. This volatile derivative was then isolated using gas chromatography and measured with a mass selective detector. Deuterated internal standards were used to correct for variations in recovery. Choline, glycerophosphocholine, phosphocholine, phosphatidylcholine, lysophosphatidylcholine, and acetylcholine were measured in rat liver, heart, muscle, kidney, plasma, red blood cells, and brain and in human plasma.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Analysis of odour compounds from scented consumer products using gas chromatography-mass spectrometry and gas chromatography-olfactometry. (United States)

    Bartsch, Jennifer; Uhde, Erik; Salthammer, Tunga


    Scented consumer products are being bought in increasing amounts and gaining more popularity. There is, however, relatively little information available about their ingredients, emissions and allergenic potential. Frequently, a mixture of different fragrance substances and not solely an individual substance contributes to the overall desired smell. The aim of this study was to investigate the odorous volatile organic compounds (OVOCs) in consumer products containing fragrances. Over 44 products were selected: various scented candles, printing products with different scent types and other products types particularly meant to be used indoors. Measurements were carried out in a desiccator. Air samples were collected on thermal desorption tubes to determine the released fragrance substances by means of gas chromatography-mass spectrometry (GC-MS). Moreover, gas chromatography-olfactometry (GC-O) was used to obtain sensory data and to ensure no important odorant was overlooked. Using both methods it was possible to distinguish between odour active and inactive compounds and subsequently to identify almost 300 different odorants across all scented products. Besides the advantage of differentiation, as the human nose is a very sensitive detector, GC-O was found to be a useful tool for detecting traces and chosen target compounds. One focus in this study lay on the 26 EU-regulated fragrance allergens to prove their relevance in scented consumer goods. In total, 18 of them were identified, with at least one substance being present in almost every product. Benzyl alcohol, cinnamaldehyde, citronellol, eugenol, linalool and limonene were the prevalently detected allergens. Particularly linalool and limonene were observed in over 50% of the products. In addition, eugenol appeared to be one of the most frequently detected compounds in trace-level concentrations in the candle emissions.

  1. Determination of alkyl amines in atmospheric aerosol particles: a comparison of gas chromatography-mass spectrometry and ion chromatography approaches

    Directory of Open Access Journals (Sweden)

    R.-J. Huang


    Full Text Available In recent years low molecular weight alkyl amines have been recognized to play an important role in particle formation and growth in the lower atmosphere. However, major uncertainties are associated with their atmospheric processes, sources and sinks, mostly due to the lack of ambient measurements and the difficulties in accurate quantification of alkyl amines at trace level. In this study, we present the evaluation and optimization of two analytical approaches, i.e., gas chromatography-mass spectrometry (GC-MS and ion chromatography (IC, for the determination of alkyl amines in aerosol particles. Alkyl amines were converted to carbamates through derivatization with isobutyl chloroformate for GC-MS determination. A set of parameters affecting the analytical performances of the GC-MS approach, including reagent amount, reaction time and pH value, was evaluated and optimized. The accuracy is 84.3–99.1%, and the limits of detection obtained are 1.8–3.9 pg. For the IC approach, a solid phase extraction (SPE column was used to separate alkyl amines from interfering cations before IC analysis. 1–2% (v/v of acetone (or 2–4% (v/v of acetonitrile was added to the eluent to improve the separation of alkyl amines on the IC column. The limits of detection obtained are 2.1–15.9 ng and the accuracy is 55.1–103.4%. The lower accuracy can be attributed to evaporation losses of amines during the sample concentration procedure. Measurements of ambient aerosol particle samples collected in Hong Kong show that the GC-MS approach is superior to the IC approach for the quantification of primary and secondary alkyl amines due to its lower detection limits and higher accuracy.

  2. Basic Principles of Planar Chromatography and Its Potential for Hyphenated Techniques (United States)

    Tuzimski, Tomasz

    Sample preparation, detection, identification, and quantitative determination of biomolecules are presented in this chapter. Advantages of planar chromatography and the basic principles (chambers, sample application, and chromatogram development) are also described. Rapid detection of biomolecules plays a strategical role in their investigation. Hyphenated techniques such as planar chromatography coupled to UV diode array detection and to mass spectrometry provide on-line extensive structural information on the metabolites prior to their isolation. In this chapter, the combination of thin-layer chromatography (TLC) with biomolecules specific detection by diode array scanning (DAD), mass spectrometry (MS), nuclear magnetic resonance (NMR), and Fourier transform infrared spectroscopy (TLC-FTIR) is discussed. In the last part of the chapter, the reader will gain useful information about a recent method of planar chromatography - multidimensional planar chromatography (MD-PC) and information on application of different modes of multidimensional planar chromatography and combination of this technique with diode array detection (MD-PC-DAD) and HPLC-DAD for separation, detection, and qualitative and quantitative determination of biomolecules. Planar chromatography as a pilot technique for HPLC is also described.

  3. Analysis of global components in Ganoderma using liquid chromatography system with multiple columns and detectors. (United States)

    Qian, Zhengming; Zhao, Jing; Li, Deqiang; Hu, Dejun; Li, Shaoping


    In present study, a multiple columns and detectors liquid chromatography system for analysis of global components in traditional Chinese medicines was developed. The liquid chromatography system was consist of three columns, including size exclusion chromatography column, hydrophilic interaction chromatography column, and reversed phase chromatography column, and three detectors, such as diode array detector, evaporative light scattering detector, and mass spectrometry detector, based on column switching technique. The developed multiple columns and detectors liquid chromatography system was successfully applied to the analysis of global components, including macromolecular (polysaccharides), high (nucleosides and sugars)-, and low (triterpenes)-polarity small molecular compounds in Ganoderma, a well-known Chinese medicinal mushroom. As a result, one macromolecular chromatographic peak was found in two Ganoderma species, 19 components were identified in Ganoderma lucidum (two sugars, three nucleosides, and 14 triterpenes), and four components (two sugars and two nucleosides) were identified in Ganoderma sinense. The developed multiple columns and detectors liquid chromatography system was helpful to understand comprehensive chemical characters in TCMs.

  4. Screening antiallergic components from Carthamus tinctorius using rat basophilic leukemia 2H3 cell membrane chromatography combined with high-performance liquid chromatography and tandem mass spectrometry. (United States)

    Han, Shengli; Huang, Jing; Cui, Ronghua; Zhang, Tao


    Carthamus tinctorius, used in traditional Chinese medicine, has many pharmacological effects, such as anticoagulant effects, antioxidant effects, antiaging effects, regulation of gene expression, and antitumor effects. However, there is no report on the antiallergic effects of the components in C. tinctorius. In the present study, we investigated the antiallergic components of C. tinctorius and its mechanism of action. A rat basophilic leukemia 2H3/cell membrane chromatography coupled online with high-performance liquid chromatography and tandem mass spectrometry method was developed to screen antiallergic components from C. tinctorius. The screening results showed that Hydroxysafflor yellow A, from C. tinctorius, was the targeted component that retained on the rat basophilic leukemia 2H3/cell membrane chromatography column. We measured the amount of β-hexosaminidase and histamine released in mast cells and the key markers of degranulation. The release assays showed that Hydroxysafflor yellow A could attenuate the immunoglobulin E induced release of allergic cytokines without affecting cell viability from 1.0 to 50.0 μM. In conclusion, the established rat basophilic leukemia 2H3 cell membrane chromatography coupled with online high-performance liquid chromatography and tandem mass spectrometry method successfully screened and identified Hydroxysafflor yellow A from C. tinctorius as a potential antiallergic component. Pharmacological analysis elucidated that Hydroxysafflor yellow A is an effective natural component for inhibiting immunoglobulin E-antigen-mediated degranulation.

  5. Analysis of lignans in Magnoliae Flos by turbulent flow chromatography with online solid-phase extraction and high-performance liquid chromatography with tandem mass spectrometry. (United States)

    Zhou, Xuan; Chen, Cen; Ye, Xiaolan; Song, Fenyun; Fan, Guorong; Wu, Fuhai


    In this study, a method coupling turbulent flow chromatography with online solid-phase extraction and high-performance liquid chromatography with tandem mass spectrometry was developed for analyzing the lignans in Magnoliae Flos. By the online pretreatment of turbulent flow chromatography solid-phase extraction, the impurities removal and analytes concentration were automatically processed, and the lignans were separated rapidly and well. Seven lignans of Magnoliae Flos including epieudesmin, magnolin, 1-irioresinol-B-dimethyl ether, epi-magnolin, fargesin aschantin, and demethoxyaschantin were identified by comparing their retention behavior, UV spectra, and mass spectra with those of reference substances or literature data. The developed method was validated, and the good results showed that the method was not only automatic and rapid, but also accurate and reliable. The turbulent flow chromatography with online solid-phase extraction and high-performance liquid chromatography with tandem mass spectrometry method holds a high potential to become an effective method for the quality control of lignans in Magnoliae Flos and a useful tool for the analysis of other complex mixtures.

  6. Ultrafiltration liquid chromatography combined with high-speed countercurrent chromatography for screening and isolating potential α-glucosidase and xanthine oxidase inhibitors from Cortex Phellodendri. (United States)

    Li, Sainan; Liu, Chunming; Guo, Liping; Zhang, Yuchi; Wang, Jing; Ma, Bing; Wang, Yueqi; Wang, Yumeng; Ren, Junqi; Yang, Xiaojing; Qin, Yao; Tang, Ying


    Cortex Phellodendri is a typical Chinese herb with a large number of alkaloids existing in all parts of it. The most common methods for screening and isolating alkaloids are mostly labor intensive and time consuming. In this study, a new assay based upon ultrafiltration liquid chromatography was developed for the rapid screening of ligands for α-glucosidase and xanthine oxidase. The C. Phellodendri extract was found to contain two alkaloids with both α-glucosidase- and xanthine oxidase binding activities and one lactone with α-glucosidase-binding activity. Subsequently, with the help of high-speed countercurrent chromatography, the specific binding ligands including palmatine, berberine, and obaculactone with purities of 97.38, 96.12, and 96.08%, respectively, were successfully separated. An optimized low-toxicity two-phase solvent system composed of ethyl acetate/n-butanol/ethanol/water (3.5:1.7:0.5:5, v/v/v/v) was used to isolate the three compounds mentioned above from C. Phellodendri. The targeted compounds were identified by liquid chromatography coupled with mass spectrometry and NMR spectroscopy. Therefore, ultrafiltration liquid chromatography combined with high-speed countercurrent chromatography is not only a powerful tool for screening and isolating α-glucosidase and xanthine oxidase inhibitors in complex samples but is also a useful platform for discovering bioactive compounds for the prevention and treatment of diabetes mellitus and gout.

  7. Adsorption and displacement effects in the gas-chromatography of metal beta-diketonates. (United States)

    Uden, P C; Jenkins, C R


    A detailed study of the gas chromatography of the aluminium(III), chromium(III) and iron(III) beta-diketonates has shown that their elution and separation characteristics arise from a number of adsorptive effects. Conditions of optimal peak shape for individual compounds are established by varying Chromatographie parameters, and the adsorption of one metal chelate on a column and its subsequent displacement by another chelate are investigated. The chromatography of iron(III) beta-diketonates is seen to be further complicated by gradual elution of a portion of the chelate before the rest of the sample.

  8. Paper SERS chromatography for detection of trace analytes in complex samples (United States)

    Yu, Wei W.; White, Ian M.


    We report the application of paper SERS substrates for the detection of trace quantities of multiple analytes in a complex sample in the form of paper chromatography. Paper chromatography facilitates the separation of different analytes from a complex sample into distinct sections in the chromatogram, which can then be uniquely identified using SERS. As an example, the separation and quantitative detection of heroin in a highly fluorescent mixture is demonstrated. Paper SERS chromatography has obvious applications, including law enforcement, food safety, and border protection, and facilitates the rapid detection of chemical and biological threats at the point of sample.

  9. Application of coupled affinity-sizing chromatography for the detection of proteolyzed HSA-tagged proteins. (United States)

    London, Anne Serdakowski; Patel, Kunal; Quinn, Lisa; Lemmerer, Martin


    Coupled affinity liquid chromatography and size exclusion chromatography (ALC-SEC) is a technique that has been shown to successfully report product quality of proteins during cell expression and prior to the commencement of downstream processing chromatography steps. This method was applied to monitoring the degradation and subsequent partial remediation of a HSA-tagged protein which showed proteolysis, allowing for rapid cell line development to address this product quality dilemma. This paper outlines the novel application of this method for measuring and addressing protease-induced proteolysis.

  10. Fluorimetric assay for ornithine decarboxylase by high-performance liquid chromatography. (United States)

    Haraguchi, K; Kai, M; Kohashi, K; Ohkura, Y


    A highly sensitive method for the assay of ornithine decarboxylase in sample solutions prepared from rat tissue homogenate is described which employs high-performance liquid chromatography with fluorescence detection. Putrescine formed from ornithine under the optimal conditions for the enzyme reaction is treated by Cellex P column chromatography for clean-up and converted into the fluorescamine derivative in the presence of cupric ion which inhibits the reaction of interfering amines with fluorescamine. The derivative is separated by reversed-phase chromatography on LiChrosorb RP-18 with linear gradient elution. The lower limit of detection for putrescine formed enzymatically is 5 pmol.

  11. Cellufine sulfate column chromatography as a simple, rapid, and effective method to purify dengue virus. (United States)

    Kanlaya, Rattiyaporn; Thongboonkerd, Visith


    Conventional method to purify/concentrate dengue virus (DENV) is time-consuming with low virus recovery yield. Herein, we applied cellufine sulfate column chromatography to purify/concentrate DENV based on the mimicry between heparan sulfate and DENV envelope protein. Comparative analysis demonstrated that this new method offered higher purity (as determined by less contamination of bovine serum albumin) and recovery yield (as determined by greater infectivity). Moreover, overall duration used for cellufine sulfate column chromatography to purify/concentrate DENV was approximately 1/20 of that of conventional method. Therefore, cellufine sulfate column chromatography serves as a simple, rapid, and effective alternative method for DENV purification/concentration.

  12. Strong cation exchange chromatography in analysis of posttranslational modifications: innovations and perspectives. (United States)

    Edelmann, Mariola J


    Strong cation exchange (SCX) chromatography has been utilized as an excellent separation technique that can be combined with reversed-phase (RP) chromatography, which is frequently used in peptide mass spectrometry. Although SCX is valuable as the second component of such two-dimensional separation methods, its application goes far beyond efficient fractionation of complex peptide mixtures. Here I describe how SCX facilitates mapping of the protein posttranslational modifications (PTMs), specifically phosphorylation and N-terminal acetylation. The SCX chromatography has been mainly used for enrichment of these two PTMs, but it might also be beneficial for high-throughput analysis of other modifications that alter the net charge of a peptide.

  13. Occurrence of turbulent flow conditions in supercritical fluid chromatography. (United States)

    De Pauw, Ruben; Choikhet, Konstantin; Desmet, Gert; Broeckhoven, Ken


    Having similar densities as liquids but with viscosities up to 20 times lower (higher diffusion coefficients), supercritical CO2 is the ideal (co-)solvent for fast and/or highly efficient separations without mass-transfer limitations or excessive column pressure drops. Whereas in liquid chromatography the flow remains laminar in both the packed bed and tubing, except in extreme cases (e.g. in a 75 μm tubing, pure acetonitrile at 5 ml/min), a supercritical fluid can experience a transition from laminar to turbulent flow in more typical operation modes. Due to the significant lower viscosity, this transition for example already occurs at 1.3 ml/min for neat CO2 when using connection tubing with an ID of 127 μm. By calculating the Darcy friction factor, which can be plotted versus the Reynolds number in a so-called Moody chart, typically used in fluid dynamics, higher values are found for stainless steel than PEEK tubing, in agreement with their expected higher surface roughness. As a result turbulent effects are more pronounced when using stainless steel tubing. The higher than expected extra-column pressure drop limits the kinetic performance of supercritical fluid chromatography and complicates the optimization of tubing ID, which is based on a trade-off between extra-column band broadening and pressure drop. One of the most important practical consequences is the non-linear increase in extra-column pressure drop over the tubing downstream of the column which leads to an unexpected increase in average column pressure and mobile phase density, and thus decrease in retention. For close eluting components with a significantly different dependence of retention on density, the selectivity can significantly be affected by this increase in average pressure. In addition, the occurrence of turbulent flow is also observed in the detector cell and connection tubing. This results in a noise-increase by a factor of four when going from laminar to turbulent flow (e.g. going

  14. Ion-Exclusion Chromatography for Analyzing Organics in Water (United States)

    Sauer, Richard; Rutz, Jeffrey A.; Schultz, John R.


    A liquid-chromatography technique has been developed for use in the quantitative analysis of urea (and of other nonvolatile organic compounds typically found with urea) dissolved in water. The technique involves the use of a column that contains an ion-exclusion resin; heretofore, this column has been sold for use in analyzing monosaccharides and food softeners, but not for analyzing water supplies. The prior technique commonly used to analyze water for urea content has been one of high-performance liquid chromatography (HPLC), with reliance on hydrophobic interactions between analytes in a water sample and long-chain alkyl groups bonded to an HPLC column. The prior technique has proven inadequate because of a strong tendency toward co-elution of urea with other compounds. Co-elution often causes the urea and other compounds to be crowded into a narrow region of the chromatogram (see left part of figure), thereby giving rise to low chromatographic resolution and misidentification of compounds. It is possible to quantitate urea or another analyte via ultraviolet- and visible-light absorbance measurements, but in order to perform such measurements, it is necessary to dilute the sample, causing a significant loss of sensitivity. The ion-exclusion resin used in the improved technique is sulfonated polystyrene in the calcium form. Whereas the alkyl-chain column used in the prior technique separates compounds on the basis of polarity only, the ion-exclusion-resin column used in the improved technique separates compounds on the basis of both molecular size and electric charge. As a result, the degree of separation is increased: instead of being crowded together into a single chromatographic peak only about 1 to 2 minutes wide as in the prior technique, the chromatographic peaks of different compounds are now separated from each other and spread out over a range about 33 minutes wide (see right part of figure), and the urea peak can readily be distinguished from the other

  15. Different Ways to On-Line Hyphenate Centrifugal Partition Chromatography and Mass Spectrometry: Application to Prenylated Xanthones from Garcinia mangostana. (United States)

    Destandau, Emilie; Michel, Thomas; Toribio, Alix; Elfakir, Claire


    Centrifugal partition chromatography is a liquid-liquid separation method well adapted for the fractionation or purification of natural compounds from plant extracts. However, following the preparative isolation, the fractions collected must be analysed by high-performance thin-layer chromatography or high-performance liquid chromatography to evaluate their composition and/or their purity. These additional steps are time-consuming and increase the risk of compound degradation. In order to get an instantaneous analysis of fraction content with structural information on the phytochemicals eluted, it is possible to hyphenate on-line centrifugal partition chromatography with mass spectrometry. Depending on the complexity of the extract, two different kinds of centrifugal partition chromatography-mass spectrometry coupling can be performed: centrifugal partition chromatography-mass spectrometry or centrifugal partition chromatography-high-performance liquid chromatography-mass spectrometry coupling. In the first case, one part of the centrifugal partition chromatography effluent is directly introduced in the mass spectrometry ionisation source to identify the eluted compounds, while in the second case, it is directed to a high-performance liquid chromatography-mass spectrometry system where compounds are first separated thanks to high-performance liquid chromatography and then identified using mass spectrometry.

  16. Mixed-mode chromatography integrated with high-performance liquid chromatography for protein analysis and separation: Using bovine serum albumin and lysozyme as the model target. (United States)

    Xia, Hai-Feng; Don, Bin-Bin; Zheng, Meng-Jie


    A type of mixed-mode chromatography was integrated with high-performance liquid chromatography for protein analysis and separation. The chromatographic behavior was tested using bovine serum albumin and lysozyme as model proteins. For the mixed-mode column, the silica beads were activated with γ-(2,3-epoxypropoxy)-propytrimethoxysilane and coupled with 4-mercaptopyridine as the functional ligand. The effects of pH, salt, and the organic solvent conditions of the mobile phase on the retention behavior were studied, which provided valuable clues for separation strategy. When eluted with a suitable pH gradient, salt concentration gradient, and acetonitrile content gradient, the separation behavior of bovine serum albumin and lysozyme could be controlled by altering the conditions of the mobile phase. The results indicated this type of chromatography might be a useful method for protein analysis and separation.

