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Sample records for interaction chromatography stationary

  1. A novel amide stationary phase for hydrophilic interaction liquid chromatography and ion chromatography.

    PubMed

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

    2013-10-15

    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. PMID:24054569

  2. Imidazoline type stationary phase for hydrophilic interaction chromatography and reversed-phase liquid chromatography.

    PubMed

    Li, Yuanyuan; Feng, Yaoyao; Chen, Tong; Zhang, Haixia

    2011-09-01

    An imidazoline was prepared by solvent-free microwave-assisted organic synthesis and immobilized on porous silica particles by polymerization. The resulting material was composed of both hydrophobic alkyl ester chains and hydrophilic imidazoline rings, which gave it both hydrophilic interaction and reversed-phase characteristics. The titration curve suggests that the new material has buffering capacity and acquires increasing positive charge over the pH range 9-4, and is "zwitterionic" in the upper part of this pH range. Through investigating the effect of column temperature, the water content, pH and ion strength of mobile phase on the retention time of polar compounds in highly organic eluents, it was found that the new material could be used as a hydrophilic interaction liquid chromatography (HILIC) stationary phase which involved a complex retention process consisting of partitioning, surface adsorption and electrostatic interactions. In addition, the retention behavior of aromatic compounds in different mobile phase conditions was also studied, which showed the new material mainly exhibited a partitioning mechanism in the reversed-phase liquid chromatography (RPLC) mode. The separation of six water-soluble vitamins and five aromatic compounds were achieved by using the new material in the HILIC and RPLC modes, respectively. PMID:21543075

  3. Glucaminium ionic liquid-functionalized stationary phase for the separation of nucleosides in hydrophilic interaction chromatography.

    PubMed

    Jiang, Qiong; Zhang, Mingliang; Wang, Xusheng; Guo, Yong; Qiu, Hongdeng; Zhang, Shusheng

    2015-10-01

    A glucaminium-based ionic liquid stationary phase was prepared via facile epoxy-amine reaction and subsequent quaternization. Successful immobilization of glucaminium-based ionic liquid onto silica surface was validated by elemental analysis and infrared spectroscopy. The new stationary phase was evaluated for the separation of nucleosides in hydrophilic interaction liquid chromatography (HILIC). Effects of various factors, such as acetonitrile concentration, salt concentration, pH value, as well as column temperature, on the chromatographic behavior toward nucleosides were studied in detail. The results indicated that this new stationary phase can be used for separation of water-soluble polar substances in HILIC mode. The retention of solutes on the stationary phase was influenced by a mixed-mode retention mechanism with a combination of adsorptive and partitioning interactions. PMID:26231689

  4. Study of surface-bonded dicationic ionic liquids as stationary phases for hydrophilic interaction chromatography.

    PubMed

    Qiao, Lizhen; Li, Hua; Shan, Yuanhong; Wang, Shuangyuan; Shi, Xianzhe; Lu, Xin; Xu, Guowang

    2014-02-21

    In the present study, several geminal dicationic ionic liquids based on 1,4-bis(3-allylimidazolium)butane and 1,8-bis(3-allylimidazolium)octane in combination with different anions bromide and bis(trifluoromethanesulphonyl)imide were prepared and then bonded to the surface of 3-mercaptopropyl modified silica materials through the "thiol-ene" click chemistry as stationary phases for hydrophilic interaction chromatography (HILIC). Compared with their monocationic analogues, the dicationic ionic liquids stationary phases presented effective retention and good selectivity for typical hydrophilic compounds under HILIC mode with the column efficiency as high as 130,000 plates/m. Moreover, the influence of different alkyl chain spacer between dications and combined anions on the retention behavior and selectivity of the dicationic ionic liquids stationary phases under HILIC mode was displayed. The results indicated that the longer linkage chain would decrease the hydrophilicity and retention on the dicationic ionic liquid stationary phase, and while differently combined anions had no difference due to the exchangeability under the common HILIC mobile phase with buffer salt. Finally, the retention mechanism was investigated by evaluating the effect of chromatographic factors on retention, including the water content in the mobile phase, the mobile phase pH and buffer salt concentration. The results showed that the dicationic ionic liquids stationary phases presented a mixed-mode retention behavior with HILIC mechanism and anion exchange. PMID:24484692

  5. Glutathione-based zwitterionic stationary phase for hydrophilic interaction/cation-exchange mixed-mode chromatography.

    PubMed

    Shen, Aijin; Li, Xiuling; Dong, Xuefang; Wei, Jie; Guo, Zhimou; Liang, Xinmiao

    2013-11-01

    As a naturally hydrophilic peptide, glutathione was facilely immobilized onto silica surface to obtain a novel hydrophilic interaction/cation-exchange mixed-mode chromatographic stationary phase (Click TE-GSH) via copper-free "thiol-ene" click chemistry. The resulting material was characterized by solid state (13)C/CP MAS NMR and elemental analysis. The measurement of ζ-potential indicated the cation-exchange characteristics and adjustable surface charge density of Click TE-GSH material. The influence of acetonitrile content and pH value on the retention of ionic compounds was investigated for understanding the chromatographic behaviors. The results demonstrated that Click TE-GSH column could provide both hydrophilic and cation-exchange interaction. Taking advantage of the good hydrophilicity and inherent cation-exchange characteristics of Click TE-GSH material, the resolution of neutral fructosan with high degree of polymerization (DP), basic chitooligosaccharides and strongly acidic carrageenan oligosaccharides was successfully realized in hydrophilic interaction chromatography (HILIC), hydrophilic interaction/cation-exchange mixed-mode chromatography (HILIC/CEX), cation-exchange chromatography (CEX) and electrostatic repulsion/hydrophilic interaction chromatography (ERLIC). On the other hand, the separation of standard peptides varying in hydrophobicity/hydrophilicity and charge was achieved in both CEX and HILIC/CEX mode with high efficiency and distinct selectivity. To further demonstrate the versatility and applicability of Click TE-GSH stationary phase, the separation of a human serum albumin (HSA) tryptic digest was performed in HILIC/CEX mode. Peptides were adequately resolved and up to 86 HSA peptides were identified with sequence coverage of 85%. The results indicated the good potential of Click TE-GSH material in glycomics and proteomics. PMID:24075460

  6. A hyperbranched polyethylenimine functionalized stationary phase for hydrophilic interaction liquid chromatography.

    PubMed

    Peng, Yahui; Hou, Yanjie; Zhang, Feifang; Shen, Guobin; Yang, Bingcheng

    2016-05-01

    A hyperbranched stationary phase for hydrophilic interaction liquid chromatography (HILIC) has been prepared by grafting polyethylenimine (PEI) onto silica gel (termed as PEI-Sil). Rich primary, secondary, and tertiary amino groups associated with PEI render its good hydrophility. More importantly, the hyperbranched structure of PEI molecule is greatly helpful in improving interaction with polar analytes. For several kinds of model polar compounds, including organic acids, nucleosides, nucleic acid bases, amino acids, cephalosporins, and non-reducing sugars, PEI-Sil demonstrated excellent separation performance in terms of running stability, reproducibility, and separation efficiency (e.g., plate count ~74,000/m). In addition, PEI-Sil also exhibited much better separation selectivity toward inorganic anions when operated in the mode of ion chromatography relative to a commercial amino propyl-bonded column. PMID:26970747

  7. Underivatized amylose and cellulose as new stationary phases for hydrophilic interaction chromatography.

    PubMed

    Lehnert, Petr; Douša, Michal; Lemr, Karel

    2013-10-01

    Two polysaccharide stationary phases have been newly suggested for application in hydrophilic interaction chromatography (HILIC). Both columns (amylose-silica, 250 × 4.6 mm, 5 μm and cellulose-silica, 250 × 4.6 mm, 5 μm) demonstrated a satisfactory retention of polar compounds. The influence of the mobile-phase composition (acetonitrile content, pH, salt concentration) on the retention was in agreement with the HILIC concept. The phases showed a very similar behavior, typical efficiency of about 50,000 plates/m, cellulose retained test compounds somewhat more strongly. Under the experimental conditions, electrostatic (non-HILIC-type) interactions due to the dissociation of silanol groups on the silica surface did not influence the retention, noticeably. The applicability of polysaccharide stationary phases for the chromatography of polar compounds was proven by the separation of mixtures of sugars (fructose, glucose, saccharose, maltose, trehalose) or vitamins (nicotinamide, pyridoxine, riboflavin, thiamine, nicotinic acid, ascorbic acid). PMID:23983151

  8. Preparation of a silica-based high-performance hydrophobic interaction chromatography stationary phase for protein separation and renaturation.

    PubMed

    Yang, Yicong; Qu, Qian; Li, Weimin; Yuan, Jie; Ren, Yi; Wang, Lili

    2016-07-01

    In this work, based on the structural characteristics of bio-membrane molecules, a novel type of high-performance hydrophobic interaction chromatography stationary phase was prepared using cholesterol as a ligand. Investigating the separation performance of this stationary phase, the effect of pH and salt concentration of the mobile phase on the retention time, the absorption capacity, and the hydrophobic ability revealed that this stationary phase had a high loading capacity and moderate hydrophobic interactions compared with four different hydrophobic interaction chromatography stationary phase ligands. Five types of standard proteins could be baseline separated with a great selection for protein separation. When 3.0 M urea was added to the mobile phase, it could be refolded with simultaneous purification of denatured lysozyme by one-step chromatography. The mass recovery of lysozyme reached 89.5%, and the active recovery was 96.8%. Compared with traditional hydrophobic interaction chromatography, this new stationary phase has a good hydrophobic ability and a significant refolding efficiency. PMID:27159821

  9. Poly(vinyl alcohol) Modified Porous Graphitic Carbon Stationary Phase for Hydrophilic Interaction Liquid Chromatography.

    PubMed

    Hou, Yanjie; Zhang, Feifang; Liang, Xinmiao; Yang, Bingcheng; Liu, Xiaodong; Dasgupta, Purnendu K

    2016-05-01

    We report a poly(vinyl alcohol) (PVA)-coated porous graphitic carbon (PGC, Hypercarb) packing as a novel stationary phase for hydrophilic interaction liquid chromatography (HILIC). The exterior and the pores of the PGC particles are coated with a thin layer of PVA by soaking the particles in a PVA solution, filtering, and thermally cross-linking the PVA. Such PVA coated PGC particles (5.7 μm diameter), hereinafter called PVA-PGC are stable at least through pH 1.0-12.7, can be made in <2 h, and exhibit different selectivity relative to six commercial HILIC phases and bare PGC. To our knowledge, this is the first fully pH-stable, completely neutral HILIC phase. Excellent efficiency stable is observed for polar analytes (∼70 000 and 118 000 plates/m for cytosine and resorcinol, respectively). Retention closely resembles standard HILIC behavior. Other substances can also be easily incorporated in the PVA layer; an anion exchange column can be readily made by incorporating diallyldimethylammonium chloride in the PVA coating solution. The ease of preparation without the requirement of synthetic skills or paraphernalia and the possibility of incorporating a variety of modifiers makes this a particularly versatile approach. PMID:27053418

  10. Chromatographic evaluation of a newly designed peptide-silica stationary phase in reverse phase liquid chromatography and hydrophilic interaction liquid chromatography: mixed mode behavior.

    PubMed

    Ray, Sudipta; Takafuji, Makoto; Ihara, Hirotaka

    2012-11-30

    The short peptide Boc-Phe-Aib-Phe-OH was synthesized and immobilized onto porous silica using grafting methodology. The resulting peptide-bonded silica was characterized using DRIFT-mode FT-IR, elemental analysis, thermogravimetric analysis, solid state C(13) NMR spectroscopy and the successful immobilization of the peptide on the silica support was confirmed. This grafted phase was packed into a stainless steel column and used for mixed-mode chromatography such as reversed-phase high-performance liquid chromatography and hydrophilic interaction liquid chromatography for the efficient separation of hydrophobic compounds, small polar molecules, and drug molecules. Compared with ODS and phenyl columns, this new stationary phase shows considerably higher molecular-planarity selectivity towards polyaromatic hydrocarbons and also available for separation of nucleo-analytes and sulfa-drug molecules in a hydrophilic interaction liquid chromatography mode. The multiple interactions induced by polar carbonyl group and hydrophobic phenyl group allow this peptide-modified silica to serve as a multi-mode stationary phase in high performance liquid chromatography. PMID:23116801

  11. Poly(L-lactic acid)-modified silica stationary phase for reversed-phase and hydrophilic interaction liquid chromatography.

    PubMed

    Ohyama, Kaname; Takasago, Shizuka; Kishikawa, Naoya; Kuroda, Naotaka

    2015-03-01

    Poly(L-lactic acid) is a linear aliphatic thermoplastic polyester that can be produced from renewable resources. A poly(L-lactic acid)-modified silica stationary phase was newly prepared by amide bond reaction between amino groups on aminopropyl silica and carboxylic acid groups at the end of the poly(L-lactic acid) chain. The poly(L-lactic acid)-silica column was characterized in reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography with the use of different mobile phase compositions. The poly(L-lactic acid)-silica column was found to work in both modes, and the retention of test compounds depending on acetonitrile content exhibited "U-shaped" curves, which was an indicator of reversed-phase liquid chromatography/hydrophilic interaction liquid chromatography mixed-mode retention behavior. In addition, carbonyl groups included into the poly(L-lactic acid) backbone work as an electron-accepting group toward a polycyclic aromatic hydrocarbon and provide π-π interactions. PMID:25546473

  12. Preparation, characterization and application of a reversed phase liquid chromatography/hydrophilic interaction chromatography mixed-mode C18-DTT stationary phase.

    PubMed

    Wang, Qing; Long, Yao; Yao, Lin; Xu, Li; Shi, Zhi-Guo; Xu, Lanying

    2016-01-01

    A mixed-mode chromatographic stationary phase, C18-DTT (dithiothreitol) silica (SiO2) was prepared through "thiol-ene" click chemistry. The obtained material was characterized by fourier transform infrared spectroscope, nitrogen adsorption analysis and contact angle analysis. Chromatographic performance of the C18-DTT was systemically evaluated by studying the effect of acetonitrile content, pH, buffer concentration of the mobile phase and column temperature. It was demonstrated that the novel stationary phase possessed reversed phase liquid chromatography (RPLC)/hydrophilic interaction liquid chromatography (HILIC) mixed-mode property. The stop-flow test revealed that C18-DTT exhibited excellent compatibility with 100% aqueous mobile phase. Additionally, the stability and column-to-column reproducibility of the C18-DTT material were satisfactory, with relative standard deviations of retention factor of the tested analytes (verapamil, fenbufen, guanine, tetrandrine and nicotinic acid) in the range of 1.82-3.72% and 0.85-1.93%, respectively. Finally, the application of C18-DTT column was demonstrated in the separation of non-steroidal anti-inflammatory drugs, aromatic carboxylic acids, alkaloids, nucleo-analytes and polycyclic aromatic hydrocarbons. It had great resolving power in the analysis of various compounds in HILIC and RPLC chromatographic conditions and was a promising RPLC/HILIC mixed-mode stationary phase. PMID:26695288

  13. Deconvoluting the effects of buffer salt concentration in hydrophilic interaction chromatography on a zwitterionic stationary phase.

    PubMed

    West, Caroline; Auroux, Emeline

    2016-08-26

    Quantitative structure-retention relationships (QSRRs) furnish a detailed and reliable description of the role and extent of different molecular interactions that can be established between the analytes and the chromatographic system. Among QSRRs, the solvation parameter model using Abraham descriptors has gained acceptance as a general tool to explore the factors affecting retention in chromatographic systems. We have previously shown how a modified version of the solvation parameter model, with two extra terms to take account of interactions occurring with ionic and ionizable species (with positive and/or negative charges), could be applied to the characterization of hydrophilic interaction chromatographic (HILIC) systems. In the present study, we will show how this methodology can be used to evaluate the effects of increasing buffer salt concentration on retention and separation in a HILIC system. A commercial stationary phase possessing a sulfobetaine zwitterionic bonded ligand (Nucleodur HILIC) was used with a mobile phase composed of 80% acetonitrile and 20% pwwH4 ammonium acetate buffer, with aqueous buffer concentrations varying from 10 to 100mM, resulting in overall concentrations ranging from 2 to 20mM in the mobile phase. Retention factors were measured for a selection of 76 probe analytes. The chosen compounds are small molecules presenting a wide diversity of molecular structures and are relevant to biomedical and pharmaceutical applications. The QSRR models obtained allow for a rationalization of the interactions contributing to retention and separation in the HILIC system considered and shed some light on the effect of varying buffer salt concentration, namely the progressive transition from ion-exchange and electrostatic-repulsion mechanisms to hydrophilic partitioning. PMID:27475992

  14. Evaluation of the properties of a superficially porous silica stationary phase in hydrophilic interaction chromatography.

    PubMed

    McCalley, David V

    2008-06-01

    The sample capacity, column efficiency (and its variation with flow) of a superficially porous unbonded silica phase (Halo) was investigated using hydrophilic interaction chromatography (HILIC), particularly for separation of basic compounds. Sample capacity compared with totally porous silica phases was somewhat reduced, broadly in line with the decreased surface area, but still favourable compared with reversed-phase separations of these solutes. Efficiencies in excess of 100,000 plates were obtained at room temperature in reasonable analysis times by using a 45 cm coupled column, while generating back pressures compatible with conventional HPLC. Shorter columns offered the possibility of fast analysis of bases, and the unfavourable mass transfer properties reported by others at high flow rate for similar reversed-phase columns, were not apparent. While excellent peak shapes were obtained for many bases on silica HILIC phases, problems may still occur for some solutes. PMID:18440010

  15. Investigation on performance of zirconia and magnesia-zirconia stationary phases in hydrophilic interaction chromatography.

    PubMed

    Wang, Qing; Li, Jing; Yang, Xin; Xu, Li; Shi, Zhi-guo; Xu, Lan-Ying

    2014-11-01

    In the current study, zirconia (ZrO2) and its composite, magnesia-zirconia (MgO-ZrO2), were prepared as the hydrophilic interaction chromatographic (HILIC) stationary phases (SPs). Different experimental variables including water content, pH and buffer concentration in the mobile phase (MP) as well as column temperature were systematically studied to permit an in-depth understanding of the chromatographic properties of the mentioned SPs and to explore the retention mechanism further on. The results were compared with a native SiO2 column. Adsorption was demonstrated as the main retention mechanism on the two ZrO2-based SPs. The transferring of the analytes from the MP to the ZrO2-based SPs was endothermic and high column temperature would facilitate the retention. In addition, the MgO-ZrO2 SP exhibited superior resolution, column efficiency as well as stronger retention in comparison to the bare ZrO2 SP, which demonstrated that the introduction of MgO could improve the structure and properties of the material. In conclusion, MgO-ZrO2 was a promising material for HILIC applications. PMID:25127617

  16. Self-assembled cyclodextrin-modified gold nanoparticles on silica beads as stationary phase for chiral liquid chromatography and hydrophilic interaction chromatography.

    PubMed

    Li, Yuanyuan; Wei, Manman; Chen, Tong; Zhu, Nan; Ma, Yulong

    2016-11-01

    A facile strategy based on self-assembly of Au nanoparticles (AuNPs) (60±10nm in size) on the surfaces of amino-functionalized porous silica spheres under mild conditions was proposed. The resulting material possessed a core-shell structure in which AuNPs were the shell and silica spheres were the core. Then, thiolated-β-cyclodextrin (SH-β-CD) was covalently attached onto the AuNPs as chiral selector for the enantioseparation. The resultant packing material was evaluated by high-performance liquid chromatography (HPLC). The separations of nine pairs of enantiomers were achieved by using the new chiral stationary phase (CSP) in the reversed-phase liquid chromatography (RPLC) mode, respectively. The results showed the new CSP have more sufficient interaction with the analytes due to the existence of AuNPs on silica surfaces, resulting in faster mass transfer rate, compared with β-CD modified silica column. The result shed light on potential usage of chemical modified NPs as chiral selector for enantioseparation based on HPLC. In addition, the new phase was also used in hydrophilic interaction liquid chromatography (HILIC) to separate polar compounds and highly hydrophilic compounds. PMID:27591589

  17. [Preparation of xylitol and maltitol modified silica as novel stationary phases for hydrophilic interaction liquid chromatography and evaluation of their separation performance].

    PubMed

    Yong, Tian; Wu, Fan; Xiao, Hongbin; Wan, Boshun

    2015-09-01

    New types of stationary phases for hydrophilic interaction liquid chromatography (HILIC) with unique selectivity are very important for the separation of various polar and hydrophilic analytes. Two novel HILIC stationary phases based on sugar alcohol modified silica were synthesized by a simple two-step reaction in which xylitol and maltitol were bonded onto the surface of silica particles via the addition reaction between -NCO and -OH. The effect of acetonitrile content on the retention indicated that the two stationary phases were of typical HILIC character and exhibited strong retention for polar and hydrophilic analytes. They succeeded in the separation of a wide range of polar and hydrophilic analytes including water soluble vitamins, salicylic acid and its analogues, nucleic acid bases and nucleosides, and icariin and its analogues with unique selectivity. Especially, the maltitol stationary phase showed unique selectivity on glycosyl group, compared to xylitol stationary phase. Furthermore, the effects of buffer pH as well as salt concentration on the retention indicated that electrostatic interaction played an important role in the separation mechanism of the two stationary phases. For sure, the efficient stationary phases are of great potential applications in HILIC. PMID:26753275

  18. Novel imidazolium-embedded N,N-dimethylaminopropyl-functionalized silica-based stationary phase for hydrophilic interaction/reversed-phase mixed-mode chromatography.

    PubMed

    Liu, Shijia; Xu, Hongxin; Yu, Jiaojiao; Li, Danyang; Li, Mingyan; Qiao, Xiaoqiang; Qin, Xinying; Yan, Hongyuan

    2015-12-01

    A novel imidazolium-embedded N,N-dimethylaminopropyl-functionalized silica-based stationary phase (Sil-ImCl) was prepared and further used for hydrophilic interaction/reversed-phase mixed-mode chromatography. The Sil-ImCl stationary phase was respectively characterized by Fourier transform infrared spectrometry, thermogravimetric analysis, and element analysis. A variety of hydrophilic or hydrophobic compounds were used to evaluate the retention mechanisms of the developed stationary phase, and the effects of buffer salt concentration and pH of mobile phase on the retention of these compounds were also investigated. The developed stationary phase was successfully applied for separation of nucleosides and nucleic acid bases, water-soluble vitamins, phenols, and positional isomers. Moreover, simultaneous separation of polar and nonpolar compounds was also achieved with high resolution, outperforming the commercially available C8 column and amino column. Furthermore, the Sil-ImCl stationary phase has been successfully applied for separation of secondary metabolites of Hansfordia sinuosae. All these results demonstrate that the Sil-ImCl stationary phase might be promising for separation of complex polar and nonpolar compounds with high efficiency, especially in biological industry. PMID:26427503

  19. Is the solvation parameter model or its adaptations adequate to account for ionic interactions when characterizing stationary phases for drug impurity profiling with supercritical fluid chromatography?

    PubMed

    Galea, Charlene; West, Caroline; Mangelings, Debby; Vander Heyden, Yvan

    2016-06-14

    Nine commercially available polar and aromatic stationary phases were characterized under supercritical fluid chromatographic (SFC) conditions. Retention data of 64 pharmaceutical compounds was acquired to generate models based on the linear solvation energy relationship (LSER) approach. Previously, adaptation of the LSER model was done in liquid chromatography by the addition of two solute descriptors to describe the influence of positive (D(+)) and negative (D(-)) charges on the retention of ionized compounds. In this study, the LSER models, with and without the ionization terms for acidic and basic solutes, were compared. The improved fits obtained for the modified models support inclusion of the D(+) and D(-) terms for pharmaceutical compounds. Moreover, the statistical significance of the new terms in the models indicates the importance of ionic interactions in the retention of pharmaceutical compounds in SFC. However, unlike characterization through the retention profiles, characterization of the stationary phases by modelling never explains the retention variance completely and thus seems less appropriate. PMID:27181639

  20. Monolithic stationary phases with incorporated fumed silica nanoparticles. Part I. Polymethacrylate-based monolithic column with incorporated bare fumed silica nanoparticles for hydrophilic interaction liquid chromatography.

    PubMed

    Aydoğan, Cemil; El Rassi, Ziad

    2016-05-01

    Fumed silica nanoparticles (FSNPs), were incorporated for the first time into a polymethacrylate monolithic column containing glyceryl monomethacrylate (GMM) and ethylene dimethacrylate (EDMA) in order to develop a new monolithic column for hydrophilic interaction high performance liquid chromatography (HILIC). When compared to poly(GMM-EDMA) monolithic column without FSNPs, the same monolithic column with incorporated FSNPs yielded important effects on HILIC separations. The effects of monomers and FSNPs content of the polymerization mixture on the performance of the monolithic column were examined in details, and the optimized stationary phase was investigated over a wide range of mobile phase composition with polar acidic, weakly basic and neutral analytes including hydroxy benzoic acids, nucleotides, nucleosides, dimethylformamide, formamide and thiourea. The retention of these analytes was mainly controlled by hydrophilic interactions with the FSNPs and electrostatic repulsion from the negatively charged silica surface in the case of hydroxy benzoic acids and nucleotides. The electrostatic repulsion was minimized by decreasing the pH of the aqueous component of the mobile phase, which in turn enhanced the retention of acidic solutes. Nucleotides were best separated using step gradient elution at decreasing pH as well as ACN concentration in the mobile phase. Improved peak shape and faster analysis of nucleosides were attained by a fast linear gradient elution with a shallow decrease in the ACN content of the ACN-rich mobile phase. The run-to-run and column-to-column reproducibility were satisfactory. The percent relative standard deviations (%RSDs) for the retention times of tested solutes were lower than 2.5% under isocratic conditions and lower than 3.5 under gradient conditions. PMID:27059399

  1. Freeze drying for gas chromatography stationary phase deposition

    DOEpatents

    Sylwester, Alan P.

    2007-01-02

    The present disclosure relates to methods for deposition of gas chromatography (GC) stationary phases into chromatography columns, for example gas chromatography columns. A chromatographic medium is dissolved or suspended in a solvent to form a composition. The composition may be inserted into a chromatographic column. Alternatively, portions of the chromatographic column may be exposed or filled with the composition. The composition is permitted to solidify, and at least a portion of the solvent is removed by vacuum sublimation.

  2. Hierarchical CaCO3 chromatography: a stationary phase based on biominerals.

    PubMed

    Sato, Kosuke; Oaki, Yuya; Takahashi, Daisuke; Toshima, Kazunobu; Imai, Hiroaki

    2015-03-23

    In biomineralization, acidic macromolecules play important roles for the growth control of crystals through a specific interaction. Inspired by this interaction, we report on an application of the hierarchical structures in CaCO3 biominerals to a stationary phase of chromatography. The separation and purification of acidic small organic molecules are achieved by thin-layer chromatography and flash chromatography using the powder of biominerals as the stationary phase. The unit nanocrystals and their oriented assembly, the hierarchical structure, are suitable for the adsorption site of the target organic molecules and the flow path of the elution solvents, respectively. The separation mode is ascribed to the specific adsorption of the acidic molecules on the crystal face and the coordination of the functional groups to the calcium ions. The results imply that a new family of stationary phase of chromatography can be developed by the fine tuning of hierarchical structures in CaCO3 materials. PMID:25677568

  3. Novel stationary phases based on asphaltenes for gas chromatography.

    PubMed

    Boczkaj, Grzegorz; Momotko, Malwina; Chruszczyk, Dorota; Przyjazny, Andrzej; Kamiński, Marian

    2016-07-01

    We present the results of investigations on the possibility of the application of the asphaltene fraction isolated from the oxidized residue from vacuum distillation of crude oil as a stationary phase for gas chromatography. The results of the investigation revealed that the asphaltene stationary phases can find use for the separation of a wide range of volatile organic compounds. The experimental values of Rohrschneider/McReynolds constants characterize the asphaltenes as stationary phases of medium polarity and selectivity similar to commercially available phases based on alkyl phthalates. Isolation of asphaltenes from the material obtained under controlled process conditions allows the production of a stationary phase having reproducible sorption properties and chromatographic columns having the same selectivity. Unique selectivity and high thermal stability make asphaltenes attractive as a material for stationary phases for gas chromatography. A low production cost from a readily available raw material (oxidized petroleum bitumens) is an important economic factor in case of application of the asphaltene stationary phases for preparative and process separations. PMID:27144876

  4. A study of separation selectivity using embedded ester-bonded stationary phases for liquid chromatography.

    PubMed

    Bocian, Szymon; Krzemińska, Katarzyna; Buszewski, Bogusław

    2016-07-01

    A new type of stationary bonded phase for liquid chromatography with various functional groups bonded to diol-modified silica via ester bond was synthesized. The structures of the proposed stationary phases contain: alkyl chains (C10, C18), phenyl, and cholesterol groups. The structures of the synthesized materials were confirmed by different physico-chemical techniques such as elemental analysis, infrared spectroscopy (FTIR), (13)C CP/MAS NMR and liquid chromatography under reversed phase conditions (RP) and with hydrophilic interaction liquid chromatography (HILIC). Depending on the type of functionalities bonded to the Diol-Ester, the stationary phases are capable of separating various groups of compounds in RP and HILC, even using pure water as a mobile phase. PMID:27170946

  5. Gold nanoparticle decorated graphene oxide/silica composite stationary phase for high-performance liquid chromatography.

    PubMed

    Liang, Xiaojing; Wang, Xusheng; Ren, Haixia; Jiang, Shengxiang; Wang, Licheng; Liu, Shujuan

    2014-06-01

    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. PMID:24723561

  6. Silica, hybrid silica, hydride silica and non-silica stationary phases for liquid chromatography.

    PubMed

    Borges, Endler M

    2015-04-01

    Free silanols on the surface of silica are the "villains", which are responsible for detrimental interactions of those compounds and the stationary phase (i.e., bad peak shape, low efficiency) as well as low thermal and chemical stability. For these reasons, we began this review describing new silica and hybrid silica stationary phases, which have reduced and/or shielded silanols. At present, in liquid chromatography for the majority of analyses, reversed-phase liquid chromatography is the separation mode of choice. However, the needs for increased selectivity and increased retention of hydrophilic bases have substantially increased the interest in hydrophilic interaction chromatography (HILIC). Therefore, stationary phases and this mode of separation are discussed. Then, non-silica stationary phases (i.e., zirconium oxide, titanium oxide, alumina and porous graphitized carbon), which afford increased thermal and chemical stability and also selectivity different from those obtained with silica and hybrid silica, are discussed. In addition, the use of these materials in HILIC is also reviewed. PMID:25234386

  7. Hydrophilic interaction liquid chromatography (HILIC) in proteomics

    PubMed Central

    Boersema, Paul J.; Mohammed, Shabaz

    2008-01-01

    In proteomics, nanoflow multidimensional chromatography is now the gold standard for the separation of complex mixtures of peptides as generated by in-solution digestion of whole-cell lysates. Ideally, the different stationary phases used in multidimensional chromatography should provide orthogonal separation characteristics. For this reason, the combination of strong cation exchange chromatography (SCX) and reversed-phase (RP) chromatography is the most widely used combination for the separation of peptides. Here, we review the potential of hydrophilic interaction liquid chromatography (HILIC) as a separation tool in the multidimensional separation of peptides in proteomics applications. Recent work has revealed that HILIC may provide an excellent alternative to SCX, possessing several advantages in the area of separation power and targeted analysis of protein post-translational modifications. Figure Artistic impression of the HILIC separation mechanism PMID:18264818

  8. Implementing stationary-phase optimized selectivity in supercritical fluid chromatography.

    PubMed

    Delahaye, Sander; Lynen, Frédéric

    2014-12-16

    The performance of stationary-phase optimized selectivity liquid chromatography (SOS-LC) for improved separation of complex mixtures has been demonstrated before. A dedicated kit containing column segments of different lengths and packed with different stationary phases is commercially available together with algorithms capable of predicting and ranking isocratic and gradient separations over vast amounts of possible column combinations. Implementation in chromatographic separations involving compressible fluids, as is the case in supercritical fluid chromatography, had thus far not been attempted. The challenge of this approach is the dependency of solute retention with the mobile-phase density, complicating linear extrapolation of retention over longer or shorter columns segments, as is the case in conventional SOS-LC. In this study, the possibilities of performing stationary-phase optimized selectivity supercritical fluid chromatography (SOS-SFC) are demonstrated with typical low density mobile phases (94% CO2). The procedure is optimized with the commercially available column kit and with the classical isocratic SOS-LC algorithm. SOS-SFC appears possible without any density correction, although optimal correspondence between prediction and experiment is obtained when isopycnic conditions are maintained. As also the influence of the segment order appears significantly less relevant than expected, the use of the approach in SFC appears as promising as is the case in HPLC. Next to the classical use of SOS for faster baseline separation of all solutes in a mixture, the benefits of the approach for predicting as wide as possible separation windows around to-be-purified solutes in semipreparative SFC are illustrated, leading to significant production rate improvements in (semi)preparative SFC. PMID:25393519

  9. Separation performance of guanidinium-based ionic liquids as stationary phases for gas chromatography.

    PubMed

    Qiao, Lizhen; Lu, Kai; Qi, Meiling; Fu, Ruonong

    2013-02-01

    Room temperature ionic liquids (RTILs) as stationary phases for gas chromatography (GC) have made great achievements in both research and applications over the last decades. Until now, all of the RTIL stationary phases reported have involved imidazolium, ammonium, pyrrolidinium, and phosphonium-based RTILs, and however, no publications are available using guanidinium-based ionic liquids (GBILs) as GC stationary phases except two preliminary reports from our group. In the present work, three hexaalkyl GBILs stationary phases, namely N, N,N',N'-tetramethyl-N″, N″-dioctylguanidinum hexafluophosphate (DOTMG-PF(6)), N,N,N',N'-tetramethyl-N″, N″-dioctylguanidinium bis (trifluoromethylsulfonyl) imide (DOTMG-NTf(2)), and N,N,N',N'-tetraoctyl-N″, N″-dimethylguanidinium bis (trifluoromethylsulfonyl) imide (TODMG-NTf(2)), were synthesized and used as stationary phases for GC separation after they were statically coated onto the inner walls of fused-silica capillary columns. The evaluation of DOTMG-PF(6) and TODMG-NTf(2) as GC stationary phases is reported here for the first time, whereas additional results on the DOTMG-NTf(2) stationary phase are added here on the basis of our previous report. In this work, McReynolds constants and Abraham solvation system constants are used to evaluate the average polarity and the solvation properties of the GBILs stationary phases for GC separation, respectively. The results show that the GBILs stationary phases exhibit medium polarity with an average polarity of 293-390, and that the major molecular interactions of the GBILs with analytes are dipole/polarizable interactions, H-bond basicity and dispersion forces, etc. After this, the separation performance and thermal stability of the GBILs stationary phases were evaluated, showing that these stationary phases achieve excellent separation for analytes of great variety covering hydrocarbons, alcohols, esters, aldehydes, ketones, amines, amides and aromatics, and exhibit

  10. Calixarene ionic liquid modified silica gel: A novel stationary phase for mixed-mode chromatography.

    PubMed

    Hu, Kai; Zhang, Wenfen; Yang, Huaixia; Cui, Yongxia; Zhang, Jingya; Zhao, Wenjie; Yu, Ajuan; Zhang, Shusheng

    2016-05-15

    A novel calixarene ionic liquid functionalized silica material was synthesized by the preparation of a new calixarene monomer and its grafting on mercaptopropyl modified silica gel. The material was characterized by infrared spectra, elemental analysis, and thermogravimetric analysis. To explore the retention mechanism of the stationary phase, linear solvation energy relationships (LSER) equation as an effective mathematical model was used. In addition to this, the distinct separation mechanisms were outlined by selected examples of chromatographic separations in the different modes. In reversed-phase liquid chromatography, this new stationary phase presented specific chromatographic performance when evaluated using alkylbenzenes, PAHs and phenols as solutes. Due to the existing polar functional groups, this stationary phase can also be used in hydrophilic interaction chromatography, six nucleosides and four ginsenosides were separated successfully in hydrophilic mode. Furthermore, anions can be separated on the column in anion exchange mode. Thus, this new material was can be applied as a new kind of mixed-mode stationary phase in liquid chromatography, which allows an exceptionally flexible adjustment of retention and selectivity by tuning the experimental conditions. PMID:26992535

  11. Surface modification of polytetrafluoroethylene column for two-stationary phase separations by counter-current chromatography.

    PubMed

    Quan, Kai-jun; Huang, Xin-yi; Li, Xiao-ting; Wang, Gao-hong; Liu, Yan-juan; Duan, Wen-da; Di, Duo-long

    2015-11-27

    To improve the separation capability of CCC, a novel solid-liquid two-stationary phases CCC (ASP-CCC) column was prepared employing graphene oxide (GO) conjugated poly-dopamine (PD) coating (GO/PD) as auxiliary stationary phase (ASP). The results of Scanning electron microscopy (SEM), contact angle and X-ray photoelectron spectroscopy (XPS) indicated that nanostructured GO and PD were successfully grafted on the inner wall of the PTFE column. Three alkaloid compounds were selected as the target analytes to evaluate the performance of the novel column. Because of the intermolecular force (hydrogen bond, electrostatic interaction and π-π interaction) between the ASP and model compounds, three analytes were well separated with this novel ASP-CCC column. Additionally, the novel column exhibited higher stationary phase retention ratio, about 8%, than original column without changing the chromatographic condition. Furthermore, the eluotropic sequence of analytes on novel column was in accordance with that in the original column. This suggested that the novel column is a CCC column with auxiliary stationary phase (ASP) in its own right, and the present separation mode is the combination of partition chromatography and adsorption chromatography. PMID:26518492

  12. Highly stabilized, polymer-lipid membranes prepared on silica microparticles as stationary phases for capillary chromatography

    PubMed Central

    Gallagher, Elyssia S.; Adem, Seid M.; Baker, Christopher A.; Ratnayaka, Saliya N.; Jones, Ian W.; Hall, Henry K.; Saavedra, S. Scott; Aspinwall, Craig A.

    2015-01-01

    The ability to rapidly screen complex libraries of pharmacological modulators is paramount to modern drug discovery efforts. This task is particularly challenging for agents that interact with lipid bilayers or membrane proteins due to the limited chemical, physical, and temporal stability of conventional lipid-based chromatographic stationary phases. Here, we describe the preparation and characterization of a novel stationary phase material composed of highly stable, polymeric-phospholipid bilayers self-assembled onto silica microparticles. Polymer lipid membranes were prepared by photochemical or redox initiated polymerization of 1,2-bis[10-(2′,4′-hexadieoyloxy)decanoyl]-sn-glycero-2-phosphocholine (bis-SorbPC), a synthetic, polymerizable lipid. The resulting polymerized bis-SorbPC (poly(bis-SorbPC)) stationary phases exhibited enhanced stability compared to particles coated with 1,2-dioleoyl-sn-glycero-phosphocholine (unpolymerized) phospholipid bilayers when exposed to chemical (50mM triton X-100 or 50% acetonitrile) and physical (15 min sonication) insults after 30 days of storage. Further, poly(bis-SorbPC)-coated particles survived slurry packing into fused silica capillaries, compared to unpolymerized lipid membranes, where the lipid bilayer was destroyed during packing. Frontal chromatographic analyses of the lipophilic small molecules acetylsalicylic acid, benzoic acid, and salicylic acid showed > 44% increase in retention times (P < 0.0001) for all analytes on poly(bis-SorbPC)-functionalized stationary phase compared to bare silica microspheres, suggesting a lipophilic retention mechanism. Phospholipid membrane-functionalized stationary phases that withstand the chemical and physical rigors of capillary LC conditions can substantially increase the efficacy of lipid membrane affinity chromatography, and represents a key advance towards the development of robust membrane protein-functionalized chromatographic stationary phases. PMID:25670414

  13. An improved classification of stationary phases for ultra-high performance supercritical fluid chromatography.

    PubMed

    West, Caroline; Lemasson, Elise; Bertin, Sophie; Hennig, Philippe; Lesellier, Eric

    2016-04-01

    Supercritical fluid chromatography (SFC) has recently benefited of new instrumentation, together with the availability of many ultra-high performance columns (sub -2μm fully porous particles or sub -3μm superficially porous particles), rendering it more attractive than ever. Most of these columns commonly used in SFC were initially developed for HPLC use, with an increasing number of stationary phases specifically designed for SFC. While the availability of different stationary phase chemistries is an advantage to achieve successful SFC separations, selecting a column for method development remains difficult. For this reason, we have previously developed a classification of stationary phases dedicated to SFC use. It is based on linear solvation energy relationships (LSER) with Abraham descriptors (for neutral species). While current interest in SFC is strong in the pharmaceutical industry, the need to take account of interactions occurring with ionisable species is pressing. We have previously shown how a modified version of the solvation parameter model, adapted to take account of ionic and ionizable species, could be applied to the characterization of SFC systems. In the present paper, based on this modified LSER model, and on the analysis of 109 neutral and ionisable species, we propose an improved classification of 31 ultra-high performance stationary phases to facilitate method development with SFC. PMID:26920664

  14. Separation of dialkyl sulfides by metallo-mesogenic stationary phases for complexation gas chromatography.

    PubMed

    Chen, Jian-Lian; Liu, Chuen-Ying

    2007-08-17

    A copper mesogenic side-chain polymer (P-C(15)CuC(18)) was cross-linked onto the capillary wall as a stationary film for gas chromatography (GC) separation of alkyl sulfides. These organic sulfides are of interest for their large health impact because of their wide range of volatiles and high reactivities toward metals. Different GC parameters for optimal separation efficiency are discussed for use with a mesogenic polymer column along with flame photometric detection (FPD). Both the carrier gas flow-rate and column temperature were studied to determine the relationship of plate height to the chemical structure of the solutes, as well as to determine the morphology of the mesogenic polymer. Van 't Hoff plots show phase transitions of the stationary mesophase as the column temperature was varied. The results reveal that the separation mechanism might be based on ligand exchange and polarity interaction between the analytes and the stationary phase, with the vapor pressure of the analytes also being important. The former interaction dominates in the lamellar crystalline phase and the latter interaction dominates in the hexagonal columnar-discotic phase. With high reproducibility for retention time (RSD< or =0.37%) and for peak area (RSD< or =5.16%), the GC-FPD system produced linear calibration graphs (r> or =0.9918) for the determination of 13 sulfides with a detection limit below 2.5 ng. PMID:17568598

  15. A sulfonic-azobenzene-grafted silica amphiphilic material: a versatile stationary phase for mixed-mode chromatography.