  17. Separation and purification of three stilbenes from the radix of Polygonum cillinerve (Nakai Ohwl by macroporous resin column chromatography combined with high-speed counter-current chromatography

    Directory of Open Access Journals (Sweden)

    Xiaofeng Chi


    Full Text Available An effective method for the rapid separation and purification of three stilbenes from the radix of Polygonum cillinerve (Nakai Ohwl by macroporous resin column chromatography combined with high-speed counter-current chromatography (HSCCC was successfully established. In the present study, a two-phase solvent system composed of chloroform-n-butanol-methanol-water (4:1:4:2, v/v/v/v was used for HSCCC separation. A one-step separation in 4 h from 150 mg of crude extract produced 26.3 mg of trans-resveratrol-3-O-glucoside, 42.0 mg of pieceid-2"-O-gallate, and 17.9 mg of trans-resveratrol with purities of 99.1%, 97.8%, and 99.4%, respectively, as determined by high-performance liquid chromatography (HPLC. The chemical structures of these compounds were identified by nuclear magnetic resonance (NMR spectroscopy.

  18. Solanidine isolation from Solanum tuberosum by centrifugal partition chromatography. (United States)

    Attoumbré, Jacques; Giordanengo, Philippe; Baltora-Rosset, Sylvie


    The aim of this investigation was the preparative isolation of solanidine (aglycone of the two main potato glycoalkaloids: α-chaconine and α-solanine) from fresh Solanum tuberosum (cv. Pompadour) material by implementing a new preparation scheme using centrifugal partition chromatography (CPC). A setup for obtaining solanidine by hydrolysis of the glycoalkaloids found in the skin and sprouts of S. tuberosum was first developed. Then its isolation was carried out by the development of CPC conditions: the solvent system used for separation was ethyl acetate/butanol/water in the ratio 42.5:7.5:50 v/v/v, 0.6 g of crude extract were separated with a 8 mL/min flow rate of mobile phase while rotating at 2500 rpm. A run yielded 98 mg of solanidine (86.7% recovery from the crude extract) in a one-step separation. The purity of the isolated solanidine was over 98%. Thus, CPC has proven to be the method of choice to get solanidine of very high purity from S. tuberosum biomass in large quantities.

  19. Establishment and application of milk fingerprint by gel filtration chromatography. (United States)

    Gao, P; Li, J; Li, Z; Hao, J; Zan, L


    Raw milk adulteration frequently occurs in undeveloped countries. It not only reduces the nutritional value of milk, but it is also harmful to consumers. In this paper, we focused on investigating an efficient method for the quality control of raw milk protein. A gel filtration chromatography (GFC) fingerprint method combined with chemometrics was developed for fingerprint analysis of raw milk. To optimize the GFC conditions, milk fat was removed by centrifugation, and GFC analysis was performed on a Superdex 75 10/300GL column (Just Scientific, Shanghai, China) with 0.2 M NaH2PO4-Na2HPO4 buffer (pH 7.0) as the mobile phase. The flow rate was 0.5mL/min, and the detection wavelength was set at 280 nm. Ten batches of 120 raw milk samples were analyzed to establish the GFC fingerprint under optimal conditions. Six major peaks common to the chromatogram of each raw milk sample were selected for fingerprint analysis, and the characteristic peaks were used to establish a standard chromatographic fingerprint. Principal component analysis was then applied to classify GFC information of adulterated milk and raw milk, allowing adulterated samples to be effectively screened out from the raw milk in principal component analysis scores plot. The fingerprint method demonstrates promising features in detecting milk protein adulteration.

  20. Electrochemically modulated chromatography-separation of catechol-containing compounds

    Energy Technology Data Exchange (ETDEWEB)

    Deinhammer, R.S.; Porter, M.D.; Ting, E.Y.; Ho, M.K. [Iowa State Univ., Ames, IA (United States)


    A novel liquid chromatographic system that incorporates electrochemical control of stationary phase composition as an additional variable for optimizing and fine tuning separations has been developed. The electrochemically-modulated liquid chromatographic (EMLC) system consists of 4 gm spherical glassy carbon (GC) particles connected as the working electrode in a three-electrode cell arrangement. Electrochemically-induced changes in the excess charge at the GC surface prior to elution are used to alter the capacity factors of analytes over a wide range. This ability also provides a variable pathway for performing a gradient type elution through modification of the excess charge at the GC surface during analyte elution. This has led to the development of novel separation approaches that are analogous to gradient elution liquid chromatography, but are accomplished without changes in mobile phase composition. Results using this system for the manipulation of the elution of a variety of aromatic analytes, pharmaceutical compounds, and amino acids will be presented. In addition, studies which examined the coating of the GC spheres with electroactive species to enhance the control over stationary phase composition will be discussed.

  1. Thin layer chromatography-ion mobility spectrometry (TLC-IMS). (United States)

    Ilbeigi, Vahideh; Tabrizchi, Mahmoud


    Ion mobility spectrometry (IMS) is a fast and sensitive analytical method which operates at the atmospheric pressure. To enhance the capability of IMS for the analysis of mixtures, it is often used with preseparation techniques, such as GC or HPLC. Here, we report for the first time the coupling of the thin-layer chromatography and IMS. A variety of coupling schemes were tried that included direct electrospray from the TLC strip tip, indirect electrospray from a needle connected to the TLC strip, introducing the moving solvent into the injection port, and, the simplest way, offline introduction of scratched or cut pieces of strips into the IMS injection port. In this study a special solvent tank was designed and the TLC strip was mounted horizontally where the solvent would flow down. A very small funnel right below the TLC tip collected the solvent and transferred it to a needle via a capillary tubing. Using the TLC-ESI-IMS technique, acceptable separations were achieved for two component mixtures of morphine-papaverine and acridine-papaverine. A special injection port was designed to host the pieces cut off the TLC. The method was successfully used to identify each spot on the TLC by IMS in a few seconds.

  2. Determination of Glimepiride in Human Plasma by Liquid Chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Ho Hyun; Lee, Hee Joo [Seoul Clinical Laboratories, Seoul (Korea, Republic of); Chang, Kyu Young [Korean Biochip Society, Seoul (Korea, Republic of); Han, Sang Beom [ChungAng University, Seoul (Korea, Republic of)


    A sensitive method for quantitation of glimepiride in human plasma has been established using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS). Glipizide was used as an internal standard. Glimepiride and internal standard in plasma sample was extracted using diethyl etherethyl acetate (1 : 1). A centrifuged upper layer was then evaporated and reconstituted with the mobile phase of acetonitrile-5 mM ammonium acetate (60:40, pH 3.0). The reconstituted samples were injected into a C{sub 18} reversed-phase column. Using MS/MS in the multiple reaction monitoring (MRM) mode, glimepiride and glipizide were detected without severe interference from human plasma matrix. Glimepiride produced a protonated precursor ion ([M+H]{sup +}) at m/z 491 and a corresponding product ion at m/z 352. And the internal standard produced a protonated precursor ion ([M+H]{sup +}) at m/z 446 and a corresponding product ion at m/z 321. Detection of glimepiride in human plasma by the LC-ESI/MS/MS method was accurate and precise with a quantitation limit of 0.1 ng/mL. The validation, reproducibility, stability, and recovery of the method were evaluated. The method has been successfully applied to pharmacokinetic studies of glimepiride in human plasma

  3. The Use of Gas Chromatography for Biogas Analysis (United States)

    Andersen, Amanda; Seeley, John; Aurandt, Jennifer


    Energy from natural gas accounts for 24 percent of energy consumed in the US. Natural gas is a robust form of energy which is rich in methane content and is low in impurities. This quality suggests that it is a very clean and safe gas; it can be used in providing heat, a source for cooking, and in powering vehicles. The downside is that it is a non-renewable resource. On the contrary, methane rich gas that is produced by the breakdown of organic material in an anaerobic environment, called biogas, is a renewable energy source. This research focuses on the gas analysis portion of the creation of the anaerobic digestion and verification laboratory where content and forensic analysis of biogas is performed. Gas Chromatography is implemented as the optimal analytical tool for quantifying the components of the biogas including methane, carbon dioxide, hydrogen sulfide and siloxanes. In addition, the problems associated with the undesirable components are discussed. Anaerobic digestion of primary sludge has consistently produced about 55 percent methane; future goals of this research include studying different substrates to increase the methane yield and decrease levels of impurities in the gas.

  4. Supercritical fluid chromatography for the separation of isoflavones. (United States)

    Ganzera, Markus


    The first protocol for the analysis of isoflavones by supercritical fluid chromatography is reported. Optimum results were obtained on an Acquity UPC(2) BEH 1.7 μm column, using a solvent gradient of supercritical carbon dioxide and methanol (with phosphoric acid as additive) for elution. The method enables the baseline separation of nine isoflavones (aglyca and glycosides) in 8 min, and is suitable for their quantitative determination in dietary supplements containing soy (Glycine max), red glover (Trifolium pratense) and kudzu (Pueraria lobata). Method validation confirmed that the assay is selective, linear (R(2)≥0.9994), accurate (recovery rates from 97.6 to 102.4%), as well as precise on the short- and long-term level (intra-day precision ≤2.1%), and shows an on-column detection limit of 0.2 ng and below. This, together with an excellent performance shown in the analysis of real samples, indicates that SFC is well suited for the fast and accurate determination of isoflavones in complex matrices. Disadvantages compared to the established approaches were not observed, so that SFC has to be considered in this case as an (at least) equivalent analytical alternative.

  5. Analysis of Cordyceps by multi-column liquid chromatography. (United States)

    Qian, Zhengming; Li, Shaoping


    Cordyceps is a famous traditional Chinese medicine (TCM) that has been used in China for hundreds of years. In the present study a multi-column liquid chromatography (MC-LC) system was developed for the qualitative analysis of macromolecules and micromolecules in Cordyceps. The MC-LC system includes a size exclusion pre-column, a size exclusion column (SEC) and a reversed phase column (RP) which were controlled by column-switching valves. The sample was separated by the size exclusion pre-column into two fractions (macromolecules and micromolecules). These fractions were further separated on SEC and RP columns, respectively. A diode array detector (DAD) and a mass spectrometer (MS) were used to detect the components. This MC-LC method was utilized for analysis of Cordyceps samples. Two macromolecular peaks and 15 micromolecular peaks were found in Cordyceps, and 11 of the micromolecular peaks were identified as adenosine-5'-monophosphate (AMP), phenylalanine, uridine, hypoxanthine, inosine, guanine, guanosine, deoxyadenosine-5'-monophosphate (dAMP), adenosine, adenine and cordycepin (or its isomer). This method is useful for quality control of Cordyceps.

  6. Analysis of Cordyceps by multi-column liquid chromatography

    Directory of Open Access Journals (Sweden)

    Zhengming Qian


    Full Text Available Cordyceps is a famous traditional Chinese medicine (TCM that has been used in China for hundreds of years. In the present study a multi-column liquid chromatography (MC-LC system was developed for the qualitative analysis of macromolecules and micromolecules in Cordyceps. The MC-LC system includes a size exclusion pre-column, a size exclusion column (SEC and a reversed phase column (RP which were controlled by column-switching valves. The sample was separated by the size exclusion pre-column into two fractions (macromolecules and micromolecules. These fractions were further separated on SEC and RP columns, respectively. A diode array detector (DAD and a mass spectrometer (MS were used to detect the components. This MC-LC method was utilized for analysis of Cordyceps samples. Two macromolecular peaks and 15 micromolecular peaks were found in Cordyceps, and 11 of the micromolecular peaks were identified as adenosine-5′-monophosphate (AMP, phenylalanine, uridine, hypoxanthine, inosine, guanine, guanosine, deoxyadenosine-5′-monophosphate (dAMP, adenosine, adenine and cordycepin (or its isomer. This method is useful for quality control of Cordyceps.

  7. Methodology for optimally sized centrifugal partition chromatography columns. (United States)

    Chollet, Sébastien; Marchal, Luc; Jérémy Meucci; Renault, Jean-Hugues; Legrand, Jack; Foucault, Alain


    Centrifugal Partition Chromatography (CPC) is a separation process based on the partitioning of solutes between two partially miscible liquid phases. There is no solid support for the stationary phase. The centrifugal acceleration is responsible for both stationary phase retention and mobile phase dispersion. CPC is thus a process based on liquid-liquid mass transfer. The separation efficiency is mainly influenced by the hydrodynamics of the phases in each cell of the column. Thanks to a visualization system, called "Visual CPC", it was observed that the mobile phase can flow through the stationary phase as a sheet, or a spray. Hydrodynamics, which directly governs the instrument efficiency, is directly affected during scale changes, and non-linear phenomena prevent the successful achievement of mastered geometrical scale changes. In this work, a methodology for CPC column sizing is proposed, based on the characterization of the efficiency of advanced cell shapes, taking into account the hydrodynamics. Knowledge about relationship between stationary phase volume, cell efficiency and separation resolution in CPC allowed calculating the optimum cell number for laboratory and industrial scale CPC application. The methodology is highlighted with results on five different geometries from 25 to 5000 mL, for two applications: the separation of alkylbenzene by partitioning with heptane/methanol/water biphasic system; and the separation of peptides by partitioning with n-butanol/acetic acid/water (4/1/5) biphasic system. With this approach, it is possible to predict the optimal CPC column length leading to highest productivity.

  8. Chromatographic analysis of olopatadine in hydrophilic interaction liquid chromatography. (United States)

    Maksić, Jelena; Jovanović, Marko; Rakić, Tijana; Popović, Igor; Ivanović, Darko; Jančić-Stojanović, Biljana


    In this paper, chromatographic analysis of active substance olopatadine hydrochloride, which is used in eye drops as antihistaminic agent, and its impurity E isomer by hydrophilic interaction liquid chromatography (HILIC) and application of design of experiments (DoE) methodology are presented. In addition, benzalkonium chloride is very often used as a preservative in eye drops. Therefore, the evaluation of its chromatographic behavior in HILIC was carried out as well. In order to estimate chromatographic behavior and set optimal chromatographic conditions, DoE methodology was applied. After the selection of important chromatographic factors, Box-Behnken design was utilized, and on the basis of the obtained models factor effects were examined. Then, multi-objective robust optimization is performed aiming to obtain chromatographic conditions that comply with several quality criteria simultaneously: adequate and robust separation of critical peak pair and maximum retention of the first eluting peak. The optimal conditions are identified by using grid point search methodology. The experimental verification confirmed the adequacy of the defined optimal conditions. Finally, under optimal chromatographic conditions, the method was validated and applicability of the proposed method was confirmed.

  9. Instrument modifications that produced reduced plate heights supercritical fluid chromatography. (United States)

    Berger, Terry A


    The concept of peak fidelity was shown to be helpful in modeling tubing and detector cell dimensions. Connection tubing and flow cell variances were modeled to determine appropriate internal ID's, lengths, and volumes. A low dispersion plumbing configuration, based on these calculations, was assembled to replace the standard plumbing and produced the reported results. The modifications made were straightforward using commercially available parts. The full theoretical efficiency of a 3×100 mm column packed with 1.8 μm totally porous particles was achieved for the first time in supercritical fluid chromatography (SFC). Peak fidelity of >0.95 was maintained to below k=2. A reduced plate height as low as 1.87 was measured. Thus, true "ultra high performance" SFC was achieved, with the results a major improvement from all previous SFC reports. Since there were no efficiency losses, none could be attributed to thermal gradients caused by the expansion of the fluid over large pressure drops, under the conditions used. Similarly, changes in diffusion coefficients caused by significant decreases in density during expansion are apparently balanced by the increase in linear velocity, keeping the ratio between the diffusion coefficient and the linear velocity a constant. Changing modifier concentration to change retention was shown to not be a significant problem. All these issues have been a concern in the past. Diffusion coefficients, and viscosity data needs to be collected at high pressures before the actual limits of SFC can be discovered.

  10. Online comprehensive two-dimensional ion chromatography × capillary electrophoresis. (United States)

    Ranjbar, Leila; Gaudry, Adam J; Breadmore, Michael C; Shellie, Robert A


    A comprehensively coupled online two-dimensional ion chromatography-capillary electrophoresis (IC × CE) system for quantitative analysis of inorganic anions and organic acids in water is introduced. The system employs an in-house built sequential injection-capillary electrophoresis instrument and a nonfocusing modulation interface comprising a tee-piece and a six-port two-position injection valve that allows comprehensive sampling of the IC effluent. High field strength (+2 kV/cm) enables rapid second-dimension separations in which each peak eluted from the first-dimension separation column is analyzed at least three times in the second dimension. The IC × CE approach has been successfully used to resolve a suite of haloacetic acids, dalapon, and common inorganic anions. Two-dimensional peak capacity for IC × CE was 498 with a peak production rate of 9 peaks/min. Linear calibration curves were obtained for all analytes from 5 to 225 ng/mL (except dibromoacetic acid (10-225 ng/mL) and tribromoacetic acid (25-225 ng/mL)). The developed approach was used to analyze a spiked tap water sample, with good measured recoveries (69-119%).

  11. Determination of cyanogenic compounds in edible plants by ion chromatography. (United States)

    Cho, Hye-Jeon; Do, Byung-Kyung; Shim, Soon-Mi; Kwon, Hoonjeong; Lee, Dong-Ha; Nah, Ahn-Hee; Choi, Youn-Ju; Lee, Sook-Yeon


    Cyanogenic glycosides are HCN-producing phytotoxins; HCN is a powerful and a rapidly acting poison. It is not difficult to find plants containing these compounds in the food supply and/or in medicinal herb collections. The objective of this study was to investigate the distribution of total cyanide in nine genera (Dolichos, Ginkgo, Hordeum, Linum, Phaseolus, Prunus, Phyllostachys, Phytolacca, and Portulaca) of edible plants and the effect of the processing on cyanide concentration. Total cyanide content was measured by ion chromatography following acid hydrolysis and distillation. Kernels of Prunus genus are used medicinally, but they possess the highest level of total cyanide of up to 2259.81 CN(-)/g dry weight. Trace amounts of cyanogenic compounds were detected in foodstuffs such as mungbeans and bamboo shoots. Currently, except for the WHO guideline for cassava, there is no global standard for the allowed amount of cyanogenic compounds in foodstuffs. However, our data emphasize the need for the guidelines if plants containing cyanogenic glycosidesare to be developed as dietary supplements.

  12. Separation of tocopherols by nano-liquid chromatography. (United States)

    Fanali, Salvatore; Camera, Emanuela; Chankvetadze, Bezhan; D'Orazio, Giovanni; Quaglia, Maria Giovanna


    Nanoliquid chromatography (nano-LC) was used for the separation of tocopherols (delta-, gamma-, alpha-TOH), alpha-tocopherol acetate (alpha-TOH-Ac) and an antioxidant compound, namely butylated hydroxytoluene (BHT) used to prevent TOHs autoxidation. The separation was carried out in a fused silica capillary of 100 microm I.D. and 375 microm O.D. packed in our laboratory with RP18 silica stationary phase of either 5- or 3-microm diameter (23-cm long). The mobile phase was composed by mixtures of methanol (MeOH), acetonitrile (MeCN) and water. Typical analyses time for the separation of all the five components of the mixture were 6-9 min depending on the composition of the mobile phase. Efficiency and resolution were strongly influenced by the particle diameter and the highest Rs and N/m values were observed using 3-microm RP18 particles. Experiments performed with capillaries packed with 3-microm RP18 particles provided good limit of detection (LOD) and limit of quantification (LOQ) (for delta-, gamma-TOH, alpha-TOH-Ac were 4 and 8 microg/ml, while for alpha-TOH were 6 and 10 microg/ml, respectively). The optimized method was applied to extracts of serum and pharmaceutical preparation containing alpha-TOH and alpha-TOH-Ac.

  13. Separation of microcystins and nodularins by ultra performance liquid chromatography. (United States)

    Spoof, Lisa; Neffling, Milla-Riina; Meriluoto, Jussi


    Four ultra performance liquid chromatography (UPLC) columns with different reversed-phase characteristics were tested in the chromatographic separation of 10 microcystins and three nodularins, cyanobacterial peptide toxins. The columns had been designed by the manufacturer to withstand the ultra-high pressure generated by sub-2microm stationary phase particles and the Waters ACQUITY UPLC system in ultra-fast separations. The gradient mobile phase consisted of water and acetonitrile, both acidified with trifluoroacetic acid, with three gradient rise times: 1, 1.5 and 2min. The UV detection of the toxins was performed by a photodiode array detector. The chromatographic performance was evaluated both visually and by calculating chromatographic parameters such as capacity factor, resolution, peak width at half height, selectivity and peak asymmetry. The best chromatographic performance as judged by visual inspection was given by the ACQUITY BEH Shield RP18 and ACQUITY BEH Phenyl columns. The BEH Shield RP18 column showed excellent selectivity and resolution of chosen peak pairs considered as critical. A further advantage of the UPLC system was the high sample throughput with a total analysis time of 3.12min (injection-to-injection) equalling to 461 separations per 24h.

  14. A comprehensive study to protein retention in hydrophobic interaction chromatography. (United States)

    Baca, Martyna; De Vos, Jelle; Bruylants, Gilles; Bartik, Kristin; Liu, Xiaodong; Cook, Ken; Eeltink, Sebastiaan


    The effect of different kosmotropic/chaotropic salt systems on retention characteristics of intact proteins has been examined in hydrophobic interaction chromatography (HIC). The performance was assessed using different column chemistries, i.e., polyalkylamide, alkylamine incorporating hydrophobic moieties, and a butyl chemistry. Selectivity in HIC is mainly governed by the salt concentration and by the molal surface tension increment of the salt. Typically, a linear relationship between the natural logarithm of the retention factor and the salt concentration is obtained. Using a 250mm long column packed with 5μm polyalkylamide functionalized silica particles and applying a 30min linear salt gradient, a peak capacity of 78 was achieved, allowing the baseline separation of seven intact proteins. The hydrophobicity index appeared to be a good indicator to predict the elution order of intact proteins in HIC mode. Furthermore, the effect of adding additives in the mobile phase, such as calcium chloride (stabilizing the 3D conformation of α-lactalbumin) and isopropanol, on retention properties has been assessed. Results indicate that HIC retention is also governed by conformational in the proteins which affect the number of accessible hydrophobic moieties.

  15. Purification of coagulation factor VIII by immobilized metal affinity chromatography. (United States)

    Rodrigues, Estela S; Verinaud, Claudia I; Oliveira, Douglas S; Raw, Isaías; Lopes, Alexandre P Y; Martins, Elizabeth A L; Cheng, Elisabeth


    Factor VIII (FVIII) is a glycoprotein that plays an essential role in blood coagulation cascade. Purification of plasma-derived coagulation FVIII by direct application of plasma to a chromatographic column is a method of choice. Anion exchange column is a very powerful method because FVIII is strongly adsorbed, resulting in good activity recovery and high purification factor. However, vitamin-K-dependent coagulation factors coelute with FVIII. In the present study, we report the separation of vitamin-K-dependent coagulation proteins from FVIII using immobilized metal affinity chromatography (IMAC) with Cu(2+) as the metal ligand. Plasma was directly loaded to a Q Sepharose Big Beads column, and FVIII was recovered with 65% activity and a purification factor of approximately 50 times. Then, the Q Sepharose eluate was applied to the IMAC-Cu(2+) column, and FVIII was eluted with 200 mM imidazole, with up to 85% recovery of activity. The mass recovery in this fraction was less than 10% of the applied mass of protein. Vitamin-K-dependent proteins elute with imidazole concentrations of lower than 60 mM. Because of the difference in affinity, FVIII could be completely separated from the vitamin-K-dependent proteins in the IMAC column.

  16. Fully Automated Portable Comprehensive 2-Dimensional Gas Chromatography Device. (United States)

    Lee, Jiwon; Zhou, Menglian; Zhu, Hongbo; Nidetz, Robert; Kurabayashi, Katsuo; Fan, Xudong


    We developed a fully automated portable 2-dimensional (2-D) gas chromatography (GC x GC) device, which had a dimension of 60 cm × 50 cm × 10 cm and weight less than 5 kg. The device incorporated a micropreconcentrator/injector, commercial columns, micro-Deans switches, microthermal injectors, microphotoionization detectors, data acquisition cards, and power supplies, as well as computer control and user interface. It employed multiple channels (4 channels) in the second dimension ((2)D) to increase the (2)D separation time (up to 32 s) and hence (2)D peak capacity. In addition, a nondestructive flow-through vapor detector was installed at the end of the (1)D column to monitor the eluent from (1)D and assist in reconstructing (1)D elution peaks. With the information obtained jointly from the (1)D and (2)D detectors, (1)D elution peaks could be reconstructed with significantly improved (1)D resolution. In this Article, we first discuss the details of the system operating principle and the algorithm to reconstruct (1)D elution peaks, followed by the description and characterization of each component. Finally, 2-D separation of 50 analytes, including alkane (C6-C12), alkene, alcohol, aldehyde, ketone, cycloalkane, and aromatic hydrocarbon, in 14 min is demonstrated, showing the peak capacity of 430-530 and the peak capacity production of 40-80/min.

  17. Electrochemical flow cell, particularly use with liquid chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Matson, W.R.


    An amperometric cell modified for high pressure operation is described. The cell is a flow-through type cell defining a flow path and has at least one active testing electrode, at least one reference electrode and at least one counter electrode operatively disposed and electrically insulated from one another within the cell flow path. In a preferred embodiment the flow cell is encapsulated within a high impact, chemically resistant, chemically insulating material, and has a pair of high pressure resistance fittings communicating with the flow path and extending in part beyond the encapsulation. The cell has particular utility for use with a liquid chromatography separation and when placed in line upstream of sample injection, will remove selected electroactive components in the carrier fluid and thereby reduce background level of contaminents reaching the column and eluting from the column. The cell may also be placed in line following sample injection whereby it may be employed to electrochemically modify selected materials in the mobile phase whereby to change their chromatographic characteristics.

  18. Determination of saffron quality by high-performance liquid chromatography. (United States)

    Valle García-Rodríguez, M; Serrano-Díaz, Jéssica; Tarantilis, Petros A; López-Córcoles, Horacio; Carmona, Manuel; Alonso, Gonzalo L


    The aim of this work was to propose a high-performance liquid chromatography with diode array detection (HPLC-DAD) method for determining the three main compounds responsible for determining the quality of saffron (crocetin esters, picrocrocin, and safranal) by preparing an aqueous extract according to the ISO 3632 standard to solve the difficulty that this standard has for aroma and taste determination by ultraviolet-visible spectroscopy. Toward this aim, laboratory-isolated picrocrocin, a safranal standard with a purity of ≥ 88%, trans-crocetin di(β-D-gentiobiosyl) ester (trans-4-GG) and trans-crocetin (β-D-glucosyl)-(β-D-gentiobiosyl) ester (trans-3-Gg) standards, both with a purity of ≥ 99%, and 50 different saffron spice samples from Italy, Iran, Greece, and Spain were used in the intralaboratory validation of the HPLC method. The analytical method proposed was adequate in terms of linearity, selectivity, sensitivity, and accuracy for determining the three foremost parameters that define the quality of saffron using only a saffron solution prepared according to the ISO 3632 standard.

  19. Electrodialytic membrane suppressors for ion chromatography make programmable buffer generators. (United States)

    Chen, Yongjing; Srinivasan, Kannan; Dasgupta, Purnendu K


    The use of buffer solutions is immensely important in a great variety of disciplines. The generation of continuous pH gradients in flow systems plays an important role in the chromatographic separation of proteins, high-throughput pK(a) determinations, etc. We demonstrate here that electrodialytic membrane suppressors used in ion chromatography can be used to generate buffers. The generated pH, computed from first principles, agrees well with measured values. We demonstrate the generation of phosphate and citrate buffers using a cation-exchange membrane (CEM) -based anion suppressor and Tris and ethylenediamine buffers using an anion-exchange membrane (AEM) -based cation suppressor. Using a mixture of phosphate, citrate, and borate as the buffering ions and using a CEM suppressor, we demonstrate the generation of a highly reproducible (avg RSD 0.20%, n = 3), temporally linear (pH 3.0-11.9, r(2) > 0.9996), electrically controlled pH gradient. With butylamine and a large concentration (0.5 M) of added NaCl, we demonstrate a similar linear pH gradient of large range with a near-constant ionic strength. We believe that this approach will be of value for the generation of eluents in the separation of proteins and other biomolecules and in online process titrations.

  20. SAM Gcms Chromatography Performed at Mars : Elements of Interpretation (United States)

    Szopa, C.; Coll, P. J.; Buch, A.; François, P.; Cabane, M.; Coscia, D.; Teinturier, S.; Navarro-Gonzalez, R.; Glavin, D. P.; Freissinet, C.; Mahaffy, P. R.


    The characterisation of the chemical and mineralogical composition of regolith samples collected with the Curiosity rover is a primary objective of the SAM experiment. These data should provide essential clues on the past habitability of Gale crater. Interpretation of the data collected after SAM pyrolysis evolved gas analysis (EGA) and gas chromatography mass spectrometry (GC-MS) experiments on the first soil samples collected by MSL at the Rocknest Aeolian Deposit in Gale Crater has been challenging due to the concomitant presence in the ovens of an oxychlorine phase present in the samples, and a derivatization agent coming from the SAM wet chemistry experiment (Glavin et al., 2013). Moreover, accurate identification and quantification, in the SAM EGA mode, of volatiles released from the heated sample, or generated by reactions occurring in the SAM pyrolysis oven, is also difficult for a few compounds due to evolution over similar temperature ranges and overlap of their MS signatures. Hence, the GC analyses, coupled with MS, enabled the separation and identification and quantification of most of the volatile compounds detected. These results can have been obtained through tests and calibration done with GC individual spare components and with the SAM testbed. This paper will present a view of the interpretation of the chromatograms obtained when analyzing the Rocknest and John Klein solid samples delivered to SAM, on sols 96 and 199 respectively, supported by laboratory calibrations.

  1. Distributed pore model for bio-molecule chromatography. (United States)

    Coquebert de Neuville, Bertrand; Tarafder, Abhijit; Morbidelli, Massimo


    One of the main peculiarities in protein chromatography is that the adsorbing proteins and the adsorbent pores have comparable sizes. This has the consequence that the pore accessibility depends not only on the solute size but also on the loading conditions of the adsorbent because protein adsorption significantly reduces the size of the pores. A model that accounts for the pore size distribution of the stationary phase and for the pore shrinkage due to protein adsorption has been developed to describe mass transport and adsorption in the porous particles. This model has been shown to be equivalent to the general rate model (GRM) in the case of processes under highly diluted conditions with little adsorption. This implies that the model parameters determination follows the same procedure as for the classical GRM. The new pore model has been applied and compared to the GRM for the simulation of lysozyme breakthrough experiments and for the prediction of 5% dynamic binding capacity values solely based on static capacity measurements.

  2. Quantitative analysis of soil chromatography. I. Water and radionuclide transport

    Energy Technology Data Exchange (ETDEWEB)

    Reeves, M.; Francis, C.W.; Duguid, J.O.


    Soil chromatography has been used successfully to evaluate relative mobilities of pesticides and nuclides in soils. Its major advantage over the commonly used suspension technique is that it more accurately simulates field conditions. Under such conditions the number of potential exchange sites is limited both by the structure of the soil matrix and by the manner in which the carrier fluid moves through this structure. The major limitation of the chromatographic method, however, has been its qualitative nature. This document represents an effort to counter this objection. A theoretical basis is specified for the transport both of the carrier eluting fluid and of the dissolved constituent. A computer program based on this theory is developed which optimizes the fit of theoretical data to experimental data by automatically adjusting the transport parameters, one of which is the distribution coefficient k/sub d/. This analysis procedure thus constitutes an integral part of the soil chromatographic method, by means of which mobilities of nuclides and other dissolved constituents in soils may be quantified.

  3. Chiral Recognition and Enantioseparation Mechanisms in Capillary Electrokinetic Chromatography (United States)

    Chankvetadze, Bezhan

    This chapter deals with the basic theory of enantiomeric separations in electrokinetic chromatography (EKC) in general and with the relationships between the recognition and the separation of enantiomers in EKC, in particular. It is important to note that the dependence between recognition and separation is not as straightforward in EKC as it is in chromatographic separation techniques. Therefore, a clear understanding of these dependences is very important for the explanation of experimentally observed results, as well as for a design of new powerful separation systems, technologies, and materials. Cyclodextrins (CDs) are mainly discussed as chiral selectors not only because the author has a long-term experience of working with these multifunctional macrocycles but also because CDs belong to the most widely used chiral selectors in EKC. In addition, these materials are quite well-characterized molecules of medium size. In addition, CDs are used for separation of enantiomers almost in all analytical separation techniques, as well as for determination of the enantiomeric excess in nonseparation techniques such as nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry. This chapter does not address applications of chiral EKC in chemistry, pharmaceutical and biomedical, environmental, and food analyses.

  4. Microfabricated planar glass gas chromatography with photoionization detection. (United States)

    Lewis, Alastair C; Hamilton, Jacqueline F; Rhodes, Christopher N; Halliday, Jaydene; Bartle, Keith D; Homewood, Philip; Grenfell, Robin J P; Goody, Brian; Harling, Alice M; Brewer, Paul; Vargha, Gergely; Milton, Martin J T


    We report the development of a microfabricated gas chromatography system suitable for the separation of volatile organic compounds (VOCs) and compatible with use as a portable measurement device. Hydrofluoric acid etching of 95x95mm Schott B270 wafers has been used to give symmetrical hemi-spherical channels within a glass substrate. Two matching glass plates were subsequently cold bonded with the channels aligned; the flatness of the glass surfaces resulted in strong bonding through van der Waals forces. The device comprised gas fluidic interconnections, injection zone and 7.5 and 1.4m long, 320microm internal diameter capillaries. Optical microscopy confirmed the capillaries to have fully circular channel profiles. Direct column heating and cooling could be achieved using a combination of resistive heaters and Peltier devices. The low thermal conductivity of glass allowed for multiple uniform temperature zones to be achieved within a single glass chip. Temperature control over the range 10-200 degrees C was achieved with peak power demand of approximately 25W. The 7.5m capillary column was static coated with a 2microm film of non-polar dimethylpolysiloxane stationary phase. A standard FID and a modified lightweight 100mW photoionization detector (PID) were coupled to the column and performance tested with gas mixtures of monoaromatic and monoterpene species at the parts per million concentration level. The low power GC-PID device showed good performance for a small set of VOCs and sub ng detection sensitivity to monoaromatics.

  5. Slow-Equilibration Approximation in Kinetic Size Exclusion Chromatography. (United States)

    Cherney, Leonid T; Krylov, Sergey N


    Kinetic size exclusion chromatography with mass spectrometry detection (KSEC-MS) is a solution-based label-free approach for studying kinetics of reversible binding of a small molecule to a protein. Extraction of kinetic data from KSEC-MS chromatograms is greatly complicated by the lack of separation between the protein and protein-small molecule complex. As a result, a sophisticated time-consuming numerical approach was used for the determination of rate constants in the proof-of-principle works on KSEC-MS. Here, we suggest the first non-numerical (analytical) approach for finding rate constants of protein-small molecule interaction from KSEC-MS data. The approach is based on the slow-equilibration approximation, which is applicable to KSEC-MS chromatograms that reveal two peaks. The analysis of errors shows that the slow-equilibration approximation guarantees that the errors in the rate constants are below 20% if the ratio between the characteristic separation and equilibration times does not exceed 0.1. The latter condition can typically be satisfied for specific interactions such as receptor-ligand or protein-drug. The suggested analytical solution equips analytical scientists with a simple and fast tool for processing KSEC-MS data. Moreover, a similar approach can be potentially developed for kinetic analysis of protein-small molecule binding by other kinetic-separation methods such as nonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM).

  6. Methacrylate-based monolithic layers for planar chromatography of polymers. (United States)

    Maksimova, E F; Vlakh, E G; Tennikova, T B


    A series of macroporous monolithic methacrylate-based materials was synthesized by in situ free radical UV-initiated copolymerization of functional monomers, such as glycidyl methacrylate (GMA), butyl methacrylate (BuMA), 2-aminoethyl methacrylate (AEMA), 2-hydroxyethyl methacrylate (HEMA) and 2-cyanoethyl methacrylate (CEMA), with crosslinking agent, namely, ethylene glycol dimethacrylate (EDMA). The materials obtained were applied as the stationary phases in simple and robust technique - planar chromatography (PLC). The method of separation layer fabrication representing macroporous polymer monolith bound to the specially prepared glass surface was developed and optimized. The GMA-EDMA and BuMA-EDMA matrixes were successfully applied for the separation of low molecular weight compounds (the mixture of several dies), as well as poly(vinylpyrrolidone) and polystyrene homopolymers of different molecular weights using reversed-phase mechanism. The materials based on copolymers AEMA-HEMA-EDMA and CEMA-HEMA-EDMA were used for normal-phase PLC separation of 2,4-dinitrophenyl amino acids and polystyrene standards.


    Directory of Open Access Journals (Sweden)

    Kaushal Ramni


    Full Text Available HPLC is the most versatile and widely used elution chromatography. The technique is used to resolve and determine species in a variety of organic, inorganic, biological, ionic and polymeric materials. Detector is the heart of an instrument and efficiency of system is dependent upon detecting techniques. Many types of HPLC detectors exist, each of which has some valuable performance feature such as refractive index detector, ultraviolet detector, fluorescent detector, electrochemical detector, electric conductivity detector, liquid light scattering detector, evaporative light scattering detector. Due to strong requirement for improvements in sensitivity, selectivity and other performance characteristics of the detector recent developments in conventional techniques and some other new technologies have been adopted such as laser light scattering detector, charged aerosol detector, nano quantity aerosol detector, chiral detector and pulsed amperometric detector. These detectors provide accurate concentration analysis, excellent sensitivity, wide dynamic range, consistent response and broad applicability of the drug components. Working of these detectors involve different principles such as optical techniques, aerosol based techniques, refractive methods, light scattering principle, amperometric and fluorescence. The present review enlightens both conventional and advanced techniques and compares their capabilities of analyzing drug components and need for new techniques for better and wide range of applicability.

  8. Microemulsion electrokinetic chromatography for analysis of phthalates in soft drinks. (United States)

    Hsieh, Sung-Yu; Wang, Chun-Chi; Wu, Shou-Mei


    Microemulsion electrokinetic chromatography (MEEKC) is proposed for analysis of di-n-butyl phthalate (DBP) and di-(2-ethylhexyl) phthalate (DEHP) in soft drinks. However, the instability of microemulsion is a critical issue. In this research, a novel material, Pluronic® F-127, which has the properties of polymer and surfactant, was added for stabilizing the microemulsion in the MEEKC system. Our data demonstrate that the presence of Pluronic® F-127 (0.05-0.30%) also helps enhance resolution of highly hydrophobic compounds, DBP and DEHP. The electrokinetic injection of sodium dodecyl sulphate (SDS) including sample (-10 kV, 20 s) was introduced in this MEEKC system and this yielded about 25-fold sensitivity enhancement compared with hydrodynamic injection (1 psi, 10 s). During method validation, calibration curves were linear (r≥0.99), within a range of 75-500 ng/mL for DBP and 150-1000 ng/mL for DEHP. As the precision and accuracy assays, absolute values of relative standard deviation (RSD) and relative error (RE) in intraday (n=3) and interday (n=5) observations were less than 4.93%. This method was further applied for analyzing six commercial soft drinks and one was found containing 453.67 ng/mL of DEHP. This method is considered feasible for serving as a tool for analysis of highly hydrophobic molecules.

  9. Ion-exchange chromatography for the characterization of biopharmaceuticals. (United States)

    Fekete, Szabolcs; Beck, Alain; Veuthey, Jean-Luc; Guillarme, Davy


    Ion-exchange chromatography (IEX) is a historical technique widely used for the detailed characterization of therapeutic proteins and can be considered as a reference and powerful technique for the qualitative and quantitative evaluation of charge heterogeneity. The goal of this review is to provide an overview of theoretical and practical aspects of modern IEX applied for the characterization of therapeutic proteins including monoclonal antibodies (Mabs) and antibody drug conjugates (ADCs). The section on method development describes how to select a suitable stationary phase chemistry and dimensions, the mobile phase conditions (pH, nature and concentration of salt), as well as the temperature and flow rate, considering proteins isoelectric point (pI). In addition, both salt-gradient and pH-gradient approaches were critically reviewed and benefits as well as limitations of these two strategies were provided. Finally, several applications, mostly from pharmaceutical industries, illustrate the potential of IEX for the characterization of charge variants of various types of biopharmaceutical products.