    PubMed

    Qiu, Hongdeng; Zhang, Mingliang; Gu, Tongnian; Takafuji, Makoto; Ihara, Hirotaka

    2013-12-23

    A novel sulfonic-azobenzene-functionalized amphiphilic silica material was synthesized through the preparation of a new sulfonic azobenzene monomer and its grafting on mercaptopropyl-modified silica by a surface-initiated radical chain-transfer reaction. The synthesis was confirmed by infrared spectra, elemental analysis, and thermogravimetric analysis. This new material was successfully applied as a new kind of mixed-mode stationary phase in liquid chromatography. This allows an exceptionally flexible adjustment of retention and selectivity by tuning the experimental conditions. The distinct separation mechanisms were outlined by selected examples of chromatographic separations in the different modes. In reversed-phase liquid chromatography, this new stationary phase presented specific chromatographic performance when evaluated using a Tanaka test mixture. Seven dinitro aromatic isomers, four steroids, and seven flavonoids were separated successfully in simple reversed-phase mode. This stationary phase can also be used in hydrophilic interaction chromatography because of the existing polar functional groups; for this, nucleosides and their bases were used as a test mixture. Interestingly, the same nucleosides and bases can also be separated in per aqueous liquid chromatography using the same stationary phase. Three ginsenosides including Rg1, Re, and Rb1 were successfully separated in hydrophilic mode. There is the potential for more applications to benefit from this useful column. PMID:24353082

  16. Retention modelling in hydrophilic interaction chromatography.

    PubMed

    Euerby, Melvin R; Hulse, Jennifer; Petersson, Patrik; Vazhentsev, Andrey; Kassam, Karim

    2015-12-01

    The retention behaviour of acidic, basic and quaternary ammonium salts and polar neutral analytes has been evaluated on acidic, basic and neutral hydrophilic interaction chromatography (HILIC) stationary phases as a function of HILIC operating parameters such as MeCN content, buffer concentration, pH and temperature. Numerous empirical HILIC retention models (existing and newly developed ones) have been assessed for their ability to describe retention as a function of the HILIC operating parameters investigated. Retention models have been incorporated into a commercially available retention modelling programme (i.e. ACD/LC simulator) and their accuracy of retention prediction assessed. The applicability of HILIC modelling using these equations has been demonstrated in the two-dimensional isocratic (i.e. buffer concentration versus MeCN content modelling) and one-dimensional gradient separations for a range of analytes of differing physico-chemical properties on the three stationary phases. The accuracy of retention and peak width prediction was observed to be comparable to that reported in reversed-phase chromatography (RPC) retention modelling. Intriguingly, our results have confirmed that the use of gradient modelling to predict HILIC isocratic conditions and vice versa is not reliable. A relative ranking of the importance of the retention and selectivity of HILIC operating parameters has been determined using statistical approaches. For retention, the order of importance was observed to be organic content > stationary phase > temperature ≈ mobile phase pH (i.e. pH 3-6 which mainly effects the ionization of the analyte) ≈ buffer concentration. For selectivity, the nature of the stationary phase > mobile phase pH > buffer concentration > temperature > organic content. PMID:26563113

  17. Separations of corticosteroids using electrochemically modulated liquid chromatography: Selectivity enhancements at a porous graphitic carbon stationary phase

    SciTech Connect

    Ting, E.Y.; Porter, M.D. |

    1997-02-15

    Electrochemically modulated liquid chromatography has been applied to the separation of a mixture of structurally similar corticosteroids (i.e., prednisone, prednisolone, cortisone, and hydrocortisone) using a porous graphitic carbon stationary phase. Changes in the voltage applied to the column markedly affected the efficiency as well as the elution order of the separation, with the mixture fully resolved at large negative values of applied potential. Mechanistic aspects in terms of the influence of changes in the applied voltage on the extent of the interactions between these analytes and the stationary phase are briefly discussed. 19 refs., 2 figs.

  18. Characterization of five chemistries and three particle sizes of stationary phases used in supercritical fluid chromatography.

    PubMed

    Khater, S; West, C; Lesellier, E

    2013-12-01

    Sub-2-microns particles employed as supporting phases are known to favor column efficiency. Recently a set of columns based on sub-2-microns particles for use with supercritical fluid mobile phases have been introduced by Waters. Five different stationary phase chemistries are available: BEH silica, BEHEthyl-pyridine, X Select CSH Fluorophenyl, HSS C18 SB and BEH Shield RP18. This paper describes the characterization of 15 stationary phases, the five different chemistries, and three particle sizes, 1.7 (or 1.8), 3.5 and 5 microns, with the same carbon dioxide–methanol mobile phase and a set of more than a hundred compounds. The interactions established in the 15 different chromatographic systems used in supercritical fluid chromatography (SFC) are assessed with linear solvation energy relationships (LSERs).The results show the good complementarity of the five column chemistries, and their comparative location inside a classification map containing today around 70 different commercial phases. Among the five different chemistries, the HSS C18 SB phase displays a rather unusual behavior in regards of classical C18 phases, as it displays significant hydrogen–bonding interactions. Besides, it appears, as expected, that the BEH Ethyl–pyridine phase has weak interactions with basic compounds. The effect of particle size was studied because smaller particles induce increased inlet and internal pressure. For compressible fluids,this pressure change modifies the fluid density, i.e. the apparent void volume and the eluting strength.These changes could modify the retention and the selectivity of compounds in the case of method trans-fer, by using different particle sizes, from 5 down to 1.7 m. A hierarchical cluster analysis shows that stationary phase clusters were based on the phase chemistry rather than on the particle size, meaning that method transfer from 5 to 1.7 microns can be achieved in the subcritical domain i.e. by using a weakly compressible fluid. PMID

  19. Fluoro-substituted tetraphenyl-phenyl grafted polysiloxanes as highly selective stationary phases for gas chromatography.

    PubMed

    Han, Xue; He, Xinxin; Wang, Huan; Wang, Bing; Wu, Bo

    2016-06-01

    In this work, two new types of polycyclic aromatic grafted polysiloxanes, namely, 3,4-bis(4-fluoro phenyl)-2,5-diphenyl polysiloxane (FPP) and 3,4-bis(3,4,5-trifluoro phenyl)-2,5-diphenyl polysiloxane (TFPP), were synthesized and statically coated onto capillary columns as stationary phases for gas chromatography (GC). Based on their McReynolds constants, both columns exhibited moderate polarity. The efficiencies of the FPP and TFPP columns were 3316 (k=3.96, naphthalene; 0.25mm inner diameter) and 3768 (k=4.14, naphthalene; 0.25mm inner diameter) plates/m, respectively. The thermostability of the polymers was tested by thermogravimetric analysis (TGA), and results revealed that both TFPP and FPP began to decompose slightly at 380°C. Separation of polyethylene pyrolysis products showed that the upper working temperature of the two columns can reach up to 360°C. Relying on their unique polarizable characteristics in combination with other types of interactions, such as H-bond acceptor, dipole-dipole, and dispersive interactions, the newly synthesized polarizable stationary phases offered unique selectivity for aromatic isomers and substituted benzenes. A slight separation difference between TPP and TFPP was observed. TFPP also exerted excellent selectivity for polycyclic aromatic hydrocarbons, fatty acid esters, and fatty alcohols. Overall, FPP and TFPP demonstrated considerable potential for further applications because of their unique structures and outstanding separation performance. PMID:27139216

  20. Pharmaceutical-enantiomers resolution using immobilized polysaccharide-based chiral stationary phases in supercritical fluid chromatography.

    PubMed

    De Klerck, Katrijn; Vander Heyden, Yvan; Mangelings, Debby

    2014-02-01

    Since their introduction on the market the applicability of immobilized polysaccharide-based chiral stationary phases in high-performance liquid chromatography has been thoroughly investigated. These immobilized phases have the benefit to be applicable with a wide range of modifiers, potentially extending the application range of the polysaccharide-based stationary phases. Because an increasing number of stationary phases are being introduced in the field of chiral chromatography it is important to evaluate their enantioselectivity in different techniques in order to get an idea about their applicability. In this study, three immobilized chiral polysaccharide-based stationary phases (Chiralpak IA, IB, and IC) are evaluated in supercritical fluid chromatography (SFC) with a test set of pharmaceutical racemates. This is done in a three-fold manner: their performance is evaluated (1) using traditional modifiers, (2) using mixtures of atypical modifiers, and (3) the results were compared to those on coated stationary phases with an equivalent chiral selector. To get a visual overview of the enantioselective patterns of the different chromatographic systems (mobile and stationary phase combinations), a Principal Component Analysis is performed, which allows determining the (dis)similarity between individual systems. To assess the complementarity cumulative success rates are determined. The immobilized chiral stationary phases prove to yield high cumulative success rates. PMID:24438871

  1. Mixed Stationary Liquid Phases for Gas-Liquid Chromatography.

    ERIC Educational Resources Information Center

    Koury, Albert M.; Parcher, Jon F.

    1979-01-01

    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)

  2. Development of a decaaza-cyclophane stationary phase for high-performance liquid chromatography.

    PubMed

    Hu, Kai; Deng, Zhifen; Wang, Bei; Cui, Yongxia; Miao, Mingsan; Liu, Wei; Jiang, Qiong; Zhao, Wenjie; Huang, Yanjie; Zhang, Shusheng

    2015-01-01

    A new stationary phase for high-performance liquid chromatography was prepared by covalently bonding a heteroatom-bridged cyclophane onto silica gel using 3-aminopropyltriethoxysilane as the coupling reagent. The structure of the new material was characterized by infrared spectroscopy, elemental analysis, and thermogravimetric analysis. The linear solvation energy relationship method was successfully employed to evaluate the new phase with a set of 25 solutes, and compared with octadecylsilyl and p-tert-butyl-calix[4]arene bonded stationary phases. The retention characteristics of the new phase are similar to the octadecylsilyl and conventional calixarene phases, and it also has distinctive features. In addition, the chromatographic behavior of the phase was illustrated by eluting alkylbenzenes and inorganic anions in the reversed-phase mode and anion-exchange mode, respectively. Thus, multi-interaction mechanisms and mixed-mode separation of the new phase can very likely guarantee its promising application in the analysis of complex samples. The column has been successfully employed for the analysis of triazines in milk, and it is demonstrated to be a competitive alternative analytical method for the determination of triazine herbicide residues. PMID:25353993

  3. Measuring protein interactions by microchip self-interaction chromatography.

    PubMed

    García, Carlos D; Hadley, DeGail J; Wilson, W William; Henry, Charles S

    2003-01-01

    The self-interaction of proteins is of paramount importance in aggregation and crystallization phenomena. Solution conditions leading to a change in the state of aggregation of a protein, whether amorphous or crystalline, have mainly been discovered by the use of trial and error screening of large numbers of solutions. Self-interaction chromatography has the potential to provide a quantitative method for determination of protein self-interactions amenable to high-throughput screening. This paper describes the construction and characterization of a microchip separation system for low-pressure self-interaction chromatography using lysozyme as a model protein. The retention time was analyzed as a function of mobile-phase composition, amount of protein injected, flow rate, and stationary-phase modification. The capacity factors (k') as a function of crystallizing agent concentration are compared with previously published values for the osmotic second virial coefficient (B(22)) obtained by static light scattering, showing the ability of the chip to accurately determine protein-protein interactions. A 500-fold reduction in protein consumption and the possibility of using conventional instrumentation and automation are some of the advantages over currently used methodologies for evaluating protein-protein interactions. PMID:12790668

  4. 4-Chloro-6-pyrimidinylferrocene modified silica gel: A novel multiple-function stationary phase for mixed-mode chromatography.

    PubMed

    Qiao, Lijun; Zhou, Xiaohua; Zhang, Yanhao; Yu, Ajuan; Hu, Kai; Zhang, Shusheng; Wu, Yangjie

    2016-06-01

    A novel multi-function and mixed-mode stationary phase based on 4-chloro-6-pyrimidinylferrocene modified silica (NFcS) was synthesized and characterized by infrared spectroscopy, elemental analysis and thermogravimetric analysis. Linear solvation energy relationship method was successfully employed to evaluate the new phase with a set of 27 solutes including aromatic and aliphatic compounds. Multiple mechanisms including hydrophobic, π-π, hydrogen-bonding, charge-transfer, acid-base equilibrium and anion-exchange interactions are involved. Based on these interactions, successful separation could be achieved among polycyclic aromatic hydrocarbons, mono-substituted benzenes, aromatic amines, phenols, quinolines, pyridines and nucleosides in reversed-phase (RP) or normal-phase (NP) chromatography. Inorganic anions were also shown to be individually separated in anion-exchange chromatography by using the same column. Moreover, the results here also demonstrated that NFcS based stationary phase could effectively reduce the adverse effect of residual silanol in the separation process. Such stationary phase with characteristics of multi-interaction mechanism and mixed-mode separation is potential for the analysis of complex samples. The NFcS column was successfully employed for the analysis of plant growth regulators in Fruit. PMID:27130083

  5. Planar gas chromatography column on aluminum plate with multi-walled carbon nanotubes as stationary phase

    NASA Astrophysics Data System (ADS)

    Platonov, I. A.; Platonov, V. I.; Pavelyev, V. S.

    2016-04-01

    The high selectivity of the adsorption layer for low-boiling alkanes is shown, the separation factor (α) couple iso-butane / butane is 1.9 at a column temperature of 50 °C.The paper presents sorption and selective properties of planar gas chromatography column on aluminum plate with multi-walled carbon nanotubes as the stationary phase.

  6. Metal-Organic Framework Thin Films as Stationary Phases in Microfabricated Gas-Chromatography Columns.

    SciTech Connect

    Read, Douglas; Sillerud, Colin Halliday

    2016-01-01

    The overarching goal of this project is to integrate Sandia's microfabricated gas-chromatography ( GC) columns with a stationary phase material that is capable of retaining high-volatility chemicals and permanent gases. The successful integration of such a material with GCs would dramatically expand the repertoire of detectable compounds for Sandia's various microanalysis systems. One such promising class of candidate materials is metal-organic frameworks (MOFs). In this report we detail our methods for controlled deposition of HKUST-1 MOF stationary phases within GC columns. We demonstrate: the chromatographic separation of natural gas; a method for determining MOF film thickness from chromatography alone; and the first-reported GC x GC separation of natural gas -- in general -- let alone for two disparate MOF stationary phases. In addition we determine the fundamental thermodynamic constant for mass sorption, the partition coefficient, for HKUST-1 and several light hydrocarbons and select toxic industrial chemicals.

  7. Amine Gradient Stationary Phases on In-House Built Monolithic Columns for Liquid Chromatography.

    PubMed

    Dewoolkar, Veeren C; Jeong, Lena N; Cook, Daniel W; Ashraf, Kayesh M; Rutan, Sarah C; Collinson, Maryanne M

    2016-06-01

    Stationary phase gradients on monolithic silica columns have been successfully and reproducibly prepared and characterized with comparisons made to uniformly modified stationary phases. Stationary phase gradients hold great potential for use in liquid chromatography (LC), both in terms of simplifying analysis as well as providing novel selectivity. In this work, we demonstrate the creation of a continuous stationary phase gradient on in-house synthesized monolithic columns by infusing an aminoalkoxysilane solution through the silica monoliths via controlled rate infusion. The presence of amine and its distribution along the length of gradient and uniformly modified columns were assessed via X-ray photoelectron spectroscopy (XPS). XPS showed a clear gradient in surface coverage along the length of the column for the gradient stationary phases while a near uniform distribution on the uniformly modified stationary phases. To demonstrate the application of these gradient stationary phases, the separations of both nucleobases and weak acids/weak bases on these gradient stationary phases have been compared to uniformly modified and unmodified silica columns. Of particular note, the retention characteristics of 11 gradient columns, 5 uniformly modified columns, and 5 unmodified columns have been tested to establish the reproducibility of the synthetic procedures. Standard deviations of the retention factors were in the range from 0.06 to 0.5, depending on the analyte species. We show that selectivity is achieved with the stationary phase gradients that are significantly different from either uniformly modified amine or unmodified columns. These results indicate the significant promise of this strategy for creating novel stationary phases for LC. PMID:27203513

  8. Chemometric study of retention on binary stationary phases in gas chromatography.

    PubMed

    Bouzouane, S; Righezza, M; Touabet, A

    2012-02-01

    Using gas chromatography, data analysis is performed on a dataset consisting of 486 retention indices, 27 standards (ramified alkanes, aliphatic alcohols, and aromatic compounds), 6 pure and binary stationary phases, and three temperatures. The behavior of the pure stationary phases (OV-3, OV-225, OV-61-OH, and OV-1701-OH) and the binary stationary phases (OV-3/OV-225 and OV-61-OH/OV-1701-OH) at different temperatures (60°C-100°C) is investigated with factor and topological analysis. The influence of temperature and the nature of the mixed stationary phases on the retention indices is studied by correspondence factor analysis (CFA). The non-additivity of the retention properties of the pure phases used as mixed phases is clearly established by CFA. The topological analysis of the substituent's effect is investigated with a DARC/PELCO procedure and shows the particular influence of the stationary phase composition on the retention. The substituent effect is measured for the pure and binary stationary phases at various temperatures. The evolution of the substituent effect from the pure stationary phases to the binary phases is discussed. PMID:22298764

  9. Preparation and evaluation of surface-bonded tricationic ionic liquid silica as stationary phases for high-performance liquid chromatography.

    PubMed

    Qiao, Lizhen; Shi, Xianzhe; Lu, Xin; Xu, Guowang

    2015-05-29

    Two tricationic ionic liquids were prepared and then bonded onto the surface of supporting silica materials through "thiol-ene" click chemistry as new stationary phases for high-performance liquid chromatography. The obtained columns of tricationic ionic liquids were evaluated respectively in the reversed-phase liquid chromatography (RPLC) mode and hydrophilic interaction liquid chromatography (HILIC) mode, and possess ideal column efficiency of 80,000 plates/m in the RPLC mode with naphthalene as the test solute. The tricationic ionic liquid stationary phases exhibit good hydrophobic and shape selectivity to hydrophobic compounds, and RPLC retention behavior with multiple interactions. In the HILIC mode, the retention and selectivity were evaluated through the efficient separation of nucleosides and bases as well as flavonoids, and the typical HILIC retention behavior was demonstrated by investigating retention changes of hydrophilic solutes with water volume fraction in mobile phase. The results show that the tricationic ionic liquid columns possess great prospect for applications in analysis of hydrophobic and hydrophilic samples. PMID:25890438

  10. Resolving Isomeric Glycopeptide Glycoforms with Hydrophilic Interaction Chromatography (HILIC).

    PubMed

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

    2016-09-01

    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. PMID:27582638

  11. Resolving Isomeric Glycopeptide Glycoforms with Hydrophilic Interaction Chromatography (HILIC)

    PubMed Central

    Huang, Yining; Nie, Yongxin; Boyes, Barry

    2016-01-01

    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. PMID:27582638

  12. Phosphatidylcholine covalently linked to a methacrylate-based monolith as a biomimetic stationary phase for capillary liquid chromatography.

    PubMed

    Moravcová, Dana; Carrasco-Correa, Enrique Javier; Planeta, Josef; Lämmerhofer, Michael; Wiedmer, Susanne K

    2015-07-10

    In this study a strategy to immobilize phospholipids onto a polymer-based stationary phase is described. Methacrylate-based monoliths in capillary format (150×0.1mm) were modified by soybean phosphatidylcholine through 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide coupling to obtain stationary phases suitable to mimic cell surface membranes. The covalent coupling reaction involves the phosphate group in phospholipids; therefore, the described methodology is suitable for all types of phospholipids. Immobilization of soy bean phosphatidylcholine on the monolith was confirmed by attenuated total reflectance Fourier transform infrared spectroscopy and gas chromatography-mass spectrometry of the fatty alcohol profile, generated upon reductive cleavage of the fatty acyl side chains of the phospholipid on the monolith surface with lithium aluminium hydride. The prepared stationary phases were evaluated through studies on the retention of low-molar mass model analytes including neutral, acidic, and basic compounds. Liquid chromatographic studies confirmed predominant hydrophobic interactions between the analytes and the synthesized stationary phase; however, electrostatic interactions contributed to the retention as well. The synthesized columns showed high stability even with fully aqueous mobile phases such as Dulbecco's phosphate-buffered saline solution. PMID:26024990

  13. Synthesis and Purification of Iodoaziridines Involving Quantitative Selection of the Optimal Stationary Phase for Chromatography

    PubMed Central

    Boultwood, Tom; Affron, Dominic P.; Bull, James A.

    2014-01-01

    The highly diastereoselective preparation of cis-N-Ts-iodoaziridines through reaction of diiodomethyllithium with N-Ts aldimines is described. Diiodomethyllithium is prepared by the deprotonation of diiodomethane with LiHMDS, in a THF/diethyl ether mixture, at -78 °Cin the dark. These conditions are essential for the stability of the LiCHI2 reagent generated. The subsequent dropwise addition of N-Ts aldimines to the preformed diiodomethyllithium solution affords an amino-diiodide intermediate, which is not isolated. Rapid warming of the reaction mixture to 0 °C promotes cyclization to afford iodoaziridines with exclusive cis-diastereoselectivity. The addition and cyclization stages of the reaction are mediated in one reaction flask by careful temperature control. Due to the sensitivity of the iodoaziridines to purification, assessment of suitable methods of purification is required. A protocol to assess the stability of sensitive compounds to stationary phases for column chromatography is described. This method is suitable to apply to new iodoaziridines, or other potentially sensitive novel compounds. Consequently this method may find application in range of synthetic projects. The procedure involves firstly the assessment of the reaction yield, prior to purification, by 1H NMR spectroscopy with comparison to an internal standard. Portions of impure product mixture are then exposed to slurries of various stationary phases appropriate for chromatography, in a solvent system suitable as the eluent in flash chromatography. After stirring for 30 min to mimic chromatography, followed by filtering, the samples are analyzed by 1H NMR spectroscopy. Calculated yields for each stationary phase are then compared to that initially obtained from the crude reaction mixture. The results obtained provide a quantitative assessment of the stability of the compound to the different stationary phases; hence the optimal can be selected. The choice of basic alumina, modified to

  14. Chromatographic methods enabling the characterization of stationary phases and retention prediction in high-performance liquid chromatography and supercritical fluid chromatography.

    PubMed

    Sykora, David; Vozka, Jiri; Tesarova, Eva

    2016-01-01

    In the scope of the present review, the current status of high-performance liquid chromatography/ultra-high performance liquid chromatography and supercritical fluid chromatography is briefly provided. These techniques and their retention mechanisms are compared. Various alternative approaches utilized for the determination and description of the retention processes in these two systems are mapped. Two frequently used concepts, linear-free energy relationships, and hydrophobic subtraction models, used for the characterization of the retention interactions, are discussed. Principles and selected applications of the both methods are also covered. Then the models applied for the prediction of retention behavior of solutes on stationary phases are outlined. The procedures utilized for the sorbent/column classification are also covered. Simple chromatographic tests frequently used for the basic characterization and mutual comparison of stationary phases are summarized and briefly commented on. The importance of a statistical evaluation of complex retention data obtained from the chromatographic measurements is outlined. Finally, computer simulations aiming at the facilitation of the quest to optimize separation conditions for a given mixture of analytes are touched upon. PMID:26497150

  15. Versatile ligands for high-performance liquid chromatography: An overview of ionic liquid-functionalized stationary phases.

    PubMed

    Zhang, Mingliang; Mallik, Abul K; Takafuji, Makoto; Ihara, Hirotaka; Qiu, Hongdeng

    2015-08-01

    Ionic liquids (ILs), a class of unique substances composed purely by cation and anions, are renowned for their fascinating physical and chemical properties, such as negligible volatility, high dissolution power, high thermal stability, tunable structure and miscibility. They are enjoying ever-growing applications in a great diversity of disciplines. IL-modified silica, transforming the merits of ILs into chromatographic advantages, has endowed the development of high-performance liquid chromatography (HPLC) stationary phase with considerable vitality. In the last decade, IL-functionalized silica stationary phases have evolved into a series of branches to accommodate to different HPLC modes. An up-to-date overview of IL-immobilized stationary phases is presented in this review, and divided into five parts according to application mode, i.e., ion-exchange, normal-phase, reversed-phase, hydrophilic interaction and chiral recognition. Specific attention is channeled to synthetic strategies, chromatographic behavior and separation performance of IL-functionalized silica stationary phases. PMID:26320781

  16. Sum of ranking differences to rank stationary phases used in packed column supercritical fluid chromatography.

    PubMed

    West, Caroline; Khalikova, Maria A; Lesellier, Eric; Héberger, Károly

    2015-08-28

    The identification of a suitable stationary phase in supercritical fluid chromatography (SFC) is a major source of difficulty for those with little experience in this technique. Several protocols have been suggested for column classification in high-performance liquid chromatography (HPLC), gas chromatography (GC), and SFC. However, none of the proposed classification schemes received general acceptance. A fair way to compare columns was proposed with the sum of ranking differences (SRD). In this project, we used the retention data obtained for 86 test compounds with varied polarity and structure, analyzed on 71 different stationary phases encompassing the full range in polarity of commercial packed columns currently available to the SFC chromatographer, with a single set of mobile phase and operating conditions (carbon dioxide-methanol mobile phase, 25°C, 150bar outlet pressure, 3ml/min). First, a reference column was selected and the 70 remaining columns were ranked based on this reference column and the retention data obtained on the 86 analytes. As these analytes previously served for the calculation of linear solvation energy relationships (LSER) on the 71 columns, SRD ranks were compared to LSER methodology. Finally, an external comparison based on the analysis of 10 other analytes (UV filters) related the observed selectivity to SRD ranking. Comparison of elution orders of the UV filters to the SRD rankings is highly supportive of the adequacy of SRD methodology to select similar and dissimilar columns. PMID:26228853

  17. Determination of solute partition behavior with room-temperature ionic liquid based micellar gas-liquid chromatography stationary phases using the pseudophase model.

    PubMed

    Lantz, Andrew W; Pino, Verónica; Anderson, Jared L; Armstrong, Daniel W

    2006-05-19

    The use of micelles in ionic liquid based gas-chromatography stationary phases was evaluated using equations derived for a "three-phase" model. This model allows the determination of all three partition coefficients involved in the system, and elucidates the micellar contribution to retention and selectivity. Four types of micellar-ionic liquid columns were examined in this study: 1-butyl-3-methylimidazolium chloride with sodium dodecylsulfate or dioctyl sulfosuccinate, and 1-butyl-3-methylimidazolium hexafluorophosphate with polyoxyethylene-100-stearyl ether or polyoxyethylene-23-lauryl ether. The partition coefficients were measured for a wide range of probe molecules capable of a variety of types and magnitudes of interactions. In general, most probe molecules preferentially partitioned to the micellar pseudophase over the bulk ionic liquid component of the stationary phase. Therefore, addition of surfactant to the stationary phase usually resulted in greater solute retention. It is also shown that the selectivity of the stationary phase is significantly altered by the presence of micelles, either by enhancing or lessening the separation. The effects of surfactant on the interaction parameters of the stationary phase are determined using the Abraham solvation parameter model. The addition of sodium dodecylsulfate and dioctyl sulfosuccinate to 1-butyl-3-methylimidazolium chloride stationary phases generally increased the phase's hydrogen bond basicity and increased the level of dispersion interaction. Polyoxyethylene-100-stearyl ether and polyoxyethylene-23-lauryl ether surfactants, however, enhanced the pi-pi/n-pi, polarizability/dipolarity, and hydrogen bond basicity interactions of 1-butyl-3-methylimidazolium hexafluorophosphate to a greater degree than the ionic surfactants with 1-butyl-3-methylimidazolium chloride. However, these nonionic surfactants appeared to hinder the ability of the stationary phase to interact with solutes via dispersion forces

  18. Evaluation of an amide-based stationary phase for supercritical fluid chromatography.

    PubMed

    Borges-Muñoz, Amaris C; Colón, Luis A

    2016-09-01

    J. Sep. Sci. 2016, 39, 3469-3476 A stationary phase containing an amide group embedded in a hydrophobic backbone (i.e., C18-amide) attached to silica particles was characterized by means of the linear solvation energy relationship model, which relates the chromatographic retention factor to specific solute interactions. The evaluationwas conducted under supercritical fluid chromatographic conditions using a mobile phase composition of carbon dioxide and methanol as co-solvent. The stationary phase showed to provide an alternate separation selectivity that is attractive to separate drug-like polar compounds in a relatively fast analysis time. PMID:27598573

  19. Separation performance of cucurbit[7]uril in ionic liquid-based sol-gel coating as stationary phase for capillary gas chromatography.

    PubMed

    Wang, Xiaogang; Qi, Meiling; Fu, Ruonong

    2014-12-01

    Here we report the separation performance of a new stationary phase of cucurbit[7]uril (CB7) incorporated into an ionic liquid-based sol-gel coating (CB7-SG) for capillary gas chromatography (GC). The CB7-SG stationary phase showed an average polarity of 455, suggesting its polar nature. Abraham system constants revealed that its major interactions with analytes include H-bond basicity (a), dipole-dipole (s) and dispersive (l) interactions. The CB7-SG stationary phase achieved baseline separation for a wide range of analytes with symmetrical peak shapes and showed advantages over the conventional polar stationary phase that failed to resolve some critical analytes. Also, it exhibited different retention behaviors from the conventional stationary phase in terms of retention times and elution order. Most interestingly, in contrast to the conventional polar phase, the CB7-SG stationary phase exhibited longer retentions for analytes of lower polarity but relatively comparable retentions for polar analytes such as alcohols and phenols. The high resolving ability and unique retention behaviors of the CB7-SG stationary phase may stem from the comprehensive interactions of the aforementioned interactions and shape selectivity. Moreover, the CB7-SG column showed good peak shapes for analytes prone to peak tailing, good thermal stability up to 280°C and separation repeatability with RSD values in the range of 0.01-0.11% for intra-day, 0.04-0.41% for inter-day and 2.5-6.0% for column-to-column, respectively. As demonstrated, the proposed coating method can simultaneously address the solubility problem with CBs for the intended purpose and achieve outstanding GC separation performance. PMID:25456602

  20. Preparation and characterization of mesoporous silicas modified with chiral selectors as stationary phase for high-performance liquid chromatography.

    PubMed

    Pérez-Quintanilla, Damián; Morante-Zarcero, Sonia; Sierra, Isabel

    2014-01-15

    New hybrid materials were prepared as novel chiral stationary phases (CSPs) for high-performance liquid chromatography (HPLC). Pure mesoporous silica (SM) and ethylene-bridged periodic mesostructured organosilica (PMO) were functionalized, by a post-synthesis method, with derivates of erythromycin and vancomycin. N2 adsorption-desorption measurements, XRD, FT-IR, MAS NMR, SEM, TEM and elemental analysis were used to characterize the physico-chemical properties of these mesostructured materials, before and after the modification process. The synthesized particles had non-symmetrical 3-D wormhole-like mesostructure, spherical morphology, and a mean pore diameter between 53 and 59 Å. CSPs prepared were tested for the separation of four chiral β-blockers (atenolol, metoprolol, pindolol and propranolol) in normal phase (NP) and polar organic phase (PO) elution modes. Much stronger chiral interaction was observed in vancomycin-modified silicas. Results obtained in these preliminary studies will permit in future works to improve the synthesis route in order to design mesoporous materials with better performance as a chiral stationary phase for HPLC. PMID:24231079

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

    SciTech Connect

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

    2015-12-29

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

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

    NASA Astrophysics Data System (ADS)

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

    2015-12-01

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

  3. Stationary phase modulation in liquid chromatography through the serial coupling of columns: A review.

    PubMed

    Alvarez-Segura, T; Torres-Lapasió, J R; Ortiz-Bolsico, C; García-Alvarez-Coque, M C

    2016-06-01

    Liquid chromatography with single columns often does not succeed in the analysis of complex samples, in terms of resolution and analysis time. A relatively simple solution to enhance chromatographic resolution is the modulation of the stationary phase through the serial coupling of columns. This can be implemented with any type of column using compatible elution conditions and conventional instruments. This review describes the key features of column coupling and published procedures, where two or more columns were coupled in series to solve separation problems. In all reports, the authors could not resolve their samples with single columns, whereas significant enhancement in chromatographic performance was obtained when the columns were combined. Particularly interesting is the reduction in the analysis time in the isocratic mode, which alleviates the "general elution problem" of liquid chromatography, and may represent a stimulus for the proposal of new procedures, especially in combination with mass spectrometric, electrochemical and refractometric detection. Developments proposed to make the serial coupling of columns useful in routine and research laboratories are outlined, including optimisation strategies that facilitate the selection of the appropriate column combination and elution conditions (solvent content, flow rate or temperature) in both isocratic and gradient modes. The availability of zero dead volume couplers, able to connect standard columns, and the commercialisation of short columns with multiple lengths, have expanded the possibilities of success. PMID:27155298

  4. Interactions between an idealized rotating string and stationary constraints

    NASA Astrophysics Data System (ADS)

    Yang, L.; Hutton, S. G.

    1995-08-01

    In this paper the dynamic interactions between an idealized rotating circular string and a stationary constraint, consisting of a spring, a damper, a mass or a frictional restraint, are discussed. The method of travelling waves is applied to develop the characteristic equation, and it is shown that this method is more general than the conventional method of separation of variables. The response of the string at its critical speed is examined, and it is shown that at the critical speed the string cannot vibrate freely. An approximate solution is generated using the Galerkin method to solve the eigenvalues effectively. The physics of the interactions between the string and the restraints is discussed on the basis of both analytical and approximate solutions.

  5. Multidimensional stationary probability distribution for interacting active particles.

    PubMed

    Maggi, Claudio; Marconi, Umberto Marini Bettolo; Gnan, Nicoletta; Di Leonardo, Roberto

    2015-01-01

    We derive the stationary probability distribution for a non-equilibrium system composed by an arbitrary number of degrees of freedom that are subject to Gaussian colored noise and a conservative potential. This is based on a multidimensional version of the Unified Colored Noise Approximation. By comparing theory with numerical simulations we demonstrate that the theoretical probability density quantitatively describes the accumulation of active particles around repulsive obstacles. In particular, for two particles with repulsive interactions, the probability of close contact decreases when one of the two particle is pinned. Moreover, in the case of isotropic confining potentials, the radial density profile shows a non trivial scaling with radius. Finally we show that the theory well approximates the "pressure" generated by the active particles allowing to derive an equation of state for a system of non-interacting colored noise-driven particles. PMID:26021260

  6. [Preparation and chromatographic performance of a eugenol-bonded silica gel stationary phase for high performance liquid chromatography].

    PubMed

    Xu, Lili; Zhong, Minghua; Chen, Xiaojing

    2015-05-01

    A eugenol-bonded silica gel stationary phase (EGSP) for high performance liquid chromatography ( HPLC) has been synthesized by the solid-liquid successive reaction method. The preparation process included two steps: firstly, γ-glycidoxypropyltrimethoxy-silane (KH-560) was covalently attached to the surface of spherical silica gel. Then the bonded silica gel continued to react with eugenol ligand, which was a plant active component, and obtained EGSP. The structure of EGSP was characterized by elemental analysis, thermogravimetric analysis and Fourier transform infrared spectroscopy. Using naphthalene as a probe, the column efficiency was tested under the mobile phase of acetonitrile-water (35:65, v/v) at a flow rate of 0.8 mL/min. The chromatographic properties and the retention mechanism of EGSP were evaluated by using neutral, basic and acidic analytes as solute probes. Meanwhile, the comparative study with C18 column and phenyl column was also carried out under the same chromatographic conditions. The result showed that the eugenol ligand was successfully bonded to the surface of silica gel with a 0.28 mmol/g of bonded amount, and the theoretical plate number of EGSP column was about 24 707 N/m. The EGSP appeared to be a kind of excellent reversed-phase stationary phase with suitable hydrophobicity and various synergistic sites. The eugenol ligand bonded on silica gel could first provide π-π interaction sites for different analytes because of its benzene ring and alkenyl. In addition, the methoxy groups of eugenol were responsible for dipole-dipole and hydrogen-bonding interactions between the ligand and solutes in the effective separation process. Comparing with traditional C18 column and phenyl column, EGSP has an advantage in the fast separation of polar compounds under simple experimental conditions. PMID:26387202

  7. Grain-grain interaction in stationary dusty plasma

    SciTech Connect

    Lampe, Martin; Joyce, Glenn

    2015-02-15

    We present a particle-in-cell simulation study of the steady-state interaction between two stationary dust grains in uniform stationary plasma. Both the electrostatic force and the shadowing force on the grains are calculated explicitly. The electrostatic force is always repulsive. For two grains of the same size, the electrostatic force is very nearly equal to the shielded electric field due to a single isolated grain, acting on the charge of the other grain. For two grains of unequal size, the electrostatic force on the smaller grain is smaller than the isolated-grain field, and the force on the larger grain is larger than the isolated-grain field. In all cases, the attractive shadowing force exceeds the repulsive electrostatic force when the grain separation d is greater than an equilibrium separation d{sub 0}. d{sub 0} is found to be between 6λ{sub D} and 9λ{sub D} in all cases. The binding energy is estimated to be between 19 eV and 900 eV for various cases.

  8. Evaluation and application of a mixed-mode chromatographic stationary phase in two-dimensional liquid chromatography for the separation of traditional Chinese medicine.

    PubMed

    Wei, Zhishen; Fu, Qing; Cai, Jianfeng; Huan, Liyun; Zhao, Jianchao; Shi, Hui; Jin, Yu; Liang, Xinmiao

    2016-06-01

    In this study, two mixed-mode chromatography stationary phases (C8SAX and C8SCX) were evaluated and used to establish a two-dimensional liquid chromatography system for the separation of traditional Chinese medicine. The chromatographic properties of the mixed-mode columns were systematically evaluated by comparing with other three columns of C8, strong anion exchanger, and strong cation exchanger. The result showed that C8SAX and C8SCX had a mixed-mode retention mechanism including electrostatic interaction and hydrophobic interaction. Especially, they were suitable for separating acidic and/or basic compounds and their separation selectivities could be easily adjusted by changing pH value. Then, several off-line 2D-LC systems based on the C8SAX in the first dimension and C8SAX, C8SCX, or C8 columns in the second dimension were developed to analyze a traditional Chinese medicine-Uncaria rhynchophylla. The two-dimensional liquid chromatography system of C8SAX (pH 3.0) × C8SAX (pH 6.0) exhibited the most effective peak distribution. Finally, fractions of U. rhynchophylla prepared from the first dimension were successfully separated on the C8SAX column with a gradient pH. Thus, the mixed-mode stationary phase could provide a platform to separate the traditional Chinese medicine in practical applications. PMID:27159545

  9. Separation of transition and heavy metals using stationary phase gradients and thin layer chromatography.

    PubMed

    Stegall, Stacy L; Ashraf, Kayesh M; Moye, Julie R; Higgins, Daniel A; Collinson, Maryanne M

    2016-05-13

    Stationary phase gradients for chelation thin layer chromatography (TLC) have been investigated as a tool to separate a mixture of metal ions. The gradient stationary phases were prepared using controlled rate infusion (CRI) from precursors containing mono-, bi-, and tri-dentate ligands, specifically 3-aminopropyltriethoxysilane, N-[3-(trimethoxysilyl)propyl] ethylenediamine, and N-[3-(trimethoxysilyl)propyl] diethylenetriamine. The presence and the extent of gradient formation were confirmed using N1s X-ray photoelectron spectroscopy (XPS). XPS results showed that the degree of modification was dependent on the aminosilane precursor, its concentration, and the rate of infusion. The separation of four transition and heavy metals (Co(2+), Pb(2+), Cu(2+), and Fe(3+)) on gradient and uniformly modified plates was compared using a mobile phase containing a stronger chelating agent, ethylenediaminetetraacetic acid (EDTA). The retention of the metal ions was manipulated by varying the surface concentration of the chelating ligands. The order of retention on unmodified plates and on plates modified with a monodentate ligand was Fe(3+)>Cu(2+)∼Pb(2+)∼Co(2+), while the order of retention on plates modified with bi- and tri-dentate ligands was Fe(3+)>Cu(2+)>Pb(2+)∼Co(2+). Fe(3+) and Cu(2+) were much more sensitive to the concentration of chelating ligand on the surface (displaying lower Rf values with increasing ligand concentration) than Pb(2+) and Co(2+). Complete separation was achieved using a high concentration of the tridentate ligand coupled with a longer time for modification, yielding a retention order of Fe(3+)>Cu(2+)>Co(2+)>Pb(2+). PMID:27090390

  10. Development of a V-shape bis(tetraoxacalix[2]arene[2]triazine) stationary phase for high performance liquid chromatography.

    PubMed

    Hu, Kai; Feng, Suxiang; Wu, Mingxia; Wang, Shuang; Zhao, Wenjie; Jiang, Qiong; Yu, Ajuan; Zhang, Shusheng

    2014-12-01

    A new stationary phase for high-performance liquid chromatography was prepared by covalently bonding a V-shape cage heteroatom-bridged calixarene onto silica gel using 3-aminopropyltriethoxysilane as coupling reagent. The structure of the new material was characterized by infrared spectroscopy, elemental analysis and thermogravimetric analysis. Linear solvation energy relationship method was successfully employed to evaluate the new phase with a set of 34 solutes. The retention characteristic of the new phase shows significant similarities with ODS, as well as distinctive features. Multiple mechanisms including hydrophobic, hydrogen bonding, π-π and n-π interaction are involved. The chromatographic behavior of the phase was illustrated by using alkylbenzenes, aromatics positional isomers and flavonoids as probes. Moreover, inorganic anions were individually separated in anion-exchange mode by using the same column. Thus, multi-interaction mechanisms and mixed-mode separation of the new phase can very likely guarantee its excellent chromatographic performance for the analysis of complex samples. The column has been successfully employed for the analysis of clenbuterol in animal urine, and it is demonstrated to be suitable and a competitive alternative analytical method for the determination of clenbuterol. PMID:25159380

  11. Fundamental and practical studies on high-performance liquid affinity chromatography of biopolymers with novel stationary phases

    SciTech Connect

    Bacolod, M.D.

    1992-01-01

    Rigid microparticulate stationary phases having surface-bound metal chelating functions were developed and evaluated in high performance metal chelate affinity chromatography of proteins. Silica- and polystyrene-divinylbenzene-based metal chelate sorbents were produced in wide pore and in non-porous type of column packings. A major effort has been placed on development of non-porous highly crosslinked polystyrene-divinylbenzene (PSDVB). These PSDVB microparticles were produced by a two-step swelling polymerization, and exhibited excellent mechanical strength over a wide range of flow-rates and composition used in high performance liquid chromatography (HPLC). Simple and reproducible hydrophilic coatings were developed for the surface modification of hydrophobic PSDVB supports. A tetradentate metal chelating ligand, ethylenediamine-N, N[prime]-diacetic acid (EDDA), was covalently bound to the surface of the various supports. Sorbents having iminodiacetic acid (IDA) metal chelating functions were also evaluated. The hydrophilic character and surface coverage of various stationary phases were assessed chromatographically. Studies concerning the effects of eluent pH as well as the nature and concentration of salts on retention and selectivity with different metal chelate stationary phases having various immobilized metal ions were carried out. Elution schemes were developed for rapid separation of proteins in metal chelate affinity chromatography. EDDA stationary phases in metal forms can be viewed as complementary to IDA stationary phases since they afforded different selectivity and retentivity toward proteins. Hydrophilic PSDVB could be functionalized with IDA or EDDA metal chelating ligands or lectins. The non-porous metal chelate stationary phases afforded rapid separation of proteins by the development of multiple gradient systems, which permitted higher column peak capacity, enabling the separation of a greater number of proteins in a single chromatographic run.

  12. Ultrahigh-pressure liquid chromatography of isoflavones and phenolic acids on different stationary phases.

    PubMed

    Klejdus, B; Vacek, J; Lojková, L; Benesová, L; Kubán, V

    2008-06-27

    Complete separation of aglycones and glucosides of selected isoflavones (genistin, genistein, daidzin, daidzein, glycitin, glycitein, ononin, sissotrin, formononetin, and biochanin A) was possible in 1.5 min using an ultrahigh-pressure liquid chromatography (U-HPLC) on a different particular chemically modified stationary phases with a particle size under 2 microm. In addition, selected separation conditions for simultaneous determination of isoflavones together with a group of phenolic acids (gallic, protocatechuic, p-hydroxybenzoic, vanillic, caffeic, syringic, p-coumaric, ferulic, and sinapic acid) allowed separation of all 19 compounds in 1.9 min. Separations were conducted on a non-polar reversed phase (C(18)) and also on more polar phases with cyanopropyl or phenyl groups using a gradient elution with a mobile phase consisting of 0.3% aqueous acetic acid and methanol. Chromatographic peaks were characterised using parameters such as resolution, symmetry, selectivity, etc. Individual substances were identified and quantified using UV-vis diode array detector at wavelength 270 nm. Limits of detection (3S/N) were in the range 200-400 pg ml(-1). Proposed U-HPLC technique was used for separation of isoflavones and phenolic acids in samples of plant materials (Trifolium pratense, Glycine max, Pisum sativum and Ononis spinosa) after acid hydrolysis of the samples and modified Soxhlet extraction. PMID:18501366

  13. Silica-Based, Hyper-Crosslinked Acid Stable Stationary Phases for High Performance Liquid Chromatography

    PubMed Central

    Zhang, Yu; Luo, Hao; Carr, Peter W.

    2011-01-01

    A new family of Hyper-Crosslinked (HC) phases has been recently introduced for use under very aggressive acid conditions including those encountered in ultra-fast, high temperature Two-Dimensional Liquid Chromatography (2DLC). This type of stationary phase showed significantly enhanced acid and thermal stability compared to the most acid stable, commercial RPLC phases. In addition, the use of “orthogonal” chemistry to make surface-confined polymer networks ensures good reproducibility and high efficiency. One of the most interesting features of the HC phases is the ability to derivatize the surface aromatic groups with various functional groups. This led to the development of a family of hyper-crosslinked phases possessing a wide variety of chromatographic selectivities by attaching hydrophobic (e.g. –C8), ionizable (e.g. -COOH, -SO3H), aromatic (e.g. –toluene) or polar (e.g. -OH) species to the aromatic polymer network. HC reversed phases with various degrees of hydrophobicity and mixed-mode HC phases with added strong and weak cation exchange sites have been synthesized, characterized and applied. These silica-based acid-stable HC phases, with their attractive chromatographic properties, should be very useful in the separations of bases or biological analytes in acidic media, especially at elevated temperatures. This work reviews the prior research on HC phases and introduces a novel HC phase made by alternative chemistry. PMID:21906745

  14. A fullerene C60-based ligand in a stationary phase for affine chromatography of membrane porphyrin-binding proteins

    NASA Astrophysics Data System (ADS)

    Amirshakhi, N.; Alyautdin, R. N.; Orlov, A. P.; Poloznikov, A. A.; Kuznetsov, D. A.

    2008-11-01

    A new affine chromatography technique is suggested for the purification of porphyrin-binding proteins (PBP) from mammal cell membranes. The procedure uses new fullerene-porphyrin ligands immobilized on agarose and bound to the polysaccharide matrix via the epoxycyclohexyl residue. A selective PBP stationary phase was used in a single-column chromatography run for the complete purification of a monomeric protein (17.6 kDa) from mitochondrial membranes of rat myocardium. This protein was characterized by high affinity for porphyrin-related structures. To separate it from other nonspecifically sorbed membrane proteins, synchronous linear pH and ionic strength gradients were used.

  15. Dynamic Behavior of Clobazam on High-Performance Liquid Chromatography Chiral Stationary Phases.

    PubMed

    Sabia, Rocchina; De Martino, Michela; Cavazzini, Alberto; Villani, Claudio

    2016-01-01

    Clobazam, a 1,5-benzodiazepin-2,4-dione, is a chiral molecule because its ground state conformation features a nonplanar seven-membered ring lacking reflection symmetry elements. The two conformational enantiomers of clobazam interconvert at room temperature by a simple ring-flipping process. Variable temperature HPLC on the Pirkle type (R)-N-(3,5-dinitronenzoyl)phenylglycine and (R,R)-Whelk-O1 chiral stationary phases (CSPs) allowed us to separate for the first time the conformational enantiomers of clobazam and to observe peak coalescence-decoalescence phenomena due to concomitant separation and interconversion processes occurring on the same time scale. Clobazam showed temperature dependent dynamic high-performance liquid chromatography (HPLC) profiles with interconversion plateaus on the two CSPs indicative of on-column enantiomer interconversion. (enantiomerization) in the column temperature range between Tcol = 10°C and Tcol = 30°C, whereas on-column interconversion was absent at temperature close to or lower than Tcol = 5°C. Computer simulation of exchange-deformed HPLC profiles using a program based on the stochastic model yielded the apparent rate constants for the on-column enantiomerization and the corresponding free energy activation barriers. At Tcol = 20°C the averaged enantiomerization barriers, ΔG(‡), for clobazam were found in the range 21.08-21.53 kcal mol(-1) on the two CSPs. The experimental dynamic chromatograms and the corresponding interconversion barriers reported in this article are consistent with the literature data measured by DNMR at higher temperatures and in different solvents. PMID:26477466

  16. Chiral stationary phases based on chitosan bis(methylphenylcarbamate)-(isobutyrylamide) for high-performance liquid chromatography.

    PubMed

    Tang, Sheng; Bin, Qin; Chen, Wei; Bai, Zheng-Wu; Huang, Shao-Hua

    2016-04-01

    A series of chitosan bis(methylphenylcarbamate)-(isobutyrylamide) derivatives were synthesized by carbamylating chitosan isobutyrylamide with different methylphenyl isocyanates. Then the prepared chitosan derivatives were coated onto 3-aminopropyl silica particles, resulting in a series of new chiral stationary phases (CSPs) for high-performance liquid chromatography. It was observed that the chiral recognition abilities of these coated-type CSPs depended very much on the substituents on the phenyl moieties of the chitosan derivatives, the eluent composition, as well as the structure of racemates. As a typical example, the eluent tolerance of the prepared CSP with the best enantioseparation ability was investigated in detail, and the results revealed that the CSP exhibited extraordinary solvent tolerance and could still work without significant loss in enantioseparation capability after being flushed with chloroform (100%), ethyl acetate (100%) and even THF/n-hexane (70/30, v/v), while the traditional coated-type CSPs based on the cellulose and amylose derivatives, such as cellulose tris(3,5-dimethylphenylcarbamate) (CDMPC) and amylose tris(3,5-dimethylphenylcarbamate) (ADMPC), might be dissolved or highly swollen in these eluents. Therefore, the application of the resultant CSPs could address the problem of the dissolution and high swelling of traditional coated-type CSPs in some unusual eluents, broadening the possibility of eluent choice. In addition, a comparison of the prepared CSPs with the well known CDMPC- and ADMPC- based CSPs concerning the chiral recognition ability was also made. Separation performances achieved on the as-prepared CSPs in different eluents were found to be even superior to CDMPC- and ADMPC-based CSPs for the tested chiral compounds. In summary, we could safely draw the conclusion that the CSPs derived from chitosan isobutyrylamide derivatives were capable of excellent chiral recognition ability, and meanwhile possessed satisfactory

  17. Hydrophilic interaction chromatography for the analysis of aminoglycosides.

    PubMed

    Kumar, Praveen; Rubies, Antoni; Companyó, Ramon; Centrich, Francesc

    2012-02-01

    The effect of mobile-phase constituents (pH and ionic strength) and chromatographic behaviour of ten aminoglycosides (streptomycin, dihydrostreptomycin, spectinomycin, apramycin, paramomycin, kanamycin A, gentamycin C1, gentamycin C2/C2a, gentamycin C1a and neomycin) in the bare silica, amino, amide and zwitterionic phases of hydrophilic interaction chromatography (HILIC) were studied systematically. Among the stationary phases studied, the zwitterionic phase provided the best separation of aminoglycosides. The effect of pH, ionic concentration and column temperature on retention time, peak shape and sensitivity was studied using a central composite design. pH affected sensitivity of the detection of analytes but not the retention time. High ionic strength in the mobile phase was necessary to control the ionic interactions between ionised aminoglycosides and the hydrophilic phase, thereby influencing peak shape and retention time. Column temperature affected sensitivity of the detection but not the retention time. During method development, crosstalk between the MS/MS channels of the analytes was observed and resolved. PMID:22282410

  18. A novel urea-functionalized surface-confined octadecylimidazolium ionic liquid silica stationary phase for reversed-phase liquid chromatography.

    PubMed

    Zhang, Mingliang; Tan, Ting; Li, Zhan; Gu, Tongnian; Chen, Jia; Qiu, Hongdeng

    2014-10-24

    One-pot synthesis of surface-confined ionic liquid functionalized silica spheres was proposed using N-(3-aminopropyl)imidazole, γ-isopropyltriethoxysilane and 1-bromooctadecane as starting materials. The surface modification of the silica spheres was successful with a high surface density of octadecylimidazolium, enabling the utilization of this new urea-functionalized ionic liquid-grafted silica material as stationary phase for high-performance liquid chromatography in reversed-phase mode. The long aliphatic chain combined with the multiple polar group embedded in the ligands imparted the new stationary phase fine selectivity towards PAH isomers and polar aromatics and higher affinity for phenolic compounds. The unique features of the new material, especially the effect of the urea group on the retention were elucidated by mathematic modeling. PMID:25249489

  19. A micro gas chromatography with separation capability enhanced by polydimethylsiloxane stationary phase functionalized by carbon nanotubes and graphene.

    PubMed

    Li, Yubo; Zhang, Runzhou; Wang, Tao; Wang, Youhao; Wang, Yonghuan; Li, Lingfeng; Zhao, Weijun; Wang, Xiaozhi; Luo, Jikui

    2016-07-01

    Polydimethylsiloxane (PDMS) stationary phases functionalized with multi-walled carbon nanotubes (MWCNTs) and graphene, respectively, for the columns in micro gas chromatography are presented in this paper. To exploit the merits of MWCNTs and graphene in terms of their high specific surface area, low surface energy and chemical inertness, experimental conditions for separation (heating rate and final temperature of temperature programming, flow rate of carrier gas and the volume of samples injection) are investigated, and separations of both polar and nonpolar compound mixtures under these conditions are performed. Compared with PDMS-only coated stationary phases, the functionalization of the phases with carbon nano-materials improves the performance of columns in separation, repeatability, stability and revolution significantly. PMID:27154654

  20. Orthogonal separation on one beta-cyclodextrin column by switching reversed-phase liquid chromatography and hydrophilic interaction chromatography.

    PubMed

    Feng, Jia-tao; Guo, Zhi-mou; Shi, Hui; Gu, Jiang-ping; Jin, Yu; Liang, Xin-miao

    2010-06-15

    A dual retention combined with reversed-phase liquid chromatography (RP-LC) and hydrophilic interaction chromatography (HILIC) has been observed on beta-cyclodextrin (beta-CD) bonded stationary phase. A typical U-shaped retention curve was achieved owing to dual retention mechanism. Based on this observation, a beta-CD column can be operated under reversed-phase liquid chromatography (RP-LC) and hydrophilic interaction chromatography (HILIC) modes. Two-dimensional liquid chromatography (2D-LC) analysis can be realized on just a beta-CD column by switching these two different separation modes. In this study, off-line 2D-LC analysis for a natural product was carried out to prove the orthogonal separation between RP-LC and HILIC modes on a Click beta-CD column. Herba Hedyotis Diffusae, the whole grass of Hedyotis Diffusae wild was extracted with water, pretreated with macroporous resin and then first separated at RP-LC mode on the Click beta-CD column to obtain successive fractions, which were then reanalyzed at HILIC mode on the same Click beta-CD column. The result proved that both separation modes on the Click beta-CD column have good retention and peak shape, and these two separation modes have good orthogonality. 2D-LC analysis revealed abundant information in the natural product. Especially numerous minor components were enriched and separated. The mobile phase used in RP-LC and HILIC modes can be same and the switch between these two separation modes is easily realized by changing the ratio of the acetonitrile and water. Hence the mobile phase in this 2D-LC system is completely compatible. This advantage makes this combination is an appropriate 2D-LC method for the solutes having retention at both separation modes. PMID:20441989

  1. Ultra high performance liquid chromatography versus high performance liquid chromatography: stationary phase selectivity for generic carotenoid screening.

    PubMed

    Bijttebier, Sebastiaan; D'Hondt, Els; Noten, Bart; Hermans, Nina; Apers, Sandra; Voorspoels, Stefan

    2014-03-01

    Aim of study was to find the most suitable LC column for generic carotenoid screening. To represent the diversity of carotenoids in nature and to optimize chromatographic separation, a set of carotenoid standards was carefully chosen to account for the various classes of carotenoids. The HPLC C30 column has since long been the 'golden standard' in the chromatographic separation of carotenoids. Since approximately one decade, new UHPLC technology has led to much shorter analysis times, smaller peak widths and higher chromatographic resolution. However, there are currently no UHPLC columns on the market containing the specific stationary phase chemistry of the HPLC C30 column. Therefore during this study, we investigated the separation of carotenoids on a set of UHPLC columns and compared it to their separation on the HPLC C30 column. Comparison of carotenoids separations on the different stationary phases with objective column comparison parameters clearly indicated that the HPLC C30 column is an overall better performer in the separation of carotenoids. This is due to the lack of UHPLC column chemistries that are adapted for carotenoid analysis. However, analysis time on the HPLC C30 column takes about four times longer compared to UHPLC analysis. Therefore, with the range of columns that are commercially available nowadays, a choice has to be made between very high selectivity (HPLC C30 column) and analysis times that are adapted to modern laboratory requirements (UHPLC technology). Therefore, carotenoid separations would be even more performing if an appropriate UHPLC C30 column would be available. PMID:24534422

  2. Thermodynamic study of molecularly imprinted polymer used as the stationary phase in high performance liquid chromatography.

    PubMed

    Denderz, Natalia; Lehotay, Jozef; Cižmárik, Jozef; Cibulková, Zuzana; Simon, Peter

    2012-04-27

    Molecularly imprinted polymer (MIP) and non-imprinted polymer (NIP) on the base of methacrylic acid prepared by a bulk polymerization were used as stationary phases for the HPLC analysis. The thermodynamic processes were carried out to investigate the temperature effects during sorption processes of potential local anaesthetics - morpholinoethyl esters of alkoxy-substituted phenylcarbamic acid (MEsP), local anaesthetic - diperodon, flavonoid - quercetin in methanol, acetonitrile and toluene (porogen) as mobile phases. Mobile phases and corresponding solvents were selected according to the solubility of each analyte. The template was chosen from the set of homologous of MEsP - 2-(morpholin-4-yl)ethyl (2-methoxyphenyl)carbamate. Values of retention factors were measured over the temperature range of 20-60°C. There were determined van't Hoff curves - dependences between logarithms of the retention factors (lnk) and the inverse value of the temperature (1/T). Observed graphs were linear directly indicating that there were no changes of interaction mechanisms in the studied range of temperature. Selectivities (evaluated by the separation factors, α) and sorption selectivities (evaluated by the imprinting factors, IFs) of the MIP and the NIP toward template, related and not-related structures with the template were evaluated chromatographically. The highest separation factors and the imprinting factors (IF=4.73 ± 0.35 for the template) were observed in methanol, not in porogen. Only in the case of quercetin the highest IF was observed in ACN (1.88 ± 0.13). Contrary to expectations, the driving force for the affinity of the target molecules for both of polymers was enthalpic term (with an average of 54%, 82% and 84% contribution of enthalpic term for MeOH, ACN and toluene, respectively on the MIP and 53%, 57% and 65% for MeOH, ACN and toluene, respectively on the NIP). The MIP and NIP were also characterized by attenuated total reflectance analysis Fourier transform

  3. Quasi-stationary states and a classification of the range of pair interactions

    SciTech Connect

    Gabrielli, A.; Joyce, M.; Marcos, B.

    2011-03-24

    Systems of long-range interacting particles present typically 'quasi-stationary' states (QSS). Investigating their lifetime for a generic pair interaction V(r{yields}{infinity}){approx}1/r{sup {gamma}} we give a classification of the range of the interactions according to the dynamical properties of the system.

  4. Size exclusion chromatography of synthetic polymers and biopolymers on common reversed phase and hydrophilic interaction chromatography columns.

    PubMed

    Caltabiano, Anna M; Foley, Joe P; Barth, Howard G

    2016-03-11

    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. PMID:26877177

  5. Quantitative analysis of molecular interaction potentials of ionic liquid anions using multi-functionalized stationary phases in HPLC.

    PubMed

    Cho, Chul-Woong; Stolte, Stefan; Ranke, Johannes; Preiss, Ulrich; Krossing, Ingo; Thöming, Jorg

    2014-08-01

    The molecular interaction potentials, including S (dipolarity/polarizability), A (hydrogen bonding acidity), and B (hydrogen bonding basicity), of anions are experimentally determined using multi-functionalized stationary phases in high-performance liquid chromatography (HPLC) systems. We employ three different multi-functionalized stationary phase columns (Obelisc R, Obelisc N, and Acclaim Trinity-P1) combined with two ingredients, namely, acetonitrile (ACN) and methanol (MeOH). These conditions can cause neutral, cationic, and anionic compounds to be retained. By using the retention characteristics of calibration compounds, including cations, anions, and neutral compounds, system parameters including the ionic interaction terms (zc Zc , za Za ) are evaluated using multiple linear regression, resulting in a standard deviation (SD) of 0.090-0.158 log units. Based on the system parameters and retention characteristics of the anions of interest, their molecular interaction potentials are characterized on the same scale for neutral and cationic molecules. Furthermore, to verify the determined molecular interaction potentials, we predict anion hydrophobicity. The results show that the determined S, A, and B, together with the computable descriptors E (excess molar refraction) and V (McGowan volume), can predict anion hydrophobicity with R(2) =0.982 and SD=0.167 (dimensionless). PMID:24850224

  6. Enhanced resolution of Mentha piperita volatile fraction using a novel medium-polarity ionic liquid gas chromatography stationary phase.

    PubMed

    Ragonese, Carla; Sciarrone, Danilo; Grasso, Elisa; Dugo, Paola; Mondello, Luigi

    2016-02-01

    The evaluation of a novel medium-polarity ionic-liquid-based gas chromatography column, SLB-IL60, towards the analysis of a complex essential oil, namely, a peppermint essential oil sample, is reported. The SLB-IL60 30 m column was subjected to bleeding measurements, by means of conventional gas chromatography with mass spectrometry. The SLB-IL60 column was then evaluated in the analysis of pure standard compounds, chosen as typical constituents of peppermint essential oil. Resolution and peak symmetry (expressed as tailing factors at 10% of peak height) were measured and the results were compared to those obtained on the most widely used columns in such an application, namely a medium-polarity [100% poly(ethyleneglycol)] stationary phase, and an apolar 5% diphenyl/95% dimethyl siloxane. The final part of the evaluation was dedicated to the gas chromatography with mass spectrometry analysis of a peppermint essential oil sample and again the data were compared to those obtained on the 100% poly(ethyleneglycol) and the 5% diphenyl/95% dimethyl siloxane phase. Linear retention indices were determined for all the identified components on the ionic liquid capillary. PMID:26613675

  7. Thermodynamics of the sorption of organic compounds on polyethylene glycol 400-permethylated β-cyclodextrin stationary phase and its enantioselectivity in gas chromatography

    NASA Astrophysics Data System (ADS)

    Kuraeva, Yu. G.; Onuchak, L. A.; Evdokimova, M. A.

    2016-08-01

    The thermodynamic characteristics of sorption of 24 organic compounds of various classes from the gas phase on the binary stationary phase based on polyethylene glycol 400 and permethylated β-cyclodextrin were determined. The influence of geometrical structure and optical activity of organic compounds on the possibility of forming sorbate-macrocycle complexes was examined. It was found that the studied stationary phase shows the enantioselectivity towards low-polar terpenes under the conditions of gas chromatography.

  8. Polystyrene-divinylbenzene-glycidyl methacrylate stationary phase grafted with poly (amidoamine) dendrimers for ion chromatography.

    PubMed

    Guo, Dandan; Lou, Chaoyan; Zhang, Peimin; Zhang, Jiajie; Wang, Nani; Wu, Shuchao; Zhu, Yan

    2016-07-22

    In this work, a novel ion exchange stationary phase based on different generations of poly (amidoamine) dendrimers (PAMAM) was developed for the determination of inorganic anions and carbohydrates. Synthesis of the PAMAM was carried out with the polymerization reaction of ethylenediamine and methyl acrylate. The synthesized PAMAM was then grafted to the polystyrene-divinylbenzene-glycidyl methacrylate (PS-GMA) to form PAMAM-based beads. These beads were finally modified with 1,4-butanediol diglycidyl ether (BDDE) to generate the anion exchanger, which were characterized by scanning electron microscopy (SEM), brunauer-emmett-teller (BET), fourier transform infrared spectroscopy (FTIR), and elemental analysis. Elemental analysis, breakthrough curves and capacity factors showed that more epoxy groups and higher PAMAM generations in stationary phase could result in higher anion exchange capacity. The efficiency, durability and stability of the proposed anion exchanger were investigated by using six inorganic anions (fluoride, chloride, nitrite, bromide, nitrate and sulfate) and four carbohydrates (trehalose, glucose, maltotriose and galacturonic acid) as analytes, respectively. The reliability of the proposed ion chromatographic stationary phase was demonstrated by determining the content of galacturonic acid in polysaccharides from Poria cocos and Atractylodes macrocephala. The relative standard deviations of retention time, peak height, and peak area for galacturonic acid were 0.39%, 1.22%, and 2.02%, respectively. The spiked recoveries were in the range of 88.29%-100.51% for plant polysaccharides. Due to the good structural homogeneity, intense internal porosity, biological compatibility and high density of active groups in PAMAM, this grafted stationary phase showed good ion-exchange characteristics, especially in biological charged molecules. PMID:27311659

  9. Preparation and characterization of a new microwave immobilized poly(2-phenylpropyl)methylsiloxane stationary phase for reversed phase high-performance liquid chromatography.

    PubMed

    Begnini, Fernanda R; Jardim, Isabel C S F

    2013-07-01

    A new reversed phase high-performance liquid chromatography (RP-HPLC) stationary phase was prepared and its chromatographic and physical-chemical properties were evaluated. The new stationary phase was prepared with a silica support and poly(2-phenylpropyl)methylsiloxane (PPPMS), a phenyl type polysiloxane copolymer. Since this is a new copolymer and there is little information in the literature, it was submitted to physical-chemical characterization by infrared spectroscopy and thermogravimetry. The chromatographic phase was prepared through sorption and microwave immobilization of the copolymer onto a silica support. The chromatographic performance was evaluated by employing test procedures suggested by Engelhardt and Jungheim, Tanaka and co-workers, Neue, and Szabó and Csató. These test mixtures provide information about the hydrophobic selectivity, silanophilic activity, ion-exchange capacity, shape selectivity and interaction with polar analytes of the new Si-PPPMS reversed phase. Stability tests were developed using accelerated aging tests under both basic and acidic conditions to provide information about the lifetime of the packed columns. PMID:23726242

  10. Nanoengineered analytical immobilized metal affinity chromatography stationary phase by atom transfer radical polymerization: Separation of synthetic prion peptides

    PubMed Central

    McCarthy, P.; Chattopadhyay, M.; Millhauser, G.L.; Tsarevsky, N.V.; Bombalski, L.; Matyjaszewski, K.; Shimmin, D.; Avdalovic, N.; Pohl, C.

    2010-01-01

    Atom transfer radical polymerization (ATRP) was employed to create isolated, metal-containing nanoparticles on the surface of non-porous polymeric beads with the goal of developing a new immobilized metal affnity chromatography (IMAC) stationary phase for separating prion peptides and proteins. Transmission electron microscopy was used to visualize nanoparticles on the substrate surface. Individual ferritin molecules were also visualized as ferritin–nanoparticle complexes. The column's resolving power was tested by synthesizing peptide analogs to the copper binding region of prion protein and injecting mixtures of these analogs onto the column. As expected, the column was capable of separating prion-related peptides differing in number of octapeptide repeat units (PHGGGWGQ), (PHGGGWGQ)2, and (PHGGGWGQ)4. Unexpectedly, the column could also resolve peptides containing the same number of repeats but differing only in the presence of a hydrophilic tail, Q → A substitution, or amide nitrogen methylation. PMID:17481564

  11. Purification of C70 using charcoal as a stationary phase in a flash chromatography column. Technical report

    SciTech Connect

    Scrivens, W.A.; Cassell, A.M.; Kinsey, K.E.; Tour, J.M.

    1995-06-07

    Described is a method for the purification of C60 and C70 using a flash chromatography column that contains charcoal as the stationary phase. A number of functionalized aromatic solvents are studied and their efficacy for extraction, NMR spectral acquisition, and chromatographic purification of fullerenes is discussed. Ortho-dichlorobenzene was chosen as the best solvent for these applications and examples of its use in the extraction of higher fullerenes (>C84) and in the rapid acquisition of (13)C NMR spectra are given. Finally, single column purification of both C60 and C70 is discussed. Starting with a typical arc-derived mixture of soluble fullerenes, 5.97 g of C60 at >99.9% purity and 1.58 g of C70 at >97% purity were produced in a single column pass.

  12. Silica-based polypeptide-monolithic stationary phase for hydrophilic chromatography and chiral separation.

    PubMed

    Zhao, Licong; Yang, Limin; Wang, Qiuquan

    2016-05-13

    Glutathione (GSH)-, somatostatin acetate (ST)- and ovomucoid (OV)-functionalized silica-monolithic stationary phases were designed and synthesized for HILIC and chiral separation using capillary electrochromatography (CEC). GSH, ST and OV were covalently incorporated into the silica skeleton via the epoxy ring-opening reaction between their amino groups and the glycidyl moiety in γ-glycidoxypropyltrimethoxysilane (GPTMS) together with polycondensation and copolymerization of tetramethyloxysilane and GPTMS. Not only could the direction and electroosmotic flow magnitude on the prepared GSH-, ST- and OV-silica hybrid monolithic stationary phases be controlled by the pH of the mobile phase, but also a typical HILIC behavior was observed so that the nucleotides and HPLC peptide standard mixture could be baseline separated using an aqueous mobile phase without any acetonitrile during CEC. Moreover, the prepared monolithic columns had a chiral separation ability to separate dl-amino acids. The OV-silica hybrid monolithic column was most effective in chiral separation and could separate dl-glutamic acid (Glu) (the resolution R=1.07), dl-tyrosine (Tyr) (1.57) and dl-histidine (His) (1.06). Importantly, the chiral separation ability of the GSH-silica hybrid monolithic column could be remarkably enhanced when using gold nanoparticles (AuNPs) to fabricate an AuNP-mediated GSH-AuNP-GSH-silica hybrid monolithic column. The R of dl-Glu, dl-Tyr and dl-His reached 1.19, 1.60 and 2.03. This monolithic column was thus applied to separate drug enantiomers, and quantitative separation of all four R/S drug enantiomers were achieved with R ranging from 4.36 to 5.64. These peptide- and protein-silica monolithic stationary phases with typical HILIC separation behavior and chiral separation ability implied their promise for the analysis of not only the future metabolic studies, but also drug enantiomers recognition. PMID:27083263

  13. Evaluation of innovative stationary phase ligand chemistries and analytical conditions for the analysis of basic drugs by supercritical fluid chromatography.

    PubMed

    Desfontaine, Vincent; Veuthey, Jean-Luc; Guillarme, Davy

    2016-03-18

    Similar to reversed phase liquid chromatography, basic compounds can be highly challenging to analyze by supercritical fluid chromatography (SFC), as they tend to exhibit poor peak shape, especially those with high pKa values. In this study, three new stationary phase ligand chemistries available in sub -2μm particle sizes, namely 2-picolylamine (2-PIC), 1-aminoanthracene (1-AA) and diethylamine (DEA), were tested in SFC conditions for the analysis of basic drugs. Due to the basic properties of these ligands, it is expected that the repulsive forces may improve peak shape of basic substances, similarly to the widely used 2-ethypyridine (2-EP) phase. However, among the 38 tested basic drugs, less of 10% displayed Gaussian peaks (asymmetry between 0.8 and 1.4) using pure CO2/methanol on these phases. The addition of 10mM ammonium formate as mobile phase additive, drastically improved peak shapes and increased this proportion to 67% on 2-PIC. Introducing the additive in the injection solvent rather than in the organic modifier, gave acceptable results for 2-PIC only, with 31% of Gaussian peaks with an average asymmetry of 1.89 for the 38 selected basic drugs. These columns were also compared to hybrid silica (BEH), DIOL and 2-EP stationary phases, commonly employed in SFC. These phases commonly exhibit alternative retention and selectivity. In the end, the two most interesting ligands used as complementary columns were 2-PIC and BEH, as they provided suitable peak shapes for the basic drugs and almost orthogonal selectivities. PMID:26895829

  14. Biotin-functionalized poly(ethylene terephthalate) capillary-channeled polymer fibers as HPLC stationary phase for affinity chromatography.

    PubMed

    Jiang, Liuwei; Marcus, R Kenneth

    2015-01-01

    Native poly(ethylene terephthalate) (PET) capillary-channeled polymer (C-CP) fibers have been used as the stationary phase for high-performance liquid chromatography (HPLC) of proteins via reversed-phase and ion-exchange processes. Functionalization can be used to bring about greater selectivity through surface modification. PET fibers were treated with ethylenediamine to generate primary amine groups on the fiber surface, enabling subsequent covalent attachment of ligands. The ninhydrin test for primary amines revealed surface densities of 13.9-60.0 μmol m(-2) for PET fibers exposed for periods of 3-12 min. Here, 8-amino-3,6-dioxaoctanoic acid was linked to the EDA-treated PET fiber surface as a hydrophilic spacer, and then D-biotin was attached on the end of the spacer as an affinity ligand. The streptavidin binding capacity and binding homogeneity were studied on the biotin-functionalized PET C-CP fiber microbore column. The selectivity of the biotin surface functionalization was assessed by spiking lysate with Texas Red-labeled streptavidin and enhanced green fluorescent protein. Greater than 99% selectivity was realized. This ligand-coupling strategy from standard solid-phase peptide synthesis used in stationary phase functionalization creates great potential for PET C-CP fiber-packed HPLC columns to perform a variety of chromatographic separations. PMID:25410640

  15. Automated screening of reversed-phase stationary phases for small-molecule separations using liquid chromatography with mass spectrometry.

    PubMed

    Appulage, Dananjaya K; Wang, Evelyn H; Carroll, Frances; Schug, Kevin A

    2016-05-01

    There are various reversed-phase stationary phases that offer significant differences in selectivity and retention. To investigate different reversed-phase stationary phases (aqueous stable C18 , biphenyl, pentafluorophenyl propyl, and polar-embedded alkyl) in an automated fashion, commercial software and associated hardware for mobile phase and column selection were used in conjunction with liquid chromatography and a triple quadrupole mass spectrometer detector. A model analyte mixture was prepared using a combination of standards from varying classes of analytes (including drugs, drugs of abuse, amino acids, nicotine, and nicotine-like compounds). Chromatographic results revealed diverse variations in selectivity and peak shape. Differences in the elution order of analytes on the polar-embedded alkyl phase for several analytes showed distinct selectivity differences compared to the aqueous C18 phase. The electron-rich pentafluorophenyl propyl phase showed unique selectivity toward protonated amines. The biphenyl phase provided further changes in selectivity relative to C18 with a methanolic phase, but it behaved very similarly to a C18 when an acetonitrile-based mobile phase was evaluated. This study shows the value of rapid column screening as an alternative to excessive mobile phase variation to obtain suitable chromatographic settings for analyte separation. PMID:26959840

  16. Toward enantioselective nano ultrahigh-performance liquid chromatography with Whelk-O1 chiral stationary phase.

    PubMed

    Ciogli, Alessia; Pierri, Giuseppe; Kotoni, Dorina; Cavazzini, Alberto; Botta, Lorenzo; Villani, Claudio; Kocergin, Jelena; Gasparrini, Francesco

    2014-10-01

    In this study, a Whelk-O1 chiral stationary phase immobilized on 2.5 μm silica particles was employed in nanoLC. Two nanocolumns (180 and 250 mm long, 75 μm id) with a single polymeric organic monolithic outlet frit were packed under high-pressure ultrasonic-assisted packing procedure. The monolithic outlet frit was prepared by thermal polymerization of methacrylate-based monomers affording high-mechanical stability and high-pressure resistance. Very efficient enantioseparations with more than 70 000 plates/m were achieved in normal phase mode by eluting (+/-) acenaphthenol. Nanocolumns were also tested in RP mode by using on-line MS detection with nano-spray ESI ion source. Kinetic performances of columns in RP mode were comparable to those in normal phase-conditions. PMID:25043154

  17. Stationary entanglement between two nanomechanical oscillators induced by Coulomb interaction

    NASA Astrophysics Data System (ADS)

    Qin, Wu; Yin, Xiao; Zhi-Ming, Zhang

    2016-01-01

    We propose a scheme for entangling two nanomechanical oscillators by Coulomb interaction in an optomechanical system. We find that the steady-state entanglement of two charged nanomechanical oscillators can be obtained when the coupling between them is stronger than a critical value which relies on the detuning. Remarkably, the degree of entanglement can be controlled by the Coulomb interaction and the frequencies of the two charged oscillators. Project supported by the Major Research Plan of the National Natural Science Foundation of China (Grant No. 91121023), the National Natural Science Foundation of China (Grant Nos. 61378012, 60978009, and 11574092), the Specialized Research Fund for the Doctoral Program of Higher Education of China (Grant No. 20124407110009), the National Basic Research Program of China (Grant Nos. 2011CBA00200 and 2013CB921804), and the Program for Changjiang Scholar and Innovative Research Team in University, China (Grant No. IRT1243).

  18. Improving selectivity in gas chromatography by using chemically modified multi-walled carbon nanotubes as stationary phase.

    PubMed

    Speltini, Andrea; Merli, Daniele; Dondi, Daniele; Paganini, Giorgio; Profumo, Antonella

    2012-05-01

    Amino-terminated alkyl MWCNTs (MWCNTs-R-NH(2)), synthesized by chemical modification of the nanotube skeleton by nucleophilic substitution with 2,2'-(ethylenedioxy)diethylamine, were successfully used as stationary phases for gas chromatographic separation of esters and chloroaromatics. The presence of alkyl chains with polar embedded groups made the functionalized MWCNTs (f-MWCNTs) a mixed-mode GC separation material able to interact in different ways with the analytes. Compared with non-functionalized MWCNTs (nf-MWCNTs), MWCNTs-R-NH(2) had higher selectivity, enhanced resolution, and optimum retention behaviour, and they were proved to perform better than the commercial stationary phase Porapak QS (PQS), claimed to be suitable for similar applications. The so-prepared stationary phase was used for analysis of a synthetic mixture containing different classes of analytes, viz. esters, ketones, alcohols, alkanes, and aromatic hydrocarbons, and finally used for investigation of similar real matrices. In particular, the constituents of a commercial paint thinner were determined by direct injection of the sample, with good reproducibility (inter-day precision RSDs from 5 to 19%). Two unknown samples of commercial white spirit were also analysed for determination of the aromatic hydrocarbon content, and their composition was profiled on the basis of the different compounds identified. PMID:22234403

  19. Aminoglycoside analysis in food of animal origin with a zwitterionic stationary phase and liquid chromatography-tandem mass spectrometry.

    PubMed

    Díez, Cristina; Guillarme, Davy; Staub Spörri, Aline; Cognard, Emmanuelle; Ortelli, Didier; Edder, Patrick; Rudaz, Serge

    2015-07-01

    In this study, fourteen highly polar aminoglycoside (AGs) antibiotics were selected. Various stationary phases were tested, including Obelisc R, ZIC-HILIC, BEH amide and aminopropyl. The nature of the stationary phase, mobile phase (water or buffer solutions and acetonitrile), pH (percentage of formic acid), gradient conditions and injection solvents were systematically studied as relevant parameters for tuning retention selectivity and detectability of AGs in liquid chromatography electrospray tandem mass spectrometry (LC-(ESI)-MS/MS). Only the two zwitterionic phases (Obelisc R and ZIC-HILIC) achieved a proper chromatographic separation considering interferences due to the crosstalk effect in low resolution mass spectrometers. The water/acetonitrile mobile phase containing 1% formic acid used with Obelisc R provided more sensitivity than the highly concentrated buffered mobile phases required for ZIC-HILIC. A solid phase extraction (SPE) clean-up procedure with polymeric weak cation exchange (WCX) cartridges was optimized for honey, milk and liver samples. Different brands of cartridges and elution solvents were tested, and the Taurus WCX offered the best recovery rate with a buffer elution at pH 3. The final optimized method was validated in these matrices according to Decision 2002/657/EC. A monitoring campaign for sixty honey, milk and liver samples was carried out at the Food Authority Control in Geneva. The concentration of dihydrostreptomycin (DSTP) found in one ovine liver exceeded the established maximum residue levels (MRLs) within the European and Swiss legislations but it was compliant taking into account the validation data. PMID:26043099

  20. Porous silica particles grafted with an amphiphilic side-chain polymer as a stationary phase in reversed-phase high-performance liquid chromatography.

    PubMed

    Shahruzzaman, Md; Takafuji, Makoto; Ihara, Hirotaka

    2015-07-01

    The amphiphilic polymer-grafted silica was newly prepared as a stationary phase in high-performance liquid chromatography. Poly(4-vinylpyridine) with a trimethoxysilyl group at one end was grafted onto porous silica particles and the pyridyl side chains were quaternized with 1-bromooctadecane. The obtained poly(octadecylpyridinium)-grafted silica was characterized by elemental analysis, diffuse reflectance infrared Fourier transform spectroscopy and Brunauer-Emmett-Teller analysis. The degree of quaternization of the pyridyl groups on the obtained stationary phase was estimated to be 70%. The selective retention behaviors of polycyclic aromatic hydrocarbons including some positional isomers were investigated using poly(octadecylpyridinium)-grafted silica as an amphiphilic polymer stationary phase in high-performance liquid chromatography and results were compared with commercially available polymeric octadecylated silica and phenyl-bonded silica columns. The results indicate that the selectivity toward polycyclic aromatic hydrocarbons exhibited by the amphiphilic polymer stationary phase is higher than the corresponding selectivity exhibited by a conventional phenyl-bonded silica column. However, compared with the polymeric octadecylated silica phase, the new stationary phase presents similar retention behavior for polycyclic aromatic hydrocarbons but different retention behavior particularly for positional isomers of disubstituted benzenes as the aggregation structure of amphiphilic polymers on the surface of silica substrate has been altered during mobile phase variation. PMID:25944152

  1. In situ synthesis of metal-organic frameworks in a porous polymer monolith as the stationary phase for capillary liquid chromatography.