  10. Solvent extraction, ion chromatography, and mass spectrometry of molybdenum isotopes. (United States)

    Dauphas, N; Reisberg, L; Marty, B


    A procedure was developed that allows precise determination of molybdenum isotope abundances in natural samples. Purification of molybdenum was first achieved by solvent extraction using di(2-ethylhexyl) phosphate. Further separation of molybdenum from isobar nuclides was obtained by ion chromatography using AG1-X8 strongly basic anion exchanger. Finally, molybdenum isotopic composition was measured using a multiple collector inductively coupled plasma hexapole mass spectrometer. The abundances of molybdenum isotopes 92, 94, 95, 96, 97, 98, and 100 are 14.8428(510), 9.2498(157), 15.9303(133), 16.6787(37), 9.5534(83), 24.1346(394), and 9.6104(312) respectively, resulting in an atomic mass of 95.9304(45). After internal normalization for mass fractionation, no variation of the molybdenum isotopic composition is observed among terrestrial samples within a relative precision on the order of 0.00001-0.0001. This demonstrates the reliability of the method, which can be applied to searching for possible isotopic anomalies and mass fractionation.

  11. Determination of inorganic anions in papermaking waters by ion chromatography

    Directory of Open Access Journals (Sweden)



    Full Text Available A suppressed ion chromatography (IC method for the determination of inorganic anions in process water from paperboard production was developed and validated. Common inorganic anions (Cl-, NO3-, PO43- and SO42- were detected in fresh and process water samples collected from a paperboard production system at 16 characteristic points. It was shown that the use of an IonPac®-AS14 column under isocratic conditions with Na2CO3/NaHCO3 as the eluent and a suppression device proved to be a reliable analytical solution for the separation of the inorganic anions present in papermaking waters. This IC method is quite satisfactory concerning selectivity and sensitivity, and enables the determination of several inorganic anions over a wide concentration range. According to the obtained results, the total amount of analyzed inorganic anions was below 0.1 g/L, i.e., below the critical value which may trigger operational problems in paper production.

  12. Application of High Performance Liquid Chromatography to Separation of Novel Chiral Tetrahedral Heterometal Clusters

    Institute of Scientific and Technical Information of China (English)

    Xin Yi ZHU; Wei Qiang ZHANG; Yu Hua ZHANG; Li Ren CHEN; Yong Min LI


    A series of novel chiral tetrahedral heterometal clusters have firstly been separated oncellulose tris-(3,5-dimethylphenylcarbamate) stationary phase by high performance liquid chrom-atography, using hexane as the mobile phase with various alcohols as modifiers.

  13. Determination of Aspartame, Caffeine, Saccharin, and Benzoic Acid in Beverages by High Performance Liquid Chromatography. (United States)

    Delaney, Michael F.; And Others


    Describes a simple and reliable new quantitative analysis experiment using liquid chromatography for the determinaiton of caffeine, saccharin, and sodium benzoate in beverages. Background information, procedures used, and typical results obtained are provided. (JN)

  14. Separation by column chromatography of cells active in delayed-onset hypersensitivities. (United States)

    Godfrey, H P; Gell, P G


    Lymph node cells from guinea-pigs contact sensitive to 1-thiocyamo-2,4-dinitrobenzene have been fractionated by affinity chromatography over modified polyacrylamide beads. Cells mediating lymphokine release in response to active sensitizer were depleted only by chromatography over dinitrophyenyl (DNP) containing substrates and could be specifically eluted with DNP-glycine. DNP rosette-forming cells (RFC) were equally well depleted by chromatography using either DNP or trinitrophenyl containing materials but could not be eluted from the columns by DNP-glycine. While the antigen receptors of cells mediating the release of macrophage agglutination factor in response to DNP-containing antigens and of DNP-RFC were found to be hapten-specific, their specificity was shown to differ using chromatography over trinitrophenyl containing polyacrylamide. PMID:776818

  15. The Binding of Biotin to Sepharose-Avidin Column: Demonstration of the Affinity Chromatography Technique (United States)

    Landman, A. D.; Landman, N. N.


    Describes a biochemistry experiment that illustrates the methodology of affinity chromatography by attaching avidin, a glycoprotein in egg white, to a Sepharose matrix in order to bind biotin-containing proteins. (MLH)

  16. Physical-chemical fundamentals and the application of fluid chromatography (SFC)

    Energy Technology Data Exchange (ETDEWEB)

    Wasen, U. van; Swaid, I.; Schneider, G.M.


    In super fluid chromatography chromatography (SFC), highly condensed gases in the critical temperature region are used as mobile phases. This method has considerable advantages in the analytical or preparative separation of particularly difficultly volatile or thermally unstable substances compared to gas chromatography. It competes with HPLC and gel chromatography in certain special applications and preparative separations, e.g. in petroleum industry and the separation of oligomers. It is also of great interest for basic investigations of fluid extraction and determination of physical-chemical parameters of fluid systems. The most important physical-chemical, methodical and apparative fundamentals of SFC are discussed in summary form; characteristic physical-chemical examples of application are the determination of capacity ratios, distribution quotients, partial mole volumes, mixed virial coefficients and diffusion coefficients.

  17. Simultaneous Determination of Chelating Agents by Ion-Suppression and Ion-Pair Chromatography in Wastewater

    Energy Technology Data Exchange (ETDEWEB)

    Dodi, Alain; Bouscarel, Maelle [Commissariat a l' energie atomique - C.E.A, Centre d' Etude de Cadarache, Laboratoire d' Analyses Radiochimiques et Chimiques, St Paul lez Durance (France)


    This article describes two methods for analysing chelating agents found in nuclear waste. First, ion-suppression chromatography using an anion exchange stationary phase and mobile phase consisting of a nitric acid solution and pure water gradient. UV detection was performed at 330 nm after the reaction with a post-column reagent composed of iron nitrate in perchloric acid. Secondly, ion-pair chromatography with a mobile phase consisting of a mixture of nitric acid, tetra-butyl-ammonium hydrogeno-sulphate, tetra-butyl-ammonium hydroxide and iron chloride. A reversed-phase material was used as a stationary phase and detection was performed by direct measurement of the UV absorption at 260 nm. The quantification limits were lower for ion-pair chromatography than for ion-suppression chromatography. Both methods were easy to implement and allow a multi-element separation in less than 30 min with low detection limits. (authors)

  18. Steroid monochloroacetates : Physical-chemical characteristics and use in gas-liquid chromatography

    NARCIS (Netherlands)

    Molen, H.J. van der; Groen, D.; Maas, J.H. van der


    Synthesis and physical-chemical characteristics (melting points, infrared-, visible- and ultraviolet spectra, paper-,thin-layer- and gas-liquid Chromatographie behaviour) of monochloroacetate derivatives of steroids representing the androstane-, pregnane-, estrane- and cholestane series are describe

  19. Selective retention of basic compounds by metal aquo-ion affinity chromatography. (United States)

    Asakawa, Yoshiki; Yamamoto, Eiichi; Asakawa, Naoki


    A novel metal aquo-ion affinity chromatography has been developed for the analysis of basic compounds using heat-treated silica gel containing hydrated metal cations (metal aquo-ions) as the packing material. The packing materials of the metal aquo-ion affinity chromatography were prepared by the immobilization of a single metal component such as Fe(III), Al(III), Ag(I), and Ni(II) on silica gel followed by extensive heat treatment. The immobilized metals form aquo-ions to present cation-exchange ability for basic analytes and the cation-exchange ability for basic analytes depends on pKa of the immobilized metal species. In the present study, to evaluate the retention characteristics of metal aquo-ion affinity chromatography, the on-line solid-phase extraction of drugs was investigated. Obtained data clearly evidence the selective retention capability of metal aquo-ion affinity chromatography for basic analytes with sufficient capacity.

  20. Refolding of detergent-denatured lysozyme using β-cyclodextrin-assisted ion exchange chromatography. (United States)

    Zhang, Li; Zhang, Qinming; Wang, Chaozhan


    Chromatography-based protein refolding is widely used. Detergent is increasingly used for protein solubilization from inclusion bodies. Therefore, it is necessary to develop a refolding method for detergent-denatured/solubilized proteins based on liquid chromatography. In the present work, sarkosyl-denatured/dithiothreitol-reduced lysozyme was used as a model, and a refolding method based on ion exchange chromatography, assisted by β-cyclodextrin, was developed for refolding detergent-denatured proteins. Many factors affecting the refolding, such as concentration of urea, concentration of β-cyclodextrin, pH and flow rate of mobile phases, were investigated to optimize the refolding conditions for sarkosyl-denatured lysozymes. The results showed that the sarkosyl-denatured lysozyme could be successfully refolded using β-cyclodextrin-assisted ion exchange chromatography.

  1. Fast methods for screening of trichothecenes in fungal cultures using gas chromatography-tandem mass spectrometry

    DEFF Research Database (Denmark)

    Nielsen, Kristian Fog; Thrane, Ulf


    The paper presents a fast method for trichothecene profiling and chemotaxonomic studies in species of Fusarium, Stachybotrys, Trichoderma and Memnoniella. Micro scale extracted crude Fusarium extracts were derivatised using pentafluoropropionic anhydride and analysed by gas chromatography...

  2. Theory of chromatography of partially cyclic polymers: Tadpole-type and manacle-type macromolecules. (United States)

    Vakhrushev, Andrey V; Gorbunov, Alexei A


    A theory of chromatography is developed for partially cyclic polymers of tadpole- and manacle-shaped topological structures. We present exact equations for the distribution coefficient K at different adsorption interactions; simpler approximate formulae are also derived, relevant to the conditions of size-exclusion, adsorption, and critical chromatography. Theoretical chromatograms of heterogeneous partially cyclic polymers are simulated, and conditions for good separation by topology are predicted. According to the theory, an effective SEC-radius of tadpoles and manacles is mostly determined by the molar mass M, and by the linear-cyclic composition. In the interactive chromatography, the effect of molecular topology on the retention becomes significant. At the critical interaction point, partial dependences K(Mlin) and K(Mring) are qualitatively different: while being almost independent of Mlin, K increases with Mring. This behavior could be realized in critical chromatography-for separation of partially cyclic polymers by the number and molar mass of cyclic elements.

  3. Purification of a Mycoplasma pneumoniae adhesin by monoclonal antibody affinity chromatography.


    Leith, D K; Baseman, J B


    A 165,000-dalton surface protein of Mycoplasma pneumoniae, designated protein P1, appears to be the major attachment ligand of the pathogen. We employed monoclonal antibody affinity chromatography to obtain purified protein P1.

  4. Utilizing ion-pairing hydrophilic interaction chromatography solid phase extraction for efficient glycopeptide enrichment in glycoproteomics

    DEFF Research Database (Denmark)

    Mysling, Simon; Palmisano, Giuseppe; Højrup, Peter;


    Glycopeptide enrichment is a prerequisite to enable structural characterization of protein glycosylation in glycoproteomics. Here we present an improved method for glycopeptide enrichment based on zwitter-ionic hydrophilic interaction chromatography solid phase extraction (ZIC-HILIC SPE...

  5. Purification of the envelope glycoproteins of western equine encephalitis virus by glass wool column chromatography.


    Yamamoto, K.; Simizu, B


    Glass wool column chromatography was used for separation of the two glycoproteins of western equine encephalitis virus. Cross-contamination of each protein separated was confirmed to be negligible by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.


    With the objective of detecting and quantitating low concentrations of perfluorinated carboxylic acids (PFCAs), including perfluorinated octanoic acid (PFOA), in soils, we compared the analytical suitability of liquid chromatography columns containing three different stationary p...

  7. Evaluation of a silicon oxynitride hydrophilic interaction liquid chromatography column in saccharide and glycoside separations. (United States)

    Wan, Huihui; Sheng, Qianying; Zhong, Hongmin; Guo, Xiujie; Fu, Qing; Liu, Yanfang; Xue, Xingya; Liang, Xinmiao


    The retention characteristics of a silicon oxynitride stationary phase for carbohydrate separation were studied in hydrophilic interaction chromatography mode. Four saccharides including mono-, di-, and trisaccharides were employed to investigate the effects of water content and buffer concentration in the mobile phase on hydrophilic interaction liquid chromatography retention. For the tested saccharides, the silicon oxynitride column demonstrated excellent performance in terms of separation efficiency, hydrophilicity, and interesting separation selectivity for carbohydrates compared to the bare silica stationary phase. Finally, the silicon oxynitride hydrophilic interaction liquid chromatography column was employed in the separation of complex samples of fructooligosaccharides, saponins, and steviol glycoside from natural products. The resulting chromatograms demonstrated good separation efficiency and longer retention compared with silica, which further confirmed the advantages and potential application of silicon oxynitride stationary phase for hydrophilic interaction liquid chromatography separation.

  8. Twin-column CaptureSMB: a novel cyclic process for protein A affinity chromatography. (United States)

    Angarita, Monica; Müller-Späth, Thomas; Baur, Daniel; Lievrouw, Roel; Lissens, Geert; Morbidelli, Massimo


    A twin-column counter-current chromatography processes, CaptureSMB, was used for the protein A affinity capture of a monoclonal antibody (mAb). By means of sequential loading, the process improves the utilization of the stationary phase by achieving loadings much closer to the static binding capacity of the resin in comparison to batch chromatography. Using a mAb capture case study with protein A affinity chromatography, the performance and product quality obtained from CaptureSMB and batch processes were compared. The effect of the flow rate, column length and titer concentration on the process performance and product quality were evaluated. CaptureSMB showed superior performance compared to batch chromatography with respect to productivity, capacity utilization, product concentration and buffer consumption. A simplified economic evaluation showed that CaptureSMB could decrease resin costs of 10-30% depending on the manufacturing scenario.


    A procedure is presented that uses a vacuum distillation/gas chromatography/mass spectrometry system for analysis of problematic matrices of volatile organic compounds. The procedure compensates for matrix effects and provides both analytical results and confidence intervals from...

  10. Two-dimensional liquid chromatography and its application in traditional Chinese medicine analysis and metabonomic investigation. (United States)

    Li, Zheng; Chen, Kai; Guo, Meng-zhe; Tang, Dao-quan


    Two-dimensional liquid chromatography has become an attractive analytical tool for the separation of complex samples due to its enhanced selectivity, peak capacity, and resolution compared with one-dimensional liquid chromatography. Recently, more attention has been drawn on the application of this separation technique in studies concerning traditional Chinese medicines, metabonomes, proteomes, and other complex mixtures. In this review, we aim to examine the application of two-dimensional liquid chromatography in traditional Chinese medicine analysis and metabonomic investigation. The classification and evaluation indexes were first introduced. Then, various switching methods were summarized when used in an on-line two-dimensional liquid chromatography system. Finally, the applications of this separation technique in traditional Chinese medicine analysis and metabonomic investigation were discussed on the basis of specific studies.

  11. Two-dimensional liquid chromatography for direct chiral separations: a review. (United States)

    León-González, María Eugenia; Rosales-Conrado, Noelia; Pérez-Arribas, Luis Vicente; Guillén-Casla, Vanesa


    Separation of enantiomers remains a challenge owing to their identical physical and chemical properties in an achiral environment, and research on specialized separation techniques such as multidimensional achiral-chiral liquid chromatography continues to resolve individual enantiomers in complex samples. Recent advances in application of multidimensional liquid chromatography applied to chiral analysis are reviewed. For this reason, benefits of achiral-chiral coupling are shown, with emphasis in applications on biological and pharmaceutical fields as well as pesticide analysis. A description of standard instrumental setup in both heart-cut and comprehensive multidimensional liquid chromatography is shown. The most broadly used chiral stationary phases for multidimensional liquid chromatography are summarized. An extensive overview of different interface designs applied to complex samples is presented.

  12. Analysis of chemical signals in red fire ants by gas chromatography and pattern recognition techniques (United States)

    The combination of gas chromatography and pattern recognition (GC/PR) analysis is a powerful tool for investigating complicated biological problems. Clustering, mapping, discriminant development, etc. are necessary to analyze realistically large chromatographic data sets and to seek meaningful relat...

  13. Comparison of microemulsion electrokinetic chromatography with high-performance liquid chromatography for fingerprint analysis of resina draconis. (United States)

    Cao, Yuhua; Gong, Wenjun; Li, Nan; Yin, Changna; Wang, Yun


    Microemulsion electrokinetic chromatography (MEEKC) has been developed for fingerprint analysis of resina draconis, a substitute for sanguis draconis in the Chinese market. The microemulsion as the running buffer was made up of 3.3% (w/v) sodium dodecyl sulfate (SDS), 6.6% (w/v) n-butanol, 0.8% (w/v) n-octane, and 10 mmol/L sodium tetraborate buffer (pH 9.2), which was also used as the solvent for ultrasonic extraction of both water- and fat-soluble compounds in the traditional Chinese medicine samples. Four batches of resina draconis obtained from different pharmaceutical factories located in different geographic regions were used to establish the electrophoretic fingerprint. MEEKC was performed using a Beckman PACE/MDQ system equipped with a diode-array detector and with monitoring at 280 nm. The fingerprint of resina draconis comprised 27 common peaks within 100 min. The relative standard deviations of the relative migration time of these common peaks were less than 2.1%. Through repetitive injection of the sample solution six times in 24 h, all relative standard deviations of the migration time and peak area of loureirin A and loureirin B were less than 2.5 and 3.8%, which demonstrated that the method had good stability and reproducibility. The relative peak areas of these common peaks in the electropherograms of four batches of resina draconis were processed with two mathematical methods, the correlation coefficient and the interangle cosine, to valuate the similarity. The values of the similarity degree of all samples were more than 0.91, which showed resina draconis samples from different origins were consistent. On the other hand, high-performance liquid chromatography (HPLC) coupled with photodiode-array detection was also applied to establish the fingerprint of resina draconis. The samples were separated with a LiChrospher C(18) column using acetonitrile (solvent A) and water containing 0.1% H(3)PO(4) (solvent B) as the mobile phase in linear gradient

  14. Enantiomeric separation of organophosphorus pesticides by high-performance liquid chromatography, gas chromatography and capillary electrophoresis and their applications to environmental fate and toxicity assays. (United States)

    Li, Ling; Zhou, Shanshan; Jin, Lixia; Zhang, Cheng; Liu, Weiping


    In recent years, the continuous evolution of the field of stereochemistry has produced a heightened awareness of the applications of pure enantiomers of agrochemicals. This review describes reports of the enantiomeric separation of commercial organophosphorus pesticides (OPs) and the applications of these methods to research on the enantioselectivity of the toxicity and environmental fate of these compounds. Chiral OPs can be analysed by high-performance liquid chromatography (HPLC), gas chromatography (GC), and capillary electrophoresis (CE). These different separation techniques for OP enantiomers are briefly discussed, and their applications are presented.

  15. Dosage direct des alcools dans les carburols par chromatographie en phase gazeuse Direct Gas Chromatographie Determination of Alcohols in Gasolines Blended by Oxygenated Compounds

    Directory of Open Access Journals (Sweden)

    Petroff N.


    Full Text Available Dans cette étude, on décrit une méthode de chromatographie en phase gazeuse utilisant une colonne remplie ; elle permet de doser, sans préfractionnement, les alcools jusqu'aux butanols dans les carburols (carburants additionnés de composés oxygénés. La phase stationnaire sélectionnée est le glycérol. This report describes a gas chromatographie method using a packed column. It allows the direct determination of alcohols up to butanols in gasolines blended with oxygenated compounds without prefractionation. Glycerol has been selected as stationary phase.

  16. The Application of High Performance Liquid Chromatography in the Analysis of Trace Rare Earth

    Institute of Scientific and Technical Information of China (English)

    WANG; Xiu-feng; DING; You-qian


    <正>Rare earth elements are very important in the field of radioanalytical chemistry, for it must be separated and determined in the measurements of burn-up and fission yield. High performance liquid chromatography has become a main method in the separation of rare earth elements due to its obvious advantages, this is, high speed of analysis, high efficiency and easy automation. The ion exchange chromatography is the main means to separate rare earth elements, especially the cation exchange

  17. Virtual Practice of Biochemistry: Ascendant Chromatography on Paper of Amino acids.


    Zanatta,G.; De Moraes, C G; E.R. Zimmer; E. Ewald; Pettenuzzo,L.F.; Salbego, C. G.; V.M.T. Trindade


    Chromatography is a method used for analyses of mixture components based on different criteria: adsorbility, solubility, molecular mass, ionic charge and affinity. It is an important subject of Biochemistry. This work reports the development and the validation of a software that simulated a laboratory activity named Ascendant Chromatography on Paper of Aminoacids. The organization and the multimedia material collection were done during the 2008/2 semester. The most representative images were ...