    PubMed

    Yang, Shengchao; Ye, Fanggui; Zhang, Cong; Shen, Shufen; Zhao, Shulin

    2015-04-21

    In this study, HKUST-1 was synthesized in situ on the porous polymer monolith as the stationary phase for capillary liquid chromatography (cLC). The unique carboxyl functionalized poly(methacrylic acid-co-ethylene dimethacrylate) (poly(MAA-co-EDMA)) monolith was used as a support to directly grow HKUST-1 by a controlled layer-by-layer self-assembly strategy. X-ray diffraction, scanning electron microscopy, energy dispersive X-ray spectrometry, and Fourier transform infrared spectroscopy of the resulting HKUST-1-poly(MAA-co-EDMA) monoliths indicated that HKUST-1 was successfully grafted onto the pore surface of the poly(MAA-co-EDMA) monolith. The column performance of HKUST-1-poly(MAA-co-EDMA) monoliths for the separation of various small molecules, such as benzenediols, xylenes, ethylbenzenes, and styrenes, was evaluated. The chromatographic performance was found to improve with increasing HKUST-1 density, and the column efficiencies and resolutions of HKUST-1-poly(MAA-co-EDMA) monoliths were 18 320-19 890 plates m(-1) and 1.62-6.42, respectively, for benzenediols. The HKUST-1-poly(MAA-co-EDMA) monolith displayed enhanced resolution for the separation of positional isomers when compared to the traditional C18 and HKUST-1 incorporated polymer monoliths. Hydrophobic, π-π, and hydrogen bonding interactions within the HKUST-1-poly(MAA-co-EDMA) monolith were observed in the separation of small molecules. The results showed that the HKUST-1-poly(MAA-co-EDMA) monoliths are promising stationary phases for cLC. PMID:25710284

  2. Titanium-scaffolded organic-monolithic stationary phases for ultra-high-pressure liquid chromatography.

    PubMed

    Vonk, Rudy J; Vaast, Axel; Eeltink, Sebastiaan; Schoenmakers, Peter J

    2014-09-12

    Organic-polymer monoliths with overall dimensions larger than one millimetre are prone to rupture - either within the monolith itself or between the monoliths and the containing wall - due to the inevitable shrinkage accompanying the formation of a cross-linked polymeric network. This problem has been addressed by creating titanium-scaffolded poly(styrene-co-divinylbenzene) (S-co-DVB) monoliths. Titanium-scaffolded monoliths were successfully used in liquid chromatography at very high pressures (up to 80MPa) and using gradients spanning the full range of water-acetonitrile compositions (0 to 100%). The kinetic-performance of (50-mm long) titanium-scaffolded monoliths was compared to that of similar monolith created in 1-mm i.d. glass-lined tubing at pressures up to 50MPa. The peak capacities obtained with the titanium-scaffolded column was about 30% lower. An increased Eddy-diffusion, due to the pillar-structure, and a decreased permeability are thought to be the main reasons for this reduced kinetic-performance. No decrease in performance was observed when the titanium-scaffolded columns were operated at pressures of 80MPa for up to 12h. The column-to-column repeatability (n=5) was acceptable in terms of observed peak widths at half heights (RSD ca. 10%) The run-to-run repeatability (n=135) in terms of retention times and peak widths at half height were found to be good. Titanium-scaffolded columns coupled in series up to a combined length of (200mm) were used for the analyses of a complex Escherichia coli protein sample. Our experiments demonstrate that columns based on titanium-scaffolded organic-polymer monolith can be operated under strenuous conditions without loss in performance. The titanium-scaffolded approach makes it feasible to create organic-polymer monoliths in wide-bore columns with accurate temperature control. PMID:25086753

  3. Polymer-coated fibrous materials as the stationary phase in packed capillary gas chromatography.

    PubMed

    Saito, Yoshihiro; Tahara, Ai; Imaizumi, Motohiro; Takeichi, Tsutomu; Wada, Hiroo; Jinno, Kiyokatsu

    2003-10-15

    Synthetic polymer filaments have been introduced as the support material in packed capillary gas chromatography (GC). The filaments of the heat-resistant polymers, Zylon, Kevlar, Nomex, and Technora, were longitudinally packed into a short fused-silica capillary, followed by the conventional coating process for open-tubular GC columns. The separation of several test mixtures such as n-alkylbenzenes and n-alkanes was carried out with these polymer-coated fiber-packed capillary columns. With the coating by various polymeric materials on the surface of these filaments, the retentivity was significantly improved over the parent fiber-packed column (without polymer coating) as well as a conventional open-tubular capillary of the same length. The results demonstrated a good combination of Zylon as the support and poly(dimethylsiloxane)-based materials as the coating liquid-phase for the successful GC separation of n-alkanes and polycyclic aromatic hydrocarbons (PAHs), while successful applications for other separations such as poly(ethylene glycol) coating for the separation of alcohols were also obtained. From the results it has been suggested that the selectivity of the fiber-packed column could be tuned by selecting different coating materials, indicating the promising possibility for a novel usage of fine fibrous polymers as the support material that can be combined with newly synthesized coating materials specially designed for particular separations. Taking advantage of good thermal stability of the fibers, the column temperature could be elevated to higher than 350 degrees C with the combination of a short metallic capillary. PMID:14710834

  4. Polarity-based fractionation in proteomics: hydrophilic interaction vs reversed-phase liquid chromatography.

    PubMed

    Jafari, M; Mirzaie, M; Khodabandeh, M; Rezadoost, H; Ghassempour, A; Aboul-Enein, H Y

    2016-07-01

    During recent decades, hydrophilic interaction liquid chromatography (HILIC) ahs been introduced to fractionate or purify especially polar solutes such as peptides and proteins while reversed-phase liquid chromatography (RPLC) is also a common strategy. RPLC is also a common dimension in multidimensional chromatography. In this study, the potential of HILIC vs RPLC chromatography was compared for proteome mapping of human peripheral blood mononuclear cell extract. In HILIC a silica-based stationary phase and for RPLC a C18 column were applied. Then separated proteins were eluted to an ion trap mass spectrometry system. Our results showed that the HILIC leads to more proteins being identified in comparison to RPLC. Among the total 181 identified proteins, 56 and 38 proteins were fractionated specifically by HILIC and RPLC, respectively. In order to demonstrate this, the physicochemical properties of identified proteins such as polarity and hydrophobicity were considered. This analysis indicated that polarity may play a major role in the HILIC separation of proteins vs RPLC. Using gene ontology enrichment analysis, it was also observed that differences in physicochemical properties conform to the cellular compartment and biological features. Finally, this study highlighted the potential of HILIC and the great orthogonality of RPLC in gel-free proteomic studies. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26555197

  5. Analytical and semipreparative chiral separation of cis-itraconazole on cellulose stationary phases by high-performance liquid chromatography.

    PubMed

    Kurka, Ondřej; Kučera, Lukáš; Bednář, Petr

    2016-07-01

    cis-Itraconazole is a chiral antifungal drug administered as a racemate. The knowledge of properties of individual cis-itraconazole stereoisomers is vital information for medicine and biosciences as different stereoisomers of cis-itraconazole may possess different affinity to certain biological pathways in the human body. For this purpose, either chiral synthesis of enantiomers or chiral separation of racemate can be used. This paper presents a two-step high-performance liquid chromatography approach for the semipreparative isolation of four stereoisomers (two enantiomeric pairs) of itraconazole using polysaccharide stationary phases and volatile organic mobile phases without additives in isocratic mode. The approach used involves the separation of the racemate into three fractions (i.e. two pure stereoisomers and one mixed fraction containing the remaining two stereoisomers) in the first run and consequent separation of the collected mixed fraction in the second one. For this purpose, combination of cellulose tris-(4-methylbenzoate) and cellulose tris-(3,5-dimehylphenylcarbamate) columns with complementary selectivity for cis-itraconazole provided full separation of all four stereoisomers (with purity of each isomer > 97%). The stereoisomers were collected, their optical rotation determined and their identity confirmed based on the results of a previously published study. Pure separated stereoisomers are subjected to further biological studies. PMID:27240968

  6. Enantioseparation using ortho- or meta-substituted phenylcarbamates of amylose as chiral stationary phases for high-performance liquid chromatography.

    PubMed

    Shen, Jun; Zhao, Yongqiang; Inagaki, Shinji; Yamamoto, Chiyo; Shen, Yue; Liu, Shuangyan; Okamoto, Yoshio

    2013-04-19

    Six ortho- and six meta-substituted phenylcarbamate derivatives of amylose were prepared and their chiral recognition abilities were evaluated as chiral stationary phases (CSPs) for high-performance liquid chromatography (HPLC). The substitution at the meta-position on the aromatic ring was more preferable than that at the ortho-position to obtain CSPs with a high chiral recognition ability, and the introduction of either an electron-withdrawing or electron-donating substituent can improve the chiral resolving power of the meta-substituted phenylcarbamates of amylose. The chiral recognition ability of the amylose phenylcarbamates and elution order of the enantiomers were significantly dependent on the position, nature and number of the substituents on the phenyl group. Correlations between the chiral recognition ability and the N-H frequencies in the IR spectra and the chemical shifts of the N-H protons in the (1)H NMR spectra of the carbamate moieties of the amylose derivatives were discussed. The structures of the amylose derivatives were also investigated by circular dichroism spectroscopy. PMID:23506702

  7. Size-exclusion chromatography system for macromolecular interaction analysis

    DOEpatents

    Stevens, Fred J.

    1988-01-01

    A low pressure, microcomputer controlled system employing high performance liquid chromatography (HPLC) allows for precise analysis of the interaction of two reversibly associating macromolecules such as proteins. Since a macromolecular complex migrates faster than its components during size-exclusion chromatography, the difference between the elution profile of a mixture of two macromolecules and the summation of the elution profiles of the two components provides a quantifiable indication of the degree of molecular interaction. This delta profile is used to qualitatively reveal the presence or absence of significant interaction or to rank the relative degree of interaction in comparing samples and, in combination with a computer simulation, is further used to quantify the magnitude of the interaction in an arrangement wherein a microcomputer is coupled to analytical instrumentation in a novel manner.

  8. Analysis of histone post translational modifications in primary monocyte derived macrophages using reverse phase×reverse phase chromatography in conjunction with porous graphitic carbon stationary phase.

    PubMed

    Minshull, Thomas C; Cole, Joby; Dockrell, David H; Read, Robert C; Dickman, Mark J

    2016-07-01

    A two dimensional-liquid chromatography (2D-LC) based approach was developed for the identification and quantification of histone post translational modifications in conjunction with mass spectrometry analysis. Using a bottom-up strategy, offline 2D-LC was developed using reverse phase chromatography. A porous graphitic carbon stationary phase in the first dimension and a C18 stationary phase in the second dimension interfaced with mass spectrometry was used to analyse global levels of histone post translational modifications in human primary monocyte-derived macrophages. The results demonstrated that 84 different histone peptide proteoforms, with modifications at 18 different sites including combinatorial marks were identified, representing an increase in the identification of histone peptides by 65% and 51% compared to two different 1D-LC approaches on the same mass spectrometer. The use of the porous graphitic stationary phase in the first dimension resulted in efficient separation of histone peptides across the gradient, with good resolution and is orthogonal to the online C18 reverse phase chromatography. Overall, more histone peptides were identified using the 2D-LC approach compared to conventional 1D-LC approaches. In addition, a bioinformatic pipeline was developed in-house to enable the high throughput efficient and accurate quantification of fractionated histone peptides. The automation of a section of the downstream analysis pipeline increased the throughput of the 2D-LC-MS/MS approach for the quantification of histone post translational modifications. PMID:27260198

  9. Chromatography.

    ERIC Educational Resources Information Center

    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)

  10. Analysis of Drug Interactions with Lipoproteins by High-Performance Affinity Chromatography

    PubMed Central

    Sobansky, Matthew R.; Hage, David S.

    2013-01-01

    Lipoproteins such as high-density lipoprotein (HDL) and low-density lipoprotein (LDL) are known to interact with drugs and other solutes in blood. These interactions have been examined in the past by methods such as equilibrium dialysis and capillary electrophoresis. This chapter describes an alternative approach that has recently been developed for examining these interactions by using high-performance affinity chromatography. In this method, lipoproteins are covalently immobilized to a solid support and used within a column as a stationary phase for binding studies. This approach allows the same lipoprotein preparation to be used for a large number of binding studies, leading to precise estimates of binding parameters. This chapter will discuss how this technique can be applied to the identification of interaction models and be used to differentiate between systems that have interactions based on partitioning, adsorption or mixed-mode interactions. It is also shown how this approach can then be used for the measurement of binding parameters for HDL and LDL with drugs. Examples of these studies are provided, with particular attention being given to the use of frontal analysis to examine the interactions of R- and S-propranolol with HDL and LDL. The advantages and possible limitations of this method are described. The extension of this approach to other types of drug-lipoprotein interactions is also considered. PMID:25392741

  11. The evaluation of 25 chiral stationary phases and the utilization of sub-2.0μm coated polysaccharide chiral stationary phases via supercritical fluid chromatography.

    PubMed

    Hamman, Chris; Wong, Mengling; Aliagas, Ignacio; Ortwine, Daniel F; Pease, Joseph; Schmidt, Donald E; Victorino, Joseph

    2013-08-30

    A rapid screening method to identify the best conditions for chiral separations is described. We analyzed a representative set of 80 racemic compounds against 25 different chiral stationary phases with three different mobile phases to identify the combination of columns and mobile phases that will separate the most compounds on the initial screen. While the OD separated the largest number of compounds, we found the best combination of six columns to be the AD, AS, AY, CC4, ID and Whelk-O1. The second team included the CCC, Cellulose-1, Cellulose-3 or OJ, IA, IE and IF. All 80 compounds were separated with a resolution range of 0.65-15.36. Screening the covalently bonded phases provided separation for 79 of the 80 compounds. We also found ethanol (0.1% NH4OH) separated more compounds than methanol (0.1% NH4OH) or isopropanol (0.1% NH4OH). As part of this study, we also compared the effectiveness of stationary phases that have the same chiral selector. Finally, we demonstrated the effectiveness of using a fast, 1.5-min screening method that utilizes a 1.7μm coated polysaccharide chiral stationary phase. PMID:23891213

  12. Application of gas-liquid chromatography to the analysis of essential oils. Part XVII. Fingerprinting of essential oils by temperature-programmed gas-liquid chromatography using capillary columns with non-polar stationary phases. Analytical methods committee.

    PubMed

    1997-10-01

    Problems in obtaining reproducible results when 'fingerprinting' essential oils by temperature-programmed gas-liquid chromatography have been reported on in Parts VII and VIII of this series. Those reports were concerned with the general problems and the use of packed columns. This report is concerned with the use of capillary columns and non-polar stationary phases. A collaborative study using capillary columns with non-polar stationary phases has resulted in a method which specifies the 'g-pack value' of a column and gives reproducible relative retention indices for the test compounds limonene, acetophenone, linalol, naphthalene, linalyl acetate and cinnamyl alcohol. The method has been applied successfully to the examination of oil of rosemary. A recommended method is given for the reproducible temperature-programmed gas-liquid chromatographic fingerprinting of essential oils using capillary columns with non-polar stationary phases. PMID:9463975

  13. Kinetic Studies of Biological Interactions By Affinity Chromatography

    PubMed Central

    Schiel, John E.; Hage, David S.

    2009-01-01

    The rates at which biological interactions occur can provide important information on the mechanism and behavior of such processes in living systems. This review will discuss how affinity chromatography can be used as a tool to examine the kinetics of biological interactions. This approach, referred to here as biointeraction chromatography, uses a column with an immobilized binding agent to examine the association or dissociation of this agent with other compounds. The use of HPLC-based affinity columns in kinetic studies has received particular attention in recent years. Advantages of using HPLC with affinity chromatography for this purpose include the ability to reuse the same ligand within a column for a large number of experiments, and the good precision and accuracy of this approach. A number of techniques are available for kinetic studies through the use of affinity columns and biointeraction chromatography. These approaches include plate height measurements, peak profiling, peak fitting, split-peak measurements, and peak decay analysis. The general principles for each of these methods are discussed in this review and some recent applications of these techniques are presented. The advantages and potential limitations of each approach are also considered. PMID:19391173

  14. Recent progress in the fundamental understanding of hydrophilic interaction chromatography (HILIC).

    PubMed

    Guo, Yong

    2015-10-01

    With the exponential growth in the application of the HILIC technique, there has been a significant progress in understanding the fundamental aspects of hydrophilic interaction chromatography. The experimental studies tend to be more extensive in terms of the number of stationary phases investigated and the number of probe compounds employed in comparison with the earlier studies; and more theoretical studies in quantitative structure retention relationship (QSRR) and molecular dynamics simulations have also been published and provide molecular-level insights into the retention mechanism. This review focuses on the recent progress in understanding the retention mechanism, retention models, selectivity, and the kinetic performance of HILIC. A better understanding of these fundamental aspects will undoubtedly facilitate more applications of this chromatographic technique in a wider range of fields. PMID:26221630

  15. Evaluation of the Kinetic Performance Differences between Hydrophilic-Interaction Liquid Chromatography and Reversed-Phase Liquid Chromatography under Conditions of Identical Packing Structure.

    PubMed

    Song, Huiying; Desmet, Gert; Cabooter, Deirdre

    2015-12-15

    A protocol using trifluoroacetic acid at a temperature of 60 °C is developed for the adequate removal of the stationary phase of reversed-phase liquid chromatography (RPLC) columns. This procedure allows for studying the same column first under RPLC and subsequently under hydrophilic-interaction liquid chromatography (HILIC) conditions to isolate intrinsic differences between mass transfer properties in HILIC and RPLC from differences in packing quality. The established procedure allows for a complete removal of the stationary phase (confirmed by retention studies and thermogravimetry analyses) while leaving the structure of the packing unaffected (witnessed by an unchanged external porosity and pressure drop). Accurate plate height analysis comparing compounds at the same zone retention factor indicates a significant difference in reduced c-term (typically 40-80% larger under HILIC conditions), despite the columns otherwise being identical. Correcting for the known contributions of longitudinal diffusion (b-term) and mass transfer (cm- and cs-term) to focus on band broadening originating from eddy dispersion, similar strong differences are observed (differences of some h = 0.3 up to 1.2). These findings show that the interior structure and retention mechanism of the particles have a very strong effect on the observed eddy dispersion, a factor typically ascribed to phenomena occurring outside the particles. This also implies that comparing the quality of packings of different particle types is virtually impossible without the availability of a sound model to correct for the intraparticle effect on the observed eddy dispersion. PMID:26595107

  16. Organic monoliths for hydrophilic interaction electrochromatography/chromatography and immunoaffinity chromatography

    PubMed Central

    Gunasena, Dilani N.; El Rassi, Ziad

    2012-01-01

    This article is aimed at providing a review of the progress made over the past decade in the preparation of polar monoliths for hydrophilic interaction liquid chromatography (HILIC)/capillary electrochromatography (HI-CEC) and in the design of immuno-monoliths for immunoaffinity chromatography (IAC) that are based on some of the polar monolith precursors used in HILIC/HI-CEC. In addition, this review article discusses some of the applications of polar monoliths by HILIC and HI-CEC, and the applications of immuno-monoliths. This article is by no means an exhaustive review of the literature; it is rather a survey of the recent progress made in the field with 83 references published in the past decade on the topics of HILIC and IAC monoliths. PMID:22147366

  17. In-depth characterization of six cellulose tris-(3,5-dimethylphenylcarbamate) chiral stationary phases in supercritical fluid chromatography.

    PubMed

    Khater, Syame; Zhang, Yingru; West, Caroline

    2013-08-16

    Since the expiration of the patent protection of Chiralcel OD, similar chiral stationary phases (CSPs), all based on the same chiral selector, have been introduced on the market with the promise to reproduce or improve the performance of the original cellulose tris-(3,5-dimethylphenylcarbamate) CSP. We report here-in an in-depth evaluation of four generic versions of Chiralcel OD (CelluCoat, RegisCell, Lux Cellulose-1, Reprosil-OM) and the immobilized version (Chiralpak IB) in comparison to the original Chiralcel OD in terms of retention and enantioselectivity, with the help of chemometrics. First of all, the CSPs are compared based on the retentions of 230 achiral compounds. Agglomerative hierarchical clustering and quantitative structure-retention relationships based on a modified version of the solvation parameter model are used to assess the differences in non-enantioselective interactions contributing to retention. Secondly, the CSPs are compared based on the separation factors measured for 130 racemates. Discriminant analysis is then used to unravel the structural features contributing to the successful enantioselective separations. Chiralcel OD is shown to be the most versatile of the six tested CSPs, and involves a unique and unequalled mechanism to achieve enantioseparation. PMID:23838300

  18. Kinetic analysis of drug-protein interactions by affinity chromatography.

    PubMed

    Bi, Cong; Beeram, Sandya; Li, Zhao; Zheng, Xiwei; Hage, David S

    2015-10-01

    Information on the kinetics of drug-protein interactions is of crucial importance in drug discovery and development. Several methods based on affinity chromatography have been developed in recent years to examine the association and dissociation rates of these processes. These techniques include band-broadening measurements, the peak decay method, peak fitting methods, the split-peak method, and free fraction analysis. This review will examine the general principles and applications of these approaches and discuss their use in the characterization, screening and analysis of drug-protein interactions in the body. PMID:26724332

  19. Simultaneous analysis of multiple neurotransmitters by hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry.

    PubMed

    Tufi, Sara; Lamoree, Marja; de Boer, Jacob; Leonards, Pim

    2015-05-22

    Neurotransmitters are endogenous metabolites that allow the signal transmission across neuronal synapses. Their biological role is crucial for many physiological functions and their levels can be changed by several diseases. Because of their high polarity, hydrophilic interaction liquid chromatography (HILIC) is a promising tool for neurotransmitter analysis. Due to the large number of HILIC stationary phases available, an evaluation of the column performances and retention behaviors has been performed on five different commercial HILIC packing materials (silica, amino, amide and two zwitterionic stationary phases). Several parameters like the linear correlation between retention and the distribution coefficient (logD), the separation factor k and the column resolution Rs have been investigated and the column performances have been visualized with a heat map and hierarchical clustering analysis. An optimized and validated HILIC-MS/MS method based on the ZIC-cHILIC column is proposed for the simultaneous detection and quantification of twenty compounds consisting of neurotransmitters, precursors and metabolites: 3-methoxytyramine (3-MT), 5-hydroxyindoleacetic acid (5-HIAA), 5-hydroxy-L-tripthophan, acetylcholine, choline, L-3,4-dihydroxyphenylalanine (L-DOPA), dopamine, epinephrine, γ-aminobutyric acid (GABA), glutamate, glutamine, histamine, histidine, L-tryptophan, L-tyrosine, norepinephrine, normetanephrine, phenylalanine, serotonin and tyramine. The method was applied to neuronal metabolite profiling of the central nervous system of the freshwater snail Lymnaea stagnalis. This method is suitable to explore neuronal metabolism and its alteration in different biological matrices. PMID:25869798

  20. Interaction Chromatography of Random Copolymers with Tunable Monomer Sequence Distributions

    NASA Astrophysics Data System (ADS)

    Ryu, Chang Y.; Han, Junwon; Jeon, Byung Ho; Semler, James J.; Jhon, Young K.; Genzer, Jan

    2008-03-01

    We demonstrate that high performance liquid chromatography (HPLC) in the interaction chromatography (IC) mode is capable of distinguishing among various comonomer sequences in random copolymers (RCPs). A series of poly(styrene-co-4-bromostyrene) (PBrxS), where x is the mole fraction of 4-BrS, RCPs have been prepared by brominating parent monodisperse polystyrene (PS). The distribution of S and 4-BrS segments in PBrxS was adjusted by varying the solvent quality for PS before the bromination reaction. We utilize both normal and reversed phase IC techniques to demonstrate that the adsorption-based retention of PBrxS RCPs is affected not only by their chemical composition, but also by the comonomer distribution in the RCP. Both IC techniques are mutually complementary; they provide information on the interplay between the macromolecular collapse and segment blockiness affected by the adsorption-based retention times in HPLC.

  1. Attractor nonequilibrium stationary states in perturbed long-range interacting systems

    NASA Astrophysics Data System (ADS)

    Joyce, Michael; Morand, Jules; Viot, Pascal

    2016-05-01

    Isolated long-range interacting particle systems appear generically to relax to nonequilibrium states ("quasistationary states" or QSSs) which are stationary in the thermodynamic limit. A fundamental open question concerns the "robustness" of these states when the system is not isolated. In this paper we explore, using both analytical and numerical approaches to a paradigmatic one-dimensional model, the effect of a simple class of perturbations. We call them "internal local perturbations" in that the particle energies are perturbed at collisions in a way which depends only on the local properties. Our central finding is that the effect of the perturbations is to drive all the very different QSSs we consider towards a unique QSS. The latter is thus independent of the initial conditions of the system, but determined instead by both the long-range forces and the details of the perturbations applied. Thus in the presence of such a perturbation the long-range system evolves to a unique nonequilibrium stationary state, completely different from its state in absence of the perturbation, and it remains in this state when the perturbation is removed. We argue that this result may be generic for long-range interacting systems subject to perturbations which are dependent on the local properties (e.g., spatial density or velocity distribution) of the system itself.

  2. Selectivity differences of water-soluble vitamins separated on hydrophilic interaction stationary phases.

    PubMed

    Yang, Yuanzhong; Boysen, Reinhard I; Hearn, Milton T W

    2013-06-01

    In this study, the retention behavior and selectivity differences of water-soluble vitamins were evaluated with three types of polar stationary phases (i.e. an underivatized silica phase, an amide phase, and an amino phase) operated in the hydrophilic interaction chromatographic mode with ESI mass spectrometric detection. The effects of mobile phase composition, including buffer pH and concentration, on the retention and selectivity of the vitamins were investigated. In all stationary phases, the neutral or weakly charged vitamins exhibited very weak retention under each of the pH conditions, while the acidic and more basic vitamins showed diverse retention behaviors. With the underivatized silica phase, increasing the salt concentration of the mobile phase resulted in enhanced retention of the acidic vitamins, but decreased retention of the basic vitamins. These observations thus signify the involvement of secondary mechanisms, such as electrostatic interaction in the retention of these analytes. Under optimized conditions, a baseline separation of all vitamins was achieved with excellent peak efficiency. In addition, the effects of water content in the sample on retention and peak efficiency were examined, with sample stacking effects observed when the injected sample contained a high amount of water. PMID:23554360

  3. Master equation approach for interacting slow- and stationary-light polaritons

    SciTech Connect

    Kiffner, M.; Hartmann, M. J.

    2010-09-15

    A master equation approach for the description of dark-state polaritons in coherently driven atomic media is presented. This technique provides a description of light-matter interactions under conditions of electromagnetically induced transparency (EIT) that is well suited for the treatment of polariton losses. The master equation approach allows us to describe general polariton-polariton interactions that may be conservative, dissipative, or a mixture of both. In particular, it enables us to study dissipation-induced correlations as a means for the creation of strongly correlated polariton systems. Our technique reveals a loss mechanism for stationary-light polaritons that has not been discussed so far. We find that polariton losses in level configurations with nondegenerate ground states can be a multiple of those in level schemes with degenerate ground states.

  4. Phenyl Functionalized Sol-gel Silica Sorbent for Capillary Microextraction and Chromia-Based Sol-gel Ucon Stationary Phase for Capillary Gas Chromatography

    NASA Astrophysics Data System (ADS)

    McLean, Michael M.

    The first chapter of this thesis presents an introduction to sol-gel methodology whose usefulness as a synthetic route will be demonstrated with two applications in chromatography. The first application involves the fabrication of a capillary micro-extraction (CME) device by coating a phenyl functionalized extracting phase on the inner surface of a fused silica capillary for analyte pre-concentration. The device was coupled on-line to a RP-HPLC system and practicality was demonstrated using allergens as target analytes. The allergens chosen as model analytes are typically found in fragrance products and food. Most of the 26 fragrance allergens that are monitored by various government authorities have a phenyl organic moiety (a strong chromophore), thus making them appropriate probes for exploring the extraction efficiency of the coating using a UV detector. The CME device showed ppt level limit of detection which makes it suitable for trace analyses of allergens and similar compounds in a variety of matrices. The second application explores the feasibility of using sol-gel derived chromia-based stationary phase in gas chromatographic columns. The organic moiety of the stationary phase was derived from Ucon 75-H-90,000 while the inorganic backbone was prepared using chromium(III) dichloride hydroxide - methacrylic acid - aqua complex, 40% in isopropanol/acetone . Usefulness of prepared chromia-based GC stationary phase was examined for petrochemical application. Promising results were obtained using aliphatic-aromatics, polyaromatic hydrocarbons, BTEX test mixture, cycloalkanes, branched alkanes and akylbenzenes. The column was able to perform without degradation despite being rinsed multiples times sequentially with the following solvents: dichloromethane, methanol, water and finally methanol again. Maximum theoretical plate number calculated is around 2,400 plates/m. The plate number clearly needs improvement but is a promising result for the newly explored

  5. Direct quantification of dimethylsulfoniopropionate (DMSP) with hydrophilic interaction liquid chromatography/mass spectrometry.

    PubMed

    Spielmeyer, Astrid; Pohnert, Georg

    2010-12-01

    A simple, derivatization free method for the direct determination of dimethylsulfoniopropionate (DMSP) using hydrophilic interaction liquid chromatography (HILIC)/mass spectrometry is introduced. DMSP is a zwitterionic osmolyte which is produced from marine plankton, macro algae and higher plants. Due to its central role in climate relevant geochemical processes as well as in plant physiology and chemical ecology there is a great interest in methods for its quantification. Since DMSP is labile and difficult to extract currently most protocols for quantification are based on indirect methods. Here we show that ultra performance liquid chromatography/mass spectrometry using a HILIC stationary phase is suitable for the direct quantification of DMSP from aqueous samples and microalgal extracts. The protocol requires minimal sample preparation and phytoplankton samples can be investigated after filtration of small volumes. The limit of detection is 20nM and the calibration curve is linear in the range of 60nM to 50μM. The use of [(2)H(6)]-DMSP as internal standard allows prolonged sample storage since it is transformed with the same kinetics as natural DMSP. This makes the method suitable for both laboratory and field studies. PMID:21030323

  6. New reversed-phase/anion-exchange/hydrophilic interaction mixed-mode stationary phase based on dendritic polymer-modified porous silica.

    PubMed

    Li, Yun; Yang, Jiajia; Jin, Jing; Sun, Xiaoli; Wang, Longxing; Chen, Jiping

    2014-04-11

    A novel dendritic polymer-modified silica (DPS) stationary phase was prepared by a divergent synthesis scheme starting from propylamine on silica by consecutive amine-epoxy reactions with 1,4-butanedioldiglycidyl ether and aniline. Both elemental analysis and infrared spectra data shows the successful growth of dendritic polymer on silica particles. The carbon and nitrogen contents increased with an increasing number of reaction cycles and achieved 25.2% and 2.1% (w/w) after 11 reaction cycles. The combination of a phenyl ring with a quaternary ammonium, or a tertiary amine at the branch point along with embedded polar functionalities (including ether and hydroxyl groups) in the branch, generated hydrophobic, electrostatic, as well as hydrophilic interactive domains. Depending on solute structure and mobile phase composition, the DPS stationary phase provided multiple retention mechanisms, including reversed phase (RP), anion-exchange (AEX), and hydrophilic interactions. The RP capability achieved separation of polycyclic aromatic hydrocarbons. Basic, neutral and acidic molecules were well separated under RP/AEX mixed mode. Effective separation of small polar compounds (such as nucleobases and nucleosides) was also obtained under hydrophilic interaction liquid chromatography (HILIC) mode. PMID:24630062

  7. Adsorption of water from aqueous acetonitrile on silica-based stationary phases in aqueous normal-phase liquid chromatography.

    PubMed

    Soukup, Jan; Jandera, Pavel

    2014-12-29

    Excess adsorption of water from aqueous acetonitrile mobile phases was investigated on 16 stationary phases using the frontal analysis method and coulometric Karl-Fischer titration. The stationary phases include silica gel and silica-bonded phases with different polarities, octadecyl and cholesterol, phenyl, nitrile, pentafluorophenylpropyl, diol and zwitterionic sulfobetaine and phosphorylcholine ligands bonded on silica, hybrid organic-silica and hydrosilated matrices. Both fully porous and core-shell column types were included. Preferential uptake of water by the columns can be described by Langmuir isotherms. Even though a diffuse rather than a compact adsorbed discrete layer of water on the adsorbent surface can be formed because of the unlimited miscibility of water with acetonitrile, for convenience, the preferentially adsorbed water was expressed in terms of a hypothetical monomolecular water layer equivalent in the inner pores. The uptake of water strongly depends on the polarity and type of the column. Less than one monomolecular water layer equivalent was adsorbed on moderate polar silica hydride-based stationary phases, Ascentis Express F5 and Ascentis Express CN column at the saturation capacity, while on more polar stationary phases, several water layer equivalents were up-taken from the mobile phase. The strongest affinity to water was observed on the ZIC cHILIC stationary phases, where more than nine water layer equivalents were adsorbed onto its surface at its saturation capacity. Columns with bonded hydroxyl and diol ligands show stronger water adsorption in comparison to bare silica. Columns based on hydrosilated silica generally show significantly decreased water uptake in comparison to stationary phases bonded on ordinary silica. Significant correlations were found between the water uptake and the separation selectivity for compounds with strong polarity differences. PMID:25544246

  8. Metal-organic framework MIL-100(Fe) as the stationary phase for both normal-phase and reverse-phase high performance liquid chromatography.

    PubMed

    Fu, Yan-Yan; Yang, Cheng-Xiong; Yan, Xiu-Ping

    2013-01-25

    Metal-organic framework MIL-100(Fe) was explored as a novel stationary phase for both normal-phase and reverse-phase high performance liquid chromatography. Two groups of analytes (benzene, toluene, ethylbenzene, naphthalene and 1-chloronaphthalene; aniline, acetanilide, 2-nitroaniline and 1-naphthylamine) were used to test the separation performance of MIL-100(Fe) in the reverse-phase mode, while the isomers of chloroaniline or toluidine were employed to evaluate its performance in the normal-phase mode. The MIL-100(Fe) packed column gave a baseline separation of all the tested analytes with good precision. The separation was controlled by negative enthalpy change and entropy change in the reverse-phase mode, but positive enthalpy change and entropy change in the normal-phase mode. The relative standard deviations of retention time, peak area, peak height, and half peak width for eleven replicate separations of the tested analytes were 0.2-0.7%, 0.5-3.6%, 0.6-2.3% and 0.8-1.7%, respectively. The mesoporous cages, accessible windows, excellent chemical and solvent stability, metal active sites and aromatic pore walls make MIL-100(Fe) a good candidate as a novel stationary phase for both normal-phase and reverse-phase high performance liquid chromatography. PMID:23290359

  9. Systematic evaluation of matrix effects in hydrophilic interaction chromatography versus reversed phase liquid chromatography coupled to mass spectrometry.

    PubMed

    Periat, Aurélie; Kohler, Isabelle; Thomas, Aurélien; Nicoli, Raul; Boccard, Julien; Veuthey, Jean-Luc; Schappler, Julie; Guillarme, Davy

    2016-03-25

    Reversed phase liquid chromatography (RPLC) coupled to mass spectrometry (MS) is the gold standard technique in bioanalysis. However, hydrophilic interaction chromatography (HILIC) could represent a viable alternative to RPLC for the analysis of polar and/or ionizable compounds, as it often provides higher MS sensitivity and alternative selectivity. Nevertheless, this technique can be also prone to matrix effects (ME). ME are one of the major issues in quantitative LC-MS bioanalysis. To ensure acceptable method performance (i.e., trueness and precision), a careful evaluation and minimization of ME is required. In the present study, the incidence of ME in HILIC-MS/MS and RPLC-MS/MS was compared for plasma and urine samples using two representative sets of 38 pharmaceutical compounds and 40 doping agents, respectively. The optimal generic chromatographic conditions in terms of selectivity with respect to interfering compounds were established in both chromatographic modes by testing three different stationary phases in each mode with different mobile phase pH. A second step involved the assessment of ME in RPLC and HILIC under the best generic conditions, using the post-extraction addition method. Biological samples were prepared using two different sample pre-treatments, i.e., a non-selective sample clean-up procedure (protein precipitation and simple dilution for plasma and urine samples, respectively) and a selective sample preparation, i.e., solid phase extraction for both matrices. The non-selective pretreatments led to significantly less ME in RPLC vs. HILIC conditions regardless of the matrix. On the contrary, HILIC appeared as a valuable alternative to RPLC for plasma and urine samples treated by a selective sample preparation. Indeed, in the case of selective sample preparation, the compounds influenced by ME were different in HILIC and RPLC, and lower and similar ME occurrence was generally observed in RPLC vs. HILIC for urine and plasma samples

  10. Chromatography

    MedlinePlus

    Chromatography is a way of separating two or more chemical compounds. Chemical compounds are chemicals that are ... of chemical compound. There are different kinds of chromatography. These include gas, high pressure liquid, or ion ...

  11. High-throughput protein precipitation and hydrophobic interaction chromatography: salt effects and thermodynamic interrelation.

    PubMed

    Nfor, Beckley K; Hylkema, Nienke N; Wiedhaup, Koenraad R; Verhaert, Peter D E M; van der Wielen, Luuk A M; Ottens, Marcel

    2011-12-01

    Salt-induced protein precipitation and hydrophobic interaction chromatography (HIC) are two widely used methods for protein purification. In this study, salt effects in protein precipitation and HIC were investigated for a broad combination of proteins, salts and HIC resins. Interrelation between the critical thermodynamic salting out parameters in both techniques was equally investigated. Protein precipitation data were obtained by a high-throughput technique employing 96-well microtitre plates and robotic liquid handling technology. For the same protein-salt combinations, isocratic HIC experiments were performed using two or three different commercially available stationary phases-Phenyl Sepharose low sub, Butyl Sepharose and Resource Phenyl. In general, similar salt effects and deviations from the lyotropic series were observed in both separation methods, for example, the reverse Hofmeister effect reported for lysozyme below its isoelectric point and at low salt concentrations. The salting out constant could be expressed in terms of the preferential interaction parameter in protein precipitation, showing that the former is, in effect, the net result of preferential interaction of a protein with water molecules and salt ions in its vicinity. However, no general quantitative interrelation was found between salting out parameters or the number of released water molecules in protein precipitation and HIC. In other words, protein solubility and HIC retention factor could not be quantitatively interrelated, although for some proteins, regular trends were observed across the different resins and salt types. PMID:21868020

  12. Enantioseparations of primary amino compounds by high-performance liquid chromatography using chiral crown ether-based chiral stationary phase.

    PubMed

    Hyun, Myung Ho

    2013-01-01

    Liquid chromatographic resolution of racemic compounds containing a primary amino group has been known to be most successful when chiral crown ether-based chiral stationary phases (CSPs) are used. Among various crown ether-based CSPs, the stationary phase based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid covalently bonded to silica gel has been successfully applied in the resolution of various racemic compounds containing primary amino groups. In this chapter, the preparation of the CSP based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid covalently bonded to silica gel and examples for the application to the enantioseparation of racemic compounds including α-amino acids, cyclic amines, amino alcohols, and chiral drugs are described. PMID:23283776

  13. Monolayer-Protected Gold Nanoparticles as an Efficient Stationary Phase for Open Tubular Gas Chromatography using a Square Capillary Model for Chip-Based Gas Chromatography in Square Cornered Microfabricated Channels

    SciTech Connect

    Gross, Gwen M.; Grate, Jay W.); Synovec, Robert E.

    2004-03-12

    The application of a dodecanethiol monolayer protected gold nanoparticle (MPN) stationary phase within a microchannel environment was explored using a square capillary column as a model for a high-speed, microfabricated gas chromatography (?GC). Successful deposition and evaluation of a dodecanethiol MPN phase within a 1.3 m long, 100?m by 100?m square capillary is reported. Depth of the MPN phase was evaluated using SEM analysis. An average thickness of 15 nm along the capillary walls was determined. While the film depth along the walls was very uniform, the corner depths were greater with the largest observed depth being 430 nm. Overall, an efficient chromatographic system was obtained with a minimum reduced plate height, hmin, of 1.2 for octane (k= 0.22). Characterization of the MPN column was completed using four compound classes (alkanes, alcohols, ketones, and aromatics) that were used to form a 7 component mixture with a 2 second separation. A mixture consisting of a nerve agent simulator in a sample containing analytes that may commonly interfere with detection was also separated in 2 seconds, much faster than a similar separation previously reported using a?GC system in 50 seconds. Application of the square capillary MPN column for a high-speed separation as the second column of a comprehensive two-dimensional gas chromatography system (GC x GC) was also explored. Comparison of the MPN stationary phase was compared to phases employed in previously reported?GC systems.

  14. Overcoming solubility limits in overloaded gradient hydrophobic interaction chromatography.

    PubMed

    Poplewska, Izabela; Piątkowski, Wojciech; Antos, Dorota

    2015-03-20

    The impact of the solubility limits on the performance of gradient protein chromatography has been studied. As a case study elution of model protein, i.e., lysozyme, on hydrophobic interaction media has been selected. A dependence of the protein solubility and crystallization kinetics on the content of cosmotropic salt in the mobile phase has been determined. Moreover, adsorption properties of the protein versus the mobile phase composition have been quantified. A model of chromatographic column dynamics has been developed which incorporated the mass transport kinetics accompanying both adsorption and crystallization processes. The model was used to study the influence of operating parameters such as flowrate and concentration loading on the solubility pattern inside the column and the separation performance. The analysis performed indicated existence of supersaturation regions for which, due to slow kinetics of crystallization, chromatographic process could be performed under conditions of strong concentration overloading while avoiding undesirable effects of flow blockage in chromatographic systems. PMID:25687455

  15. Use of protein-protein interactions in affinity chromatography.

    PubMed

    Muronetz, V I; Sholukh, M; Korpela, T

    2001-10-30

    Biospecific recognition between proteins is a phenomenon that can be exploited for designing affinity-chromatographic purification systems for proteins. In principle, the approach is straightforward, and there are usually many alternative ways, since a protein can be always found which binds specifically enough to the desired protein. Routine immunoaffinity chromatography utilizes the recognition of antigenic epitopes by antibodies. However, forces involved in protein-protein interactions as well the forces keeping the three-dimensional structures of proteins intact are complicated, and proteins are easily unfolded by various factors with unpredictable results. Because of this and because of the generally high association strength between proteins, the correct adjustment of binding forces between an immobilized protein and the protein to be purified as well as the release of bound proteins in biologically active form from affinity complexes are the main problem. Affinity systems involving interactions like enzyme-enzyme, subunit-oligomer, protein-antibody, protein-chaperone and the specific features involved in each case are presented as examples. This article also aims to sketch prospects for further development of the use of protein-protein interactions for the purification of proteins. PMID:11694271

  16. Direct stereochemical resolution of aspartame stereoisomers and their degradation products by high-performance liquid chromatography on a chiral crown ether based stationary phase.

    PubMed

    Motellier, S; Wainer, I W

    1990-09-21

    The direct stereochemical resolution of the four stereoisomers of aspartame (N-DL-alpha-aspartyl-DL-phenylalanine methyl ester) and their degradation products was achieved on a high-performance liquid chromatography chiral stationary phase based upon a chiral crown ether. The chromatographic conditions included a mobile phase composed of aqueous perchloric acid adjusted to a pH of 2.8 and modified with 1.5% of 2-propanol and a temperature gradient. The active L,L-isomer (sold under the brand name NutraSweet) was measured in a diet cola and coffee sweetened with NutraSweet. Degradation products of NutraSweet were also detected but no racemization of stereochemical contamination was observed. PMID:2150410

  17. Characterization and utilization of a novel triflate ionic liquid stationary phase for use in comprehensive two-dimensional gas chromatography.

    PubMed

    Reid, Vanessa R; Crank, Jeffery A; Armstrong, Daniel W; Synovec, Robert E

    2008-10-01

    A novel triflate (trifluoromethylsulfonate) ionic liquid (IL) thin film (0.08 microm) stationary phase was implemented for use within the second column of a comprehensive GC x GC configuration. The first column in the configuration had a 5% phenyl/95% dimethyl polysiloxane (DMPS) stationary phase with a 0.4 microm film. The DMPS x IL column configuration was used to separate a mixture of 32 compounds of various chemical functional classes. The GC x GC results for the IL column were compared with a commercially available polar column (with a 0.1 microm PEG stationary phase film) used as the second column instead. Additional studies focused on the rapid and selective separation of four phosphorous-oxygen (P-O) containing compounds from the 32-compound matrix: dimethyl methylphosphonate (DMMP), diethyl methylphosphonate (DEMP), diisopropyl methylphosphonate (DIMP), and triethyl phosphate (TEP). van't Hoff plots (plots of ln k vs. 1/T) demonstrated the difference in retention between the P-O containing compounds (with DMMP reported in detail) and other classes of compounds (i. e., 2-pentanol and n-dodecane as representative) using either the IL column or the commercial PEG column. The selectivity (alpha) of the triflate IL column and the commercially available PEG column were also compared. The IL column provided significantly larger selectivities between DMMP and the other two compounds (2-pentanol and n-dodecane) than the commercial PEG column. The alpha for DMMP relative to n-dodecane was 3.0-fold greater for the triflate IL column, and the alpha for DMMP relative to 2-pentanol was 1.7-fold greater for the triflate IL column than for the PEG column. PMID:18798215

  18. Use of a Novel Sub-2 µm Silica Hydride Vancomycin Stationary Phase in Nano-Liquid Chromatography. II. Separation of Derivatized Amino Acid Enantiomers.

    PubMed

    Rocchi, Silvia; Fanali, Chiara; Fanali, Salvatore

    2015-11-01

    A novel vancomycin silica hydride stationary phase was synthesized and the particles of 1.8 µm were packed into fused silica capillaries of 75 µm internal diameter (I.D.). The chiral stationary phase (CSP) was tested for the separation of some derivatized amino acid enantiomers by using nano-liquid chromatography (nano-LC). Some experimental parameters such as the type and the content of organic modifier, the pH, and the concentration of the buffer added to the mobile phase were modified and the effect on enantioselectivity, retention time, and enantioresolution factor was studied. The separation of selected dansyl amino acids (Dns-AAs), e.g., Asp, Glu, Leu, and Phe in their enantiomers was initially achieved utilizing a mobile phase containing 85% (v/v) methanol (MeOH) and formate buffer measuring the enantioresolution factor and enantioselectivity in the range 1.74-4.17 and 1.39-1.59, respectively. Better results were obtained employing a more polar organic solvent as acetonitrile (ACN) in the mobile phase. Optimum results (Rs 1.41-6.09 and α 1.28-2.36) were obtained using a mobile phase containing formate buffer pH 2.5/water/MeOH/ACN 6:19:12.5:62.5 (v/v/v/v) in isocratic elution mode at flow rate of 130 nL/min. PMID:26335144

  19. Two-dimensional hydrophilic interaction chromatography × reversed-phase liquid chromatography for the preparative isolation of potential anti-hepatitis phenylpropanoids from Salvia prattii.

    PubMed

    Dang, Jun; Shao, Yun; Zhao, Jianqiang; Mei, Lijuan; Tao, Yanduo; Wang, Qilan; Zhang, Li

    2016-09-01

    Traditional Tibetan medicine is important for discovery of drug precursors. However, knowledge of the chemical composition of traditional Tibetan medicines is very limited due to the lack of appropriate chromatographic purification methods. In the present work, Salvia prattii was taken as an example, and an off-line hydrophilic interaction liquid chromatography/reversed-phase liquid chromatography preparative method was developed for the purification of phenylpropanoids with high purity from a crude sample of Salvia prattii. Based on the separation results of four different chromatographic stationary phases, the first-dimensional preparation was performed on an XAmide preparative column with the crude sample concentration of 62.0 mg/mL, and five main fractions were obtained from the 12.4 g crude sample with a recovery of 54.8%. An XCharge C18 preparative column was applied in the second-dimensional preparation to further isolate the phenylpropanoids from the redissolved first-dimensional fractions with concentration of approximately 50.0 mg/mL. The purities of the phenylpropanoids isolated from the crude sample of Salvia prattii were higher than 98%, indicating that the method was efficient for the purification of phenylpropanoids with high purity from Salvia prattii. Additionally, this method showed great potential in the preparation of phenylpropanoids and can serve as a good example for the purification of phenylpropanoids from other plant materials. PMID:27390114

  20. Novel supports in chiral stationary phase development for liquid chromatography. Preparation, characterization and application of ordered mesoporous silica particles.

    PubMed

    Sierra, Isabel; Pérez-Quintanilla, Damián; Morante, Sonia; Gañán, Judith

    2014-10-10

    Recent advances in the development of new materials are having a major impact on analytical chemistry. For example, the unique properties of ordered mesoporous silicas (OMSs) have been shown to enhance the analytical performance of many existing techniques or allow new, exciting ones to be developed. Likewise, the introduction of organo-functional groups makes OMSs highly versatile and enables them to perform specialized tasks, such as the separation of chiral compounds. This review provides an overview with the most relevant achievements in the preparation of OMS particles functionalized with chiral selectors. In addition, some examples from the last fifteen years regarding the analytical applications of functionalized OMS for chiral separations by high-performance liquid chromatography, ultra-high pressure high-performance liquid chromatography and capillary electrochromatography have been reviewed. PMID:25015243

  1. Evidence of nonlinear interaction between quasi 2 day wave and quasi-stationary wave

    NASA Astrophysics Data System (ADS)

    Gu, Sheng-Yang; Liu, Han-Li; Li, Tao; Dou, Xiankang; Wu, Qian; Russell, James M.

    2015-02-01

    The nonlinear interaction between the westward quasi 2 day wave (QTDW) with zonal wave number s = 3 (W3) and stationary planetary wave with s = 1 (SPW1) is first investigated using both Thermosphere, Ionosphere, and Mesosphere Electric Dynamics (TIMED) satellite observations and the thermosphere-ionosphere-mesosphere electrodynamics general circulation model (TIME-GCM) simulations. A QTDW with westward s = 2 (W2) is identified in the mesosphere and lower thermosphere (MLT) region in TIMED/Sounding of the Atmosphere using Broadband Emission Radiometry (SABER) temperature and TIMED/TIMED Doppler Imager (TIDI) wind observations during 2011/2012 austral summer period, which coincides with a strong SPW1 episode at high latitude of the northern winter hemisphere. The temperature perturbation of W2 QTDW reaches a maximum amplitude of ~8 K at ~30°S and ~88 km in the Southern Hemisphere, with a smaller amplitude in the Northern Hemisphere at similar latitude and minimum amplitude at the equator. The maximum meridional wind amplitude of the W2 QTDW is observed to be ~40 m/s at 95 km in the equatorial region. The TIME-GCM is utilized to simulate the nonlinear interactions between W3 QTDW and SPW1 by specifying both W3 QTDW and SPW1 perturbations at the lower model boundary. The model results show a clear W2 QTDW signature in the MLT region, which agrees well with the TIMED/SABER temperature and TIMED/TIDI horizontal wind observations. We conclude that the W2 QTDW during the 2011/2012 austral summer period results from the nonlinear interaction between W3 QTDW and SPW1.

  2. Penetrable silica microspheres for immobilization of bovine serum albumin and their application to the study of the interaction between imatinib mesylate and protein by frontal affinity chromatography.

    PubMed

    Ma, Liyun; Li, Jing; Zhao, Juan; Liao, Han; Xu, Li; Shi, Zhi-guo

    2016-01-01

    In the current study, novel featured silica, named penetrable silica, simultaneously containing macropores and mesopores, was immobilized with bovine serum albumin (BSA) via Schiff base method. The obtained BSA-SiO2 was employed as the high-performance liquid chromatographic (HPLC) stationary phase. Firstly, D- and L-tryptophan were used as probes to investigate the chiral separation ability of the BSA-SiO2 stationary phase. An excellent enantioseparation factor was obtained up to 4.3 with acceptable stability within at least 1 month. Next, the BSA-SiO2 stationary phase was applied to study the interaction between imatinib mesylate (IM) and BSA by frontal affinity chromatography. A single type of binding site was found for IM with the immobilized BSA, and the hydrogen-bonding and van der Waals interactions were expected to be contributing interactions based on the thermodynamic studies, and this was a spontaneous process. Compared to the traditional silica for HPLC stationary phase, the proposed penetrable silica microsphere possessed a larger capacity to bond more BSA, minimizing column overloading effects and enhancing enantioseparation ability. In addition, the lower running column back pressure and fast mass transfer were meaningful for the column stability and lifetime. It was a good substrate to immobilize biomolecules for fast chiral resolution and screening drug-protein interactions. PMID:26573171

  3. Synthesis of cis-C-Iodo-N-Tosyl-Aziridines using Diiodomethyllithium: Reaction Optimization, Product Scope and Stability, and a Protocol for Selection of Stationary Phase for Chromatography

    PubMed Central

    2013-01-01

    The preparation of C-iodo-N-Ts-aziridines with excellent cis-diastereoselectivity has been achieved in high yields by the addition of diiodomethyllithium to N-tosylimines and N-tosylimine–HSO2Tol adducts. This addition-cyclization protocol successfully provided a wide range of cis-iodoaziridines, including the first examples of alkyl-substituted iodoaziridines, with the reaction tolerating both aryl imines and alkyl imines. An ortho-chlorophenyl imine afforded a β-amino gem-diiodide under the optimized reaction conditions due to a postulated coordinated intermediate preventing cyclization. An effective protocol to assess the stability of the sensitive iodoaziridine functional group to chromatography was also developed. As a result of the judicious choice of stationary phase, the iodoaziridines could be purified by column chromatography; the use of deactivated basic alumina (activity IV) afforded high yield and purity. Rearrangements of electron-rich aryl-iodoaziridines have been promoted, selectively affording either novel α-iodo-N-Ts-imines or α-iodo-aldehydes in high yield. PMID:23738857

  4. Ionic liquid-based zwitterionic organic polymer monolithic column for capillary hydrophilic interaction chromatography.

    PubMed

    Wang, Tingting; Chen, Yihui; Ma, Junfeng; Zhang, Xiaodan; Zhang, Lihua; Zhang, Yukui

    2015-08-21

    In the current study, a novel ionic liquid-based zwitterionic organic polymer monolithic column was developed by copolymerizing 1-vinyl-3-(butyl-4-sulfonate) imidazolium, acrylamide and N,N'-methylenebisacrylamide in a quaternary porogenic solvent consisting of formamide, dimethyl sulphoxide, polyethylene glycol 8000 and polyethylene glycol 10,000 for capillary hydrophilic interaction chromatography. The monolithic stationary phase was optimized by adjusting the amount of monomer in the polymerization solution along with the composition of porogenic solvent. The optimized monolith exhibited excellent selectivity and favorable retention for nucleosides and benzoic acid derivatives. The primary factors affecting the separation efficiency of the monolithic column (including acetonitrile content, pH, and buffer salt concentration in the mobile phase) have been thoroughly evaluated. Excellent reproducibility of the retention times for five nucleosides was achieved, with relative standard deviations of run-to-run (n = 3), column-to-column (n = 3) and batch-to-batch (n = 3) in the range of 0.18-0.48%, 2.33-4.20% and 3.07-6.50%, respectively. PMID:26114194

  5. Comparison of peak shape in hydrophilic interaction chromatography using acidic salt buffers and simple acid solutions.

    PubMed

    Heaton, James C; Russell, Joseph J; Underwood, Tim; Boughtflower, Robert; McCalley, David V

    2014-06-20

    The retention and peak shape of neutral, basic and acidic solutes was studied on hydrophilic interaction chromatography (HILIC) stationary phases that showed both strong and weak ionic retention characteristics, using aqueous-acetonitrile mobile phases containing either formic acid (FA), ammonium formate (AF) or phosphoric acid (PA). The effect of organic solvent concentration on the results was also studied. Peak shape was good for neutrals under most mobile phase conditions. However, peak shapes for ionised solutes, particularly for basic compounds, were considerably worse in FA than AF. Even neutral compounds showed deterioration in performance with FA when the mobile phase water concentration was reduced. The poor performance in FA cannot be entirely attributed to the negative impact of ionic retention on ionised silanols on the underlying silica base materials, as results using PA at lower pH (where their ionisation is suppressed) were inferior to those in AF. Besides the moderating influence of the salt cation on ionic retention, it is likely that salt buffers improve peak shape due to the increased ionic strength of the mobile phase and its impact on the formation of the water layer on the column surface. PMID:24813934

  6. Assessment of the retention properties of poly(vinyl alcohol) stationary phase for lipid class profiling in liquid chromatography.

    PubMed

    Deschamps, F S; Chaminade, P; Ferrier, D; Baillet, A

    2001-09-14

    Potentialities of polymerized vinyl alcohol on silica gel were assessed for class separation of simple lipids, sphingolipids, glyceroglycolipids and phospholipids by high-performance liquid chromatography. A screening of pure solvents in binary gradient elution and a chemometric approach was used to define a rugged two segment linear gradient formed from four solvents for total lipid class separation. Triethylamine and formic acid were added in all mobile phase components for acidic phospholipid separation and evaporative light scattering response enhancement. Simple analytical procedures are described for the analysis of complex lipid materials. PMID:11587330

  7. Chromatography

    MedlinePlus

    ... a way of separating two or more chemical compounds. Chemical compounds are chemicals that are bonded together. For example, ... and hydrogen. Proteins are another type of chemical compound. There are different kinds of chromatography. These include ...

  8. Characterization of post-translationally modified peptides by hydrophilic interaction and reverse phase liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry.

    PubMed

    Hernandez-Hernandez, Oswaldo; Quintanilla-Lopez, Jesus Eduardo; Lebron-Aguilar, Rosa; Sanz, Maria Luz; Moreno, F Javier

    2016-01-01

    This work explores the use of both hydrophilic interaction liquid chromatography (HILIC) and reverse phase liquid chromatography (RPLC) for the separation and subsequent characterization of bovine caseinomacropeptide (CMP) phosphopeptides and O-glycopeptides using a quadrupole-time-of-flight (QTOF) mass spectrometer with electrospray ionization. Two neutral, ethylene bridged hybrid (BEH) amide and polyhydroxyethyl aspartamide (PHEA), and a zwitterionic, sulfobetaine (ZIC), stationary phases were used for the HILIC mode, whilst an octadecylsilane (C18) stationary phase was employed for the RPLC separation. Overall, developed HILIC-QTOF method using the ZIC or BEH amide stationary phases resulted to be the most efficient methods to separate and characterize post-translationally modified (PTM) peptides without the need of any previous fractionation or derivatization step. The separation of phosphopeptides and differently sialylated O-glycopeptides in the ZIC stationary phase was dominated by an electrostatic repulsion interaction mechanism between the negatively charged phosphate groups or sialic acid moieties and the negatively charged terminal sulfonate group of the stationary phase, whereas the separation of either non-modified peptides or neutral O-glycopeptides both free of basic amino acids was based on a partitioning mechanism. In neutral amide columns, the separation was mainly dominated by hydrophilic partitioning, leading to a higher retention of the post-translationally modified peptides than the unmodified counterparts due to the hydrophilicity provided by the phosphate groups and/or O-glycans. As a consequence, HILIC-ESI-QTOF MS operating in the positive ion mode is a powerful tool for the characterization of underivatized O-glycopeptides and phosphopeptides. PMID:26278355

  9. Thiol-ene click chemistry derived cationic cyclodextrin chiral stationary phase and its enhanced separation performance in liquid chromatography.

    PubMed

    Yao, Xiaobin; Tan, Timothy Thatt Yang; Wang, Yong

    2014-01-24

    This work is the first demonstration of a simple thiol-ene click chemistry to anchor vinyl imidazolium β-CD onto thiol silica to form a novel cationic native cyclodextrin (CD) chiral stationary phase (CSP). The CSP afforded high enantioseparation ability towards dansyl (Dns) amino acids, carboxylic aryl compounds and flavonoids in chiral HPLC. The current CSP demonstrates the highest resolving ability (selectivity >1.1, resolution >1.5) towards Dns amino acids in a mobile phase buffered at pH=6.5, with the resolution of Dns-dl-leucine as high as 6.97. 2,4-dichloride propionic acid (2,4-ClPOPA) was well resolved with the selectivity and resolution of 1.37 and 4.88, respectively. Compared to a previously reported native CD-CSP based on a triazole linkage, the current cationic CD-CSP shows a stronger retention and higher resolution towards acidic chiral compounds, ascribed to the propitious strong electrostatic attraction. Stability evaluation results indicated that thiol-ene reaction can provide a facile and robust approach for the preparation of positively charged CD CSPs. PMID:24411138

  10. Some factors that can lead to poor peak shape in hydrophilic interaction chromatography, and possibilities for their remediation.

    PubMed

    Heaton, James C; McCalley, David V

    2016-01-01

    Some factors which present difficulties for obtaining good peak shape in hydrophilic interaction chromatography (HILIC) were studied. The effect of injection solvent composition and volume was systematically investigated using a selection of weak and stronger basic compounds on a hybrid bare silica phase. Increasing the mismatch between the injection solvent (range 95-0% ACNv/v) and the mobile phase (maintained at 95% ACNv/v) gave increasing deterioration in peak shape. With the 2.1mm ID columns used, injections in the mobile phase of increasing volume (1-20 μL) gave poorer peak shape, but the magnitude of the effect was considerably smaller than that of solvent mismatch over this range. Some solute structural features such as galloyl (trihydroxy benzene), catechol (benzene diol) and phosphate (in nucleotides) gave serious peak tailing, attributed to interactions with metals in the stationary phase or the chromatographic hardware. These undesirable effects can be moderated by including complexing agents in the mobile phase, by changing the stationary phase chemistry, or by altering the mobile phase pH. PMID:26689823

  11. Fabrication of ZIF-8@SiO2 core-shell microspheres as the stationary phase for high-performance liquid chromatography.

    PubMed

    Fu, Yan-Yan; Yang, Cheng-Xiong; Yan, Xiu-Ping

    2013-09-27

    The unique features of high porosity, shape selectivity, and multiple active sites make metal-organic frameworks (MOFs) promising as novel stationary phases for high-performance liquid chromatography (HPLC). However, the wide particle size distribution and irregular shape of conventional MOFs lead to lower column efficiency of such MOF-packed columns. Herein, the fabrication of monodisperse MOF@SiO2 core-shell microspheres as the stationary phase for HPLC to overcome the above-mentioned problems is reported. Zeolitic imidazolate framework 8 (ZIF-8) was used as an example of MOFs due to its permanent porosity, uniform pore size, and exceptional chemical stability. Unique carboxyl-modified silica spheres were used as the support to grow the ZIF-8 shell. The fabricated monodisperse ZIF-8@SiO2 packed columns (5 cm long × 4.6 mm i.d.) show high column efficiency (23,000 plates m(-1) for bisphenol A) for the HPLC separation of endocrine-disrupting chemicals (bisphenol A, β-estradiol, and p-(tert-octyl)phenol) and pesticides (thiamethoxam, hexaflumuron, chlorantraniliprole, and pymetrozine) within 7 min with good relative standard deviations for 11 replicate separations of the analytes (0.01-0.39, 0.65-1.7, 0.70-1.3, and 0.17-0.91% for retention time, peak area, peak height, and half peak width, respectively). The ZIF-8@SiO2 microspheres combine the advantages of the good column packing properties of the uniform monodisperse silica microspheres and the separation ability of the ZIF-8 crystals. PMID:23955766

  12. Comparison between polymerized ionic liquids synthesized using chain-growth and step-growth mechanisms used as stationary phase in gas chromatography.

    PubMed

    Roeleveld, Kevin; David, Frank; Lynen, Frédéric

    2016-06-17

    In this study the merits of polymerized imidazolium based ionic liquid (PIL) stationary phases obtained via condensation and free radical polymerizations are compared as stationary phases in gas chromatography (GC). Poly(1-vinyl-3-butyl-imidazolium - bis(trifluoromethane)sulfonamide) (poly(ViC4Im(+) NTf2(-))) was obtained via a chain-growth mechanism while poly(propylimidazolium-NTf2) (poly(C3Im(+) NTf2(-))) was synthesized via a step-growth polymerization. The thermal stability of both polymers was assessed using thermal gravimetric analysis and compared with bleeding profiles obtained from the statically coated GC columns (30m×0.25mm×0.25μm). The performance was compared to what could be obtained on commercially available 1,5-di(2,3-dimethylimidazolium)pentane(2+) 2NTf2(-) (SLB-IL111) ionic liquid based columns. It was observed that the step-growth polymer was more thermally stable, up to 325°C, while the chain-growth polymer showed initial degradation at 250°C. Both polymers allowed reaching minimal plate heights of 0.400-0.500mm for retained solutes such as benzaldehyde, acetophenone, 1-methylnaphthalene and aniline. Assessment of the McReynolds constants illustrated that the polarity of the step-growth polymer was similar to the SLB-IL111 column, while displaying improved column stability. The PIL phases and particularly the so far little studied condensation based polymer shows particular retention and satisfactory column performance for polar moieties such as esters, amine and carbonyl functionalities. PMID:27189433

  13. Aqueous size exclusion chromatography in semimicro and micro-columns by newly synthesized monodisperse macroporous hydrophilic beads as a stationary phase.

    PubMed

    Gölgelioğlu, Ciğdem; Bayraktar, Aslıhan; Celebi, Bekir; Uğuzdoğan, Erdal; Tuncel, Ali

    2012-02-10

    A new class of monodisperse macroporous beads in the hydrophilic form were synthesized by seeded microsuspension copolymerization of two acrylic crosslinking agents, glycerol dimethacrylate (GDMA) and glycerol-1,3-diglycerolate diacrylate (GDGDA). The monodisperse porous poly(glycerol dimethacrylate-co-glycerol-1,3-diglycerolate diacrylate), poly(GDMA-co-GDGDA) beads were highly hydrophilic in nature due to hydroxyl functionality resulting from both crosslinking agents. The beads with different particle sizes between 4.5 and 6.7 μm and with different porous properties were obtained by changing the seed latex to monomer ratio. The bead size decreased, the average pore size increased and the specific surface area decreased with increasing seed latex to monomer ratio. Poly(GDMA-co-GDGDA) beads were slurry packed in microbore and semimicro-HPLC columns and successfully used as a stationary phase in aqueous size exclusion chromatography (SEC) mode in a micro-liquid chromatography system. The aqueous SEC runs were performed by using dextran standards in the molecular weight range of 1000-670,000 Da. SEC calibration curves exhibiting linearity in a wider range of molecular weight were obtained with the semi-micro and micro-HPLC columns packed with the poly(GDMA-co-GDGDA) beads synthesized with the seed latex to monomer ratios of 0.038 and 0.058 g/mL. The dextran standards could be eluted in an analysis time shorter than 2 min using micro or semi-micro columns packed with poly(GDMA-co-GDGDA) beads as stationary medium. These packings are suitable for molecular weight determination between 5×10(3) and 5×10(5) Da in the aqueous medium by using mobile phase flow rates in the range of 25-250 μL/min. The average molecular weight determinations of different water soluble polymers, an ionic polymer, poly(2-dimethylaminoethyl methacrylate), a zwitterionic polymer, poly([2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide), and a non-ionic polymer, poly

  14. Chromatographic performance of synthetic polycrystalline diamond as a stationary phase in normal phase high performance liquid chromatography.

    PubMed

    Peristyy, Anton; Paull, Brett; Nesterenko, Pavel N

    2015-04-24

    The chromatographic properties of high pressure high temperature synthesised diamond (HPHT) are investigated in normal phase mode of high performance liquid chromatography. Purified nonporous irregular shape particles of average particles size 1.2 μm and specific surface area 5.1 m(2) g(-1) were used for packing 100×4.6 mm ID or 50×4.6 mm ID stainless steel columns. The retention behaviour of several classes of compounds including alkyl benzenes, polyaromatic hydrocarbons (PAH), alkylphenylketones, phenols, aromatic acids and bases were studied using n-hexane-2-propanol mixtures as mobile phase. The results are compared with those observed for microdispersed sintered detonation nanodiamond (MSDN) and porous graphitic carbon (PGC). HPHT diamond revealed distinctive separation selectivity, which is orthogonal to that observed for porous graphitic carbon; while selectivities of HPHT diamond and microdispersed sintered detonation nanodiamonds are similar. Owing to non-porous particle nature, columns packed with high pressure high temperature diamond exhibited excellent mass transfer and produce separations with maximum column efficiency of 128,200 theoretical plates per meter. PMID:25777051

  15. Determination of pore size distributions in capillary-channeled polymer fiber stationary phases by inverse size-exclusion chromatography and implications for fast protein separations.

    PubMed

    Wang, Zhengxin; Marcus, R Kenneth

    2014-07-18

    Capillary-channeled polymer (C-CP) fibers have been utilized as liquid chromatography stationary phases, primarily for biomacromolecule separations on the analytical and preparative scales. The collinear packing of the eight-channeled C-CP fibers provides for very efficient flow, allowing operation at high linear velocity (u>100mm s(-1)) and low backpressure (<2000psi) in analytical-scale separations. To take advantage of these fluid transport properties, there must not be mass transfer limitations as would be imposed by having an appreciably porous phase, wherein solute diffusion limits the overall mass transport rates. To better understand the physical nano-/micro- structure of C-CP fibers, inverse size exclusion chromatography (iSEC) has been employed to determine the pore size distribution (PSD) within C-CP fibers. A diversity of test species (from metal ions to large proteins) was used as probes under non-retaining conditions to obtain a response curve reflecting the apparent partition coefficient (Kd) versus hydrodynamic radii (rm). A mean pore radius (rp) of 4.2nm with standard deviation (sp) of ±1.1nm was calculated by fitting the Kd versus rm data to model equations with a Gaussian pore size distribution, and a pore radius of 4.0±0.1nm was calculated based on a log-normal distribution. The derived mean pore radius is much smaller than traditional support materials, with the standard deviation showing a relatively uniform pore distribution. van Deemter plots were analyzed to provide practical confirmation of the structural implications. Large molecules (e.g., proteins) that are fully excluded from pores have no significant C-terms in the van Deemter plots whereas small molecules that can access the pore volumes display appreciable C-terms, as expected. Fitting of retention data to the Knox equation suggests that the columns operate with a characteristic particle diameter (dp) of ∼53μm. PMID:24877979

  16. Evaluation of stationary phases packed with superficially porous particles for the analysis of pharmaceutical compounds using supercritical fluid chromatography.

    PubMed

    Perrenoud, Alexandre Grand-Guillaume; Farrell, William P; Aurigemma, Christine M; Aurigemma, Nicole C; Fekete, Szabolcs; Guillarme, Davy

    2014-09-19

    Superficially porous particles (SPP), or core shell particles, which consist of a non-porous silica core surrounded by a thin shell of porous silica, have gained popularity as a solid support for chromatography over the last decade. In the present study, five unbonded silica, one diol, and two ethylpyridine (2-ethyl and 4-ethyl) SPP columns were evaluated under SFC conditions using two mixtures, one with 17 drug-like compounds and the other one with 7 drug-like basic compounds. Three of the SPP phases, SunShell™ 2-ethylpyridine (2-EP), Poroshell™ HILIC, and Ascentis(®) Express HILIC, exhibited superior performances relative to the others (reduced theoretical plate height (hmin) values of 1.9-2.5 for neutral compounds). When accounting for both achievable plate count and permeability of the support using kinetic plot evaluation, the Cortecs™ HILIC 1.6μm and Ascentis(®) Express HILIC 2.7μm phases were found to be the best choices among tested SPPs to reach efficiencies up to 30,000 plates in the minimum amount of time. For desired efficiencies ranging from 30,000 to 60,000 plates, the SunShell™ 2-EP 2.6μm column clearly outperformed all other SPPs. With the addition of a mobile phase additive such as 10mM ammonium formate, which was required to elute the basic components with sharp peaks, the Poroshell™ HILIC, SunShell™ Diol and SunShell™ 2-EP phases represent the most orthogonal SPP columns with the highest peak capacities. This study demonstrates the obvious benefits of using columns packed with SPP on current SFC instrumentation. PMID:25129391

  17. Synthesis of chitosan 3,6-diphenylcarbamate-2-urea derivatives and their applications as chiral stationary phases for high-performance liquid chromatography.

    PubMed

    Zhang, Lili; Shen, Jun; Zuo, Wenli; Okamoto, Yoshio

    2014-10-24

    Fourteen chitosan 3,6-diphenylcarbamate-2-urea derivatives were synthesized using well-deacetylated chitosan and the corresponding phenyl isocyanates. After coating them on silica gel, their chiral recognition abilities were evaluated as the chiral stationary phases (CSPs) for high-performance liquid chromatography. These coated-type CSPs exhibited different chiral recognitions depending on the position, nature, and number of the substituents introduced on the phenyl group, and the introduction of either an electron-withdrawing or an electron-donating substituent improved the chiral recognition of the CSPs. Among the CSPs, the 2-substituted CSPs showed low chiral recognition abilities, while those with 3,5-dimethyl and 3,5-dichloro substituents showed relatively higher chiral recognition abilities, which enabled the baseline separation of some racemates. The CSPs could be used with some eluents containing chloroform, which cannot be used for other polysaccharide-based CSPs. Some racemates were more efficiently resolved with these nonstandard eluents. The correlation between the chiral recognition ability and the chemical shifts of the N-H protons in the (1)H NMR spectra of the chitosan derivatives or the N-H frequencies in the IR spectra of the carbamate moieties was discussed. PMID:25262030

  18. Evaluation of new cellulose-based chiral stationary phases Sepapak-2 and Sepapak-4 for the enantiomeric separation of pesticides by nano liquid chromatography and capillary electrochromatography.

    PubMed

    Pérez-Fernández, Virginia; Dominguez-Vega, Elena; Chankvetadze, Bezhan; Crego, Antonio L; García, Maria Ángeles; Marina, Maria Luisa

    2012-04-20

    Two novel polysaccharide-based chiral stationary phases (CSPs), known as Sepapak-2 (cellulose tris(3-chloro-4-methylphenylcarbamate)) and Sepapak-4 (cellulose tris(4-chloro-3-methylphenylcarbamate)), have been evaluated in this work for the chiral separation of a group of 16 pesticides including herbicides, insecticides and fungicides. The optimization of the mobile phase employed in nano-liquid chromatography (nano-LC) enabled the chiral separation of seven pesticides on Sepapak-2 and of nine pesticides on Sepapak-4. Due to the fact that Sepapak-4 gave better results, this column was selected to compare nano-LC and capillary electrochromatography (CEC) under the same conditions that consisted in the use of a 90/9/1 (v/v/v) ACN/H₂O/ammonium formate (pH 2.5) background electrolyte (BGE). As expected, both the efficiency and the chiral resolution obtained in CEC experiments were higher than in nano-LC for all the analyzed compounds. The analytical characteristics of the CEC developed methodology were evaluated in terms of linearity, LODs, LOQs, precision, selectivity, and accuracy allowing its application to the quantitation of metalaxyl and its enantiomeric impurity in a commercial fungicide product marketed as enantiomerically pure (metalaxyl-M) and in soil and tap water samples after solid phase extraction (SPE). The determined amount of metalaxyl-M was found to be a 26% above the labeled content and it contained an enantiomeric impurity of a 3.7% of S-metalaxyl was determined. PMID:22321947

  19. Application of a cholesterol stationary phase in the analysis of phosphorothioate oligonucleotides by means of ion pair chromatography coupled with tandem mass spectrometry.

    PubMed

    Studzińska, Sylwia; Krzemińska, Katarzyna; Szumski, Michał; Buszewski, Bogusław

    2016-07-01

    The main aim of this study was the investigation of the influence of several ion pair reagents towards both the retention and the mass spectrometry sensitivity of phosphorothioate oligonucleotides. A cholesterol stationary phase was applied for the first time in the analysis of this group of compounds. The mobile phase composition was modified by changing the concentration and the type of amines and acetates or 1,1,1,3,3,3-hexafluoroisopropanol. It has been shown that the increase of amines concentration results in the retention factor increase for each oligonucleotide, on each adsorbent. The only exception was the mobile phase composed of triethylamine and 1,1,1,3,3,3-hexafluoroisopropanol. This is a consequence of interactions taking place between a cholesterol molecule and an alcohol. This effect was convenient when the mass spectrometry detection was applied, since it allowed an increase in the sensitivity. Moreover, optimization of the mobile phase composition and its impact on the efficiency of ionization process and on the sensitivity in mass spectrometry were also presented. The optimization of this new method, based on cholesterol stationary phase coupled with mass spectrometry detection, was finally applied for the determination of phosphorothioate oligonucleotides impurity in a real sample. PMID:27154674

  20. Overdamped motion of interacting particles in general confining potentials: time-dependent and stationary-state analyses

    NASA Astrophysics Data System (ADS)

    Ribeiro, M. S.; Nobre, F. D.; Curado, E. M. F.

    2012-12-01

    By comparing numerical and analytical results, it is shown that a system of interacting particles under overdamped motion is very well described by a nonlinear Fokker-Planck equation, which can be associated with nonextensive statistical mechanics. The particle-particle interactions considered are repulsive, motivated by three different physical situations: (i) modified Bessel function, commonly used in vortex-vortex interactions, relevant for the flux-front penetration in disordered type-II superconductors; (ii) Yukawa-like forces, useful for charged particles in plasma, or colloidal suspensions; (iii) derived from a Gaussian potential, common in complex fluids, like polymer chains dispersed in a solvent. Moreover, the system is subjected to a general confining potential, φ( x) = ( α| x| z )/ z ( α > 0 , z > 1), so that a stationary state is reached after a sufficiently long time. Recent numerical and analytical investigations, considering interactions of type (i) and a harmonic confining potential ( z = 2), have shown strong evidence that a q-Gaussian distribution, P( x,t), with q = 0, describes appropriately the particle positions during their time evolution, as well as in their stationary state. Herein we reinforce further the connection with nonextensive statistical mechanics, by presenting numerical evidence showing that: (a) in the case z = 2, different particle-particle interactions only modify the diffusion parameter D of the nonlinear Fokker-Planck equation; (b) for z ≠ 2, all cases investigated fit well the analytical stationary solution P st( x), given in terms of a q-exponential (with the same index q = 0) of the general external potential φ( x). In this later case, we propose an approximate time-dependent P( x,t) (not known analytically for z ≠ 2), which is in very good agreement with the simulations for a large range of times, including the approach to the stationary state. The present work suggests that a wide variety of physical phenomena

  1. Group separation of transplutonium and rare-earth elements by liquid chromatography with a free stationary phase using 2,4,6-Tris[ditolylphosphoryl]-1,3,5-triazine

    SciTech Connect

    Chmutova, M.K.; Ivanova, L.A.; Bodrin, G.B.

    1995-03-01

    Methods are developed for group separation of trace quantities of transplutonium (TPE) and weighable amounts of rare-earth elements (REE) by liquid chromatography with a free stationary phase in systems based on bifunctional neutral organophosphorus compounds. Using a stationary phase of 2,4,6-tris(ditolylphosphoryl)-1,3,5-triazine in CHCl{sub 3}, REE are first eluted by 0.5 M NH{sub 4}SCN-1 M HCl and then TPE by 0.025 M hydroxyethylidenediphosphonic acid in H{sub 2}O. The fractions contained {approximately} 100% of one of the groups without an impurity of the other. Use of the same eluents and a CHCl{sub 3} solution of tetraphenyl-methylenediphosphine dioxide as the stationary phase gave 95.4% pure REE and 97.5% pure TPE.

  2. Cationic Ionic Liquids Organic Ligands Based Metal-Organic Frameworks for Fabrication of Core-Shell Microspheres for Hydrophilic Interaction Liquid Chromatography.

    PubMed

    Dai, Qian; Ma, Junqian; Ma, Siqi; Wang, Shengyu; Li, Lijun; Zhu, Xianghui; Qiao, Xiaoqiang

    2016-08-24

    In this study, new metal-organic frameworks (MOFs) nanocrystals modified SiO2 core-shell microspheres were designed with cationic ionic liquids (ILs) 1,3-bis(4-carboxybutyl)imidazolium bromide (ILI) as organic ligands. By further adjustment the growth cycles, the new ILI-01@SiO2 core-shell stationary phase was facilely fabricated. The developed stationary phase was respectively characterized via element analysis, thermogravimetric analysis, scanning electron microscopy, X-ray diffraction, and Fourier transform infrared spectrometry. Because the introduction of cationic imidazolium-based ILs ILI for fabrication of the MOFs nanocrystals shell, the new stationary phase exhibits the retention mechanism of hydrophilic interaction liquid chromatography (HILIC). Many polar samples, such as amides, vitamins, nucleic acid bases, and nucleosides, were utilized to investigate the performance of the prepared ILI-01@SiO2 column. Compared to the conventional aminosilica column, the new ILI-01@SiO2 column displays high separation selectivity in a shorter separation time. Furthermore, the new ILI-01@SiO2 column was also used for detection of illegal melamine addition in the baby formula. All the above results demonstrate the new ILI-01@SiO2 core-shell stationary phase is of good potentials for high-selectivity separation the polar samples. PMID:27483161

  3. Hydrophilic interaction chromatography of seized drugs and related compounds with sub 2 μm particle columns.

    PubMed

    Lurie, Ira S; Li, Li; Toske, Steven G

    2011-12-30

    The use of hydrophilic interaction chromatography (HILIC) with sub 2 μm particle columns for the analysis of drugs and related compounds of forensic interest is described. This technique uses a high organic/low aqueous buffered mobile phase with a polar stationary phase, and is excellent for the separation of many of the charged solutes that are found in forensic drug exhibits. In this study, HILIC is investigated for 11 solutes of forensic interest, including weak bases, weak acids, and a neutral solute. In addition, for columns containing either ethylene bridged hybrid particles with or without an amide bonded phase, the effects of acetonitrile concentration, buffer type, buffer concentration, linear velocity, and sample concentration were studied. Based on these studies, HILIC with sub 2 μm particle columns can offer highly efficient, selective, and rapid isocratic separations of drugs and related compounds of forensic interest, with excellent peak shapes and low back pressures. This is in contrast to reverse phase chromatography (RPLC), where gradient elution is usually required, which can result in extensive overlap between acidic, neutral, and basic solutes. In addition, since HILIC exhibits a much greater loading capacity than RPLC, it could be a preferred technique for drug profiling. Furthermore, because high organic content mobile phases are highly amenable to mass spectrometric detection, the use of HILIC with tandem mass spectrometric detection for the analysis of seized drugs is described. PMID:22098930

  4. Theory of stationary solitary waves generated by optical parametric interactions in the presence of Kerr-type nonlinearities and dissipations

    NASA Astrophysics Data System (ADS)

    Hayata, K.; Koshiba, M.

    1995-11-01

    We show analytically that inclusion of contributions from third-order nonlinearities in a theoretical model for optical parametric interactions derived from second-order nonlinearities makes possible the prediction of various kinds of stationary solitary-wave solution. Specifically these waves consist of hyperbolic (bright and dark types), algebraic (bright and dark types), and kink/antikink types. In the limit of vanishing third-order nonlinearities the first solitary-wave family (hyperbolic type) is reduced to solitary waves already reported. Effects of dissipations including one- and two-photon absorption are discussed as well. Copyright (c) 1995 Optical Society of America

  5. Adsorption mechanism in reversed-phase liquid chromatography. Effect of the surface coverage of a monomeric C18-silica stationary phase

    SciTech Connect

    Gritti, Fabrice; Guiochon, Georges A

    2006-04-01

    The effect of the bonding density of the octadecyl chains onto the same silica on the adsorption and retention properties of low molecular weight compounds (phenol, caffeine, and sodium 2-naphthalene sulfonate) was investigated. The same mobile phase (methanol:water, 20:80, v/v) and temperature (T = 298 K) were applied and two duplicate columns (A and B) from each batch of packing material (neat silica, simply endcapped or C{sub 1} phase, 0.42, 1.01, 2.03, and 3.15 {micro}mol/m{sup 2} of C{sub 18} alkyl chains) were tested. Adsorption data of the three compounds were acquired by frontal analysis (FA) and the adsorption energy distributions (AEDs) were calculated using the expectation-maximization method. Results confirmed earlier findings in linear chromatography of a retention maximum at an intermediate bonding density. From a general point of view, the saturation capacity of the adsorbent tends to decrease with increasing bonding density, due to the vanishing space intercalated between the C{sub 18} bonded chains and to the decrease of the specific surface area of the stationary phase. The equilibrium constants are maximum for an intermediary bonding density ({approx}2 {micro}mol/m{sup 2}). An enthalpy-entropy compensation was found for the thermodynamic parameters of the isotherm data. Weak equilibrium constants (small {Delta}H) and high saturation capacities (large {Delta}S) were observed at low bonding densities, higher equilibrium constants and lower saturation capacities at high bonding densities, the combinations leading to similar apparent retention in RPLC. The use of a low surface coverage column is recommended for preparative purposes.

  6. Hydrophilic interaction liquid chromatography for the separation of acidic agricultural compounds.

    PubMed

    Yang, Peilin; Pursch, Matthias

    2015-07-01

    Organic acids with very low pKa require extremely low pH conditions to achieve adequate retention in reversed-phase liquid chromatography, but an extremely low pH mobile phase can cause instrument reliability problems and limit the choice of columns. Hydrophilic interaction chromatography is a potential alternative to reversed-phase liquid chromatography for the separation of organic acids using more moderate conditions. However, the hydrophilic interaction chromatography separation mechanism is known to be very complex and involves multiple competing mechanisms. In the present study, a hydrophilic interaction chromatography column packed with bare silica core-shell particles was used as the separation column and six agricultural organic acids were used as model analytes to evaluate the effects of buffer concentration, buffer pH, and temperature on sample loading capacity, selectivity, retention, and repeatability. It was found that using a higher concentration of buffer can lead to a significant improvement in the overall performance and reproducibility of the separation. Investigation of column equilibration time revealed that a very long equilibration time is needed when changing mobile phase conditions in between runs. This limitation needs to be acknowledged in hydrophilic interaction chromatography method development and sufficient equilibration time needs to be allowed in method scouting. PMID:25907680

  7. A chromatographic estimate of the degree of surface heterogeneity of reversed-phase liquid chromatography packing materials II-Endcapped monomeric C18-bonded stationary phase

    SciTech Connect

    Gritti, Fabrice; Guiochon, Georges A

    2006-01-01

    In a previous report, the heterogeneity of a non-endcapped C{sub 30}-bonded stationary phase was investigated, based on the results of the measurements of the adsorption isotherms of two neutral compounds (phenol and caffeine) and two ionizable compounds (sodium naphthalene sulfonate and propranololium chloride) by frontal analysis (FA). The same method is applied here for the characterization of the surface heterogeneity of two new brands of endcapped C{sub 18}-bonded stationary phases (Gemini and Sunfire). The adsorption isotherms of the same four chemicals were measured by FA and the results confirmed by the independent calculation of the adsorption energy distribution (AED), using the expectation-maximization (EM) method. The effect of the length of the bonded alkyl chain was investigated. Shorter alkyl-bonded-chains (C{sub 18} versus C{sub 30}) and the end-capping of the silica surface contribute to decrease the surface heterogeneity under the same experimental conditions (30% methanol, 25 mM NaCl). The AEDs of phenol and caffeine are bimodal with the C{sub 18}-bonded columns while they are trimodal and quadrimodal, respectively, with a non-endcapped C{sub 30}-bonded column. The 'supersites' (adsorption energy >20 kJ/mol) found on the C{sub 30}-Prontosil column and attributed to a cation exchange mechanism completely disappear on the C{sub 18}-Gemini and C{sub 18}-Sunfire, probably because the end-capping of the silica surface eliminates most if not all the ionic interactions.