  18. Separation of mAbs molecular variants by analytical hydrophobic interaction chromatography HPLC: overview and applications


    Haverick, Mark; Mengisen, Selina; Shameem, Mohammed; Ambrogelly, Alexandre


    Hydrophobic interaction chromatography-high performance liquid chromatography (HIC-HPLC) is a powerful analytical method used for the separation of molecular variants of therapeutic proteins. The method has been employed for monitoring various post-translational modifications, including proteolytic fragments and domain misfolding in etanercept (Enbrel®); tryptophan oxidation, aspartic acid isomerization, the formation of cyclic imide, and α amidated carboxy terminus in recombinant therapeutic...

  19. Copolymerization preparation of cationic cyclodextrin chiral stationary phases for drug enantioseparation in chromatography




    Authors: Ren-Qi Wang, Teng-Teng Ong, Ke Huang, Weihua Tang & Siu-Choon Ng ### Abstract We described a facile and effective protocol wherein radical copolymerization is employed to covalently bond cationic β-cyclodextrin (β-CD) onto silica particles with extended linkage, resulting in a chiral stationary phase (IMPCSP) that can be used for the enantioseparation of racemic drugs in both high-performance liquid chromatography (HPLC) and supercritical fluid chromatography (SFC). Start...

  20. If You Were a Molecule in a Chromatography Column, What Would You See? (United States)

    Mattice, John


    To visualize what takes place in a chromatography column, enlarge the molecules to human size and expand the columns to keep the ratio of size of molecule to size of column the same. If we were molecules, what would the columns be like? A typical gas chromatography (GC) capillary column would be 50 x 10 [superscript 6] 6 km (31 million mi) long,…



    Michaela Vieriková; Miriam Vlčáková


     Acrylamide in food was determined by gas chromatography - mass spectrometry (GC-MS) after bromination of acrylamide and underivatized acrylamide was quantified by ultra performance liquid chromatography -mass spectrometry (UPLC-MS). Two different sample preparation methods were used and optimised. The GC-MS method was used for various food matrices like breads, potato crisps, potato crackers, french fries. The UPLC-MS method was used for analysis of coffee. The limit of detect...

  2. [Development of methods for determining acrylamide in food products by gas-liquid chromatography]. (United States)

    Bessonov, V V; Malinkin, A D; Perederiaev, O I; Bogachuk, M N; Volkovich, S V; Medvedev, Iu V


    The method of determination of acrylamide in various food (milk powder, potato chips, instant coffee) by gas-liquid chromatography after pre-bromination was developed. Studies have shown the possibility of using bromination of acrylamide to give it the necessary properties for better extraction, purification and detection. Also revealed the possibility of qualitative and quantitative determine a acrylamide in food by gas-liquid chromatography with detection by electron capture detector.

  3. A Simple and Efficient Method for Purification of Egg White Major Proteins Using Ion Exchange Chromatography


    Sh. Veisi; A. Mostafaie; Z. Mohammad Hasan


    Introduction & Objective: Egg white contains four high-quantity proteins which have numerous applications. In this research, a simple and efficient method for the purification of those proteins was designed and performed based on ion exchange chromatography.Materials & Methods: In this experimental study egg white was initially separated from insoluble substances by acidic pH. The resulting extract was isolated after two steps of ion exchange chromatography using CM-Sepharose and DEAE-Sepharo...

  4. Sample displacement chromatography as a method for purification of proteins and peptides from complex mixtures


    Gajdosik, Martina Srajer; Clifton, James; Josic, Djuro


    Sample displacement chromatography (SDC) in reversed-phase and ion-exchange modes was introduced approximately twenty years ago. This method takes advantage of relative binding affinities of components in a sample mixture. During loading, there is a competition among different sample components for the sorption on the surface of the stationary phase. SDC was first used for the preparative purification of proteins. Later, it was demonstrated that this kind of chromatography can also be perform...

  5. The analysis of aqueous mixtures using liquid chromatography-electrospray mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, Steven [Iowa State Univ., Ames, IA (United States)


    The focus of this dissertation is the use of chromatographic methods coupled with electrospray mass spectrometry (ES-MS) for the determination of both organic and inorganic compounds in aqueous solutions. The combination of liquid chromatography (LC) methods and ES-MS offers one of the foremost methods for determining compounds in complex aqueous solutions. In this work, LC-ES-MS methods are devised using ion exclusion chromatography, reversed phase chromatography, and ion exchange chromatography, as well as capillary electrophoresis (CE). For an aqueous sample, these LC-ES-MS and CE-ES-MS techniques require no sample preparation or analyte derivatization, which makes it possible to observe a wide variety of analytes as they exist in solution. The majority of this work focuses on the use of LC-ES-MS for the determination of unknown products and intermediates formed during electrochemical incineration (ECI), an experimental waste remediation process. This report contains a general introduction to the project and the general conclusions. Four chapters have been removed for separate processing. Titles are: Chapter 2: Determination of small carboxylic acids by ion exclusion chromatography with electrospray mass spectrometry; Chapter 3: Electrochemical incineration of benzoquinone in aqueous media using a quaternary metal oxide electrode in the absence of a soluble supporting electrolyte; Chapter 4: The determination of electrochemical incineration products of 4-chlorophenol by liquid chromatography-electrospray mass spectrometry; and Chapter 5: Determination of small carboxylic acids by capillary electrophoresis with electrospray mass spectrometry.

  6. Triticonazole enantiomers: Separation by supercritical fluid chromatography and the effect of the chromatographic conditions. (United States)

    He, Jianfeng; Fan, Jun; Yan, Yilun; Chen, Xiaodong; Wang, Tai; Zhang, Yaomou; Zhang, Weiguang


    Enantiomeric pairs of triticonazole have been successfully separated by supercritical fluid chromatography coupled with a tris(3,5-dimethylphenylcarbamoyl) cellulose-coated chiral stationary phase in this work. The effects of co-solvent, dissolution solvent, flow rate, backpressure, and column temperature have been studied in detail with respect to retention, selectivity, and resolution of triticonazole. As indicated, the co-solvents mostly affected the retention factors and resolution, due to the different molecular structure and polarity. In addition, the dissolution solvents, namely, chloromethanes and alcohols, have been also important for enantioseparation because of the different interaction with stationary phase. Higher flow rate and backpressure led to faster elution of the triticonazole molecules, and the change of column temperature showed slight effect on the resolution of triticonazole racemate. Moreover, a comparative separation experiment between supercritical fluid chromatography and high performance liquid chromatography revealed that chiral supercritical fluid chromatography gave the 3.5 times value of Rs /tR2 than high performance liquid chromatography, which demonstrated that supercritical fluid chromatography had much higher separation efficiency.

  7. Applications of hydrophilic interaction chromatography to amino acids, peptides, and proteins. (United States)

    Periat, Aurélie; Krull, Ira S; Guillarme, Davy


    This review summarizes the recent advances in the analysis of amino acids, peptides, and proteins using hydrophilic interaction chromatography. Various reports demonstrate the successful analysis of amino acids under such conditions. However, a baseline resolution of the 20 natural amino acids has not yet been published and for this reason, there is often a need to use mass spectrometry for detection to further improve selectivity. Hydrophilic interaction chromatography is also recognized as a powerful technique for peptide analysis, and there are a lot of papers showing its applicability for proteomic applications (peptide mapping). It is expected that its use for peptide mapping will continue to grow in the future, particularly because this analytical strategy can be combined with reversed-phase liquid chromatography, in a two-dimensional setup, to reach very high resolving power. Finally, the interest in hydrophilic interaction chromatography for intact proteins analysis is less evident due to possible solubility issues and a lack of suitable hydrophilic interaction chromatography stationary phases. To date, it has been successfully employed only for the characterization of membrane proteins, histones, and the separation of glycosylated isoforms of an intact glycoprotein. From our point of view, the number of hydrophilic interaction chromatography columns compatible with intact proteins (higher upper temperature limit, large pore size, etc.) is still too limited.

  8. Nonspecific native elution of proteins and mumps virus in immunoaffinity chromatography. (United States)

    Brgles, Marija; Sviben, Dora; Forčić, Dubravko; Halassy, Beata


    Immunoaffinity chromatography, based on the antigen-antibody recognition, enables specific purification of any antigen (protein, virus) by its antibody. The problem with immunoaffinity chromatography is the harsh elution conditions required for disrupting strong antigen-antibody interactions, such as low pH buffers, which are often deleterious for the immobilized protein and the protein to be isolated since they can also disrupt the intramolecular forces. Therefore, immunoaffinity chromatography can only be partially used for protein and virus purification. Here we report on a nonspecific elution in immunoaffinity chromatography using native conditions by elution with amino acid solution at physiological pH for which we suppose possible competing mechanism of action. Elution potential of various amino acid solutions was tested using immunoaffinity columns specific for ovalbumin and mumps virus, and protein G affinity column. Results have shown that the most successful elution solutions were those containing imidazole and arginine of high molarity. Imidazole represents aromatic residues readily found at the antigen-antibody interaction surface and arginine is most frequently found on protein surface in general. Therefore, results on their eluting power in immunoaffinity chromatography, which increases with increasing molarity, are in line with the competing mechanism of action. Virus immunoaffinity chromatography resulted in removal on nonviable virus particles, which is important for research and biotechnology purposes. In addition, amino acids are proven stabilizers for proteins and viruses making approach presented in this work a very convenient purification method.

  9. Construction of a new interface for comprehensive supercritical fluid chromatography x reversed phase liquid chromatography (SFC x RPLC). (United States)

    François, Isabelle; Pereira, Alberto dos Santos; Lynen, Frédéric; Sandra, Pat


    The design of a new interface to hyphen high efficiency supercritical fluid chromatography (SFC) and fast RPLC in a comprehensive configuration is described. SFC x RPLC is a viable alternative to normal phase (NP) LC x RPLC and is characterized by high orthogonality. Compared to NPLC x RPLC an additional advantage is the expansion of supercritical carbon dioxide (CO(2)) when exposed to atmospheric pressure leading to fractions consisting of solvents that are miscible with the second dimension RPLC mobile phase. The interface consists of a two-position/ten-port switching valve equipped with two packed octadecyl silica (C(18)) loops for effective trapping and focusing of the analytes after elution from the SFC dimension. The addition of a water make-up flow to the SFC effluent prior to entering the loops is of fundamental importance to efficiently focus the solutes on the C(18) material and to reduce interferences of expanded CO(2) gas on the second dimension separation. The features of the system are illustrated with the analysis of a lemon oil sample.

  10. Predicting drug penetration across the blood-brain barrier: comparison of micellar liquid chromatography and immobilized artificial membrane liquid chromatography. (United States)

    De Vrieze, Mike; Lynen, Frédéric; Chen, Kai; Szucs, Roman; Sandra, Pat


    Several in vitro methods have been tested for their ability to predict drug penetration across the blood-brain barrier (BBB) into the central nervous system (CNS). In this article, the performance of a variety of micellar liquid chromatographic (MLC) methods and immobilized artificial membrane (IAM) liquid chromatographic approaches were compared for a set of 45 solutes. MLC measurements were performed on a C18 column with sodium dodecyl sulfate (SDS), polyoxyethylene (23) lauryl ether (Brij35), or sodium deoxycholate (SDC) as surfactant in the micellar mobile phase. IAM liquid chromatography measurements were performed with Dulbecco's phosphate-buffered saline (DPBS) and methanol as organic modifier in the mobile phase. The corresponding retention and computed descriptor data for each solute were used for construction of models to predict transport across the blood-brain barrier (log BB). All data were correlated with experimental log BB values and the relative performance of the models was studied. SDS-based models proved most suitable for prediction of log BB values, followed closely by a simplified IAM method, in which it could be observed that extrapolation of retention data to 0% modifier in the mobile phase was unnecessary.

  11. Contribution to the chromatography of atmospheric gases (1963); Contribution a la chromatographie des gaz de l'air (1963)

    Energy Technology Data Exchange (ETDEWEB)

    Ghalamsiah, A. [Commissariat a l' Energie Atomique, Saclay (France). Centre d' Etudes Nucleaires


    In the first part, the author studies the gas-phase chromatographic separation of the atmospheric rare gases, of hydrogen, and of some gaseous compounds of carbon (CO, CO{sub 2}, CH{sub 4}) using inactive gases to obtain the most favourable operational conditions far this separation. In the second part, the optimum conditions for detecting non-active gases using an ionisation chamber and a {sup 239}Pu radioactive source emitting 5.15 MeV {alpha} particles are determined. (author) [French] Dans une premiere partie, l'auteur etudie la separation par chromatographie en phase gazeuse des gaz rares de l'air, de l'hydrogene, et de quelques composes gazeux du carbone (CO, CO{sub 2}, CH{sub 4}) en utilisant des gaz inactifs an vue d'obtenir les conditions experimenales les plus favorables en vue de cette separation. Dans une deuxieme partie, les conditions optimales de detection de gaz non actifs a l'aide d'une chambre d'ionisation, en utilisant comme source radioactive du {sup 239}Pu qui emet des particules {alpha} de 5,15 MeV, sont determinees. (auteur)

  12. Lipophilic properties of anti-Alzheimer's agents determined by micellar electrokinetic chromatography and reversed-phase thin-layer chromatography. (United States)

    Godyń, Justyna; Hebda, Michalina; Więckowska, Anna; Więckowski, Krzysztof; Malawska, Barbara; Bajda, Marek


    Lipophilicity as one of the most important physicochemical properties of the biologically active compounds is closely related to their pharmacokinetic parameters and therefore, it is taken into account at the design stage of new drugs. Among the novel, fast and reliable methods for determination of the lipophilicity of compounds micellar electrokinetic chromatography (MEKC) is considered to be an appropriate one for bioactive molecules, as it closely mimics the physiological conditions. In this paper MEKC was used for the estimation of log P values for 49 derivatives of phthalimide, tetrahydroisochinoline and indole, designed and synthesized as potential anti-Alzheimer's agents with cholinesterase inhibitory activity. RP-TLC method was applied for determination of another lipophilicity descriptor - RM0 . The results of both experimental methods were compared with each other giving satisfactory correlation (R = 0.784), and with computational methods (Marvin, ChemOffice Software) resulting in weaker correlation (R = 0.466 - 0.687). The lipophilicity-activity relationship was finally established, showing significant influence of lipophilicity on cholinesterase inhibition in some subgroups of phthalimide derivatives. This article is protected by copyright. All rights reserved.

  13. Enhanced methodology for porting ion chromatography retention data. (United States)

    Park, Soo Hyun; Shellie, Robert A; Dicinoski, Greg W; Schuster, Georg; Talebi, Mohammad; Haddad, Paul R; Szucs, Roman; Dolan, John W; Pohl, Christopher A


    Porting is a powerful methodology to recalibrate an existing database of ion chromatography (IC) retention times by reflecting the changes of column behavior resulting from either batch-to-batch variability in the production of the column or the manufacture of new versions of a column. This approach has been employed to update extensive databases of retention data of inorganic and organic anions forming part of the "Virtual Column" software marketed by Thermo Fisher Scientific, which is the only available commercial optimization tool for IC separation. The current porting process is accomplished by performing three isocratic separations with two representative analyte ions in order to derive a porting equation which expresses the relationship between old and new data. Although the accuracy of retention prediction is generally enhanced on new columns, errors were observed on some columns. In this work, the porting methodology was modified in order to address this issue, where the porting equation is now derived by using six representative analyte ions (chloride, bromide, iodide, perchlorate, sulfate, and thiosulfate). Additionally, the updated porting methodology has been applied on three Thermo Fisher Scientific columns (AS20, AS19, and AS11HC). The proposed approach showed that the new porting methodology can provide more accurate and robust retention prediction on a wide range of columns, where average errors in retention times for ten test anions under three eluent conditions were less than 1.5%. Moreover, the retention prediction using this new approach provided an acceptable level of accuracy on a used column exhibiting changes in ion-exchange capacity.

  14. Carnosol purification. Scaling-up centrifugal partition chromatography separations. (United States)

    Bouju, Elodie; Berthod, Alain; Faure, Karine


    This paper illustrates the application of a recently proposed protocol allowing the scale-up prediction on hydrostatic countercurrent chromatography columns (centrifugal partition chromatographs or CPC). A commercial extract of rosemary (Rosmarinus officinalis L.) was used as the starting material containing 0.48% of carnosol, an active pharmaceutical ingredient with great potential. After a rapid method development on a small-scale 35-mL CPC instrument that allowed for the determination of the solvent system and maximum sample concentration and volume, the purification was transferred on two larger instruments using the "free space between peaks" method. The method takes into account the technical limitations of the larger instruments, such as pressure and/or maximum centrifugal field, and allows, by simply running an analytical-sized injection on the large scale rotor, to give an accurate prediction of the maximum sample load and best throughput. The 0.27g of rosemary extract maximum load on a 35-mL CPC was transferred as a 1.9g load on a 254-mL medium size CPC and 9g load on a 812-mL CPC. The maximum process efficiency of 3.1mg of carnosol per hour obtained on the small 35-mL column was transferred on the 254-mL CPC giving 8.3mg/h and, on the larger 812-mL column 49.4mg of carnosol could be obtained per hour. If the scaling-up in CPC instruments is not directly homothetic, it can be highly predictable through a few simple experiments.

  15. Liquid chromatography method to determine polyamines in thermosetting polymers

    Energy Technology Data Exchange (ETDEWEB)

    Dopico-Garcia, M.S. [Laboratorio de Quimica - Centro de Investigacions Tecnoloxicas, Universidade da Coruna, Campus de Esteiro s/n, 15403 Ferrol (Spain); Centro Galego do Plastico, A Cabana s/n, 15590 Ferrol (Spain); Lopez-Vilarino, J.M. [Laboratorio de Quimica - Centro de Investigacions Tecnoloxicas, Universidade da Coruna, Campus de Esteiro s/n, 15403 Ferrol (Spain); Fernandez-Martinez, G. [Unidad de Tecnicas Cromatograficas, Servizos de Apoio a Investigacion, Edificio Servizos Centrais de Investigacion, Universidade da Coruna, Campus de Elvina s/n, 15071 A Coruna (Spain); Gonzalez-Rodriguez, M.V., E-mail: [Dpto. de Quimica Analitica - E.U. Politecnica, Universidade da Coruna, Avda. 19 de Febrero s/n, 15405 Ferrol (Spain)


    A simple, robust and sensitive analytical method to determine three polyamines commonly used as hardeners in epoxy resin systems and in the manufacture of polyurethane is reported. The studied polyamines are: one tetramine, TETA (triethylenetetramine), and two diamines, IPDA (Isophorone diamine) and TCD-diamine (4,7-methano-1H-indene-5,?-dimethanamine, octahydro-). The latter has an incompletely defined structure, and, as far as we know, has not been previously determined by other methods. All three polyamines contain primary amines; TETA also contains secondary amines. The analytical method involves derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate, used for the first time for these compounds, followed by high performance liquid chromatography (HPLC) analysis with a fluorescence (FL) detector ({lambda} excitation 248 nm, {lambda} emision 395 nm). The HPLC-DAD-LTQ Orbitrap MS was used in order to provide structural information about the obtained derivatized compounds. The hybrid linear ion trap LTQ Orbitrap mass spectrometer has been introduced in recent years and provides a high mass accuracy. The structures of the derivatized analytes were identified from the protonated molecular ions [M+H]{sup +} and corresponded to the fully labelled analytes. The following analytical parameters were determined for the method using the HPLC-FL: linearity, precision (2.5-10%), instrumental precision intraday (0.8-1.5%) and interday (2.9-6.3%), and detection limits (0.02-0.14 mg L{sup -1}). The stability of stock solutions and derivatized compounds was also investigated. The method was applied to determine the amine free content in epoxy resin dust collected in workplaces.