  8. Frontal affinity chromatography: A unique research tool for biospecific interaction that promotes glycobiology

    PubMed Central

    KASAI, Kenichi

    2014-01-01

    Combination of bioaffinity and chromatography gave birth to affinity chromatography. A further combination with frontal analysis resulted in creation of frontal affinity chromatography (FAC). This new versatile research tool enabled detailed analysis of weak interactions that play essential roles in living systems, especially those between complex saccharides and saccharide-binding proteins. FAC now becomes the best method for the investigation of saccharide-binding proteins (lectins) from viewpoints of sensitivity, accuracy, and efficiency, and is contributing greatly to the development of glycobiology. It opened a door leading to deeper understanding of the significance of saccharide recognition in life. The theory is also concisely described. PMID:25169774

  9. Frontal affinity chromatography: a unique research tool for biospecific interaction that promotes glycobiology.

    PubMed

    Kasai, Kenichi

    2014-01-01

    Combination of bioaffinity and chromatography gave birth to affinity chromatography. A further combination with frontal analysis resulted in creation of frontal affinity chromatography (FAC). This new versatile research tool enabled detailed analysis of weak interactions that play essential roles in living systems, especially those between complex saccharides and saccharide-binding proteins. FAC now becomes the best method for the investigation of saccharide-binding proteins (lectins) from viewpoints of sensitivity, accuracy, and efficiency, and is contributing greatly to the development of glycobiology. It opened a door leading to deeper understanding of the significance of saccharide recognition in life. The theory is also concisely described. PMID:25169774

  10. On the interaction of stationary crossflow vortices and Tollmien-Schlichting waves in the boundary layer on a rotating disc

    NASA Technical Reports Server (NTRS)

    Bassom, Andrew P.; Hall, Philip

    1989-01-01

    There are many fluid flows where the onset of transition can be caused by different instability mechanisms which compete among themselves. The interaction is considered of two types of instability mode (at an asymptotically large Reynolds number) which can occur in the flow above a rotating disc. In particular, the interaction is examined between lower branch Tollmien-Schlichting (TS) waves and the upper branch, stationary, inviscid crossflow vortex whose asymptotic structure has been described by Hall (1986). This problem is studied in the context of investigating the effect of the vortex on the stability characteristics of a small TS wave. Essentially, it is found that the primary effect is felt through the modification to the mean flow induced by the presence of the vortex. Initially, the TS wave is taken to be linear in character and it is shown (for the cases of both a linear and a nonlinear stationary vortex) that the vortex can exhibit both stabilizing and destabilizing effects on the TS wave and the nature of this influence is wholly dependent upon the orientation of this latter instability. Further, the problem is examined with a larger TS wave, whose size is chosen so as to ensure that this mode is nonlinear in its own right. An amplitude equation for the evolution of the TS wave is derived which admits solutions corresponding to finite amplitude, stable, traveling waves.

  11. On the interaction of stationary crossflow vortices and Tollmien-Schlichting waves in the boundary layer on a rotating disc

    NASA Technical Reports Server (NTRS)

    Bassom, Andrew P.; Hall, Philip

    1990-01-01

    There are many fluid flows where the onset of transition can be caused by different instability mechanisms which compete among themselves. The interaction is considered of two types of instability mode (at an asymptotically large Reynolds number) which can occur in the flow above a rotating disc. In particular, the interaction is examined between lower branch Tollmien-Schlichting (TS) waves and the upper branch, stationary, inviscid crossflow vortex whose asymptotic structure has been described by Hall (1986). This problem is studied in the context of investigating the effect of the vortex on the stability characteristics of a small TS wave. Essentially, it is found that the primary effect is felt through the modification to the mean flow induced by the presence of the vortex. Initially, the TS wave is taken to be linear in character and it is shown (for the cases of both a linear and a nonlinear stationary vortex) that the vortex can exhibit both stabilizing and destabilizing effects on the TS wave and the nature of this influence is wholly dependent upon the orientation of this latter instability. Further, the problem is examined with a larger TS wave, whose size is chosen so as to ensure that this mode is nonlinear in its own right. An amplitude equation for the evolution of the TS wave is derived which admits solutions corresponding to finite amplitude, stable, traveling waves.

  12. Retentivity, selectivity and thermodynamic behavior of polycyclic aromatic hydrocarbons on charge-transfer and hypercrosslinked stationary phases under conditions of normal phase high performance liquid chromatography.

    PubMed

    Jiang, Ping; Lucy, Charles A

    2016-03-11

    Charge-transfer and hypercrosslinked polystyrene phases offer retention and separation for polycyclic aromatic hydrocarbons (PAHs) and thus have potential for petroleum analysis. The size, shape and planarity selectivity for PAH standards on charge-transfer (DNAP column) and hypercrosslinked polystyrene (HC-Tol and 5HGN columns) phases are different under normal phase liquid chromatography (NPLC). The HC-Tol column behaves like a conventional NPLC column with low retention of PAHs. Retention of PAHs on the DNAP and 5HGN are strong and increases with the number of aromatic rings. The main retention mechanism is through π-π interactions and dipole-induced dipole interaction. Thermodynamics indicates that the retention mechanism of PAHs remains unchanged over the temperature range 20-60°C. In addition, on either DNAP or 5HGN column, both linear and bent PAHs are retained through the same mechanism. But DNAP possesses smaller π-π interaction and higher planarity selectivity than 5HGN for PAHs. This is suggestive that DNAP interacts with PAHs through a disordered phase arrangement, while 5HGN behaves as an ordered adsorption phase. PMID:26879454

  13. Preparation of micron-sized spherical particles of mesoporous silica from a triblock copolymer surfactant, usable as a stationary phase for liquid chromatography

    NASA Astrophysics Data System (ADS)

    Mesa, Monica; Sierra, Ligia; López, Betty; Ramirez, Alejandro; Guth, Jean-Louis

    2003-09-01

    Spherical particles (∅>3 μm, with 5-10 nm pore size) of SBA15-type mesoporous silica, usable as stationary phase for HPLC, were prepared with tetraethoxysilane (TEOS) as silica source, triblock copolymer Pluronic P123 (EO 20PO 70EO 20) as surfactant S 0 and cetyltrimethylammonium bromide (CTMABr) as co-surfactant S +. The synthesis mechanism involves a surfactant-silica species self-assembly process in acidic medium under quiescent conditions, where S 0H 3O +X -I + and S +X -I + interactions occur. The syntheses were carried out using two procedures: (a) with one heating step and (b) with two heating steps. Synthesis conditions, such as the dilution, temperature and acidity of the reaction mixture and the temperature and duration of the reaction, influence the porous characteristics as well as the morphology and size of the particles. Spherical particles were obtained with one heating step under conditions that weaken the surfactants-silicate interactions such as high dilution, high temperature and low acidity. The modification of the micelle volume with the temperature and acidity allowed the adjustment of the pore size.

  14. [Preparation of triterpene saponins from Platycodon grandiflorum by two-dimensional hydrophilic interaction liquid chromatography-reversed phase liquid chromatography].

    PubMed

    Xing, Qianqian; Fu, Qing; Jin, Yu; Liang, Xinmiao

    2014-07-01

    A two-dimensional (2-D) preparative liquid chromatography method was developed for the preparation of triterpene saponins from Platycodon grandiflorum using hydrophilic interaction liquid chromatography (HILIC) coupled with reversed phase liquid chromatography (RPLC). At first, the crude extract was obtained from Platycodon grandiflorum by boiled alcohol precipitation. Then, the concentrated crude extracts were continuously pretreated using solid phase extraction (SPE) under reversed-phase and hydrophilic-phase modes to remove the impurities. Subsequently, XAmide column (150 mm x 20 mm, 5 microm) was selected to separate the triterpene saponin constituents under HILIC mode using water and acetonitrile as mobile phases. From 6 min to 25 min, each fraction was collected per minute under time-triggered mode and 20 fractions were collected. The 18th fraction (JG23) was selected for further purification. The column of Atlantis PrepT3 (100 mm x 30 mm, 5 microm) was chosen and two monomeric compounds were obtained. The two compounds with over 90% purity were identified as deapiplatycoside E and platycoside E with mass spectrometry (MS) and nuclear magnetic resonance (NMR). This 2-D HILIC-RPLC method with high orthogonality can be used in the preparation of triterpene saponins from natural products. PMID:25255571

  15. A fast, simple, and reliable hydrophilic interaction liquid chromatography method for the determination of ascorbic and isoascorbic acids.

    PubMed

    Barros, Ana I R N A; Silva, Ana P; Gonçalves, Berta; Nunes, Fernando M

    2010-03-01

    A reliable method for the determination of total vitamin C must be able to resolve ascorbic acid (AA) and the epimeric isoascorbic acid (IAA) and determine the sum of AA and its oxidized form dehydroascorbic acid. AA and IAA are polar molecules with a low retention time in conventional reversed phase systems, and hence of difficult resolution. Hydrophilic interaction chromatography using a TSKgel Amide-80 stationary phase with isocratic elution was successful in resolving the two epimers. The column was compatible with injections of high concentrations of metaphosphoric acid, tris(2-carboxyethyl)-phosphine, and EDTA without drift of baseline and retention time. Total AA and IAA were extracted, stabilized, and reduced in one step at 40 °C, using 5% m-phosphoric acid, 2 mM of EDTA, and 2 mM of tris(2-carboxyethyl)-phosphine as reducing agent. This simple, fast, and robust hydrophilic interaction chromatography-DAD method was applied for the analysis of food products namely fruit juices, chestnut, and ham and also in pharmaceutical and multivitamin tablets. Method validation was performed on the food products, including parameters of precision, accuracy, linearity, limit of detection, and quantification (LOQ). The absence of matrix interferences was assessed by the standard addition method and Youden calibration. The method was fast, accurate, and precise with a LOQ(AA) of 1.5 mg/L and LOQ(IAA) of 3.7 mg/L. The simple experimental procedure, completed in 1 h, the possibility of using IAA as an internal standard, and low probability of artifacts are the major advantages of the proposed method for the routine determination of these compounds in a large number of samples. PMID:20091158

  16. Comparison of the adsorption mechanisms of pyridine in hydrophilic interaction chromatography and in reversed-phase aqueous liquid chromatography.

    PubMed

    Gritti, Fabrice; Pereira, Alberto dos Santos; Sandra, Pat; Guiochon, Georges

    2009-11-27

    The adsorption isotherms of pyridine were measured by frontal analysis (FA) on a column packed with shell particles of neat porous silica (Halo), using water-acetonitrile mixtures as the mobile phase at 295K. The isotherm data were measured for pyridine concentrations covering a dynamic range of four millions. The degree of heterogeneity of the surface was characterized by the adsorption energy distribution (AED) function calculated from the raw adsorption data, using the expectation-maximization (EM) procedure. The results showed that two different retention mechanisms dominate in Per aqueous liquid chromatography (PALC) at low acetonitrile concentrations and in hydrophilic interaction chromatography (HILIC) at high acetonitrile concentrations. In the PALC mode, the adsorption mechanism of pyridine on the silica surface is controlled by hydrophobic interactions that take place on very few and ultra-active adsorption sites, which might be pores on the irregular and rugose surface of the porous silica particles. The surface is seriously heterogeneous, with up to five distinct adsorption sites and five different energy peaks on the AED of the packing material. In contrast, in the HILIC mode, the adsorption behavior is quasi-homogeneous and pyridine retention is governed by its adsorption onto free silanol groups. For intermediate mobile phase compositions, the siloxane and the silanol groups are both significantly saturated with acetonitrile and water, respectively, causing a minimum of the retention factor of pyridine on the Halo column. PMID:19853257

  17. Grafting of silica with a hydrophilic triol acrylamide polymer via surface-initiated "grafting from" method for hydrophilic-interaction chromatography.

    PubMed

    Peng, Xi-Tian; Yuan, Bi-Feng; Feng, Yu-Qi

    2011-11-01

    A novel hydrophilic polymer-coated silica sorbent has been prepared using the radical "grafting from" polymerization method through surface-bound azo initiators for hydrophilic-interaction chromatography (HILIC). The azo groups were introduced to the surface of silica gel through the reaction with amino groups on the surface of silica gel with 4,4'-azobis(4-cyanopentanoic acid chloride) (ACVC). The resultant azo-immobilized silica gel served as surface initiator to polymerize hydrophilic triol acrylamide monomer N-acryloyltris(hydroxymethyl) aminomethane (NA) in methanol to get hydrophilic polymer-coated silica sorbent. The obtained poly(NA)-coated silica (pNA-sil) was characterized by Fourier transform infrared spectroscopy (FT-IR), elemental analysis (EA), and nitrogen sorption porosimetry (NSP). Then the pNA-sil was packed into the stainless-steel column and evaluated in high-performance liquid chromatography (HPLC). Good chromatographic performance for the separation of peptides and nucleosides was obtained under HILIC mode. The results indicated that the pNA-sil stationary phase behaved as mixed-mode retention mechanisms of hydrophilic and ionic interactions. Furthermore, the pNA-sil phase was used to separate tryptic digest of β-casein and our results showed that more than 12 peptides peaks were resolved and well distributed within the elution window. Finally, the pNA-sil stationary phase was demonstrated to possess remarkable reproducibility and stability. Taken together, the pNA-sil stationary phase prepared in the current study offers a potential application in proteomics study. PMID:21998034

  18. A new method for separation and determination of Cr(III) and Cr(VI) in water samples by high-performance liquid chromatography based on anion exchange stationary phase of ionic liquid modified silica.

    PubMed

    Sadeghi, Susan; Moghaddam, Ali Zeraatkar

    2015-12-01

    In this work, N-methylimidazolium-chloride ionic liquid functionalized silica was prepared and used as an anion-exchange stationary phase for separation of chromium species by high-performance liquid chromatography (HPLC) with UV detection at 200 nm. The Cr(VI) as HCr2O7(-) and chelated Cr(III) with potassium hydrogen phthalate (PHP) as Cr(PHP)2 (-) was retained on the prepared column and separated using a mobile phase composed of 5% methanol in 25 mM phosphate buffer at pH 6.5. Several variables affecting the chelation/separation steps were modeled by response surface methodology (RSM) using Box-Behnken (BBD) design. The significance of the independent variables and their interactions were tested by the analysis of variances (ANOVA) with 95% confidence limit. Under the optimized conditions, the Cr(III) and Cr(VI) anionic species were well separated with a single peak for each Cr species at retention times of 2.3 and 4.3 min, respectively. The relationship between the peak area and concentration was linear in the range of 0.025-30 for Cr(III) and 0.5-20 mg L(-1) for Cr(VI) with detection limits of 0.010 and 0.210 mg L(-1) for Cr(III) and Cr(VI), respectively. The proposed method was validated by simultaneous separation and determination of the Cr species in tap and underground water samples without impose to any pretreatment. PMID:26526699

  19. Investigating interactions between phospholipase B-Like 2 and antibodies during Protein A chromatography.

    PubMed

    Tran, Benjamin; Grosskopf, Vanessa; Wang, Xiangdan; Yang, Jihong; Walker, Don; Yu, Christopher; McDonald, Paul

    2016-03-18

    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. PMID:26896920

  20. Identification of proteins interacting with ammodytoxins in Vipera ammodytes ammodytes venom by immuno-affinity chromatography.

    PubMed

    Brgles, Marija; Kurtović, Tihana; Kovačič, Lidija; Križaj, Igor; Barut, Miloš; Lang Balija, Maja; Allmaier, Günter; Marchetti-Deschmann, Martina; Halassy, Beata

    2014-01-01

    In order to perform their function, proteins frequently interact with other proteins. Various methods are used to reveal protein interacting partners, and affinity chromatography is one of them. Snake venom is composed mostly of proteins, and various protein complexes in the venom have been found to exhibit higher toxicity levels than respective components separately. Complexes can modulate envenomation activity of a venom and/or potentiate its effect. Our previous data indicate that the most toxic components of the Vipera ammodytes ammodytes (Vaa) venom isolated so far-ammodytoxins (Atxs)-are contributing to the venom's toxicity only moderately; therefore, we aimed to explore whether they have some interacting partner(s) potentiating toxicity. For screening of possible interactions, immuno-affinity chromatography combined with identification by mass spectrometry was used. Various chemistries (epoxy, carbonyldiimidazole, ethylenediamine) as well as protein G functionality were used to immobilize antibodies on monolith support, a Convective Interaction Media disk. Monoliths have been demonstrated to better suit the separation of large biomolecules. Using such approach, several proteins were indicated as potential Atx-binding proteins. Among these, the interaction of Atxs with a Kunitz-type inhibitor was confirmed by far-Western dot-blot and surface plasmon resonance measurement. It can be concluded that affinity chromatography on monolithic columns combined with mass spectrometry identification is a successful approach for screening of protein interactions and it resulted with detection of the interaction of Atx with Kunitz-type inhibitor in Vaa venom for the first time. PMID:24217948

  1. Hydrophilic interaction liquid chromatography coupled to high-resolution mass spectrometry to determine artificial sweeteners in environmental waters.

    PubMed

    Salas, Daniela; Borrull, Francesc; Fontanals, Núria; Marcé, Rosa Maria

    2015-06-01

    Artificial sweeteners are food additives employed as sugar substitutes which are now considered to be emerging organic contaminants. In the present study, a method is developed for the determination of a group of artificial sweeteners in environmental waters. Considering the polar and hydrophilic character of these compounds, hydrophilic interaction liquid chromatography is proposed for their separation as an alternative to traditional reversed-phase liquid chromatography. Two stationary phases with different chemistry were compared for this purpose. For the detection of the analytes, high-resolution mass spectrometry (Orbitrap) was employed to take advantage of its benefits in terms of reliable quantification and confirmation for the measurement of accurate masses. Solid-phase extraction was chosen as the sample treatment, in which the extract in a mixture of NH4OH:MeOH:ACN (1:4:15) was directly injected into the chromatographic system, simplifying the analytical procedure. The optimized method was validated on river and waste water samples. For example, in the case of effluent water samples, limits of detection ranged from 0.002 to 0.7 μg/L and limits of quantification ranged from 0.004 to 1.5 μg/L. Apparent (whole method) recoveries ranged from 57 to 74% with intra-day precision (%RSD, n = 5) ranging from 6 to 25%. The method was successfully applied to water samples from different rivers in Catalonia and different waste water treatment plants in Tarragona. Acesulfame, cyclamate, saccharine and sucralose were found in several samples. PMID:25428455

  2. Distinction of synthetic dl-α-tocopherol from natural vitamin E (d-α-tocopherol) by reversed-phase liquid chromatography. Enhanced selectivity of a polymeric C18 stationary phase at low temperature and/or at high pressure.

    PubMed

    Yui, Yuko; Miyazaki, Shota; Ma, Yan; Ohira, Masayoshi; Fiehn, Oliver; Ikegami, Tohru; McCalley, David V; Tanaka, Nobuo

    2016-06-10

    Separation of diastereomers of dl-α-tocopherol was studied by reversed-phase liquid chromatography using three types of stationary phases, polymeric ODS, polymeric C30, and monomeric ODS. Polymeric ODS stationary phase (Inertsil ODS-P, 3mmID, 20cm) was effective for the separation of the isomers created by the presence of three chiral centers on the alkyl chain of synthetic dl-α-tocopherol. Considerable improvement of the separation of isomers was observed on ODS-P phase at high pressure and at low temperature. Complete separation of four pairs of diastereomers was achieved at 12.0°C, 536bar, while three peaks were observed when the separation was carried out either at 12.0°C at low pressure or at 20°C at 488bar. Higher temperature (30.0°C) with the ODS-P phase resulted in only partial separation of the diastereomers even at high pressure. Only slight resolution was observed for the mixture of diastereomers with the C30 stationary phase (Inertsil C30) at 12.0°C and 441bar, although the stationary phase afforded greater resolution for β- and γ-tocopherol than ODS-P. A monomeric C18 stationary phase did not show any separation at 12.0°C and 463bar. The results suggest that the binding site of the polymeric ODS-P phase is selective for flexible alkyl chains that provided the longest retention for the natural form, (R,R,R) form, and the enantiomer, (S,S,S) form, of dl-α-tocopherol. PMID:27157422

  3. Synthesis of poly(N-[tris(hydroxymethyl)methyl]acrylamide) functionalized porous silica for application in hydrophilic interaction chromatography.

    PubMed

    Bui, Nhat Thi Hong; Jiang, Wen; Sparrman, Tobias; Irgum, Knut

    2012-12-01

    Porous silica coated by a highly hydrophilic and nonionic tentacle-type polymeric layer was synthesized by free radical "grafting from" polymerization of N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]-2-propenamide (TRIS-acrylamide) in partly aqueous solutions. The radical initiator sites were incorporated on the silica surfaces via a two-step reaction comprising thionyl chloride activation and subsequent reaction with tert-butyl hydroperoxide. The surface-bound tert-butylperoxy groups were then used as thermally triggered initiators for graft polymerization of TRIS-acrylamide. The synthesized materials were characterized by diffusive reflectance Fourier transform infrared specotroscopy, X-ray photoelectron spectroscopy, and CHN elemental analysis. Photon correlation spectroscopy was used to determine changes in ζ-potentials resulting from grafting, (29)Si magic angle spinning nuclear magnetic resonance spectroscopy (MAS-NMR) spectroscopy was used to assess the ratio of silanol to siloxane groups in the substrate and the grafted material, and the changes in surface area and mesopore distribution were determined by nitrogen cryosorption. Chromatographic evaluation in hydrophilic interaction chromatography (HILIC) mode showed that the materials were suitable for use as stationary phases, featuring good separation efficiency, a comparatively high retention, and a selectivity that differed from most commercially available HILIC phases. A comparison of this neutral phase with a previously reported N-(2-hydroxypropyl)-linked TRIS-type hydrophilic tentacle phase with weak anion exchange functionality revealed substantial differences in retention patterns. PMID:23184369

  4. Simulation of Alpha Particles in Rotating Plasma Interacting with a Stationary Ripple

    SciTech Connect

    Abraham J. Fetterman and Nathaniel J. Fisch

    2011-01-11

    Superthermal ExB rotation can provide magnetohydrodynamic (MHD) stability and enhanced confinement to axisymmetric mirrors. However, the rotation speed has been limited by phenomena at end electrodes. A new prediction is that rotation might instead be produced using a magnetic ripple and alpha particle kinetic energy, in an extension of the alpha channeling concept. The interaction of alpha particles with the ripple results in visually interesting and practically useful orbits.

  5. Effects of the surface concentration of fixed charges in C18-bonded stationary phases on the adsorption process and on the preparative chromatography of small ionizable compounds.

    PubMed

    Gritti, Fabrice; Guiochon, Georges

    2014-11-01

    The effects of the surface concentration of positive charges attached to the surface of research BEH-C18 hybrid particles on the overloaded band profiles and the adsorption isotherms of a neutral (caffeine) and a positively charged (nortryptilinium hydrochloride) compounds were measured and investigated. The inverse method (IM) of chromatography was used to determine the isotherm parameters. Three columns were packed with endcapped BEH-C18 particles doped with three different charge densities on their surfaces (LOW, MEDIUM and HIGH). Two other columns packed with unbonded, non-endcapped, and endcapped BEH-C18 particles served as standard reference materials. Minor disturbance method (MDM) experiments were conducted with acetonitrile/water mixtures in order to assess qualitatively the surface densities of the fixed positive charges. A more quantitative approach based on the solution of the linearized Poisson-Boltzmann equation and the decrease of the experimental Henry constant was also applied. The results show that the surface concentrations of the fixed charges in the LOW, MEDIUM and HIGH columns were 0.029, 0.050, and 0.064μmol/m(2), e.g., close to two orders of magnitude smaller than the surface density of bonded C18 chains (2.1μmol/m(2)). The adsorption isotherm of the ionizable compound nortryptilinium onto the BEH-C18 columns is consistent with a two-sites adsorption model. The density of the high energy sites correlates directly to the total amount of the fixed charges and isolated silanols amidst the C18-bonded chains. The amount of low energy sites reflects the specific surface area of the adsorbent. The binding constants on the high- and low-energy adsorption sites are respectively ten and two times lower on the HIGH column than on the reference endcapped column. The active sites are closer to the adsorbent surface than the weak adsorption sites. Finally, a higher production rate of ionizable compounds can be achieved in preparative chromatography with

  6. Determination of Oxidized Phosphatidylcholines by Hydrophilic Interaction Liquid Chromatography Coupled to Fourier Transform Mass Spectrometry

    PubMed Central

    Sala, Pia; Pötz, Sandra; Brunner, Martina; Trötzmüller, Martin; Fauland, Alexander; Triebl, Alexander; Hartler, Jürgen; Lankmayr, Ernst; Köfeler, Harald C.

    2015-01-01

    A novel liquid chromatography-mass spectrometry (LC-MS) approach for analysis of oxidized phosphatidylcholines by an Orbitrap Fourier Transform mass spectrometer in positive electrospray ionization (ESI) coupled to hydrophilic interaction liquid chromatography (HILIC) was developed. This method depends on three selectivity criteria for separation and identification: retention time, exact mass at a resolution of 100,000 and collision induced dissociation (CID) fragment spectra in a linear ion trap. The process of chromatography development showed the best separation properties with a silica-based Kinetex column. This type of chromatography was able to separate all major lipid classes expected in mammalian samples, yielding increased sensitivity of oxidized phosphatidylcholines over reversed phase chromatography. Identification of molecular species was achieved by exact mass on intact molecular ions and CID tandem mass spectra containing characteristic fragments. Due to a lack of commercially available standards, method development was performed with copper induced oxidation products of palmitoyl-arachidonoyl-phosphatidylcholine, which resulted in a plethora of lipid species oxidized at the arachidonoyl moiety. Validation of the method was done with copper oxidized human low-density lipoprotein (LDL) prepared by ultracentrifugation. In these LDL samples we could identify 46 oxidized molecular phosphatidylcholine species out of 99 possible candidates. PMID:25874761

  7. [Analysis of carbapenems by hydrophilic interaction chromatography and its application].

    PubMed

    Zhu, Yinfang; Ji, Shunli; Li, Shaohui; Li, Cheng; Zhang, Feifang; Liang, Xinmiao

    2015-09-01

    A hydrophilic interaction chromatographic (HILIC) method has been developed for the determination of the four carbapenems in human urine and tap water. The parameters including acetonitrile amount, buffer concentration and pH on the retention behavior of the four carbapenem antibiotics on an XAmide column were explored and the possible HILIC retention mechanism was proposed. Good linearities were obtained over the mass concentration ranges of 0.1-250 mg/L for biapenem, doripenem and ertapenem with correlation coefficients (R2) = 0.999 9 and while it was 0.5-250 mg/L with R2 = 0.999 8 for meropenem. The limits of quantification (LOQs) of all carbapenems were 0.1-0.5 mg/L. The spiked recoveries were within 100.4%-111.9% (RSD < 1%) for urine samples and 79.6%-107.4% (RSD < 5%) for tap water samples all at the spiked levels of 5 mg/L and 25 mg/L. The proposed method is accurate, sensitive, simple and suitable for the determination of the four carbapenems in human urine samples and tap water samples. PMID:26753282

  8. Novel highly hydrophilic zwitterionic monolithic column for hydrophilic interaction chromatography.

    PubMed

    Jiang, Zhengjin; Smith, Norman W; Ferguson, Paul D; Taylor, Mark R

    2009-08-01

    A novel zwitterionic hydrophilic porous poly(SPV-co-MBA) monolithic column was prepared by thermal co-polymerisation of 1-(3-sulphopropyl)-4-vinylpyridinium-betaine (4-SPV) and N,N'-methylenebisacrylamide (MBA). An HILIC/RP dual separation mechanism was observed on this optimised poly(SPV-co-MBA) monolithic column and the composition of the mobile phase corresponding to the transition from the HILIC to the RP mode was around 30% ACN in water. Higher hydrophilicity was achieved on this novel monolithic column compared to the poly(N,N-dimethyl-N-methacryloxyethyl-N-(3-sulphopropyl)ammonium betaine-co-ethylene dimethacrylate) monolithic column. Permeability studies showed slight swelling and/or shrinking with mobile phases of different polarity. As might be anticipated, a weak electrostatic interaction for charged analytes was also observed by studying the influence of mobile phase pH and salt concentration on their retention on the poly(SPV-co-MBA) monolithic column. The final optimised poly(SPV-co-MBA) monolith showed comparable selectivities to commercial ZIC-pHILIC phases for polar test analytes. Fast separation of five pyrimidines and purines was achieved in less than 1 min due to the high permeability of the monolithic column. Additionally, baseline separation of nine benzoic acid derivatives was also observed using either a pH or ACN gradient. PMID:19606441

  9. Monolithic stationary phases with incorporated fumed silica nanoparticles. Part II. Polymethacrylate-based monolithic column with "covalently" incorporated modified octadecyl fumed silica nanoparticles for reversed-phase chromatography.

    PubMed

    Aydoğan, Cemil; El Rassi, Ziad

    2016-05-01

    This study is concerned with the incorporation of surface modified fumed silica nanoparticles (FSNPs) into polymethacrylate based monolithic columns for use in reversed phase chromatography (RPC) of small solutes and proteins. First, FSNPs were modified with 3-(trimethoxysilyl)propylmethacrylate (TMSPM) to yield the "hybrid" methacryloyl fumed silica nanoparticle (MFSNP) monomer. The resulting MFSNP was then mixed with glyceryl monomethacrylate (GMM) and ethylene dimethacrylate (EDMA) in a binary porogenic solvent composed of cyclohexanol and dodecanol, and the in situ copolymerization of MFSNP, GMM and EDMA was performed in a stainless steel column of 4.6 mm i.d. The silanol groups of the hybrid monolith thus obtained were grafted with octadecyl ligands by perfusing the hybrid monolithic column with a solution of 4% w/v of dimethyloctadecylchlorosilane (DODCS) in toluene while the column was maintained at 110°C for 6h (in a heated HPLC oven). One of the originalities of this study was to demonstrate MFSNP as a novel derivatized "hybrid monomer" in making RPC monolithic columns with surface bound octadecyl ligands. In this respect, the RPC behavior of the monolithic column with "covalently" incorporated FNSPs having surface grafted octadecyl ligands was evaluated with alkylbenzenes, aniline derivatives and phenolic compounds. The results showed that the hybrid poly(GMA-EDMA-MFSNP) having surface bound octadecyl ligands exhibited hydrophobic interactions under reversed phase elution conditions. Furthermore, six standard proteins were baseline separated on the column using a 10min linear gradient elution at increasing ACN concentration in the mobile phase at a flow rate of 1.0mL/min using a 10 cm×4.6mm i.d. column. The relative standard deviations (RSDs) for the retention times of the tested solutes were lower than 2.1% and 2.4% under isocratic elution and gradient elution conditions, respectively. PMID:27059396

  10. Measurement of supersonic plasma interacting with stationary plasma by electric probes

    NASA Astrophysics Data System (ADS)

    Lee, Dong Han; Kang, In Je; Bae, Min Keun; Cho, Soon-Gook; Kim, Sang-You; Choi, Heung-Gyoon; Hong, Sung-Hoon; Lho, Tae-Hyup; Chung, Kyu-Sun

    2015-09-01

    Supersonic plasma is generally related to the formation of young star object (YSO), active galactic nuclei (AGN) and new galaxies via plasma bubble expansion during the event of super nova. Capacitive coupled plasma (CCP) is produced by RF power of 13.56 MHz and the plasma is accelerated by negatively biased cascade grid to produce supersonic flow. Electron temperature, plasma density and Mach number are measured by using a single probe and a Mach probe. Electron temperature and plasma density of CCP are 0.8 eV and 1.8 × 109 cm-3, respectively. Mach number of supersonic plasma flow is about 2 and 50 W RF power at 52 mTorr. Ambient plasma is generated by DC filament discharge and its electron temperature and plasma density are 0.5 eV and 3 × 1010 cm-3, respectively. When the supersonic plasma flow interact with ambient plasma, electron temperature is increased higher than ambient plasma up to 4 eV, and plasma density is decreased from 4 × 1010 cm-3 to 1 × 1010 cm-3. Density contrast η of supersonic plasma flow of our experiment is about 0.04, while AGN jets in universe are observed to have density contrast η of lower than 10-2.

  11. Quasi-normal phase chromatography of nitrogen-containing adamantane derivatives

    NASA Astrophysics Data System (ADS)

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

    2013-01-01

    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.

  12. Protein losses in ion-exchange and hydrophobic interaction high-performance liquid chromatography

    SciTech Connect

    Goheen, Steven C.; Gibbins, Betty M.

    2000-01-01

    Protein losses in ion-exchange and hydrophobic interaction HPLC were examined. The supports were allnon-porous, packed in columns of identical dimensions. Two ion-exchange chromatography (IEC), anion and cation, as well as a hydrophobic interaction chromatography (HIC) columns were tested. Proteins included cytochrome c, bovine serum albumin (BSA), immunoglobulin G and fibrinogen. Temperature effects on HIC supports were studied for cytochrome c and BSA. Both retention times and recoveries of the proteins were measured. The influence of column residence time on the recovery of proteins were also investigated. We found a linear relationship between the amount of protein recovered and the log of the molecular mass. Retention times also generally increased with temperature for both HIC and IEC. Other trends in retention behavior and recoveries are discussed.

  13. Hydrophilic interaction chromatography versus reversed phase liquid chromatography coupled to mass spectrometry: effect of electrospray ionization source geometry on sensitivity.

    PubMed

    Periat, Aurélie; Kohler, Isabelle; Bugey, Aurélie; Bieri, Stefan; Versace, François; Staub, Christian; Guillarme, Davy

    2014-08-22

    In this study, the influence of electrospray ionization (ESI) source design on the overall sensitivity achieved in hydrophilic interaction chromatography (HILIC) and reversed phase liquid chromatography (RPLC), was investigated. State-of-the-art triple quadrupole mass analyzers from AB Sciex, Agilent Technologies and Waters equipped with brand specific source geometries were tested with various mobile phase pH on 53 pharmaceutical compounds. The design of the ESI source showed to strongly influence the gain in sensitivity that can be achieved in HILIC compared to RPLC mode. The 6460 Triple Quadrupole LC/MS system from Agilent Technologies was particularly affected by mobile phase settings. Indeed, compared to RPLC conditions, 92% of the compounds had an increased signal-to-noise ratio at a flow rate of 300 μL/min in HILIC mode at pH 6, while this percentage dropped to only 7% at 1000 μL/min and pH 3. In contrast, the influence of flow rate and mobile phase pH on the gain in sensitivity between RPLC and HILIC was found very limited with the API 5000 LC/MS/MS system from AB Sciex, as only 15 to 36% of the tested compounds showed an enhanced sensitivity in HILIC mode. With the Xevo TQ-S instrument from Waters, superior sensitivity in HILIC was noticed for 85% of the compounds with optimal conditions (i.e., pH 3 and 1000 μL/min), whereas at sub-optimal conditions (i.e. pH 6 and 300 μL/min), it represented less than 50%. The gain in sensitivity observed in HILIC was found less significant with the recent LC-MS platforms used in this study than for old-generation instruments. Indeed, the improved ESI sources equipping the recent mass analyzers allow for enhanced evaporation efficiency, mainly for RPLC mobile phases containing high proportion of water and this even at high flow rates. PMID:25017394

  14. Measurement of interactions between polysaccharides and flavour compounds by exclusion size chromatography: advantages and limits.

    PubMed

    Guichard, E; Etiévant, P

    1998-12-01

    Interactions between flavour compounds and polysaccharides have been studied by exclusion size chromatography, the Hummel and Dreyer method. Hydrogen bonding was found between 2-acetyl thiazole and dextrines of different degrees of polymerisation. The number of binding sites and the affinity constant increase by increasing the degree of polymerisation. Hydrogen bonding was also responsible for the interactions between xanthane and 1-octen-3-ol or 2-acetyl pyrazine, with 1 mole of 1-octen-3-ol bound per pentasaccharide repeating unit. Unfortunately, the number of flavour compounds, which can be studied with this method, is limited due to their low water solubility and their low UV absorption. PMID:9881364

  15. Comparison of zonal elution and nonlinear chromatography in determination of the interaction between seven drugs and immobilised β(2)-adrenoceptor.

    PubMed

    Li, Qian; Wang, Jing; Zheng, Yuqing Yuan; Yang, Lingjian; Zhang, Yajun; Bian, Liujiao; Zheng, Jianbin; Li, Zijian; Zhao, Xinfeng; Zhang, Youyi

    2015-07-01

    Zonal elution and nonlinear chromatography are two mainstream models for the determination of drug-protein interaction in affinity chromatography. This work intended to compare the results by zonal elution with that by nonlinear chromatography when it comes to the analysis of the interaction between seven drugs and immobilised β2-adrenoceptor (β2-AR). The results of the zonal elution showed that clorprenaline, clenbuterol, methoxyphenamine, salbutamol, terbutaline, tulobuterol and bambuterol have only one type of binding site on immobilised β2-AR, while nonlinear chromatography confirmed the existence of at least two types of binding sites between β2-AR and clorprenaline, clenbuterol and bambuterol. On these sites, both zonal elution and nonlinear chromatography presented the same rank order for the association constants of the seven drugs. Compared with the data from zonal elution, the association constants calculated using nonlinear chromatography gave a good linear response to the corresponding values by radio-ligand binding assay. The sampling efficiencies of nonlinear chromatography were clearly higher than zonal elution. Nonlinear chromatography will probably become a powerful alternative for the high throughput determination of drug-protein interaction. PMID:26002106

  16. Screening for low molecular weight compounds in fish meal solubles by hydrophilic interaction liquid chromatography coupled to mass spectrometry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A simple analytical method using hydrophilic interaction liquid chromatography coupled with mass spectrometry was developed to screen for low molecular weight compounds in enzyme treated and untreated Alaskan pollock (Theragra chalcogramma) stickwater (SW) generated from processing fish meal with po...

  17. Separation of mAbs molecular variants by analytical hydrophobic interaction chromatography HPLC: overview and applications

    PubMed Central

    Haverick, Mark; Mengisen, Selina; Shameem, Mohammed; Ambrogelly, Alexandre

    2014-01-01

    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 monoclonal antibodies; and carboxy terminal heterogeneity and serine fucosylation in Fc and Fab fragments. HIC-HPLC is also a powerful analytical technique for the analysis of antibody-drug conjugates. Most current analytical columns, methods, and applications are described, and critical method parameters and suitability for operation in regulated environment are discussed, in this review. PMID:24751784

  18. Compositional and monomer sequence distribution analysis of monodisperse brominated-polystyrenes using interaction chromatography

    NASA Astrophysics Data System (ADS)

    Han, Junwon; Semler, James J.

    2005-03-01

    High performance liquid chromatography techniques have been developed for characterizing complex polymers that are often heterogeneous in molecular weight, molecular architecture, and chemical composition. Recently, interaction chromatography (IC) techniques have been developed, which facilitate separation of polymers based on enthalpic ``attraction'' difference among the chemical constituents of the molecule. Here, we use IC for characterizing the composition and monomer sequence distribution in statistical copolymers of poly(styrene-co-4-bromostyrene) (PBrxS). The PBrxS copolymers were synthesized by brominating monodisperse polystyrenes; the degree of bromination (x) and the sequence distribution have been adjusted by varying the bromination time and the solvent quality, respectively. Both normal-phase (bare silica) and reversed-phase (C18-bonded silica) columns are used at different combinations of solvents and non-solvents to monitor the content of the 4-bromostyrene units in the copolymer and their average monomer sequence distribution.

  19. Detailed insights into the retention mechanism of caffeine metabolites on the amide stationary phase in hydrophilic interaction chromatography.

    PubMed

    Guo, Yong; Shah, Rajan

    2016-09-01

    The amide phase was investigated using a wide range of acetonitrile content in the mobile phase in both the HILIC and RPLC modes. Using caffeine metabolites as the model compounds, the retention, thermodynamic and kinetic data was obtained under various mobile phase conditions and supported the previous postulation that there might be a transition of the predominant retention mechanism in relation to the acetonitrile content in HILIC. On the amide phase, hydrophilic partitioning seemed to be the predominant retention mechanism below 85% acetonitrile; and a different retention mechanism (presumably surface adsorption) made more and more significant contributions to the overall retention when the acetonitrile content reached above 85%. This study also provided more direct evidences to explain the effect of salt concentration on the retention of non-charged solutes in HILIC. In addition, the retention, thermodynamic and kinetic data suggest that the amide phase behaved very differently from the conventional C18 phase in the RPLC mode. PMID:27522153

  20. A reversed-phase/hydrophilic interaction mixed-mode C18-Diol stationary phase for multiple applications.

    PubMed

    Wang, Qing; Ye, Mao; Xu, Li; Shi, Zhi-guo

    2015-08-12

    A mixed-mode chromatographic packing material, C18 and diol groups modified silica (C18-Diol), was prepared with controllable hydrophobicity and hydrophilicity. It demonstrated excellent aqueous compatibility and stability in aqueous mobile phase; compared to the traditional C18 column, improved peak shape of basic analytes was also obtained. Additionally, it exhibited both reversed-phase liquid chromatographic (RPLC) and hydrophilic interaction chromatographic (HILIC) performance; the analyte separation scope was thus enlarged, demonstrated by simultaneous separation of twenty acids, bases and neutrals. More interestingly, a novel on-line two-dimensional liquid chromatography on the single column (2D-LC-1C) was established by modifying the high performance liquid chromatographic instrument only with the addition of an extra six-port two-position valve. The early co-eluted components of the extract of Lonicera japonica on the 1st-dimension (RPLC) were collected for the online re-injection to the 2nd-dimension (HILIC) by conveniently varying the mobile phase components. Six more peaks were obtained. The established system was simple, easy operation and low cost, which had advantages in analyzing complicated samples. PMID:26320974

  1. Development of an achiral supercritical fluid chromatography method with ultraviolet absorbance and mass spectrometric detection for impurity profiling of drug candidates. Part II. Selection of an orthogonal set of stationary phases.

    PubMed

    Lemasson, Elise; Bertin, Sophie; Hennig, Philippe; Boiteux, Hélène; Lesellier, Eric; West, Caroline

    2015-08-21

    Impurity profiling of organic products that are synthesized as possible drug candidates requires complementary analytical methods to ensure that all impurities are identified. Supercritical fluid chromatography (SFC) is a very useful tool to achieve this objective, as an adequate selection of stationary phases can provide orthogonal separations so as to maximize the chances to see all impurities. In this series of papers, we have developed a method for achiral SFC-MS profiling of drug candidates, based on a selection of 160 analytes issued from Servier Research Laboratories. In the first part of this study, focusing on mobile phase selection, a gradient elution with carbon dioxide and methanol comprising 2% water and 20mM ammonium acetate proved to be the best in terms of chromatographic performance, while also providing good MS response [1]. The objective of this second part was the selection of an orthogonal set of ultra-high performance stationary phases, that was carried out in two steps. Firstly, a reduced set of analytes (20) was used to screen 23 columns. The columns selected were all 1.7-2.5μm fully porous or 2.6-2.7μm superficially porous particles, with a variety of stationary phase chemistries. Derringer desirability functions were used to rank the columns according to retention window, column efficiency evaluated with peak width of selected analytes, and the proportion of analytes successfully eluted with good peak shapes. The columns providing the worst performances were thus eliminated and a shorter selection of columns (11) was obtained. Secondly, based on 160 tested analytes, the 11 columns were ranked again. The retention data obtained on these columns were then compared to define a reduced set of the best columns providing the greatest orthogonality, to maximize the chances to see all impurities within a limited number of runs. Two high-performance columns were thus selected: ACQUITY UPC(2) HSS C18 SB and Nucleoshell HILIC. PMID:26195036

  2. Determination of trantinterol enantiomers in human plasma by high-performance liquid chromatography - tandem mass spectrometry using vancomycin chiral stationary phase and solid phase extraction and stereoselective pharmacokinetic application.

    PubMed

    Qin, Feng; Wang, Yanjuan; Wang, Lijuan; Zhao, Longshan; Pan, Li; Cheng, Maosheng; Li, Famei

    2015-05-01

    A sensitive and enantioselective vancomycin chiral stationary phase high-performance liquid chromatography-tandem mass spectrometry method was developed for the determination of trantinterol enantiomers in human plasma. Baseline resolution was achieved using the vancomycin chiral stationary phase known as Chirobiotic V with polar ionic mobile phase consisting of acetonitrile-methanol (60:40, v/v) containing 0.01% ammonia and 0.02% acetic acid at a flow rate of 1.0 mL/min. Waters Oasis HLB C18 solid phase extraction cartridges were used in the sample preparation of trantinterol samples from plasma. The detection was performed on a triple-quadrupole tandem mass spectrometer by multiple reaction monitoring mode via electrospray ionization. The calibration curve was linear in a concentration range from 0.0606 to 30.3 ng/mL in plasma, with the lower limit of quantification of 0.0606 ng/mL. The intra- and interday precision (relative standard deviation) values were within 9.7% and the accuracy (relative error) was from -6.6 to 7.2% at all quality control levels. The method was successfully applied to a study of stereoselective pharmacokinetics in human. PMID:25782043

  3. Liquid chromatography with mass spectrometry enantioseparation of pomalidomide on cyclodextrin-bonded chiral stationary phases and the elucidation of the chiral recognition mechanisms by NMR spectroscopy and molecular modeling.

    PubMed

    Szabó, Zoltán-István; Szőcs, Levente; Horváth, Péter; Komjáti, Balázs; Nagy, József; Jánoska, Ádám; Muntean, Daniela-Lucia; Noszál, Béla; Tóth, Gergő

    2016-08-01

    A sensitive and validated liquid chromatography with mass spectrometry method was developed for the enantioseparation of the racemic mixture of pomalidomide, a novel, second-generation immunomodulatory drug, using β-cyclodextrin-bonded stationary phases. Four cyclodextrin columns (β-, hydroxypropyl-β-, carboxymethyl-β-, and sulfobutyl-β-cyclodextrin) were screened and the effects of eluent composition, flow rate, temperature, and organic modifier on enantioseparation were studied. Optimized parameters, offering baseline separation (resolution = 2.70 ± 0.02) were the following: β-cyclodextrin stationary phase, thermostatted at 15°C, and mobile phase consisting of methanol/0.1% acetic acid 10:90 v/v, delivered with 0.8 mL/min flow rate. For the optimized parameter at multiple reaction monitoring mode 274.1-201.0 transition with 20 eV collision energy and 100 V fragmentor voltage the limit of detection and limit of quantitation were 0.75 and 2.00 ng/mL, respectively. Since enantiopure standards were not available, elution order was determined upon comparison of the circular dichroism signals of the separated pomalidomide enantiomers with that of enantiopure thalidomide. The mechanisms underlying the chiral discrimination between the enantiomers were also investigated. Pomalidomide-β-cyclodextrin inclusion complex was characterized using nuclear magnetic resonance spectroscopy and molecular modeling. The thermodynamic aspects of chiral separation were also studied. PMID:27279456

  4. Studies of a pyridino-crown ether-based chiral stationary phase on the enantioseparation of biogenic chiral aralkylamines and α-amino acid esters by high-performance liquid chromatography.

    PubMed

    Lévai, Sándor; Németh, Tamás; Fődi, Tamás; Kupai, József; Tóth, Tünde; Huszthy, Péter; Balogh, György Tibor

    2015-11-10

    This paper reports the enantioseparation ability of a pyridino-18-crown-6 ether-based chiral stationary phase [(S,S)-CSP-1]. The enantiomeric discrimination of chiral stationary phase (S,S)-CSP-1 was evaluated by HPLC using the mixtures of enantiomers of various protonated primary aralkylamines [1-phenylethylamine hydrogen perchlorate (PEA), 2,3-dihydro-1H-inden-1-amine (1-aminoindan), 2,2'-(1,2-diaminoethane-1,2-diyl) diphenol (HPEN)] and perchlorate salts of α-amino acid esters [alanine benzyl ester (Ala-OBn), phenylalanine benzyl ester (Phe-OBn), phenylalanine methyl ester (Phe-OMe), phenylglycine methyl ester (PhGly-OMe), glutamic acid dibenzyl ester (Glu-diOBn), and valine benzyl ester (Val-OBn)]. The best enantioseparation was achieved in the case of PEA. The high enantioselectivity was rationalized by the strong π-π interaction of the extended π system of the aryl-substituted pyridine unit. PMID:26218505

  5. Immunoglobulins acquire the ability to interact with DNA after chromatography on QAE-Sephadex

    SciTech Connect

    Sulaeva, N.I.; Lekakh, I.V.; Poverennyi, A.M.

    1986-10-10

    It was established that IgG isolated from the sera of healthy humans contains a substantial number of antibodies that react with native DNA. Their ability to interact with DNA is manifested only after chromatography on an anion exchange resin, as a result of which IgG is divided into two portions - acid and basic immunoglobulins. The peculiarities of the interaction of both fractions with DNA and the specificity of this reaction were investigated. It was shown that the investigated IgG can react with native and denatured DNA, dextran sulfate, poly(G), and poly(I). The question of the possibility of the interaction of the antibodies studied with the charged structures of the cell and that of the role of these antibodies in the normal and pathological states are discussed.

  6. Preparation of poly(glycidylmethacrylate-divinylbenzene) weak acid cation exchange stationary phases with succinic anhydride, phthalic anhydride, and maleic anhydride for ion chromatography.

    PubMed

    Liu, Junwei; Wang, Yong; Wu, Shuchao; Zhang, Peimin; Zhu, Yan

    2016-08-01

    In this work, poly(glycidylmethacrylate-divinylbenzene) microspheres were prepared and applied for the preparation of weak acid cation exchange stationary phases. Succinic anhydride, phthalic anhydride, and maleic anhydride were selected as carboxylation reagents to prepare three weak acid cation exchangers by direct chemical derivatization reaction without solvent or catalyst. The diameters and dispersity of the microspheres were characterized by scanning electron microscopy; the amount of accessible epoxy groups and mechanical stability were also measured. The weak acid cation exchangers were characterized by Fourier transform infrared spectroscopy; the content of carboxyl groups was measured by traditional acid base titration method. The chromatographic properties were characterized and compared by separating alkali, alkaline earth metal ions and ammonium and polar amines. The separation properties enhanced in the order of succinic anhydride, phthalic anhydride, and maleic anhydride modified poly(glycidylmethacrylate-divinylbenzene) cation exchangers. PMID:27288092

  7. Similar interaction chromatography of proteins: A cross interaction chromatographic approach to estimate the osmotic second virial coefficient.

    PubMed

    Quigley, A; Williams, D R

    2016-08-12

    Self-interaction chromatography (SIC) has established itself as an important experimental technique for the measurement of the second osmotic virial coefficients B22. B22 data are critical for understanding a range of protein solution phenomena, particularly aggregation and crystallisation. A key limitation to the more extensive use of SIC is the need to develop a method for immobilising each specific protein of interest onto a chromatographic support. This requirement is both a time and protein consuming constraint, which means that SIC cannot be used as a high throughput method for screening a wide range of proteins and their variants. Here an experimental framework is presented for estimating B22 values using Similar Interaction Chromatography (SimIC). This work uses experimental B23 and B32 data for lysozyme, lactoferrin, catalase and concanavalin A to reliably estimate B22 using arithmetic mean field approximations and is demonstrated to give good agreement with SIC measurements of B22 for the same proteins. SimIC could form the basis of a rapid protein variant screening methods to assess the developability of protein therapeutic candidates for industrial and academic researchers with respect to aggregation behaviour by eluting target proteins through a series of well-characterised protein immobilized reference columns. PMID:27417064

  8. Advancement in stationary phase for peptide separation helps in protein identification: application to atheroma plaque proteomics using nano-chip liquid chromatography and mass spectrometry.

    PubMed

    Delporte, Cédric; Noyon, Caroline; Raynal, Pierre; Dufour, Damien; Nève, Jean; Abts, Frederic; Haex, Martin; Zouaoui Boudjeltia, Karim; Van Antwerpen, Pierre

    2015-03-13

    In the last decades, proteomics has largely progressed. Mass spectrometry and liquid chromatography (LC) are generally used in proteomics. These techniques enable proper separation of peptides and good identification and/or quantification of them. Later, nano-scaled liquid chromatography, improvements of mass spectrometry resolution and sensitivity brought huge advancements. Enhancements in chemistry of chromatographic columns also brought interesting results. In the present work, the potency of identification of proteins by different nano-chip columns was studied and compared with classical LC column. The present study was applied to cardiovascular field where proteomics has shown to be highly helpful in research of new biomarkers. Protein extracts from atheroma plaques were used and proteomics data were compared. Results show that fewer spectra were acquired by the mass spectrometer when nano-chip columns were used instead of the classical ones. However, approximately 40% more unique peptides were identified by the recently optimized chip named Polaris-HR-chip-3C18 column, and 20% more proteins were identified. This fact leads to the identification of more low-abundance proteins. Many of them are involved in atheroma plaque development such as apolipoproteins, ceruloplasmin, etc. In conclusion, present data shows that recent developments of nanoLC column chemistry and dimensions enabled the improved detection and identification of low-abundance proteins in atheroma plaques. Several of them are of major interest in the field of cardiovascular disease. PMID:25680550

  9. Interactions of delta shock waves and stability of Riemann solutions for nonlinear chromatography equations

    NASA Astrophysics Data System (ADS)

    Zhang, Qingling

    2016-03-01

    This paper is devoted to studying the simplified nonlinear chromatography equations by introducing the change of state variables. The Riemann solutions containing delta shock waves are presented. In order to study wave interactions of delta shock waves with elementary waves, the global structure of solutions is constructed completely when the initial data are taken as three pieces of constants and the delta shock waves are included. In particular, the strength of delta shock wave is expressed explicitly and the delta contact discontinuity is discovered during the process of wave interactions. Moreover, by analyzing the limits of the solutions as the middle region vanishes, we observe that the Riemann solutions are stable for such a local small perturbation of the Riemann initial data.

  10. Enantiomeric separation of volatile organics by gas chromatography for the in situ analysis of extraterrestrial materials: kinetics and thermodynamics investigation of various chiral stationary phases.

    PubMed

    Freissinet, C; Buch, A; Szopa, C; Sternberg, R

    2013-09-01

    The performances of several commercial chiral capillary columns have been evaluated with the aim of determining the one most suitable for enantiomeric separation in a gas chromatograph onboard a space probe. We compared the GC-MS response of three capillary columns coated with different chiral stationary phases (CSP) using volatile chiral organic molecules which are potential markers of a prebiotic organic chemistry. The three different chiral capillary columns are Chirasil-Val, with an amino acid derivative CSP, ChiralDex-β-PM, with a CSP composed of dissolved permethylated β-cyclodextrins in polysiloxane, and Chirasil-Dex, with a CSP made of modified cyclodextrins chemically bonded to the polysiloxane backbone. Both kinetics and thermodynamics studies have been carried out to evaluate the chiral recognition potential in these different types of columns. The thermodynamic parameters also allow a better understanding of the driving forces affecting the retention and separation of the enantiomers. The Chirasil-Dex-CSP displays the best characteristics for an optimal resolution of the chiral compounds, without preliminary derivatization. This CSP had been chosen to be the only chiral column in the Sample Analysis at Mars (SAM) experiment onboard the current Mars Science Laboratory (MSL) mission, and is also part of the Mars Organic Molecules Analyzer (MOMA) gas chromatograph onboard the next Martian mission ExoMars. The use of this column could also be extended to all space missions aimed at studying chirality in space. PMID:23921265

  11. Use of contactless conductivity detection for non-invasive characterisation of monolithic stationary-phase coatings for application in capillary ion chromatography.

    PubMed

    Gillespie, Eoin; Connolly, Damian; Macka, Miroslav; Nesterenko, Pavel N; Paull, Brett

    2007-12-01

    A capacitively-coupled contactless conductivity detector (C4D) has been utilised as an on-capillary detector within a capillary ion chromatograph, incorporating a reversed-phase monolithic silica capillary column semi-permanently modified with a suitable ionic surfactant. The monolithic capillary column (150 x 0.1 mm i.d.) was modified using sodium dioctyl sulfosuccinate (DOSS), an anionic surfactant, for the separation of small inorganic and organic cations. With the use of the on-capillary conductivity detector, the longitudinal homogeneity and temporal stability of the coating were investigated. The approach allowed a detailed non-invasive observation of the nature of the ion-exchange coating over time, and an example of an application of the technique to produce a longitudinal stationary-phase charge gradient is shown. An investigation of the basis of the measured on-capillary conductivity was carried out with a counter ion study, clearly showing the on-capillary detection technique could also distinguish between chemical forms of the immobilised ion exchanger. The above method was used to produce a stable and homogeneously-modified monolithic ion-exchange capillary column, for application to the separation of inorganic alkaline earth cations and amino acids. PMID:18318285

  12. Extensive database of liquid phase diffusion coefficients of some frequently used test molecules in reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography.

    PubMed

    Song, Huiying; Vanderheyden, Yoachim; Adams, Erwin; Desmet, Gert; Cabooter, Deirdre

    2016-07-15

    Diffusion plays an important role in all aspects of band broadening in chromatography. An accurate knowledge of molecular diffusion coefficients in different mobile phases is therefore crucial in fundamental column performance studies. Correlations available in literature, such as the Wilke-Chang equation, can provide good approximations of molecular diffusion under reversed-phase conditions. However, these correlations have been demonstrated to be less accurate for mobile phases containing a large percentage of acetonitrile, as is the case in hydrophilic interaction liquid chromatography. A database of experimentally measured molecular diffusion coefficients of some 45 polar and apolar compounds that are frequently used as test molecules under hydrophilic interaction liquid chromatography and reversed-phase conditions is therefore presented. Special attention is given to diffusion coefficients of polar compounds obtained in large percentages of acetonitrile (>90%). The effect of the buffer concentration (5-10mM ammonium acetate) on the obtained diffusion coefficients is investigated and is demonstrated to mainly influence the molecular diffusion of charged molecules. Diffusion coefficients are measured using the Taylor-Aris method and hence deduced from the peak broadening of a solute when flowing through a long open tube. The validity of the set-up employed for the measurement of the diffusion coefficients is demonstrated by ruling out the occurrence of longitudinal diffusion, secondary flow interactions and extra-column effects, while it is also shown that radial equilibration in the 15m long capillary is effective. PMID:27240944

  13. Automated hydrophobic interaction chromatography column selection for use in protein purification.

    PubMed

    Murphy, Patrick J M; Stone, Orrin J; Anderson, Michelle E

    2011-01-01

    In contrast to other chromatographic methods for purifying proteins (e.g. gel filtration, affinity, and ion exchange), hydrophobic interaction chromatography (HIC) commonly requires experimental determination (referred to as screening or "scouting") in order to select the most suitable chromatographic medium for purifying a given protein (1). The method presented here describes an automated approach to scouting for an optimal HIC media to be used in protein purification. HIC separates proteins and other biomolecules from a crude lysate based on differences in hydrophobicity. Similar to affinity chromatography (AC) and ion exchange chromatography (IEX), HIC is capable of concentrating the protein of interest as it progresses through the chromatographic process. Proteins best suited for purification by HIC include those with hydrophobic surface regions and able to withstand exposure to salt concentrations in excess of 2 M ammonium sulfate ((NH(4;))(2;)SO(4;)). HIC is often chosen as a purification method for proteins lacking an affinity tag, and thus unsuitable for AC, and when IEX fails to provide adequate purification. Hydrophobic moieties on the protein surface temporarily bind to a nonpolar ligand coupled to an inert, immobile matrix. The interaction between protein and ligand are highly dependent on the salt concentration of the buffer flowing through the chromatography column, with high ionic concentrations strengthening the protein-ligand interaction and making the protein immobile (i.e. bound inside the column) (2). As salt concentrations decrease, the protein-ligand interaction dissipates, the protein again becomes mobile and elutes from the column. Several HIC media are commercially available in pre-packed columns, each containing one of several hydrophobic ligands (e.g. S-butyl, butyl, octyl, and phenyl) cross-linked at varying densities to agarose beads of a specific diameter (3). Automated column scouting allows for an efficient approach for determining

  14. Chromatographic comparison of atenolol separation in reaction media on cellulose tris-(3,5-dimethylphenylcarbamate) chiral stationary phase using ultra fast liquid chromatography.

    PubMed

    Agustian, Joni; Kamaruddin, Azlina Harun; Aboul-Enein, Hassan Y

    2012-05-01

    Because chiral liquid chromatography (LC) could become a powerful tool to estimate racemic atenolol quantity, excellent enantiomeric separation should be produced during data acquisition for satisfactory observation of atenolol concentrations throughout the racemic resolution processes. Selection of chiral LC column and analytical protocol that fulfill demands of the ultra fast LC analysis is essential. This article describes the characteristics of atenolol chromatographic separation that resulted from different resolution media and analytical protocols with the use of a Chiralcel® OD column. The chromatograms showed quite different characteristics of the separation process. The single enantiomer and racemic atenolol could be recognized by the Chiralcel® OD column in less than 20 min. Symmetrical peaks were obtained; however, several protocols produced peaks with wide bases and slanted baselines. Observations showed that efficient enantioresolution of racemic atenolol was obtained at slow mobile phase flow rate, decreased concentration of amine-type modifier but increased alcohol content in mobile phase and highest ultraviolet detection wavelength were required. The optimal ultra fast LC protocol enables to reduce and eliminate the peaks of either the atenolol solvent or the buffers and provided the highest peak intensities of both atenolol enantiomers. PMID:22517322

  15. Enantioselective determination of selfotel in human urine by high-performance liquid chromatography on a chiral stationary phase after derivatization with 9-fluorenylmethyl chloroformate.

    PubMed

    Knoche, B; Milosavljev, S; Gropper, S; Brunner, L A; Powell, M L

    1997-08-01

    An analytical method for the enantioselective determination of selfotel in human urine has been developed and validated. The method is based on high-performance liquid chromatography and utilizes CGS 20005 (a selfotel analog) as the internal standard. Urine samples were derivatized in situ with o-phthalic dicarboxaldehyde-3-mercaptopropionic acid and 9-fluorenylmethyl chloroformate (FMOC). Chromatographic separations of the FMOC derivatives of selfotel enantiomers and the internal standard were achieved using a column switching system consisting of an Inertsil ODS-2 column (75x4.6 mm I.D., 5 microm) and a Chiralcel OD-R column (250x4.6 mm I.D., 10 microm). The composition of the mobile phase was acetonitrile-0.1 M phosphate buffer, pH 2.50 (35:65) for the Inertsil ODS-2 column and acetonitrile-0.1 M phosphate buffer, pH 2.00 (35:65) for the Chiralcel OD-R column. The analytes were monitored using fluorescence detection at an excitation wavelength of 262 nm and an emission wavelength of 314 nm. The limit of quantification (LOQ) for this method is 0.25 microg/ml for each selfotel enantiomer. The method was successfully utilized to determine preliminary selfotel stereospecific pharmacokinetics. PMID:9300872

  16. Hydrophobic interaction chromatography for purification of monoPEGylated RNase A.

    PubMed

    Mayolo-Deloisa, Karla; Lienqueo, Ma Elena; Andrews, Barbara; Rito-Palomares, Marco; Asenjo, Juan A

    2012-06-15

    The chromatographic methods used for the purification of PEGylated proteins are mainly Size Exclusion (SEC) and Ion Exchange Chromatography (IEX). Although the PEGylation affects the protein hydrophobicity, Hydrophobic Interaction Chromatography (HIC) has not been extensively applied for the separation of these proteins. Purification of monoPEGylated Ribonuclease A (RNase A) using HIC is studied in this work. The products of the PEGylation reaction of RNase A with 20 kDa methoxy-poly(ethylene glycol) were separated using three resins with different degrees of hydrophobicity: Butyl, Octyl and Phenyl sepharose. The effects of resin type, concentration and salt type (ammonium sulphate or sodium chloride), and gradient length on the separation performance were evaluated. Yield and purity were calculated using the plate model. Under all conditions assayed the native protein was completely separated from PEGylated species. The best conditions for the purification of monoPEGylated RNase A were: Butyl sepharose, 1 M ammonium sulphate and 35 column volumes (CVs); this resulted in a yield as high as 85% with a purity of 97%. The purity of monoPEGylated RNase A is comparable to that obtained when the separation is performed using SEC, but the yield increases from 65% with SEC to ~85% with HIC. This process represents a viable alternative for the separation of PEGylated proteins. PMID:22579360

  17. Ethyl glucuronide determination in meconium and hair by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Tarcomnicu, Isabela; van Nuijs, Alexander L N; Aerts, Katrien; De Doncker, Mireille; Covaci, Adrian; Neels, Hugo

    2010-03-20

    Ethyl glucuronide (EtG) detection in non-conventional matrices, such as hair and meconium, can provide useful information on alcohol abuse over a long time frame, for example during pregnancy or after a withdrawal treatment. This study reports on the development, validation and application of a new hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for the analysis of EtG in meconium and hair. For each matrix, the sample preparation and the chromatographic separation were thoroughly optimised. Additionally, experiments with reversed-phase liquid chromatography were also performed in the development stages. Analyses were carried out using a Phenomenex Luna HILIC column (150 mm x 3 mm, 5 microm) and a mobile phase composed by ammonium acetate 2mM and acetonitrile, in gradient. Different SPE cartridges (Oasis MAX, Oasis WAX, aminopropyl silica) and solvents were tested in order to obtain the highest recoveries and cleanest extracts. Optimal results were obtained for meconium with aminopropyl cartridges, while for hair an incubation of 16 h with 2 mL of water and acetonitrile (50/50, v/v) provided good results. The analytical method was validated for both matrices (meconium and hair) by assessing linearity, precision, accuracy, recovery and limit of quantification. The calibration curve concentrations ranged from 50 to 1200 pg/mg for meconium and from 20 to 1000 pg/mg for hair. Real meconium and hair samples were analyzed and results were consistent with literature. PMID:20061101

  18. Application of Hydrophilic Interaction Liquid Chromatography for the Quantification of Flavonoids in Genista tinctoria Extract

    PubMed Central

    Biesaga, Magdalena; Pyrzynska, Krystyna

    2016-01-01

    Hydrophilic interaction chromatography (HILIC) was employed to investigate chromatographic behavior of selected flavonoids from their different subgroups differing in polarity. Chromatographic measurements were performed on two different HILIC columns: unmodified silica (Atlantis-HILIC) and zwitterionic sulfoalkylbetaine (SeQuant ZIC-HILIC). Separation parameters such as content and type of organic modifier were studied. On ZIC column retention factors were observed to be inversely proportional to the buffer content in the mobile phase, which is the typical partitioning mechanism. In the case of bare silica column more or less apparent dual retention mechanism was observed, depending on the water component content in the mobile phase. ZIC-HILIC showed better selectivity (in comparison to silica column) with the detection limit of 0.01 mg/L (only for rutin was 0.05 mg/L). Finally, this chromatographic procedure was validated and applied for the determination of some flavonoids in Genista tinctoria L. extract. PMID:27433372

  19. Separation of proteins from human plasma by sample displacement chromatography in hydrophobic interaction mode

    PubMed Central

    Josic, Djuro; Breen, Lucas; Clifton, James; Gajdosik, Martina Srajer; Gaso-Sokac, Dajana; Rucevic, Marijana; Müller, Egbert

    2013-01-01

    Sample displacement chromatography (SDC) in reversed-phase and ion-exchange modes was introduced approximately twenty years ago. This method was first used for the preparative purification of peptides and proteins. Recently, SDC in ion-exchange mode was also successfully used for enrichment of low abundance proteins from human plasma. In this paper, the use of SDC for the separation of plasma proteins in hydrophobic interaction mode is demonstrated. By use of two or more columns coupled in series during sample application, and subsequent elution of detached columns in parallel, additional separation of bound proteins was achieved. Further low-abundance, physiologically active proteins could be highly enriched and detected by ESI-MS/MS. PMID:22740472

  20. Analytical method for determining iminoctadine triacetate by LC/ESI/MS using hydrophilic interaction chromatography.

    PubMed

    Kawamoto, Tatsuhiko; Yano, Miho; Makihata, Nobuko

    2006-04-01

    A target value for iminoctadine triacetate residues in tap water was set at 6 microg/l in Japan. We have developed a highly selective and sensitive analytical method for iminoctadine triacetate by solid phase extraction LC/ESI/MS using hydrophilic interaction chromatography. The recovery rates at concentration of 0.06, 0.6, and 6 microg/l in distilled water, tap water, and raw water were 77.1 - 96.7%, and CV were 3.7 - 13.2%. The quantitation limit of the present method was 0.04 microg/l, and it was able to measure even one-hundredth of the target value of iminoctadine triacetate quantitatively. PMID:16760588

  1. Efficient method for preparation of highly purified lipopolysaccharides by hydrophobic interaction chromatography.

    PubMed

    Muck, A; Ramm, M; Hamburger, M

    1999-09-10

    A method for the efficient preparation of highly purified lipopolysaccharides (LPSs) by hydrophobic interaction chromatography (HIC) has been developed. The procedure can be used for the purification of cell wall bound LPSs after hot phenol-water extraction and for the isolation of extracellular LPSs from the supernatant, respectively. The method described has been tested with artificial mixtures containing LPSs, polysaccharide, protein and RNA and subsequently employed for the preparative purification of two LPSs of different origin, namely the extracellular LPS secreted by Escherichia coli E49 into the culture medium, and the cell wall bound LPS from Pseudomonas aeruginosa VA11465/1. Compared to currently used methods for LPS purification such as enzymatic digestion and ultracentrifugation, the chromatographic separation reported here combines superior purity with minimal loss of LPS, high reproducibility and simple handling. The removal of contaminants such as protein, RNA and polysaccharides and the recovery of LPSs were monitored by appropriate assays. PMID:10517220

  2. Separation of type IX collagen from other cartilage collagens by hydrophobic interaction chromatography.

    PubMed

    Macek, J; Lichý, A; Tesarová, E; Adam, M

    1988-12-30

    Collagen type IX was separated from other cartilage collagens (types II and XI) by hydrophobic interaction chromatography on a 25 cm X 8 mm I.D. stainless-steel column packed with Separon HEMA 1000 Bio. The mobile phase was 0.84 M ammonium sulphate with 0.1 M potassium dihydrogenphosphate (pH 6.5). Under these conditions only collagen type IX was eluted from the column; it could be monitored with UV detection (218 nm) or selectively with fluorescence detection (excitation 330 nm, emission filter 389 nm). The method can be used for the isolation and quantitation of collagen type IX. The assay was linear in the range 0-10 micrograms, the correlation coefficient was 0.99, precision 5.5% and accuracy 13%. The detection limit was about 0.6 microgram. PMID:3246532

  3. Quantification of homocysteine and cysteine by derivatization with pyridoxal 5'-phosphate and hydrophilic interaction liquid chromatography.

    PubMed

    Głowacki, Rafał; Stachniuk, Justyna; Borowczyk, Kamila; Jakubowski, Hieronim

    2016-03-01

    A simple and rapid assay using pyridoxal 5'-phosphate (PLP) as a derivatizing reagent was developed for the simultaneous determination of homocysteine (Hcy) and cysteine (Cys) in human plasma. Derivatization with PLP affords UV-absorbing tetrahydrothiazine and thiazolidine derivatives of Hcy and Cys, respectively. Separation of these derivatives was achieved in 5 min using a hydrophilic interaction liquid chromatography, followed by UV detection at 330 nm. Linearity in detector response was observed over the range of 0.25-20 μM for Hcy and 10-300 μM for Cys. The limit of quantification (LOQ) values for Hcy and Cys were 0.25 and 2.5 μM, respectively. The method was successfully applied to plasma samples donated by apparently healthy volunteers. Graphical Abstract A flow chart diagram of analytical procedure for total plasma Hcy and Cys determination. PMID:26794212

  4. Determination of major phlorotannins in Eisenia bicyclis using hydrophilic interaction chromatography: seasonal variation and extraction characteristics.

    PubMed

    Kim, Sang Min; Kang, Suk Woo; Jeon, Je-Seung; Jung, Yu-Jin; Kim, Woo-Ri; Kim, Chul Young; Um, Byung-Hun

    2013-06-15

    In this study, a hydrophilic interaction chromatography (HILIC) condition was developed for the simultaneous determination of five major phlorotannins from an extract of Eisenia bicyclis (Kjellman) Setchell with good linearity (r(2)>0.999). Based on this method, the seasonal variations and extraction characteristics, in terms of total extraction yield and the content of the phlorotannins, were investigated under various extraction conditions. In results, the yields and phlorotannins were increased two-to-four times in summer (June-October) and then, were decreased to normal levels in winter (November-March). In the extraction of E. bicyclis, ethanol percentage in water, extraction time and washing time significantly affected the yield of the extract and the phlorotannins, whereas the temperature and the sample/solvent ratio impacted the extraction to a lesser degree. These results will be useful information in the application of this macroalga in the commercial areas related to nutraceuticals, pharmaceuticals, and cosmeceuticals. PMID:23497901

  5. Effect of content of chiral selector and pore size of core-shell type silica support on the performance of amylose tris(3,5-dimethylphenylcarbamate)-based chiral stationary phases in nano-liquid chromatography and capillary electrochromatography.

    PubMed

    Rocchi, Silvia; Fanali, Salvatore; Farkas, Tivadar; Chankvetadze, Bezhan

    2014-10-10

    In this study the separation performance of various chiral stationary phases (CSPs) made of polysaccharide-based chiral selectors coated onto superficially porous (core-shell or fused-core) silica supports were evaluated. The CSPs obtained by coating of various amounts of chiral selector (1-5%) onto supports of various pore size (100 and 300 Å) were studied. Their evaluation was pursued in both chiral nano-liquid chromatography (nano-LC) and chiral capillary electrochromatography (CEC). Among the goals of this study was to re-examine our previous unexpected finding of better performance of superficially porous CSP under CEC conditions compared to nano-LC conditions for a new set of chiral compounds, as well as to study the effect of varying the chiral selector content and nominal pore size of supporting silica on the performance of core-shell silica-based polysaccharide-type CSPs. Based on the results of this study it can be seen that CSPs based on superficially porous silica can successfully be used for the separation of enantiomers in both nano-LC and CEC mode. Only a slight advantage of CEC over nano-LC mode was observed in this study from the viewpoint of plate numbers, especially at higher mobile phase flow rates. It must also be noted that the optimal theoretical plate height is still too high and further optimization of superficially porous CSPs is necessary for both nano-LC and CEC applications. PMID:24908153

  6. Estimation of polyurethane-carbon black interactions by means of inverse gas chromatography.

    PubMed

    Strzemiecka, Beata; Voelkel, Adam; Donate-Robles, Jessica; Martín-Martínez, José Miguel

    2013-11-01

    The properties of composites depend mainly on the interfacial interactions between filler and matrix that can be related to the adhesion between filler and polymer matrix. In this study the work of cohesion between the carbon black particles - Wcoh - and the thermodynamic work of adhesion - Wa - between four carbon blacks of different specific surface area and surface chemistry (nature and content of carbon-oxygen functional groups) and thermoplastic polyurethane were calculated by means of inverse gas chromatography (IGC) at infinite dilution. IGC derived data indicated that the work of adhesion increased by increasing the surface area of the carbon black, but the opposite trend was found in Wa/Wcoh and work of cohesion. According to the Wa/Wcoh values the filler particles should be well dispersed into the polyurethane matrix giving homogenous composites. The carbon black-thermoplastic polyurethane interactions determined by plate-plate rheology showed the same trend than that for the Wa/Wcoh values. However, the thermodynamic work of adhesion values derived from IGC were not in agreement with the carbon black-polyurethane interfacial interactions, likely due to the dominant effect of the carbon black in reducing the crystallinity and increasing the degree of phase separation of the thermoplastic polyurethane. PMID:24063983

  7. CHARACTERIZATION OF DRUG INTERACTIONS WITH SERUM PROTEINS BY USING HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Hage, David S.; Anguizola, Jeanethe; Barnaby, Omar; Jackson, Abby; Yoo, Michelle J.; Papastavros, Efthimia; Pfaunmiller, Erika; Sobansky, Matt; Tong, Zenghan

    2011-01-01

    The binding of drugs with serum proteins can affect the activity, distribution, rate of excretion, and toxicity of pharmaceutical agents in the body. One tool that can be used to quickly analyze and characterize these interactions is high-performance affinity chromatography (HPAC). This review shows how HPAC can be used to study drug-protein binding and describes the various applications of this approach when examining drug interactions with serum proteins. Methods for determining binding constants, characterizing binding sites, examining drug-drug interactions, and studying drug-protein dissociation rates will be discussed. Applications that illustrate the use of HPAC with serum binding agents such as human serum albumin, α1-acid glycoprotein, and lipoproteins will be presented. Recent developments will also be examined, such as new methods for immobilizing serum proteins in HPAC columns, the utilization of HPAC as a tool in personalized medicine, and HPAC methods for the high-throughput screening and characterization of drug-protein binding. PMID:21395530

  8. ANALYSIS OF DRUG INTERACTIONS WITH HIGH DENSITY LIPOPROTEIN BY HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Chen, Sike; Sobansky, Matthew R.; Hage, David S.

    2009-01-01

    Columns containing immobilized lipoproteins were prepared for the analysis of drug interactions with these particles by high-performance affinity chromatography. This approach was evaluated by using it to examine the binding of high density lipoprotein (HDL) to the drugs propranolol or verapamil. HDL was immobilized by the Schiff base method onto silica and gave HPLC columns with reproducible binding to propranolol over four to five days of continuous operation at pH 7.4. Frontal analysis experiments indicated that two types of interactions were occurring between R/S-propranolol and HDL at 37°C: saturable binding with an association equilibrium constant (Ka) of 1.1–1.9 × 105 M−1, and non-saturable binding with an overall affinity constant (n Ka) of 3.7–4.1 × 104 M−1. Similar results were found at 4 and 27°C. Verapamil also gave similar behavior, with a Ka of 6.0 × 104 M−1 at 37°C for the saturable sites and a n Ka value for the non-saturable sites of 2.5 × 104 M−1. These measured affinities gave good agreement with solution-phase values. The results indicated HPAC can be used to study drug interactions with HDL, providing information that should be valuable in obtaining a better description of how drugs are transported within the body. PMID:19833090

  9. Characterization of goat colostrum oligosaccharides by nano-liquid chromatography on chip quadrupole time-of-flight mass spectrometry and hydrophilic interaction liquid chromatography-quadrupole mass spectrometry.

    PubMed

    Martín-Ortiz, A; Salcedo, J; Barile, D; Bunyatratchata, A; Moreno, F J; Martin-García, I; Clemente, A; Sanz, M L; Ruiz-Matute, A I

    2016-01-01

    A detailed qualitative and quantitative characterization of goat colostrum oligosaccharides (GCO) has been carried out for the first time. Defatted and deproteinized colostrum samples, previously treated by size exclusion chromatography (SEC) to remove lactose, were analyzed by nanoflow liquid chromatography-quadrupole-time of flight mass spectrometry (Nano-LC-Chip-Q-TOF MS). Up to 78 oligosaccharides containing hexose, hexosamine, fucose, N-acetylneuraminic acid or N-glycolylneuraminic acid monomeric units were identified in the samples, some of them detected for the first time in goat colostra. As a second step, a hydrophilic interaction liquid chromatography coupled to mass spectrometry (HILIC-MS) methodology was developed for the separation and quantitation of the main GCO, both acidic and neutral carbohydrates. Among other experimental chromatographic conditions, mobile phase additives and column temperature were evaluated in terms of retention time, resolution, peak width and symmetry of target carbohydrates. Narrow peaks (wh: 0.2-0.6min) and good symmetry (As: 0.8-1.4) were obtained for GCO using an acetonitrile:water gradient with 0.1% ammonium hydroxide at 40°C. These conditions were selected to quantify the main oligosaccharides in goat colostrum samples. Values ranging from 140 to 315mgL(-1) for neutral oligosaccharides and from 83 to 251mgL(-1) for acidic oligosaccharides were found. The combination of both techniques resulted to be useful to achieve a comprehensive characterization of GCO. PMID:26427327

  10. Characterization of goat colostrum oligosaccharides by nano-liquid chromatography on chip quadrupole time-of-flight mass spectrometry and hydrophilic interaction liquid chromatography-quadrupole mass spectrometry

    PubMed Central

    Martín-Ortiz, A.; Salcedo, J.; Barile, D.; Bunyatratchata, A.; Moreno, F.J.; Martin-García, I.; Clemente, A.; Sanz, M.L.; Ruiz-Matute, A.I.

    2016-01-01

    A detailed qualitative and quantitative characterization of goat colostrum oligosaccharides (GCO) has been carried out for the first time. Defatted and deproteinized colostrum samples, previously treated by size exclusion chromatography (SEC) to remove lactose, were analyzed by nanoflow liquid chromatography-quadrupole-time of flight mass spectrometry (Nano-LC-Chip-Q-TOF MS). Up to 78 oligosaccharides containing hexose, hexosamine, fucose, N-acetylneuraminic acid or N-glycolylneuraminic acid monomeric units were identified in the samples, some of them detected for the first time in goat colostra. As a second step, a hydrophilic interaction liquid chromatography coupled to mass spectrometry (HILIC-MS) methodology was developed for the separation and quantitation of the main GCO, both acidic and neutral carbohydrates. Among other experimental chromatographic conditions, mobile phase additives and column temperature were evaluated in terms of retention time, resolution, peak width and symmetry of target carbohydrates. Narrow peaks (wh: 0.2–0.6 min) and good symmetry (As: 0.8–1.4) were obtained for GCO using an acetonitrile:water gradient with 0.1% ammonium hydroxide at 40 °C. These conditions were selected to quantify the main oligosaccharides in goat colostrum samples. Values ranging from 140 to 315 mg L−1 for neutral oligosaccharides and from 83 to 251 mg L−1 for acidic oligosaccharides were found. The combination of both techniques resulted to be useful to achieve a comprehensive characterization of GCO. PMID:26427327

  11. Comprehensive analysis of pharmaceutical products using simultaneous mixed-mode (ion-exchange/reversed-phase) and hydrophilic interaction liquid chromatography.

    PubMed

    Kazarian, Artaches A; Nesterenko, Pavel N; Soisungnoen, Phimpha; Burakham, Rodjana; Srijaranai, Supalax; Paull, Brett

    2014-08-01

    Liquid chromatographic assays were developed using a mixed-mode column coupled in sequence with a hydrophilic interaction liquid chromatography column to allow the simultaneous comprehensive analysis of inorganic/organic anions and cations, active pharmaceutical ingredients, and excipients (carbohydrates). The approach utilized dual sample injection and valve-mediated column switching and was based upon a single high-performance liquid chromatography gradient pump. The separation consisted of three distinct sequential separation mechanisms, namely, (i) ion-exchange, (ii) mixed-mode interactions under an applied dual gradient (reversed-phase/ion-exchange), and (iii) hydrophilic interaction chromatography. Upon first injection, the Scherzo SS C18 column (Imtakt) provided resolution of inorganic anions and cations under isocratic conditions, followed by a dual organic/salt gradient to elute active pharmaceutical ingredients and their respective organic counterions and potential degradants. At the top of the mixed-mode gradient (high acetonitrile content), the mobile phase flow was switched to a preconditioned hydrophilic interaction liquid chromatography column, and the standard/sample was reinjected for the separation of hydrophilic carbohydrates, some of which are commonly known excipients in drug formulations. The approach afforded reproducible separation and resolution of up to 23 chemically diverse solutes in a single run. The method was applied to investigate the composition of commercial cough syrups (Robitussin®), allowing resolution and determination of inorganic ions, active pharmaceutical ingredients, excipients, and numerous well-resolved unknown peaks. PMID:24890905

  12. Analysis of drugs of abuse in wastewater by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    van Nuijs, Alexander L N; Tarcomnicu, Isabela; Bervoets, Lieven; Blust, Ronny; Jorens, Philippe G; Neels, Hugo; Covaci, Adrian

    2009-10-01

    The simultaneous analysis of nine drugs of abuse (DOAs) and their metabolites (amphetamine, methamphetamine, methylenedioxymethamphetamine, methadone, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine, cocaine, benzoylecgonine, ecgonine methyl ester and 6-monoacetylmorphine) in wastewater based on hydrophilic interaction liquid chromatography (HILIC) coupled to tandem mass spectrometry (MS/MS) was optimised and validated. For each analyte, the deuterated analogue was used for quantification. The separation by HILIC showed good performance for all compounds, especially for the hydrophilic compounds, which elute early (amphetamine-like stimulants) or show no retention (ecgonine methyl ester) in reversed-phase liquid chromatography. Sample preparation based on solid-phase extraction was optimised by comparing Oasis HLB and Oasis MCX sorbents for various parameters such as sample pH, amount of sorbent bed and washing solvent. The method was validated for each compound by assessing the following parameters (following International Conference on Harmonisation guidelines): specificity, limit of quantification (LOQ), linearity, accuracy, precision, recovery and matrix effects. LOQs were 2 ng/L for 6-monoacetylmorphine, ecgonine methyl ester and amphetamine and 1 ng/L for the rest of the compounds, corresponding with the lowest point in the calibration curve. Except for 6-monoacetylmorphine, all compounds were detected from 1 to 819 ng/L in influent wastewater samples (n = 12) collected from 11 different wastewater treatment plants across Belgium. The presence of ecgonine methyl ester in wastewater could be demonstrated for the first time. In the future, the new HILIC-MS/MS method will be applied to assess the use of DOAs in Belgium using the "sewage epidemiology" approach. PMID:19685341

  13. Characteristics of the interaction of calcium with casein submicelles as determined by analytical affinity chromatography

    SciTech Connect

    Jang, H.D.; Swaisgood, H.E. )

    1990-12-01

    Interaction of calcium with casein submicelles was investigated in CaCl2 and calcium phosphate buffers and with synthetic milk salt solutions using the technique of analytical affinity chromatography. Micelles that had been prepared by size exclusion chromatography with glycerolpropyl controlled-pore glass from fresh raw skim milk that had never been cooled, were dialyzed at room temperature against calcium-free imidazole buffer, pH 6.7. Resulting submicelles were covalently immobilized on succinamidopropyl controlled-pore glass (300-nm pore size). Using 45Ca to monitor the elution retardation, the affinity of free Ca2+ and calcium salt species was determined at temperatures of 20 to 40 degrees C and pH 6.0 to 7.5. Increasing the pH in this range or increasing the temperature strengthened the binding of calcium to submicelles, similar to previous observations with individual caseins. However, the enthalpy change obtained from the temperature dependence was considerably greater than that reported for alpha s1- and beta-caseins. Furthermore, the elution profiles for 45Ca in milk salt solutions were decidedly different from those in CaCl2 or calcium phosphate buffers and the affinities were also greater. For example, at pH 6.7 and 30 degrees C the average dissociation constant for the submicelle-calcium complex is 0.074 mM for CaCl2 and calcium phosphate buffers, vs 0.016 mM for the milk salt solution. The asymmetric frontal boundaries and higher average affinities observed with milk salts may be due to binding of calcium salts with greater affinity in addition to the binding of free Ca2+ in these solutions.