  16. Chiral separation of amides using supercritical fluid chromatography. (United States)

    Xiang, Yanqiao; Dunetz, Joshua R; Lovdahl, Michael


    Nine amide derivatives bearing α-stereocenters as well as different substitutions on the amide nitrogen were synthesized via an n-propanephosphonic acid cyclic anhydride (T3P)-mediated coupling, and their enantiomeric pairs were separated using supercritical fluid chromatography (SFC). Five polysaccharide-based chiral stationary phases (CSPs), Chiralcel OD-H, and OJ-H, and Chiralpak AD-H, AS-H and IC columns were explored for the chiral separation of these compounds. None of the compounds could be resolved on all five columns, and no single column could separate all nine pairs of enantiomers. Comparatively, the IC and OD-H columns showed the best results for this group of amides, yielding baseline separations for eight of nine pairs. The type of polar functional group and aromatic substitution in the CSPs and the substitutions on the amide nitrogen had a significant impact on the enantiomeric resolution of the compounds in the interaction between the analyte and the stationary phases. The potential separation mechanism and the effect of substitutions in the CSPs and amide solutes on the separation are discussed. The effects of the organic modifiers, modifier composition, mobile phase additives, and temperature were investigated for the separation of these amides on the IC or the OD-H column. Baseline resolution was achieved under optimized chromatographic conditions using an IC or an OD-H column. Linearity, reproducibility, and limit of quantitation were also demonstrated for the compound 9. Approximately three-fold improvement in signal-to-noise was observed using a SFC system with better instrument design.

  17. Purification of chimeric heavy chain monoclonal antibody EG2-hFc using hydrophobic interaction membrane chromatography: an alternative to protein-A affinity chromatography. (United States)

    Sadavarte, Rahul; Spearman, Maureen; Okun, Natalie; Butler, Michael; Ghosh, Raja


    Heavy chain monoclonal antibodies are being considered as alternative to whole-IgG monoclonal antibodies for certain niche applications. Protein-A chromatography which is widely used for purifying IgG monoclonal antibodies is also used for purifying heavy chain monoclonal antibodies as these molecules possess fully functional Fc regions. However, the acidic conditions used to elute bound antibody may sometimes also leach protein-A, which is immunotoxic. Low pH conditions also tend to make the mAb molecules unstable and prone to aggregation. Moreover, protein-A affinity chromatography does not remove aggregates already present in the feed. Hydrophobic interaction membrane chromatography (or HIMC) has already been studied as an alternative to protein-A chromatography for purifying whole-IgG monoclonal antibodies. This paper describes the use of HIMC for capturing a humanized chimeric heavy chain monoclonal antibody (EG2-hFC). Binding and eluting conditions were suitably optimized using pure EG2-hFC. Based on this, an HIMC method was developed for capture of EG2-hFC directly from cell culture supernatant. The EG2-hFc purity obtained in this single-step process was high. The glycan profiles of protein-A and HIMC purified monoclonal antibody samples were similar, clearly demonstrating that both techniques captured similarly glycosylated population of EG2-hFc. Moreover, this technique was able to resolve aggregates from monomeric form of the EG2-hFc.

  18. Immunoaffinity chromatography for the sample pretreatment of Taxus plant and cell extracts prior to analysis of taxanes by high-performance liquid chromatography

    NARCIS (Netherlands)

    Theodoridis, G; Haasnoot, W; Schilt, R; Jaziri, M; Diallo, B; Papadoyannis, IN; de Jong, GJ; Cazemier, G.


    The application of immunoaffinity chromatography for the purification of Taxus plant and cell extracts prior to the HPLC analysis is described. Polyclonal antibodies raised against 10-deacetylbaccatin III (10-DAB III), paclitaxel's main precursor in plant, were characterised by enzymed-linked immuno

  19. [Rapid determination of pesticide multiresidues in porphyra by dispersive solid-phase extraction coupled with online gel permeation chromatography-gas chromatography-mass spectrometry]. (United States)

    Sun, Xiaojie; Guo, Mengmeng; Wang, Suyue; Tan, Zhijun; Li, Zhaoxin; Zhai, Yuxiu


    A rapid method for the simultaneous identification and quantification of pesticide multiresidues in porphyra was developed using gel permeation chromatography (GPC) coupled to gas chromatography-mass spectrometry (GPC-GC/MS). Nineteen pesticides (organochlorines, organophosphoruses, triazines and pyrethroids) were selected as the target analytes. The pretreatment method was applied consisting of organic solvent extraction followed by dispersive solid-phase extraction with graphitized carbon black (GCB) and primary secondary amine (PSA) adsorbents. GPC was also employed online to remove the large molecules such as pigments and lipids. The quantitative analysis was carried out by external standard method using gas chromatography coupled with mass spectrometry in selective ion monitoring (SIM) mode. Moreover, a large volume of sample was allowed to be injected using the program of GPC programmed-temperature vaporizer of gas chromatography to improve the sensitivity of measurements. The results showed that the calibration curves were linear (r > 0.995) in the range of 10-1,000 μg/L for all the pesticides. The limits of detection (LODs) for the pesticides in porphyra were from 0.005 to 0.03 mg/kg, and the average recoveries were between 70% and 120%. The advantages of the method are simple, sensitive and shorter operation time for analysis of pesticide residues in porphyra samples.

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

    Ishihara, Takashi; Kadoya, Toshihiko; Yamamoto, Shuichi


    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.

  1. Size exclusion chromatography of synthetic polymers and biopolymers on common reversed phase and hydrophilic interaction chromatography columns. (United States)

    Caltabiano, Anna M; Foley, Joe P; Barth, Howard G


    This work describes the applicability of common reversed phase and HILIC columns for size exclusion chromatography of synthetic and natural polymers. Depending on the nature of the solute and column stationary phase, a "non-retention" condition must be created with the aid of the mobile phase to achieve a unique size-based separation in isocratic mode. The various bonded phases show remarkable differences in size separations that are controlled by mobile phase conditions. Polymer-mobile phase and column-mobile phase solvation interactions determine polymer hydrodynamic volume (or solute bulkiness) and polymer-column steric interaction. Solvation interactions in turn depend on polymer, mobile phase and stationary phase polarities. Column-mobile phase solvation interactions determine the structural order of the bonded ligands that can vary from ordered (extended, aligned away from the silica substrate) to disordered (folded, pointing toward the silica substrate). Chain order increases with increased solvent penetration into the bonded phase. Increased chain order reduces pore volume, and therefore decreases the size-separation efficiency of a column. Conversely, decreased chain order increases pore volume and therefore increases the size-separation efficiency. The thermodynamic quality of the mobile phase also plays a significant role in the separation of polymers. "Poor" solvents can significantly reduce the hydrodynamic diameter of a solute and thus change their retention behavior. Medium polarity stationary phases, such as fluoro-phenyl and cyano, exhibit a unique retention behavior. With an appropriate polarity mobile phase, polar and non-polar synthetic polymers of the same molecular masses can be eluted at the same retention volumes.

  2. Separation of Nitration By-Products in Commercial-Grade Trinitro-Toluene by High Performance Liquid Chromatography. (United States)



  3. Direct Injection of Seawater for the Analysis of Nitroaromatic Explosives and their Degradation Products by Micellar Electrokinetic Chromatography (United States)


    micellar electrokinetic chromatography Braden C. Giordanoa,∗, Dean S. Burgib, Greg E. Collinsa a Uanited States Naval Research Laboratory, Chemistry...threats to our coastal regions. Micellar electrokinetic chromatography (MEKC) has been demonstrated to be a useful analytical tool in the anal- ysis of...injection of seawater for the analysis of nitroaromatic explosives and their degradation products by micellar electrokinetic chromatography 5a. CONTRACT

  4. Fabricating electrospun cellulose nanofibre adsorbents for ion-exchange chromatography. (United States)

    Dods, Stewart R; Hardick, Oliver; Stevens, Bob; Bracewell, Daniel G


    Protein separation is an integral step in biopharmaceutical manufacture with diffusion-limited packed bed chromatography remaining the default choice for industry. Rapid bind-elute separation using convective mass transfer media offers advantages in productivity by operating at high flowrates. Electrospun nanofibre adsorbents are a non-woven fibre matrix of high surface area and porosity previously investigated as a bioseparation medium. The effects of compression and bed layers, and subsequent heat treatment after electrospinning cellulose acetate nanofibres were investigated using diethylaminoethyl (DEAE) or carboxylate (COO) functionalisations. Transbed pressures were measured and compared by compression load, COO adsorbents were 30%, 70% and 90% higher than DEAE for compressions 1, 5 and 10MPa, respectively, which was attributed to the swelling effect of hydrophilic COO groups. Dynamic binding capacities (DBCs) at 10% breakthrough were measured between 2000 and 12,000CV/h (2s and 0.3s residence times) under normal binding conditions, and DBCs increased with reactant concentration from 4 to 12mgBSA/mL for DEAE and from 10 to 21mglysozyme/mL for COO adsorbents. Comparing capacities of compression loads applied after electrospinning showed that the lowest load tested, 1MPa, yielded the highest DBCs for DEAE and COO adsorbents at 20mgBSA/mL and 27mglysozyme/mL, respectively. At 1MPa, DBCs were the highest for the lowest flowrate tested but stabilised for flowrates above 2000CV/h. For compression loads of 5MPa and 10MPa, adsorbents recorded lower DBCs than 1MPa as a result of nanofibre packing and reduced surface area. Increasing the number of bed layers from 4 to 12 showed decreasing DBCs for both adsorbents. Tensile strengths were recorded to indicate the mechanical robustness of the adsorbent and be related to packing the nanofibre adsorbents in large scale configurations such as pleated cartridges. Compared with an uncompressed adsorbent, compressions of 1, 5

  5. Development of Analyses of Biological Steroids Using Chromatography--Special Reference to Vitamin D Compounds and Neurosteroids--

    Institute of Scientific and Technical Information of China (English)

    Kazutake Shimada; Tatsuya Higashi; Kuniko Mitamura


    Steroids comprise a large group of natural substances that must frequently be monitored in various biological materials. Due to the metabolic versatility of steroid molecules, extremely complex mixtures are often encountered, necessitating the use of a chromatographic procedure prior to measurement. In this article we present our work, that is, the development of analyses of biological steroids (especially vitamin D compounds and neurosteroids) using gas chromatography/mass spectrometry, high-performance liquid chromatography (including inclusion chromatography using cyclodextrin) and liquid chromatography/mass spectrometry.

  6. Monitoring the synthesis of new pitches from coal tar and its fractions by chromatography and related techniques. (United States)

    Bermejo, J; Fernández, A L; Prada, V; Granda, M; Menéndez, R


    The aim of this paper is to study the applications of chromatography and related techniques in the transformation of coal-derived products into pitches for specific uses. Anthracene oil, tar and pitch were thermally treated in the presence of air (and sulfur in the case of anthracene oil) in order to cause the polymerization/condensation of their components. The evolution of the components of the parent materials during the treatments was monitored by gas chromatography-mass spectrometry, gas chromatography-atomic emission detection, probe mass spectrometry and size-exclusion chromatography. From the results obtained, possible mechanisms for the transformation of coal-derived products into new pitches were established.

  7. Isolation and purification of uremic middle molecules by multi-step liquid chromatography

    Institute of Scientific and Technical Information of China (English)

    储结根; 刘晓航; 袁直; 何炳林


    Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and reversed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular compounds in subfraction A-3 came true by our method.

  8. Quality evaluation of moluodan concentrated pill using high-performance liquid chromatography fingerprinting coupled with chemometrics. (United States)

    Tao, Lingyan; Zhang, Qing; Wu, Yongjiang; Liu, Xuesong


    In this study, a fast and effective high-performance liquid chromatography method was developed to obtain a fingerprint chromatogram and quantitative analysis simultaneously of four indexes including gallic acid, chlorogenic acid, albiflorin and paeoniflorin of the traditional Chinese medicine Moluodan Concentrated Pill. The method was performed by using a Waters X-bridge C18 reversed phase column on an Agilent 1200S high-performance liquid chromatography system coupled with diode array detection. The mobile phase of the high-performance liquid chromatography method was composed of 20 mmol/L phosphate solution and acetonitrile with a 1 mL/min eluent velocity, under a detection temperature of 30°C and a UV detection wavelength of 254 nm. After the methodology validation, 16 batches of Moluodan Concentrated Pill were analyzed by this high-performance liquid chromatography method and both qualitative and quantitative evaluation results were achieved by similarity analysis, principal component analysis and hierarchical cluster analysis. The results of these three chemometrics were in good agreement and all indicated that batch 10 and batch 16 showed significant differences with the other 14 batches. This suggested that the developed high-performance liquid chromatography method could be applied in the quality evaluation of Moluodan Concentrated Pill.

  9. Investigating interactions between phospholipase B-Like 2 and antibodies during Protein A chromatography. (United States)

    Tran, Benjamin; Grosskopf, Vanessa; Wang, Xiangdan; Yang, Jihong; Walker, Don; Yu, Christopher; McDonald, Paul


    Purification processes for therapeutic antibodies typically exploit multiple and orthogonal chromatography steps in order to remove impurities, such as host-cell proteins. While the majority of host-cell proteins are cleared through purification processes, individual host-cell proteins such as Phospholipase B-like 2 (PLBL2) are more challenging to remove and can persist into the final purification pool even after multiple chromatography steps. With packed-bed chromatography runs using host-cell protein ELISAs and mass spectrometry analysis, we demonstrated that different therapeutic antibodies interact to varying degrees with host-cell proteins in general, and PLBL2 specifically. We then used a high-throughput Protein A chromatography method to further examine the interaction between our antibodies and PLBL2. Our results showed that the co-elution of PLBL2 during Protein A chromatography is highly dependent on the individual antibody and PLBL2 concentration in the chromatographic load. Process parameters such as antibody resin load density and pre-elution wash conditions also influence the levels of PLBL2 in the Protein A eluate. Furthermore, using surface plasmon resonance, we demonstrated that there is a preference for PLBL2 to interact with IgG4 subclass antibodies compared to IgG1 antibodies.

  10. Pressure-flow relationships for packed beds of compressible chromatography media at laboratory and production scale. (United States)

    Stickel, J J; Fotopoulos, A


    Pressure drop across chromatography beds employing soft or semirigid media can be a significant problem in the operation of large-scale preparative chromatography columns. The shape or aspect ratio (length/diameter) of a packed bed has a significant effect on column pressure drop due to wall effects, which can result in unexpectedly high pressures in manufacturing. Two types of agarose-based media were packed in chromatography columns at various column aspect ratios, during which pressure drop, bed height, and flow rate were carefully monitored. Compression of the packed beds with increasing flow velocities was observed. An empirical model was developed to correlate pressure drop with the aspect ratio of the packed beds and the superficial velocity. Modeling employed the Blake-Kozeny equation in which empirical relationships were used to predict bed porosity as a function of aspect ratio and flow velocity. Model predictions were in good agreement with observed pressure drops of industrial scale chromatography columns. A protocol was developed to predict compression in industrial chromatography applications by a few laboratory experiments. The protocol is shown to be useful in the development of chromatographic methods and sizing of preparative columns.

  11. DEAE-Affi-Gel Blue chromatography of human serum: use for purification of native transferrin. (United States)

    Werner, P A; Galbraith, R M; Arnaud, P


    Human serum was subjected to chromatography on DEAE-Affi-Gel Blue which combines ion-exchange and pseudo-ligand-affinity chromatography in a 0.02 M phosphate buffer, pH 7.0. All serum proteins were bound with the exception of transferrin, IgG (immunoglobulin G) and trace amounts of IgA. After a second step of Sephadex G-100 gel chromatography, or affinity chromatography against goat anti-human IgG F(ab')2 coupled to AH-Sepharose 4B, IgG and IgA were removed. The transferrin obtained was homogeneous and of high yield (greater than 80%), and was unaltered as judged by analyses of molecular weight, isoelectric point, iron-binding capacity, antigenicity, and ability to bind to high-affinity specific cellular receptors. Thus, DEAE-Affi-Gel Blue chromatography may be used as the basis for a simple, rapid, two-step method for the purification of large amounts of native transferrin from serum.

  12. Quantification of Lipophilicity of 1,2,4-Triazoles Using Micellar Chromatography. (United States)

    Janicka, Małgorzata; Stępnik, Katarzyna; Pachuta-Stec, Anna


    High-performance liquid chromatography (HPLC), over-pressured-layer chromatography (OPLC) and thin-layer chromatography (TLC) techniques with micellar mobile phases were proposed to evaluate the lipophilicity of 21 newly synthesized 1,2,4-triazoles, compounds of potential importance in medicine or agriculture as fungicides. Micellar parameters log k(m) were compared with extrapolated R(M0) values determined from reversed-phase (RP) TLC experimental data obtained on RP-8 stationary phases as well as with log P values (Alog Ps, AClog P, Alog P, Mlog P, KowWin, xlog P2 and xlog P3) calculated from molecular structures of solutes tested. The results obtained by applying principal component analysis (PCA) and linear regression showed considerable similarity between partition and retention parameters as alternative lipophilicity descriptors, and indicated micellar chromatography as a suitable technique to study lipophilic properties of organic substances. In micellar HPLC, RP-8e column (Purospher) was applied, whereas in OPLC and TLC, RP-CN plates were applied, which was the novelty of this study and allowed the use of micellar effluents in planar chromatography measurements.

  13. Purification of prenylated proteins by affinity chromatography on cyclodextrin-modified agarose (United States)

    Chung, Jinhwa A.; Wollack, James W.; Hovlid, Marisa L.; Okesli, Ayse; Chen, Yan; Mueller, Joachim D.; Distefano, Mark D.; Taton, T. Andrew


    Although protein prenylation is widely studied, there are few good methods for isolating prenylated proteins from their non-prenylated relatives. We report that crosslinked agarose (e.g., Sepharose) chromatography media that has been chemically functionalized with β-cyclodextrin (β-CD) is extremely effective in affinity chromatography of prenylated proteins. In this study, a variety of proteins with C-terminal prenylation target (“CAAX box”) sequences were enzymatically prenylated in vitro with natural and non-natural prenyl diphosphate substrates. The prenylated protein products could then be isolated from starting materials by gravity chromatography or fast protein liquid chromatography (FPLC) on a β-CD-Sepharose column. One particular prenylation reaction—farnesylation of a mCherry-CAAX fusion construct—was studied in detail. In this case, purified farnesylated product was unambiguously identified by electrospray mass spectrometry. In addition, when mCherry-CAAX was prenylated with a non-natural, functional isoprenoid substrate, the functional group was maintained by chromatography on β-CD-Sepharose, such that the resulting protein could be selectively bound at its C terminus to complementary functionality on a solid substrate. Finally, β-CD-Sepharose FPLC was used to isolate prenylated mCherry-CAAX from crude HeLa cell lysate, as a model for purifying prenylated proteins from cell extracts. We propose that this method could be generally useful to the community of researchers studying protein prenylation. PMID:18834849

  14. Characterization of sodium carboxymethyl cellulose by comprehensive two-dimensional liquid chromatography. (United States)

    Shakun, Maria; Heinze, Thomas; Radke, Wolfgang


    Two series of sodium carboxymethyl celluloses (NaCMC) with average degrees of substitution (DS) ranging from 0.45 to 1.55 were synthesized from low molecular mass Avicel cellulose (Avicel samples) and from high molecular mass cotton linters (BWL samples). The samples were characterized by online two-dimensional liquid chromatography using gradient liquid adsorption chromatography in the first and size exclusion chromatography (SEC) in the second dimension. This method allows the simultaneous determination of the chemical composition (DS) and the molar mass distribution within the individual samples. Moreover information was obtained on the dependence of the elution volume in gradient chromatography on molar mass. As expected, evidence for a stronger influence of molar mass on gradient elution volume was found for the low molecular mass NaCMC as compared to the high molecular mass BWL samples. Finally the applicability of the method for the simultaneous separation of blends heterogeneous with respect to chemical composition (DS) and molar mass was demonstrated. Such blends cannot be efficiently separated by either SEC or gradient chromatography alone, nor by simply combining the results of both methods. Only the complete two-dimensional chromatogram can reveal the complexity of such blends, since it reveals the correlations between molar mass and chemical composition.

  15. 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. (United States)

    Wang, Xiaohong; Liang, Yong; Zhu, Licai; Xie, Huichun; Li, Hang; He, Junting; Pan, Man; Zhang, Tianyou; Ito, Yoichiro


    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-MS(n). 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.

  16. Detailed analysis of petroleum cuts by multidimensional gas chromatography; Analyse detaillee de coupes petrolieres par chromatographie en phase gazeuse multidimensionnelle

    Energy Technology Data Exchange (ETDEWEB)

    Vendeuvre, C.


    The limitations of petroleum resources implying a better valorisation of crude oil through the optimisation of production, refinery and petrochemistry processes, as well as the environmental regulations have strengthened the necessity of more detailed characterisation of petroleum products. In order to take up this challenge, efficient analytical tools have to be developed. This work demonstrates that comprehensive two-dimensional gas chromatography (GCxGC) constitutes a major advance compared to GC owing to its improved resolution power and to the structured chromatograms indicating the polarity and the volatility of hydrocarbons. The principle of GCxGC is based on the analysis of a whole sample in two independent dimensions of separation achieved using two GC columns of different selectivities; between the two columns a modulator device samples, focuses and re-injects small portions of the effluent from the first column into the second one. Since its introduction in 1991, GCxGC has known a rapid growth and has received a wide acceptance by the analytical science community. The competitive situation has considerably evolved during this thesis with the introduction of commercial systems and the two first sessions of an international symposium dedicated to this technique (Volendam, 2003 and Atlanta, 2004). The key points of the thesis concern the development of a GCxGC prototype system using dual jets CO{sub 2} technology and a data processing program; the evaluation of a retention model allowing a rational choice of operating conditions; and the application of this technique to various and complex issues. Thus, effluents from petrochemistry, refinery or pollution areas have been analysed according to the chemical classes of hydrocarbons and to their number of carbon atoms; a new method for obtaining distillation curves for each chemical group was also presented. Furthermore, the hyphenation of GCxGC with a specific sulphur detector revealed a great interest for

  17. Systematic comparison of sensitivity between hydrophilic interaction liquid chromatography and reversed phase liquid chromatography coupled with mass spectrometry. (United States)

    Periat, Aurélie; Boccard, Julien; Veuthey, Jean-Luc; Rudaz, Serge; Guillarme, Davy


    Hydrophilic interaction liquid chromatography (HILIC) appears as a promising strategy to increase sensitivity with electrospray ionization source (ESI/MS). In the present study, peak heights, background noises and signal-to-noise ratios (S/N) obtained with HILIC-MS/MS and RPLC-MS/MS conditions were systematically compared using a dataset of 56 basic drugs possessing diverse physico-chemical properties. Various mobile phase conditions were investigated, including different pH (3 and 6 in HILIC; 3, 6 and 9 in RPLC) and flow rates (300, 600 and 1000μL/min). The average gain in sensitivity obtained between HILIC and RPLC was equal to 7 and 10 at pH 3 and 6, respectively. However, this value was not reliable, since it was altered by a few compounds possessing an "extreme" behaviour (gain in sensitivity from 100-fold to >8000-fold better). Then, the median gain in sensitivity, equal to 4 in our case, whatever the pH, should be considered. For about 90% of the tested compounds and analytical conditions, the best S/N was systematically attained under HILIC mode. Thanks to PCA representation, it was shown that the basic compounds with pKa between 6 and 8 generally had the best sensitivity in HILIC at pH 6, while the best sensitivity for basic analytes possessing pKa higher than 8 was usually obtained in HILIC at pH 3. As previously reported, the sensitivity gain in HILIC vs. RPLC was explained by the difference in acetonitrile concentration at elution (in average 29% ACN in RPLC and 82% ACN in HILIC at pH 6) leading to better analytes' desolvation. However, it seems that this high proportion of solvent also favourably influenced the ionization by modifying pH and pKa. Indeed the weakest bases of our training set of compounds (pKa between 2 and 5) showed an unexpectedly strong gain in sensitivity, between 20 and 100-fold in comparison to RPLC. These results prove that the ionic character of analytes in solution (i.e., pKa and pH) and the ionization mechanism (i.e., proton

  18. Gold nanoparticle decorated graphene oxide/silica composite stationary phase for high-performance liquid chromatography. (United States)

    Liang, Xiaojing; Wang, Xusheng; Ren, Haixia; Jiang, Shengxiang; Wang, Licheng; Liu, Shujuan


    In the initial phase of this study, graphene oxide (GO)/silica was fabricated by assembling GO onto the silica particles, and then gold nanoparticles (GNPs) were used to modify the GO/silica to prepare a novel stationary phase for high-performance liquid chromatography. The new stationary phase could be used in both reversed-phase chromatography and hydrophilic interaction liquid chromatography modes. Good separations of alkylbenzenes, isomerides, amino acids, nucleosides, and nucleobases were achieved in both modes. Compared with the GO/silica phase and GNPs/silica phase, it is found that except for hydrophilicity, large π-electron systems, hydrophobicity, and coordination functions, this new stationary phase also exhibited special separation performance due to the combination of 2D GO with zero-dimensional GNPs.

  19. Separation of Quadruplex Polymorphism in DNA Sequences by Reversed-Phase Chromatography (United States)

    Miller, M. Clarke; Ohrenberg, Carl J.; Kuttan, Ashani; Trent, John O.


    This unit describes a method for the separation of a mixture of quadruplex conformations formed from the same parent sequence via reversed-phase chromatography (RPC). Polymorphism is inherent to quadruplex formation and even relatively simple quadruplex-forming sequences can fold into a cornucopia of possible conformations and topologies. Isolation of a specific conformation for study can be problematic. This is especially true for conformations of the human telomere sequence d(GGG(TTAGGG)3), High Performance Liquid Chromatography (HPLC), especially reversed-phase chromatography, has been a mainstay of nucleic acids research and purification for many decades. We have successfully applied this method to the problem of separating individual quadruplex species in the ensemble from the same parent sequence. PMID:26344226

  20. Quantitative analysis of complex casein hydrolysates based on chromatography and membrane

    Institute of Scientific and Technical Information of China (English)

    Qi Wei; Yu Yanjun; He Zhimin


    The enzymatic hydrolysates of casein are so complex that there is no effective method to do quantitative analysis.The common techniques,such as high performance chromatography and SDS-PAGE,can only carry out qualitative analysis.On the basis of membrane separation and high performance size exclusion chromatography (HPSEC),standard peptides with different molecular mass range were prepared,and the linear relationships between mass concentration of the standard peptides and the ultraviolet absorption of corresponding peak areas were established.Consequently,mass concentration of the different hydrolysates at different reaction times could be accurately calculated.The combination of chromatography and membrane separation is of great importance to the quantitative analysis of the complex hydrolysates,which can also be applied to the other macromolecular systems,such as carbohydrates.

  1. Large scale chiral chromatography for the separation of an enantiomer to accelerate drug development. (United States)

    Hsu, Leo C; Kim, Hyunjung; Yang, Xiqin; Ross, David


    There are several approaches to produce enantiomerically pure drug substances, such as recrystallization, catalytic process (ligand and enzyme), indirect chromatographic resolution, and direct chromatographic resolution. However, the use of preparative chromatography with chiral stationary phases seems to be most effective for early phase projects, where the time and resources on the developments need to be minimized to get the drug candidates into the clinical studies. We showed that by following a well-defined process, chiral chromatography can be easily scaled up from an analytical system to a pilot plant system. We also used the results from a multicolumn continuous chromatography (MCC) study to conclude that MCC can be a cost-effective production method for chiral manufacturing.

  2. Self-interaction chromatography as a tool for optimizing conditions for membrane protein crystallization. (United States)

    Gabrielsen, Mads; Nagy, Lisa A; DeLucas, Lawrence J; Cogdell, Richard J


    The second virial coefficient, or B value, is a measurement of how well a protein interacts with itself in solution. These interactions can lead to protein crystallization or precipitation, depending on their strength, with a narrow range of B values (the 'crystallization slot') being known to promote crystallization. A convenient method of determining the B value is by self-interaction chromatography. This paper describes how the light-harvesting complex 1-reaction centre core complex from Allochromatium vinosum yielded single straight-edged crystals after iterative cycles of self-interaction chromatography and crystallization. This process allowed the rapid screening of small molecules and detergents as crystallization additives. Here, a description is given of how self-interaction chromatography has been utilized to improve the crystallization conditions of a membrane protein.

  3. Adsorption and Step Elution of Urokinase Using, Affinity Chromatography -Comparison of Data with Rate Model Simulation

    Institute of Scientific and Technical Information of China (English)

    MohammadRezaAboudzadehRovais; JiawenZhu; BinWu


    A non-equilibrium chromatographic rate model was employed to simulate the affinity chromatography of urokinase. The chromatography process was developed to a yield of high purity product of urokinase from crude materials. The affinity gel used in the process was prepared by an epichlorohydrin-activation method using epichlorohydrin activated Sepharose 4B as a matrix and p-aminobenzamidine as a ligand. The chromatographic process were numerically simulated and analyzed with the aid of VERSE-LC computer simulator. Considering the basic principles, rate model with the back mixing in column inlet was utilized in simulating and studying the effect of the column inlet pattern on other parameters. Comparison of the simulation results with the experimental data showed that the rate model can be used to describe the affinity chromatography of urokinase in a fixed bed column with satisfactory accuracy.

  4. Impedance Analysis of Colloidal Gold Nanoparticles in Chromatography Paper for Quantitation of an Immunochromatographic Assay. (United States)

    Hori, Fumitaka; Harada, Yuji; Kuretake, Tatsumi; Uno, Shigeyasu


    A detection method of gold nanoparticles in chromatography paper has been developed for a simple, cost-effective and reliable quantitation of immunochromatographic strip test. The time courses of the solution resistance in chromatography paper with the gold nanoparticles solution are electrochemically measured by chrono-impedimetry. The dependence of the solution resistance on the concentration of gold nanoparticles has been successfully observed. The main factor to increase the solution resistance may be obstruction of the ion transport due to the presence of gold nanoparticles. The existence of gold nanoparticles with 1.92 × 10(9) particles/mL in an indistinctly-colored chromatography paper is also identified by a solution resistance measurement. This indicates that the solution resistance assay has the potential to lower the detection limit of the conventional qualitative assay.

  5. Ion-exchange chromatography separation applied to mineral recycle in closed systems (United States)

    Ballou, E.; Spitze, L. A.; Wong, F. W.; Wydeven, T.; Johnson, C. C.


    As part of the controlled ecological life support system (CELSS) program, a study is being made of mineral separation on ion-exchange columns. The purpose of the mineral separation step is to allow minerals to be recycled from the oxidized waste products of plants, man, and animals for hydroponic food production. In the CELSS application, relatively large quantities of minerals in a broad concentration range must be recovered by the desired system, rather than the trace quantities and very low concentrations treated in analytical applications of ion-exchange chromatography. Experiments have been carried out to assess the parameters pertinent to the scale-up of ion-exchange chromatography and to determine feasibility. Preliminary conclusions are that the column scale-up is in a reasonable size range for the CELSS application. The recycling of a suitable eluent, however, remains a major challenge to the suitability of using ion exchange chromatography in closed systems.

  6. Analysis of insecticidal proteins from Bacillus thuringiensis and recombinant Escherichia coli by capillary electrokinetic chromatography. (United States)

    Luong, John H T; Male, Keith B; Mazza, Alberto; Masson, Luke; Brousseau, Roland


    Bacillus thuringiensis and recombinant Escherichia coli proteinaceous protoxins were subject to proteolysis and analyzed by capillary electrokinetic chromatography. Three resulting toxins (65 kDa) were baseline-resolved within 22 min using a 10 mM borate, pH 11 separation buffer consisting of 25 mM sodium dodecyl sulfate (SDS) and 30 mM phytic acid. The toxins displayed differential interactions with the SDS and phytic acid phases to effect their separation. The ion-pairing interaction between the analyte and phytic acid was also useful in preventing adsorption to the capillary walls and thus enhanced separation resolution and efficiency. The use of electrokinetic chromatography allows achievement of the separation in a significantly shorter time than conventional high-performance liquid chromatography (HPLC) using a diethylaminoethyl (DEAE) weak-anion exchanger.

  7. Demonstrating Chemical and Analytical Concepts in the Undergraduate Laboratory Using Capillary Electrophoresis and Micellar Electrokinetic Chromatography (United States)

    Palmer, Christopher P.


    This paper describes instrumental analysis laboratory exercises that utilize capillary electrophoresis and micellar electrokinetic chromatography to demonstrate several analytical and chemical principles. Alkyl parabens (4-hydroxy alkyl benzoates), which are common ingredients in cosmetic formulations, are separated by capillary electrophoresis. The electrophoretic mobilities of the parabens can be explained on the basis of their relative size. 3-Hydroxy ethylbenzoate is also separated to demonstrate the effect of substituent position on the acid dissociation constant and the effect this has on electrophoretic mobility. Homologous series of alkyl benzoates and alkyl phthalates (common plasticizers) are separated by micellar electrokinetic chromatography at four surfactant concentrations. This exercise demonstrates the separation mechanism of micellar electrokinetic chromatography, the concept of chromatographic phase ratio, and the concepts of micelle formation. A photodiode array detector is used in both exercises to demonstrate the advantages and limitations of the detector and to demonstrate the effect of pH and substituent position on the spectra of the analytes.

  8. Determination of Nicotine in Smoke Condensate by Ion Chromatography Coupled to Ultraviolet Detection

    Directory of Open Access Journals (Sweden)

    Geiss O


    Full Text Available A new method for the determination of the nicotine content of tobacco smoke condensate is described. The smoke condensate of the mainstream smoke was collected in a glass fibre filter trap and dissolved in isopropanol. The nicotine content of an aliquot of this solution was determined by ion chromatography (IC coupled with ultraviolet (UV detection against external calibration and the nicotine content of the whole smoke condensate was calculated. The nicotine content determined in each run by IC was compared with the results obtained by gas chromatography (GC according to the International Standard (ISO method 10315. The present method represents a potential alternative to the GC method. A sensitive and rapid method for the determination of nicotine in smoke condensate using IC with a standard ion chromatography column was developed.

  9. Rotation planar extraction and rotation planar chromatography of oak (Quercus robur L.) bark. (United States)

    Vovk, Irena; Simonovska, Breda; Andrensek, Samo; Vuorela, Heikki; Vuorela, Pia


    The versatile novel instrument for rotation planar extraction and rotation planar chromatography was exploited for the investigation of oak bark (Quercus robur L.). The same instrument enabled extraction of the bark, analytical proof of (+)-catechin directly in the crude extract and also its fractionation. Additionally, epimeric flavan-3-ols, (+)-catechin and (-)-epicatechin were separated by analytical ultra-micro rotation planar chromatography on cellulose plates with pure water as developing solvent. A comparison of the extraction of oak bark with 80% aqueous methanol by rotation planar extraction and medium pressure solid-liquid extraction was carried out and both techniques were shown to be suitable for the efficient extraction of oak bark. The raw extracts and fractions on thin-layer chromatography showed many compounds that possessed antioxidant activity after spraying with 1,1-diphenyl-2-picrylhydrazyl. Rotation planar fractionation of 840 mg of crude oak bark extract on silica gel gave 6.7 mg of pure (+)-catechin in one run.

  10. Analysis of acrylamide in cooked foods by liquid chromatography tandem mass spectrometry. (United States)

    Rosén, Johan; Hellenäs, Karl-Erik


    A method using liquid chromatography tandem mass spectrometry (LC-MS-MS) with electrospray for the analysis of acrylamide in foods is reported. The method comprises the addition of deuterium-labelled acrylamide-d3, extraction with water, mixed mode solid phase extraction, ultrafiltration and a graphitised carbon column for chromatography. The transitions m/z 72 > 55, 72 > 54, 72 > 44, 72 > 27, 72 > 72 and 75 > 58 were recorded in multiple reaction monitoring mode for identification and quantification. In-house validation data for products from potatoes and cereals (30 to 10,000 microg kg(-1)) are presented (accuracy 91 to 102%, relative standard deviation 3 to 21%). Interlaboratory validation data (comparison with gas chromatography mass spectrometry, 25 to 2000 microg kg(-1)) showed excellent results (r2 = 0.998).

  11. Strong Cation Exchange Chromatography in Analysis of Posttranslational Modifications: Innovations and Perspectives

    Directory of Open Access Journals (Sweden)

    Mariola J. Edelmann


    Full Text Available Strong cation exchange (SCX chromatography has been utilized as an excellent separation technique that can be combined with reversed-phase (RP chromatography, which is frequently used in peptide mass spectrometry. Although SCX is valuable as the second component of such two-dimensional separation methods, its application goes far beyond efficient fractionation of complex peptide mixtures. Here I describe how SCX facilitates mapping of the protein posttranslational modifications (PTMs, specifically phosphorylation and N-terminal acetylation. The SCX chromatography has been mainly used for enrichment of these two PTMs, but it might also be beneficial for high-throughput analysis of other modifications that alter the net charge of a peptide.

  12. Novel liquid chromatography tandem mass spectrometry (LC-MS/MS) methods for measuring steroids. (United States)

    Keevil, Brian G


    Liquid chromatography tandem mass spectrometry (LC-MS/MS) is increasingly becoming the method of choice for steroid hormone measurements due to small sample volumes, fast analysis times and improved specificity compared to immunoassays. Achievement of demanding analytical targets for steroid analysis is now becoming possible because of improvements in sample preparation technology, liquid chromatography column technology and mass spectrometer design. The most popular sample treatment strategies comprise protein precipitation (PP), solid-phase extraction (SLE) and liquid-liquid extraction (LLE). Modern liquid chromatography columns can ensure the adequate separation of isobaric compounds e.g. 21 Deoxycortisol, 11 Deoxycortisol and Corticosterone. The most appropriate method may be chosen to improve assay sensitivity by reducing matrix effects (LLE, SPE) or simplicity and speed (PP). Specific examples of some clinically important steroids including oestradiol, aldosterone, renin, serum cortisol, salivary cortisol and salivary testosterone will be described.

  13. Modeling of closed-loop recycling liquid-liquid chromatography: Analytical solutions and model analysis. (United States)

    Kostanyan, Artak E


    In closed-loop recycling (CLR) chromatography, the effluent from the outlet of a column is directly returned into the column through the sample feed line and continuously recycled until the required separation is reached. To select optimal operating conditions for the separation of a given feed mixture, an appropriate mathematical description of the process is required. This work is concerned with the analysis of models for the CLR separations. Due to the effect of counteracting mechanisms on separation of solutes, analytical solutions of the models could be helpful to understand and optimize chromatographic processes. The objective of this work was to develop analytical expressions to describe the CLR counter-current (liquid-liquid) chromatography (CCC). The equilibrium dispersion and cell models were used to describe the transport and separation of solutes inside a CLR CCC column. The Laplace transformation is applied to solve the model equations. Several possible CLR chromatography methods for the binary and complex mixture separations are simulated.

  14. Plasma lipid analysis by hydrophilic interaction liquid chromatography coupled with electrospray ionization tandem mass spectrometry. (United States)

    Sonomura, Kazuhiro; Kudoh, Shinobu; Sato, Taka-Aki; Matsuda, Fumihiko


    A novel method for the analysis of endogenous lipids and related compounds was developed employing hydrophilic interaction liquid chromatography with electrospray ionization tandem mass spectrometry. A hydrophilic interaction liquid chromatography with carbamoyl stationary phase achieved clear separation of phosphatidylcholine, lysophosphatidylcholine, sphingomyelin, ceramide, and mono-hexsosyl ceramide groups with good peak area repeatability (RSD% 0.99). The established method was applied to human plasma assays and a total of 117 endogenous lipids were successfully detected and reproducibly identified. In addition, we investigated the simultaneous detection of small polar metabolites such as amino and organic acids co-existing in the same biological samples processed in a single analytical run with lipids. Our results show that hydrophilic interaction liquid chromatography is a useful tool for human plasma lipidome analysis and offers more comprehensive metabolome coverage.

  15. Purification of proteins specifically binding human endogenous retrovirus K long terminal repeat by affinity elution chromatography. (United States)

    Trubetskoy, D O; Zavalova, L L; Akopov, S B; Nikolaev, L G


    A novel affinity elution procedure for purification of DNA-binding proteins was developed and employed to purify to near homogeneity the proteins recognizing a 21 base pair sequence within the long terminal repeat of human endogenous retroviruses K. The approach involves loading the initial protein mixture on a heparin-agarose column and elution of protein(s) of interest with a solution of double-stranded oligonucleotide containing binding sites of the protein(s). The affinity elution has several advantages over conventional DNA-affinity chromatography: (i) it is easier and faster, permitting to isolate proteins in a 1 day-one stage procedure; (ii) yield of a target protein is severalfold higher than that in DNA-affinity chromatography; (iii) it is not necessary to prepare a special affinity support for each factor to be isolated. Theaffinity elution could be a useful alternative to conventional DNA-affinity chromatography.

  16. Quantitation of sulfate and thiosulfate in clinical samples by ion chromatography. (United States)

    Cole, D E; Evrovski, J


    For assay of serum sulfate, quantitation by ion conductimetry after separation by anion-exchange chromatography is the method of choice. In comparison to classical barium precipitation methods, chromatographic methods demonstrate increased precision, specificity and sensitivity, and they may be superior to spectrophotometric methods that rely on organic cation precipitation of sulfate. The increased sensitivity and specificity, as well as the inherent capacity of chromatographic methods for simultaneous determination of other anions, has led to its increasing use in the determination of excreted sulfate in clinical profiles of urinary anion composition. Ion chromatography can also be used to quantitate free sulfate in other clinical samples, including cerebrospinal fluid, sweat, saliva, breast milk and human tissues. Finally, ion chromatography shows promise as a more precise and sensitive method for measurement of total acid-labile sulfoesters and thiosulfate.

  17. Diagonal reverse-phase chromatography applications in peptide-centric proteomics: ahead of catalogue-omics? (United States)

    Gevaert, Kris; Van Damme, Petra; Martens, Lennart; Vandekerckhove, Joël


    Diagonal electrophoresis/chromatography was described 40 years ago and was used to isolate specific sets of peptides from simple peptide mixtures such as protease digests of purified proteins. Recently, we have adapted the core technology of diagonal chromatography so that the technique can be used in so-called gel-free, peptide-centric proteome studies. Here we review the different procedures we have developed over the past few years, sorting of methionyl, cysteinyl, amino terminal, and phosphorylated peptides. We illustrate the power of the technique, termed COFRADIC (combined fractional diagonal chromatography), in the case of a peptide-centric analysis of a sputum sol phase sample of a patient suffering from chronic obstructive pulmonary disease (COPD). We were able to identify an unexpectedly high number of intracellular proteins next to known biomarkers.

  18. Determination of muscimol and ibotenic acid in Amanita mushrooms by high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry. (United States)

    Tsujikawa, Kenji; Kuwayama, Kenji; Miyaguchi, Hajime; Kanamori, Tatsuyuki; Iwata, Yuko; Inoue, Hiroyuki; Yoshida, Takemi; Kishi, Tohru


    A reliable analytical method was developed for the quantification and identification of muscimol (MUS) and ibotenic acid (IBO), the toxic constituents of Amanita muscaria and Amanita pantherina. MUS and IBO were extracted from mushrooms by aqueous methanol and derivatized with dansyl chloride (DNS-Cl). After extraction with ethyl acetate and evaporation of the solvent, the residue was ethylated with 1.25 M hydrogen chloride in ethanol. The resulting derivatives were quantified by high-performance liquid chromatography with UV detection and identified by liquid chromatography electrospray ionization tandem mass spectrometry. Calibration curves were linear in the range of 25-2500 ppm for MUS and 40-2500 ppm for IBO, respectively. This method was successfully applied to identify and quantify MUS and IBO in Amanita mushrooms naturally grown and circulated in the drug market.

  19. The use of gas chromatography/matrix isolation infrared spectroscopy (GC/MIIR) and supercritical fluid chromatography (SFC) for waste characterization

    Energy Technology Data Exchange (ETDEWEB)

    Raphaelian, L.A.; Green, D.W.


    In processes involving coal and other fossil fuels, wastes are generated that are normally biologically or chemically treated before disposal. To assist in determining the best treatment for obtaining material suitable for disposal, a knowledge of the chemical composition of the wastes is required. Typically, these wastes contain a complex mixture of organic compounds. One objective of this project is to enhance the capabilities of analyzing complex mixtures by using gas chromatography/matrix isolation infrared spectroscopy. The second objective is to simplify the mixtures found in solid wastes into less complex mixtures so that a better separation can be made by the gas chromatograph portion of GC/MS and GC/MIIR. The technique used for this is supercritical fluid chromatography.

  20. Ion exclusion chromatography for the purification of L-glutamine; Ion haijo chromatography ni yoru L-glutamine no seiseiho no kenkyu

    Energy Technology Data Exchange (ETDEWEB)

    Ito, H.; Nishi, A.; Naruse, M. [Ajinomoto Co. Inc., Kawasaki (Japan)


    Ion exclusion chromatography for the purification of L-glutamine is studied. L-glutamine is usually produced by fermentation and used in pharmaceuticals. By using a model solution of L-glutamine and L-glutamic acid, the optimum cation exchange resin is examined. As a result of the experiments, it is found that a cation exchange resin which has smaller crosslinkage and smaller diameter is better. Ammonium sulfate, L-glutamic acid and pyrrolidonecarboxylic acid, which are usually contained in fermentation broth as impurities, are effectively separated by this method. Moreover, the experimental data of the chromatography is expressed fairly well by the differential equations which express the mass transfer in the fixed bed. 8 refs., 5 figs., 2 tabs.

  1. Purification of six lignans from the stems of Schisandra chinensis by using high-speed counter-current chromatography combined with preparative high-performance liquid chromatography. (United States)

    Zhu, Lijie; Li, Bin; Liu, Xiuying; Huang, Guohui; Meng, Xianjun


    A method for the preparative purification of lignans from Schisandra chinensis was established using a combination of high-speed counter-current chromatography (HSCCC) and preparative high-performance liquid chromatography (HPLC). The crude extracts obtained from S. chinensis by using 70% ethanol were separated on a macroporous resin column and then eluted with a graded ethanol series. A two-phase solvent system consisting of n-hexane-ethyl acetate-methanol-water (1:1:1:1, v/v) was used for HSCCC, and a mobile phase of acetonitrile-water (50:50, v/v) was used for preparative HPLC. The results obtained using HSCCC were compared with those obtained using preparative HPLC, and their advantages were further integrated to improve the separation efficiency. Six known lignans were identified by electrospray ionisation mass spectrometry and (1)H nuclear magnetic resonance (NMR) and (13)C NMR analyses; the purities of all the compounds were more than 91%.

  2. Separation of three anthraquinone glycosides including two isomers by preparative high-performance liquid chromatography and high-speed countercurrent chromatography from Rheum tanguticum Maxim. ex Balf. (United States)

    Chen, Tao; Li, Hongmei; Zou, Denglang; Liu, Yongling; Chen, Chen; Zhou, Guoying; Li, Yulin


    Anthraquinone glycosides, such as chrysophanol 1-O-β-d-glucoside, chrysophanol 8-O-β-d-glucoside, and physion 8-O-β-d-glucoside, are the accepted important active components of Rheum tanguticum Maxim. ex Balf. due to their pharmacological properties: antifungal, antimicrobial, cytotoxic, and antioxidant activities. However, an effective method for the separation of the above-mentioned anthraquinone glycosides from this herb is not currently available. Especially, greater difficulty existed in the separation of the two isomers chrysophanol 1-O-β-d-glucoside and chrysophanol 8-O-β-d-glucoside. This study demonstrated an efficient strategy based on preparative high-performance liquid chromatography and high-speed countercurrent chromatography for the separation of the above-mentioned anthraquinone glycosides from Rheum tanguticum Maxim.ex Balf.

  3. On-line comprehensive two-dimensional normal-phase liquid chromatography×reversed-phase liquid chromatography for preparative isolation of toad venom. (United States)

    Li, Jia-Fu; Fang, Hua; Yan, Xia; Chang, Fang-Rong; Wu, Zhen; Wu, Yun-Long; Qiu, Ying-Kun


    An on-line comprehensive preparative two-dimensional normal-phase liquid chromatography×reversed-phase liquid chromatography (2D NPLC×RPLC) system was constructed with a newly developed vacuum evaporation assisted adsorption (VEAA) interface, allowing fast removal of NPLC solvent in the vacuum condition and successfully solving the solvent incompatibility problem between NPLC and RPLC. The system achieved on-line solvent exchange within the two dimensions and its performance was illustrated by gram-scale isolation of crude extract from the venom of Bufo bufo gargarizans. Within separation time of ∼20h, 19 compounds were obtained with high purity in a single run. With the VEAA interface, the 2D system exhibited apparent advantages in separation efficiency and automation compared with conventional methods, indicating its promising application in the routine separation process for complicated natural products.

  4. Characterization of aroma-active compounds in three Chinese Moutai liquors by gas chromatography-olfactometry, gas chromatography-mass spectrometry and sensory evaluation. (United States)

    Niu, Yunwei; Chen, Xiaomei; Xiao, Zuobing; Ma, Ning; Zhu, Jiancai


    The aroma-active compounds in three Chinese Moutai liquors, aged 1 year, 15 years and 30 years were investigated in this study. The aroma compounds were analysed by gas chromatography-olfactometry (GC-O) coupled with gas chromatography-mass spectrometry (GC-MS). A total of 79 aroma compounds were identified. Aroma extract dilution analysis (AEDA) was further employed to identify the aroma-active compounds. A total of 35 aroma-active compounds with flavour dilution (FD) values ≧ 64 simultaneously in three Chinese Moutai liquors were quantitated. Among them, ethyl acetate, ethyl lactate and acetic acid appeared with the highest concentrations. They were all >1000 mg/L. Then, the relationships between the aroma-active compounds and seven sensory attributes were studied.

  5. Biosynthesis of Cd-bound phytochelatins by Phaeodactylum tricornutum and their speciation by size-exclusion chromatography and ion-pair chromatography coupled to ICP-MS. (United States)

    Loreti, Valeria; Toncelli, Daniel; Morelli, Elisabetta; Scarano, Gioacchino; Bettmer, Jörg


    Cd-bound phytochelatins (Cd-PCs) have been synthesised by incubation of Phaeodactylum tricornutum cell cultures with Cd and purified by size-exclusion chromatography-UV-Vis. These complexes, which were identified in previous work, have now been used as model substances to develop and optimise ion-pair chromatography (IPC) coupled to inductively coupled plasma-mass spectrometry (ICP-MS) for analysis of Cd-PCs. Subsequent analysis of samples taken from Silene vulgaris plants cultivated under heavy metal stress conditions revealed Cd signals but no Cd-PC signals. By use of isotopically enriched (116)Cd-PCs the sample preparation steps were verified to determine the stability of the analytes. We observed species transformation between Cd-PCs and other unidentified Cd complexes. Consequently, the kinetic and thermodynamic lability of Cd-PCs are decisive factors in their detection.

  6. On-line coupled high performance liquid chromatography-gas chromatography for the analysis of contamination by mineral oil. Part 1: method of analysis. (United States)

    Biedermann, Maurus; Grob, Koni


    For the analysis of mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH), on-line coupled high performance liquid chromatography-gas chromatography-flame ionization detection (HPLC-GC-FID) offers important advantages: it separates MOSH and MOAH in robust manner, enables direct injection of large aliquots of raw extracts (resulting in a low detection limit), avoids contamination of the sample during preparation and is fully automated. This review starts with an overview of the technology, particularly the fundamentals of introducing large volumes of solvent into GC, and their implementation into various transfer techniques. The main part deals with the concepts of MOSH and MOAH analysis, with a thorough discussion of the choices made. It is followed by a description of the method. Finally auxiliary tools are summarized to remove interfering components, enrich the sample in case of a high fat content and obtain additional information about the MOSH and MOAH composition.


    NARCIS (Netherlands)



    Two methods for the separation of cholesterol esters, based on the number of double bonds in their fatty acid moieties, are presented. Silver ion chromatography, usually performed on thin-layer chromatographic plates, was made suitable for high-performance liquid chromatography (HPLC) and solid-phas

  8. High-performance liquid chromatography of oligoguanylates at high pH (United States)

    Stribling, R.; Deamer, D. (Principal Investigator)


    Because of the stable self-structures formed by oligomers of guanosine, standard high-performance liquid chromatography techniques for oligonucleotide fractionation are not applicable. Previously, oligoguanylate separations have been carried out at pH 12 using RPC-5 as the packing material. While RPC-5 provides excellent separations, there are several limitations, including the lack of a commercially available source. This report describes a new anion-exchange high-performance liquid chromatography method using HEMA-IEC BIO Q, which successfully separates different forms of the guanosine monomer as well as longer oligoguanylates. The reproducibility and stability at high pH suggests a versatile role for this material.

  9. Determination of 1-hydroxypyrene in human urine by high-performance liquid chromatography

    DEFF Research Database (Denmark)

    Hansen, Åse Marie; Poulsen, O M; Christensen, J M


    A high-performance liquid chromatography (HPLC)/fluorescence method for quantitative analysis of 1-hydroxypyrene in urine was developed. The method validation analysis showed the method to be in analytical control. No significant systematical errors could be demonstrated. The entire run time...... of chromatography was 10 min using isocratic elution (acetonitrile-water, 70:30), and the retention time for 1-hydroxypyrene was 3.5 min. The short run time in combination with the low limit detection (1.37 nmol/L) makes the method potentially applicable for surveillance of pyrene exposure in work environments...

  10. High-speed countercurrent chromatography for purification of single-walled carbon nanotubes

    Institute of Scientific and Technical Information of China (English)

    Ying Cai; Zhi Hong Yan; Ying Chun Lv; Min Zi; Li Ming Yuan


    A new chromatographic purification of single-walled carbon nanotubes using high-speed countercurrent chromatography is reported.The purification was accomplished on the basis of experiment that dispersed the single-walled carbon nanotubes with sodium dodecyl sulfate,and the result mixture was separated using the two phase system composed of n-butanol/water=1/1 (v/v).The sizes of SWNTs separated were observed by scanning electron microscopy.The results demonstrated that the high-speed countercurrent chromatography possessed a good efficency for purification of single-walled carbon nanotubes.

  11. Preparation of novel chiral stationary phase based on click chemistry for ligand exchange chromatography

    Institute of Scientific and Technical Information of China (English)

    Chun Mei Fu; Hong Yu Shi; Guang Sheng Qian; Zhang Wan Li


    Click chemistry was applied to immobilize L-proline derivative onto azide-modified silica gel to give a novel chiral stationary phase (denoted as click-CSP) for ligand exchange chromatography. The developed protocol combines the benefits of operational simplicity, exceptionally mild conditions and high surface loadings. The enantioselectivity α of some DE-amino acids on the click-CSP were found to be in the range from 1.13 to 3.46. The chromatographic resolutions of some DL-amino acids and the stability study firmly illustrate the potential of click chemistry for preparation chiral stationary phase for ligand exchange chromatography.

  12. A New Glow on the Chromatography of M&M Candies (United States)

    Birdwhistell, Kurt R.; Spence, Thomas G.


    This lab experiment for undergraduate or high school students presents the separation and analysis of dyes in the candy shell of M&M chocolate candies. The dyes in the shells of M&Ms are separated using paper chromatography and are identified by comparison of the Rf values, color, and fluorescence properties with those of standard FD&C ([U.S. Federal] Food, Drug, and Cosmetic Act) dyes. The experiment introduces students to chromatography of mixtures, compound identification by using Rf values, and fluorescence. They learn about fluorescence and how it appears in everyday substances and can be used to help identify compounds.

  13. Cromatografia gasosa bidimensional abrangente (GC × GC Comprehensive two-dimensional gas chromatography (GC × GC

    Directory of Open Access Journals (Sweden)

    Marcio Pozzobon Pedroso


    Full Text Available This paper presents the fundamental principles, instrumentation and selected applications of comprehensive two-dimensional gas chromatography (GC × GC. In this technique, introduced in 1991, two capillary columns are coupled and proper modulating interfaces continuously collect the eluate from the first column, transferring it to the second column. The result is a geometric increment in the chromatographic resolution, ensuring separation of extremely complex mixtures in time periods shorter or comparable to those of analysis using conventional gas chromatography and with better detectabilities and sensitivities.

  14. Application of micellar electrokinetic capillary chromatography for routine analysis of different materials

    Directory of Open Access Journals (Sweden)

    Injac Rade


    Full Text Available Micellar electrokinetic capillary chromatography (MEKC has become a popular mode among the several capillary electro-migration techniques. Most drug analysis can be performed by using MEKC because of its wide applicability. Separation of very complex mixtures, determination of drugs in the biological materials, etc., can be successfully achieved by MEKC. This review surveys typical applications of MEKC analysis. Recent advances in MEKC, especially with solid-phase extraction and large-volume sample stacking, are described. Modes of electrokinetic chromatography including MEKC, a separation theory of MEKC, environmental friendly analysis, and selectivity manipulation in MEKC are also briefly mentioned.

  15. Preparative isolation of polymerase chain reaction products using mixed-mode chromatography. (United States)

    Matos, T; Silva, G; Queiroz, J A; Bülow, L


    The polymerase chain reaction (PCR) has become one of the most useful techniques in molecular biology laboratories around the world. The purification of the target DNA product is often challenging, however, and most users are restricted to employing available commercial kits. The recent developments in mixed-mode chromatography have shown higher selectivity for a variety of nucleic acid-containing samples. Capto Adhere is a mixed-mode chromatography resin that offers a high-selectivity ligand and is here applied for the purification of amplified DNAs from PCR mixtures in a 10-min single step, with yields above 95%, high linearity, and high precision for different concentrations.

  16. Enrichment and characterization of phosphopeptides by immobilized metal affinity chromatography (IMAC) and mass spectrometry

    DEFF Research Database (Denmark)

    Thingholm, Tine E; Jensen, Ole N


    The combination of immobilized metal affinity chromatography (IMAC) and mass spectrometry is a widely used technique for enrichment and sequencing of phosphopeptides. In the IMAC method, negatively charged phosphate groups interact with positively charged metal ions (Fe3+, Ga3+, and Al3+) and thi......The combination of immobilized metal affinity chromatography (IMAC) and mass spectrometry is a widely used technique for enrichment and sequencing of phosphopeptides. In the IMAC method, negatively charged phosphate groups interact with positively charged metal ions (Fe3+, Ga3+, and Al3...

  17. Analysis of biodiesel conversion using thin layer chromatography and nonlinear calibration curves

    DEFF Research Database (Denmark)

    Fedosov, Sergey; Brask, Jesper; Xu, Xuebing


    acquisition of a large body of data. The two well-established methods of gas chromatography (GC) and HPLC are time consuming and expensive when analyzing multiple samples. Additionally, it is not always possible to record all the reactants on one elution profile. We examined applicability of thin layer...... chromatography (TLC) for this purpose, where the detection was based on either flame ionization detector (FID) or a modified staining procedure. The suggested staining method gave no background and appeared well suited for quantitative analysis. The relevant calibrations are presented, and the general principles...

  18. Analysis of glyphosate residues in cereals using liquid chromatography-mass spectrometry (LC-MS/MS)

    DEFF Research Database (Denmark)

    Granby, Kit; Johannesen, S.; Gabrielsen, Martin Vahl


    A fast and specific method for the determination of glyphosate in cereals is described. The method is based on extraction with water by ultrasonication. The samples are cleaned up and separated by high-performance liquid chromatography on a polystyrene-based reverse-phase column (clean......-up) in series with an ion chromatography column (separation) using NaHCO3 as eluent. A micro-membrane suppressor was inserted after the separator column to remove the Na + ions before detection by electrospray ionization mass spectrometry in the negative-ion mode. In MS/MS, mode the following transitions were...

  19. [Determination of dimethyl ether correction factors in gas chromatography with TCD and FID]. (United States)

    Chen, J; Zhang, L; Yang, L; Cai, G


    Dimethyl ether (DME) correction factors in gas chromatography with thermal conductivity detector (TCD) and flame ionization detector (FID) by using H2 as carrier gas were determined in this work. The homemade DME gas was quantitatively absorbed in ice-cold water. With ethanol as standard, the aqueous mixture was injected into a gas chromatograph, equipped with serially-connected TCD and FID. The weight correction factors of DME based on methanol were 0.86 and 0.55 for TCD and FID respectively. The result for TCD was also confirmed by calculation based on the stoichiometrical transformation of methanol into DME in reaction gas chromatography.


    Institute of Scientific and Technical Information of China (English)

    SuZhengquan; FengHuixia; 等


    The tetravalent metal salts of monoalkyl phosphates [M(O3POR)2]are a new kind of stationary phases of Chromatography-homogeneous bonded phases.This paper deals with the application of cerium bis(monomyristylphosphate)as support to reversed phase liquid chromatography.The results show that the best mobil phase is CH3CN:H2O=95:5.The good separation to the mixture containing six aromatic hydrocarbons and the determination of naphthalene in a group samples have been achieved.The regression analysis shows that detect limits,linearities and precision for six aromatic hydrocarbons are good.