  14. Hydrophobic interaction chromatography in dual salt system increases protein binding capacity.

    PubMed

    Senczuk, Anna M; Klinke, Ralph; Arakawa, Tsutomu; Vedantham, Ganesh; Yigzaw, Yinges

    2009-08-01

    Hydrophobic interaction chromatography (HIC) uses weakly hydrophobic resins and requires a salting-out salt to promote protein-resin interaction. The salting-out effects increase with protein and salt concentration. Dynamic binding capacity (DBC) is dependent on the binding constant, as well as on the flow characteristics during sample loading. DBC increases with the salt concentration but decreases with increasing flow rate. Dynamic and operational binding capacity have a major raw material cost/processing time impact on commercial scale production of monoclonal antibodies. In order to maximize DBC the highest salt concentration without causing precipitation is used. We report here a novel method to maintain protein solubility while increasing the DBC by using a combination of two salting-out salts (referred to as dual salt). In a series of experiments, we explored the dynamic capacity of a HIC resin (TosoBioscience Butyl 650M) with combinations of salts. Using a model antibody, we developed a system allowing us to increase the dynamic capacity up to twofold using the dual salt system over traditional, single salt system. We also investigated the application of this novel approach to several other proteins and salt combinations, and noted a similar protein solubility and DBC increase. The observed increase in DBC in the dual salt system was maintained at different linear flow rates and did not impact selectivity. PMID:19382248

  15. Analysis of Lidocaine Interactions with Serum Proteins Using High-Performance Affinity Chromatography

    PubMed Central

    Soman, Sony; Yoo, Michelle J.; Jang, Yoon Jeong; Hage, David S.

    2010-01-01

    High-performance affinity chromatography was used to study binding by the drug lidocaine to human serum albumin (HSA) and α1–acid glycoprotein (AGP). AGP had strong binding to lidocaine, with an association equilibrium constant (Ka) of 1.1-1.7 × 105 M-1 at 37 °C and pH 7.4. Lidocaine had weak-to-moderate binding to HSA, with a Ka in the range of 103 to 104 M-1. Competitive experiments with site selective probes showed that lidocaine was interacting with Sudlow site II of HSA and the propranolol site of AGP. These results agree with previous observations in the literature and provide a better quantitative understanding of how lidocaine binds to these serum proteins and is transported in the circulation. This study also demonstrates how HPAC can be used to examine the binding of a drug with multiple serum proteins and provide detailed information on the interaction sites and equilibrium constants that are involved in such processes. PMID:20138813

  16. Introducing enantioselective ultrahigh-pressure liquid chromatography (eUHPLC): theoretical inspections and ultrafast separations on a new sub-2-μm Whelk-O1 stationary phase.

    PubMed

    Kotoni, Dorina; Ciogli, Alessia; Molinaro, Carmela; D'Acquarica, Ilaria; Kocergin, Jelena; Szczerba, Ted; Ritchie, Harald; Villani, Claudio; Gasparrini, Francesco

    2012-08-01

    A new chiral stationary phase for ultrahigh-pressure liquid chromatography (UHPLC) applications was prepared by covalent attachment of the Whelk-O1 selector to spherical, high-surface-area 1.7-μm porous silica particles. Columns of varying dimensions (lengths of 50, 75, 100, and 150 mm and internal diameters of 3.0 or 4.6 mm) were packed and characterized in terms of permeability, efficiency, retention, and enantioselectivity, using both organic and water-rich mobile phases. A conventional HPLC Whelk-O1 column based on 5.0-μm porous silica particles and packed in a 250 mm × 4.6 mm column was used as a reference. Van Deemter curves, generated with low-molecular-weight solutes on a 100 mm × 4.6 mm column packed with the 1.7-μm particles, showed H(min) (μm) and μ(opt) (mm/s) values of 4.10 and 5.22 under normal-phase and 3.74 and 4.34 under reversed-phase elution conditions. The flat C term of the van Deemter curves observed with the 1.7-μm particles allowed the use of higher-than-optimal flow rates without significant efficiency loss. Kinetic plots constructed from van Deemter data confirmed the ability of the column packed with the 1.7-μm particles to afford subminute separations with good efficiency and its superior performances in the high-speed regime, compared to the column packed with 5.0-μm particles. Resolutions in the time scale of seconds were obtained using a 50-mm-long column in the normal phase or polar organic mode. The intrinsic kinetic performances of 1.7-μm silica particles are retained in the Whelk-O1 chiral stationary phase, clearly demonstrating the potentials of enantioselective UHPLC in terms of high speed, throughput, and resolution. PMID:22725676

  17. Fractionation and analysis of lipopolysaccharide-derived oligosaccharides by zwitterionic-type hydrophilic interaction liquid chromatography coupled with electrospray ionisation mass spectrometry.

    PubMed

    Man-Kupisinska, Aleksandra; Bobko, Ewelina; Gozdziewicz, Tomasz K; Maciejewska, Anna; Jachymek, Wojciech; Lugowski, Czeslaw; Lukasiewicz, Jolanta

    2016-06-01

    Lipopolysaccharide (LPS, endotoxin) is a main surface antigen and virulence factor of Gram-negative bacteria. Regardless of the source of LPS, this molecule, isolated from the smooth forms of bacteria, is characterised by a general structural layout encompassing three regions: (i) an O-specific polysaccharide (O-PS) - a polymer of repeating oligosaccharide units, (ii) core oligosaccharide (OS), and (iii) the lipid A anchoring LPS in the outer membrane of the cell envelope of Gram-negative bacteria. Structural analysis usually requires degradation of LPS and further efficient separation of various poly- and oligosaccharide glycoforms. The hydrophilic interaction liquid chromatography (HILIC) was shown as an efficient technique for separation of labelled or native neutral and acidic glycans, glycopeptides, sialylated glycans, glycosylated and nonglycosylated peptides. Herein we adopted ZIC(®) (zwitterionic stationary phase covalently attached to porous silica)-HILIC technology in combination with electrospray ionisation mass spectrometry to separate different LPS-derived oligosaccharides. As a result three effective procedures have been developed: (i) to separate different core oligosaccharides of Escherichia coli R1 LOS, (ii) to separate RU-[Hep]-Kdo oligosaccharides from core OS glycoforms of Hafnia alvei PCM 1200 LPS, and (iii) to separate Hep and Kdo-containing mono, di-, tri- and tetrasaccharides of H. alvei PCM 1200 LPS. Moreover, some of developed analytical procedures were scaled to semi-preparative protocols and used to obtain highly-purified fractions of the interest in larger quantities required for future evaluation, analysis, and biological applications. PMID:27085741

  18. Characterization of grape seed procyanidins by comprehensive two-dimensional hydrophilic interaction × reversed phase liquid chromatography coupled to diode array detection and tandem mass spectrometry.

    PubMed

    Montero, Lidia; Herrero, Miguel; Prodanov, Marin; Ibáñez, Elena; Cifuentes, Alejandro

    2013-05-01

    In this work, the development and optimization of a new methodology to analyze grape seed procyanidins based on the application of two-dimensional comprehensive LC is presented. This two-dimensional method involves the use of a microbore column containing a diol stationary phase in the first dimension coupled to either a C18 partially porous short column or a C18 monolithic column in the second dimension. The orthogonal hydrophilic interaction × reversed phase liquid chromatography (HILIC×RP-LC) system is interfaced through a ten-port two-position switching valve. The optimized HILIC×RP-LC separation followed by diode array and tandem mass spectrometry detection (HILIC×RP-LC-DAD-MS/MS) made possible the direct analysis of a complex grape seed extract and allowed the tentative identification of 43 flavan-3-ols, including monomers and procyanidin oligomers till a polymerization degree of 7 units with different galloylation degrees. To the best of our knowledge, this is the first time that this powerful analytical technique is employed to characterize complex procyanidin samples. This work successfully demonstrates the great capabilities of the HILIC×RP-LC-DAD-MS/MS coupling for the direct analysis of very complex natural samples like grape seeds. PMID:23224621

  19. Simultaneous analysis of acetaminophen, p-aminophenol and aspirin metabolites by hydrophilic interaction and strong anion exchange capillary liquid chromatography coupled to amperometric detection.

    PubMed

    Zheng, Minmin; Wu, Yimin; Lu, Lanxiang; Ding, Kang; Tang, Fengxiang; Lin, Zian; Wu, Xiaoping

    2011-08-01

    A simple and sensitive method has been developed for the simultaneous determination of polar nonsteroidal pharmaceuticals and metabolites, including acetaminophen, p-aminophenol and several aspirin metabolites (salicylic acid, gentisic acid, salicyluric acid and 2,3-dihydroxybenzoic acid), by capillary liquid chromatography with amperometric detection. Using a capillary monolithic column with mixed mode stationary phases and a mobile phase composed of acetonitrile and Tris buffer, rapid separation of six polar analytes was achieved within 8 min, and a hydrophilic interaction and strong anion exchange separation mechanism were exhibited. Method detection limits of six analytes ranged from 10 to 50 ng/mL. In terms of precision, the intra- and interday relative standard deviation values in all analytes never exceeded 3.1% for migration time and 8.9% for peak areas, respectively. This method provided a simple, rapid and cost-effective approach for the analysis of polar pharmaceuticals. The applicability of the method in pharmacokinetics was verified by spiking human serum samples with the compounds and analyzing the recoveries. PMID:21644253

  20. Separation and analysis of mono-glucosylated lipids in brain and skin by hydrophilic interaction chromatography based on carbohydrate and lipid moiety.

    PubMed

    Nakajima, Kazuki; Akiyama, Hisako; Tanaka, Kaori; Kohyama-Koganeya, Ayako; Greimel, Peter; Hirabayashi, Yoshio

    2016-09-15

    Mono-glycosylated sphingolipids and glycerophospholipids play important roles in diverse biological processes and are linked to a variety of pathologies, such as Parkinson disease. The precise identification of the carbohydrate head group of these lipids is complicated by their isobaric nature and by substantial differences in concentration in different biological samples. To overcome these obstacles, we developed a zwitterionic (ZIC)-hydrophilic interaction chromatography (HILIC) electrospray ionization tandem mass spectrometry method. ZIC-HILIC preferentially retains inositol, followed by glucose- and galactose-featuring lipids. Comparison with unmodified silica gel HILIC stationary phase revealed different retention specificity. To evaluate the precision of ZIC-HILIC, we quantified glucosyl- (GlcCer) and galactosylceramides (GalCer) in seven different regions of the mouse brain and discovered that GlcCer and GalCer concentrations are inversely related. The highest GalCer (lowest GlcCer) content was found in the medulla oblongata and hippocampus, whereas the highest GlcCer (lowest GalCer) content was found in other regions. With a neutral loss scan, ZIC-HILIC resolved glucosylceramide species featuring non-hydroxylated fatty acid, hydroxylated fatty acid, and trihydroxy sphingoid bases in mouse epidermis samples. This demonstrates that our ZIC-HILIC-based approach is a valuable tool for characterizing the structural diversity of mono-glucosylated lipids in biological material and for quantifying these important lipids. PMID:27485395

  1. Insight into the retention processes of phthalate metabolites on different liquid chromatography stationary phases for the development of improved separation methods.

    PubMed

    Gómara, B; Lebrón-Aguilar, R; González, M J; Quintanilla-López, J E

    2015-12-01

    The retention behavior of nine MPAEs has been studied, using commercial LC columns with octadecylsilane (ODS), phenyl, and amide-type SPs. First, it was found that the use of methanol in the mobile phase is not advisable, because induce a transesterification reaction of MPAEs in the electrospray ion source, regardless of the SP used. On the other hand, different responses were observed when representing the logarithm of retention factors (k) vs. the volume fraction of ACN (φ) in the mobile phase, for the three SPs tested. A quite linear trend was obtained for ODS (at φ values below 0.80) and Phenyl columns. On the contrary, the Amide column shows a striking U-shape trend, typical of both hydrophobic and hydrophilic retention mechanisms. Therefore, the separation process was mainly hydrophobic in the ODS and phenyl SPs, but in the amide-type a dual retention mechanism was found, showing zones with predominant hydrophobic or hydrophilic interactions, depending on both the compound and the experimental conditions. A high content of acetonitrile (>75%) and low concentration of formic acid in the mobile phase promote the hydrophilic separation mechanism for MPAEs on the amide SP. So, this dual separation mechanism can be modulated modifying the pH and content of organic modifier in the mobile phase, allowing greater flexibility to develop improved methods. Taking advantage of this, a separation method was optimized in this amide column using a Box-Wilson Central Composite experimental design, which allows separating the studied MPAEs with a time-saving of around 40% comparing to the conventional phenyl SP. PMID:26553955

  2. Stationary phase deposition based on onium salts

    DOEpatents

    Wheeler, David R.; Lewis, Patrick R.; Dirk, Shawn M.; Trudell, Daniel E.

    2008-01-01

    Onium salt chemistry can be used to deposit very uniform thickness stationary phases on the wall of a gas chromatography column. In particular, the stationary phase can be bonded to non-silicon based columns, especially microfabricated metal columns. Non-silicon microfabricated columns may be manufactured and processed at a fraction of the cost of silicon-based columns. In addition, the method can be used to phase-coat conventional capillary columns or silicon-based microfabricated columns.

  3. Expanding the potential of chiral chromatography for high-throughput screening of large compound libraries by means of sub-2μm Whelk-O 1 stationary phase in supercritical fluid conditions.

    PubMed

    Sciascera, Luca; Ismail, Omar; Ciogli, Alessia; Kotoni, Dorina; Cavazzini, Alberto; Botta, Lorenzo; Szczerba, Ted; Kocergin, Jelena; Villani, Claudio; Gasparrini, Francesco

    2015-02-27

    With the aim of exploring the potential of ultra-fast chiral chromatography for high-throughput analysis, the new sub-2 micron Whelk-O 1 chiral stationary phase (CSP) has been employed in supercritical fluid conditions to screen 129 racemates, mainly of pharmaceutical interest. By using a 5-cm long column (0.46cm internal diameter), a single co-solvent (MeOH) and a 7-min gradient elution, 85% of acidic and neutral analytes considered in this work have been successfully resolved, with resolution (Rs) larger than 2 in more than 65% of cases. Moreover, almost a half of basic samples that, for their own characteristics, are known to be difficult to separate on Whelk-O 1 CSP, have shown Rs greater than 0.3. The screening of the entire library could be accomplished in less than 24h (single run) with 63% of positive score. For well-resolved enantiomers (Rs roughly included between 1 and 3), we show that method transfer from gradient to isocratic conditions is straightforward. In many cases, isocratic ultra-fast separations (with analysis time smaller than 60s) have been achieved by simply employing, as isocratic mobile phase, the eluent composition at which the second enantiomer was eluted in gradient mode. By considering the extension and variety of the library in terms of chemico-physical and structural properties of compounds and numerousness, we believe that this work demonstrates the real potential of the technique for high-throughput enantioselective screening. PMID:25650355

  4. Study of Low Molecular Weight Impurities in Pluronic Triblock Copolymers using MALDI, Interaction Chromatography, and NMR

    NASA Astrophysics Data System (ADS)

    Helming, Z.; Zagorevski, D.; Ryu, C. Y.

    2014-03-01

    Poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) triblock copolymers are a group of commercial macromolecular amphiphilic surfactants that have been widely studied for their applications in polymer-based nanotechnology and drug-delivery. It has been well-established that the synthesis of commercial Pluronic triblocks results in low molecular weight ``impurities,'' which are generally disregarded in the applications and study of these polymers. These species have been shown to have significant effects on the rheological properties of the material, as well as altering the supramolecular ``micellar'' structures for which the polymers are most often used. We have isolated the impurities from the bulk Pluronic triblock using Interaction Chromatography (IC) techniques, and subjected them to analysis by H1 NMR and MALDI (Matrix-Assisted Laser Desorption Ionization) Mass Spectrometry to identify relative block composition and molecular weight information. We report significant evidence of at least two polymeric components: a low-molecular-weight homopolymer of poly(ethylene oxide) and a ``blocky'' copolymer of both poly(ethylene oxide) and poly(propylene oxide). This has significant implications, not only for the applied usage of Pluronic triblock copolymers, but for the general scientific acceptance of the impurities and their effects on Pluronic micelle and hydrogel formation.

  5. Direct determination of amino acids by hydrophilic interaction liquid chromatography with charged aerosol detection.

    PubMed

    Socia, Adam; Foley, Joe P

    2016-05-13

    A chromatographic analytical method for the direct determination of amino acids by hydrophilic interaction liquid chromatography (HILIC) was developed. A dual gradient simultaneously varying the pH 3.2 ammonium formate buffer concentration and level of acetonitrile (ACN) in the mobile phase was employed. Using a charged aerosol detector (CAD) and a 2(nd) order regression analysis, the fit of the calibration curve showed R(2) values between 0.9997 and 0.9985 from 1.5mg/mL to 50μg/mL (600ng to 20ng on column). Analyte chromatographic parameters such as the sensitivity of retention to the water fraction in the mobile phase values (mHILIC) were determined as part of method development. A degradation product of glutamine (5-pyrrolidone-2-carboxylic acid; pGlu) was observed and resolved chromatographically with no method modifications. The separation was used to quantitate amino acid content in acid hydrolysates of various protein samples. PMID:27059400

  6. Lipidomic profiling of dried seahorses by hydrophilic interaction chromatography coupled to mass spectrometry.

    PubMed

    Shen, Qing; Dai, Zhiyuan; Huang, Yao-Wen; Cheung, Hon-Yeung

    2016-08-15

    Dried seahorse is a precious raw food material for cooking soups. In this study, a lipidomics strategy using the techniques of solid-phase extraction (SPE) and hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-QTOF/MS) was developed for extraction, visualization, and quantification of phospholipids in dried seahorses. The parameters of SPE were optimized, and 1 mL of sample and chloroform/methanol (1:2, v/v) were found to be the best loading volume and eluting solvent, respectively. Afterwards, each phospholipid class was successfully separated on a HILIC column and analyzed by mass spectrometry. A total of 50 phospholipid molecular species were identified and determined, including 15 phosphatidylcholines (PCs), 14 phosphatidylethanolamines (PEs), 12 phosphatidylinositols (PIs) and 9 phosphatidylserines (PSs). In comparison to previously methods, this strategy was robust and efficient in extraction, characterization, and determination of phospholipids. The dried seahorse was found to contain large amounts of polyunsaturated fatty acyl phospholipids which are beneficial to human health. PMID:27006218

  7. Effect of electric field on the partitioning behavior of solutes in entropic interaction chromatography.

    PubMed

    Shi, Qing-Hong; Jia, Guo-Dong; Xu, Liang; Sun, Yan

    2013-09-01

    In this study, a novel column design with a round cross-section was proposed to be suitable for a transverse electric field (EF). Additionally, two beads for entropic interaction chromatography (EIC) were prepared by grafting glycidyl methacrylate onto Toyopearl HW-65F (T65F) beads. Solute partitioning was then investigated to elucidate the role of graft polymerization with and without an EF. In a T65F column, solute partitioning was attributed to the distinct pore structure in the beads and was governed by pore flow. Under EF, partition coefficients (Kp) for solutes decreased with increasing EF strength. In the two EIC columns, a decrease of Kp was also observed without an EF while the fractionation windows were extended. It was more pronounced in the EIC column with a high grafting density (T65F-H). This was explained by the decrease in the effective pore size of solutes caused by the steric hindrance of polymer chains. Under an EF, the solutes showed different partitioning behaviours in the T65F-H column. With increasing EF strength, Kp for vitamin B12 and myoglobin was decreased. In contrast, Kp for large solutes increased as a result of concentration polarization on the bead surface. Both behaviors were related to the modulation of graft polymerization to residual charge on the matrix and the pore size of the solutes. PMID:23857725

  8. CHARACTERIZATION OF DRUG-PROTEIN INTERACTIONS IN BLOOD USING HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Hage, David S.; Jackson, Abby; Sobansky, Matt; Schiel, John E.; Yoo, Michelle J.; Joseph, K. S.

    2009-01-01

    The binding of drugs with proteins in blood, serum or plasma is an important process in determining the activity, distribution, rate of excretion, and toxicity of drugs in the body. High-performance affinity chromatography (HPAC) has received a great deal of interest as a means for studying these interactions. This review examines the various techniques that have been used in HPAC to examine drug-protein binding and discusses the types of information that can be obtained through this approach. A comparison of these techniques with traditional methods for binding studies (e.g., equilibrium dialysis and ultrafiltration) will also be presented. The use of HPAC with specific serum proteins and binding agents will then be discussed, including human serum albumin and α1-acid glycoprotein. Several examples from the literature are provided to illustrate the applications of such research. Recent developments in this field are also described, such as the use of improved immobilization techniques, new data analysis methods, techniques for working for directly with complex biological samples, and work with immobilized lipoproteins. The relative advantages and limitations of the methods that are described will be considered and the possible use of these techniques in the high-throughput screening or characterization of drug-protein binding will be discussed. PMID:19278006

  9. Determination of ethyl glucuronide in human hair by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Yaldiz, Fadile; Daglioglu, Nebile; Hilal, Ahmet; Keten, Alper; Gülmen, Mete Korkut

    2013-10-01

    Ethyl glucuronide (EtG) is a direct metabolite of ethanol and has been utilized as a marker for alcohol intake. This study presents development, validation and application of a new hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for the analysis of EtG in human hair samples. The linearity was assessed in the range of 5-2000 pg/mg hair, with a correlation coefficient of >0.99. The method was selective and sensitive, with a limit of detection (LOD) and limit of quantitation (LOQ) of 0.05 pg/mg and 0.18 pg/mg in hair, respectively. Differently from the extraction procedures in the literature, a fast and simple liquid-liquid method was used and highest recoveries and cleanest extracts were obtained. The method was successfully applied to 30 human hair samples which were taken from those who state they consume alcohol. EtG concentrations in the hair samples of alcohol users participated in this study, ranged between 1.34 and 82.73 pg/mg. From the concentration of EtG in hair strands 20 of the 30 subjects can be considered regular moderate drinkers. PMID:24112322

  10. Studies of drug interactions with glycated human serum albumin by high-performance affinity chromatography

    PubMed Central

    Matsuda, Ryan; Kye, So-Hwang; Anguizola, Jeanethe; Hage, David S.

    2015-01-01

    Diabetes is a health condition associated with elevated levels of glucose in the bloodstream and affects 366 million people worldwide. Type II diabetes is often treated with sulfonylurea drugs, which are known to bind tightly in blood to the transport protein human serum albumin (HSA). One consequence of the elevated levels of glucose in diabetes is the non-enzymatic glycation of proteins such as HSA. Several areas of HSA are now known to be affected by glycation-related modifications, which may in turn affect the binding of sulfonylurea drugs and other solutes to this protein. This review discusses some recent studies that have examined these changes in drug-protein binding by employing high-performance affinity chromatography (HPAC). A description of the theoretical and experimental techniques that were used in these studies is given. The information on drug interactions with glycated HSA, as obtained through this method, is also summarized. In addition, the potential advantages of this approach in the areas of biointeraction analysis and personalized medicine are considered. PMID:26526139

  11. ANALYSIS OF DRUG INTERACTIONS WITH VERY LOW DENSITY LIPOPROTEIN BY HIGH PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Sobansky, Matthew R.; Hage, David S.

    2014-01-01

    High-performance affinity chromatography (HPAC) was utilized to examine the binding of very low density lipoprotein (VLDL) with drugs, using R/S-propranolol as a model. These studies indicated that two mechanisms existed for the binding of R- and S-propranolol with VLDL. The first mechanism involved non-saturable partitioning of these drugs with VLDL, which probably occurred with the lipoprotein's non-polar core. This partitioning was described by overall affinity constants of 1.2 (± 0.3) × 106 M-1 for R-propranolol and 2.4 (± 0.6) × 106 M-1 for S-propranolol at pH 7.4 and 37 °C. The second mechanism occurred through saturable binding by these drugs at fixed sites on VLDL, such as represented by apolipoproteins on the surface of the lipoprotein. The association equilibrium constants for this saturable binding at 37 °C were 7.0 (± 2.3) × 104 M-1 for R-propranolol and 9.6 (± 2.2) × 104 M-1 for S-propranolol. Comparable results were obtained at 20 °C and 27 °C for the propranolol enantiomers. This work provided fundamental information on the processes involved in the binding of R- and S-propranolol to VLDL, while also illustrating how HPAC can be used to evaluate relatively complex interactions between agents such as VLDL and drugs or other solutes. PMID:25103529

  12. Studies of drug interactions with glycated human serum albumin by high-performance affinity chromatography.

    PubMed

    Matsuda, Ryan; Kye, So-Hwang; Anguizola, Jeanethe; Hage, David S

    2014-08-01

    Diabetes is a health condition associated with elevated levels of glucose in the bloodstream and affects 366 million people worldwide. Type II diabetes is often treated with sulfonylurea drugs, which are known to bind tightly in blood to the transport protein human serum albumin (HSA). One consequence of the elevated levels of glucose in diabetes is the non-enzymatic glycation of proteins such as HSA. Several areas of HSA are now known to be affected by glycation-related modifications, which may in turn affect the binding of sulfonylurea drugs and other solutes to this protein. This review discusses some recent studies that have examined these changes in drug-protein binding by employing high-performance affinity chromatography (HPAC). A description of the theoretical and experimental techniques that were used in these studies is given. The information on drug interactions with glycated HSA, as obtained through this method, is also summarized. In addition, the potential advantages of this approach in the areas of biointeraction analysis and personalized medicine are considered. PMID:26526139

  13. [Deep eutectic solvent: a new kind of mobile phase modifier for hydrophilic interaction liquid chromatography].

    PubMed

    Tan, Ting; Qiao, Xin; Wan, Yiqun; Qiu, Hongdeng

    2015-09-01

    Deep eutectic solvents (DESs) were used as a new kind of mobile phase modifier in hydrophilic interaction liquid chromatography (HILIC). In our experiment, a SiO2 column (150 mm x 4.6 mm, 3 µm) was selected to separate several nucleobases and nucleosides by using the mixed solution of acetonitrile and DES (choline chloride-ethylene glycol (1:3, mol/mol) ) as mobile phase. Subsequently, the concentrations of DESs in acetonitrile and the column temperature on the effect of separation were investigated. According to the experimental results, better separation of nucleobases and nucleosides was obtained by using acetonitrile and DESs mixed solution as mobile phase than that using traditional water-based solution. For example, a baseline separation between cytosine and cytidine cannot be achieved by HILIC with water-based mobile phase, however, greater improvement was gained by HILIC with modified DES-acetonitrile mobile phase. Meanwhile, the retention times of nucleobases and nucleosides decreased as the proportion of DESs in acetonitrile increased, the most significant decrease of which was with cytidine. Similar retention behavior took place with the effect of column temperature. Decreased retention times of the analytes were observed as column temperature increased. The experimental results indicated that this new method may solve some separation difficulties in traditional water-based HILIC, which also successfully verify the feasibility of DESs as mobile phase modifiers. PMID:26753279

  14. Analytical approach to determining human biogenic amines and their metabolites using eVol microextraction in packed syringe coupled to liquid chromatography mass spectrometry method with hydrophilic interaction chromatography column.

    PubMed

    Konieczna, Lucyna; Roszkowska, Anna; Synakiewicz, Anna; Stachowicz-Stencel, Teresa; Adamkiewicz-Drożyńska, Elżbieta; Bączek, Tomasz

    2016-04-01

    Analysis of biogenic amines (BAs) in different human samples provides insight into the mechanisms of various biological processes, including pathological conditions, and thus may be very important in diagnosing and monitoring several neurological disorders and cancerous tumors. In this work, we developed a simple and fast procedure using a digitally controlled microextraction in packed syringe (MEPS) coupled to liquid chromatography mass spectrometry (LC-MS) method for simultaneous determination of biogenic amines, their precursors and metabolites in human plasma and urine samples. The separation of 12 low molecular weight and hydrophilic molecules with a wide range of polarities was achieved with hydrophilic interaction chromatography (HILIC) column without derivatization step in 12 min. MEPS was implemented using the APS sorbent in semi-automated analytical syringe (eVol(®)) and small volume of urine and plasma samples, 5 0µL and 100 μL, respectively. We evaluated important parameters influencing MEPS efficiency, including stationary phase selection, sample pH and volume, number of extraction cycles, and washing and elution volumes. In optimized MEPS conditions, the analytes were eluted by 3 × 50 μL of methanol with 0.1% formic acid. The chromatographic separation of analytes was performed on XBridge Amide™ BEH analytical column (3.0mm × 100 mm, 3.5 µm) using gradient elution with mobile phase consisting of phase A: 10mM ammonium formate buffer in water pH 3.0 and phase B: 10mM ammonium formate buffer in acetonitrile pH 3.0. The LC-HILIC-MS method was validated and, in optimum conditions, presented good linearity in concentration range within 10-2000 ng/mL for all the analytes with a determination coefficient (r(2)) higher than 0.999 for plasma and urine samples. Method recovery ranged within 87.6-104.3% for plasma samples and 84.2-98.6% for urine samples. The developed method utilizing polar APS sorbent along with polar HILIC column was applied for

  15. Ion-exchange and hydrophobic interactions affecting selectivity for neutral and charged solutes on three structurally similar agglomerated ion-exchange and mixed-mode stationary phases.

    PubMed

    Kazarian, Artaches A; Taylor, Mark R; Haddad, Paul R; Nesterenko, Pavel N; Paull, Brett

    2013-11-25

    The nature and extent of mixed-mode retention mechanisms evident for three structurally related, agglomerated, particle-based stationary phases were evaluated. These three agglomerated phases were Thermo Fisher ScientificIon PacAS11-HC - strong anion exchange, Thermo Fisher Scientific IonPac CS10--strong cation-exchange PS-DVB, and the Thermo Fisher Scientific Acclaim Trinity P1silica-based substrate, which is commercially marketed as a mixed-mode stationary phase. All studied phases can exhibit zwitterionic and hydrophobic properties, which contribute to the retention of charged organic analytes. A systematic approach was devised to investigate the relative ion-exchange capacities and hydrophobicities for each of the three phases, together with the effect of eluent pH upon selectivity, using a specifically selected range of anionic, cationic and neutral aromatic compounds. Investigation of the strong anion-exchange column and the Trinity P1 mixed-mode substrate, in relation to ion-exchange capacity and pH effects, demonstrated similar retention behaviour for both the anionic and ampholytic solutes, as expected from the structurally related phases. Further evaluation revealed that the ion-exchange selectivity of the mixed-mode phase exhibited properties similar to that of the strong anion-exchange column, with secondary cation-exchange selectivity, albeit with medium to high anion-exchange and cation-exchange capacities, allowing selective retention for each of the anionic, cationic and ampholytic solutes. Observed mixed-mode retention upon the examined phases was found to be a sum of anion- and cation-exchange interactions, secondary ion-exchange and hydrophobic interactions, with possible additional hydrogen bonding. Hydrophobic evaluation of the three phases revealed logP values of 0.38-0.48, suggesting low to medium hydrophobicity. These stationary phases were also benchmarked against traditional reversed-phase substrates namely, octadecylsilica YMC-Pac Pro C18

  16. Analysis of space-time non-stationary patterns of rainfall-groundwater interactions by integrating empirical orthogonal function and cross wavelet transform methods

    NASA Astrophysics Data System (ADS)

    Yu, Hwa-Lung; Lin, Yuan-Chien

    2015-06-01

    Rainfall-groundwater interactions are complex and can depend upon a variety of hydro-geological and geographical conditions and can commonly present high nonlinearity. This study proposed an integration of cross wavelet transform and empirical orthogonal function (EOF) analysis to analyze the space-time nonlinear relationships between the precipitation and groundwater changes. The proposed method was applied to investigating the non-stationary and nonlinear precipitation-groundwater relationships in Pingtung Plain aquifer during 2005-2010. We analyzed daily groundwater and rainfall observations at 47 wells and 27 stations across the study area operated by Taiwan Water Resources Agency and Central Weather Bureau respectively. Results show that EOF method revealed three major space-time patterns of the groundwater levels. The cross wavelet transform further identified the lagged effects between precipitation and groundwater changes. The temporal lags can vary not only with respect to the temporal scales of groundwater processes, but also the different identified groundwater regions, which can associate with distinct physical mechanisms. In general, the lagged periods between high frequency signals of precipitation and groundwater can range from 0 to 18 days, showing how rainfall events perturbed the groundwater levels across space. The long-term lags for the three identified underlying groundwater processes were 3.71, 56.6, and 72.07 days, respectively, showing seasonal recharge patterns can depend upon the geographical and geological conditions. Our results distinguished the space-time recharge processes in different frequencies, which present nonlinear and non-stationary rainfall-recharge interactions of a groundwater system, which can be spatially and frequency dependent.

  17. Efficient enrichment of glycopeptides using metal-organic frameworks by hydrophilic interaction chromatography.

    PubMed

    Ji, Yongsheng; Xiong, Zhichao; Huang, Guang; Liu, Jing; Zhang, Zhang; Liu, Zheyi; Ou, Junjie; Ye, Mingliang; Zou, Hanfa

    2014-10-01

    Selective enrichment of glycopeptides from complicated biological samples is critical for glycoproteomics to obtain the structure and glycosylation information of glycoproteins using mass spectrometry (MS), which still remains a great challenge. Hydrophilic interaction chromatography (HILIC)-based strategies have been proposed for selective isolation of glycopeptides via the interactions between the glycan of glycopeptides and the matrices. However, the application of these methods is limited by the medium selectivity of HILIC matrices. In this study, hydrophilic metal-organic frameworks (MOFs) were fabricated and used as a HILIC matrix. The cross-linked CD-MOFs (LCD-MOFs) were facilely prepared with γ-cyclodextrin as ligand and possessed nano-sized cubic structure, superior hydrophilicity, and bio-compatibility. The LCD-MOFs performance for the selective enrichment of glycopeptides from the complex biological samples were investigated with a digested mixture of human immunoglobulin G (IgG) that was used as standard samples. In the selectivity assessment, the non-glycopeptides causing ion suppression to the glycopeptides were effectively removed, the signal of glycopeptides were enhanced significantly by LCD-MOFs, and twenty glycopeptides were identified with 67 fmol of IgG digest. In addition, the resulting LCD-MOFs demonstrated the lower detection limit (3.3 fmol) with a satisfactory recovery yield (84-103%) for glycopeptide enrichment from a digest of IgG. Furthermore, a promising protocol was developed for the selective enrichment of glycopeptides from mouse liver, and 344 unique N-glycosylation sites that mapped to 290 different glycoproteins were identified in a single MS run. The results clearly demonstrated that when used in a HILIC matrix, LCD-MOFs have great potential for identifying and enriching low-abundant glycopeptides in complex biological samples. PMID:25110774

  18. Analysis of drug-protein interactions by high-performance affinity chromatography: interactions of sulfonylurea drugs with normal and glycated human serum albumin.

    PubMed

    Matsuda, Ryan; Anguizola, Jeanethe; Hoy, Krina S; Hage, David S

    2015-01-01

    High-performance affinity chromatography (HPAC) is a type of liquid chromatography that has seen growing use as a tool for the study of drug-protein interactions. This report describes how HPAC can be used to provide information on the number of binding sites, equilibrium constants, and changes in binding that can occur during drug-protein interactions. This approach will be illustrated through recent data that have been obtained by HPAC for the binding of sulfonylurea drugs and other solutes to the protein human serum albumin (HSA), and especially to forms of this protein that have been modified by non-enzymatic glycation. The theory and use of both frontal analysis and zonal elution competition studies in such work will be discussed. Various practical aspects of these experiments will be presented, as well as factors to consider in the extension of these methods to other drugs and proteins or additional types of biological interactions. PMID:25749961

  19. Drug-to-antibody ratio (DAR) and drug load distribution by hydrophobic interaction chromatography and reversed phase high-performance liquid chromatography.

    PubMed

    Ouyang, Jun

    2013-01-01

    Hydrophobic interaction chromatography (HIC) is the method of choice for determination of the drug-to-antibody ratio (DAR) and drug load distribution for cysteine (Cys)-linked antibody-drug conjugates (ADCs). The drug-loaded species are resolved based on the increasing hydrophobicity with the least hydrophobic, unconjugated form eluting first and the most hydrophobic, 8-drug form eluting last. The area percentage of a peak represents the relative distribution of the particular drug-loaded ADC species. The weighted average DAR is then calculated using the percentage peak area information and the drug load numbers. Reversed phase high-performance liquid chromatography (RP-HPLC) offers an orthogonal method to obtain DAR for Cys-linked ADCs. The method involves, first, a reduction reaction to completely dissociate the heavy and light chains of the ADC, then separation of the light and heavy chains and their corresponding drug-loaded forms on an RP column. The percentage peak area from integration of the light chain and heavy chain peaks, combined with the assigned drug load for each peak, is used to calculate the weighted average DAR. PMID:23913154

  20. Rapid determination of glyphosate, glufosinate, bialaphos, and their major metabolites in serum by liquid chromatography-tandem mass spectrometry using hydrophilic interaction chromatography.

    PubMed

    Yoshioka, Naoki; Asano, Migiwa; Kuse, Azumi; Mitsuhashi, Takao; Nagasaki, Yasushi; Ueno, Yasuhiro

    2011-06-10

    We developed a simple and rapid method for the simultaneous determination of phosphorus-containing amino acid herbicides (glyphosate, glufosinate, bialaphos) and their major metabolites, aminomethylphosphonic acid (AMPA) and 3-methylphosphinicopropionic acid (MPPA), in human serum. Serum samples were filtrated through an ultrafiltration membrane to remove proteins. The filtrate was then washed with chloroform, and injected into a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. Chromatographic separation was achieved on a hydrophilic interaction chromatography (HILIC) column. Determination of the target herbicides and metabolites was successfully carried out without derivatization or solid phase extraction (SPE) cartridge clean-up. The recoveries of these compounds, added to human serum at 0.2μg/mL, ranged from 94% to 108%, and the relative standard deviations (RSDs) were within 5.9%. The limits of detection (LODs) were 0.01μg/mL for MPPA, 0.02μg/mL for AMPA, 0.03μg/mL for both glyphosate and glufosinate, and 0.07μg/mL for bialaphos, respectively. PMID:21530973

  1. Isolation and Identification of Saponins from the Natural Pasturage Asterothamnus centrali-asiaticus Employing Preparative Two-Dimensional Reversed-Phase Liquid Chromatography/Hydrophilic Interaction Chromatography.

    PubMed

    Wang, Yan-Ming; Zhao, Jian-Qiang; Yang, Jun-Li; Tao, Yan-Duo; Mei, Li-Juan; Shi, Yan-Ping

    2016-06-22

    Asterothamnus centrali-asiaticus, a kind of characteristic shrub abundant in grassland and desert areas, has been used as forage fodder for camels and goats in Central Asia, and this plant also plays a critical role in the maintenance of desert grassland ecosystems as a result of its tolerance to poor soils and sand burial. However, its chemical composition has been rarely reported. In this study, phytochemical investigation of this pasturage was performed and three new triterpenoid saponins (1-3) were isolated together with nine known triterpenoid saponins (4-12) using preparative two-dimensional reversed-phase liquid chromatography/hydrophilic interaction chromatography (2D RPLC/HILIC). Their structures were elucidated via diverse spectroscopic analyses, including infrared (IR) spectrometry, high-resolution electrospray ionization mass spectrometry (HR-ESIMS), and one-dimensional (1D) and 2D nuclear magnetic resonance (NMR). All isolated triterpenoid saponins (1-12) were reported from this genus for the first time, and they were further evaluated for their cytotoxicity against four cancer cell lines (A549, HepG2, MGC-803, and MFC), which indicated that compound 11 showed potent cytotoxicity against the HepG2 cell line, with an IC50 value of 6.85 μg/mL. PMID:27231806

  2. [Preparation and chromatographic performance of a silica-bonded (4-cyclopentadienyl benzoic acid-iron-toluene) hexafluorophosphoric acid stationary phase].

    PubMed

    Cao, Aijuan; Li, Xiaole; Qiao, Lijun; Zhou, Xiaohua; Yu, Ajuan; Zhang, Shusheng; Wu, Yangjie

    2016-02-01

    Based on the unique molecular structure of ferrocene and its potential as a new liquid chromatography separation medium, a new silica-bonded (4-cyclopentadienyl benzoic acid-iron-toluene) hexafluorophosphoric acid stationary phase was prepared. The structure of this new material was characterized by infrared spectroscopy, elemental analysis, thermogravimetric analysis et al. The chromatographic performance and retention mechanism of this new stationary phase were evaluated using different solute probes, including polycyclic aromatic hydrocarbons (PAHs), positional isomers of naphthylamine, positional isomers of nitro-aniline, nitroimidazoles, organic phosphorus et al. It could provide various action sites for different solutes in normal-phase chromatography such as π electron transfer, π-π electron interactions, dipole-dipole interactions, and electrostatic interactions with the substrates. And the possible separation mechanisms are discussed. PMID:27382719

  3. Immobilized metal ion affinity chromatography.

    PubMed

    Yip, T T; Hutchens, T W

    1992-01-01

    Immobilized metal ion affinity chromatography (IMAC) (1,2) is also referred to as metal chelate chromatography, metal ion interaction chromatography, and ligand-exchange chromatography. We view this affinity separation technique as an intermediate between highly specific, high-affinity bioaffinity separation methods, and wider spectrum, low-specificity adsorption methods, such as ion exchange. The IMAC stationary phases are designed to chelate certain metal ions that have selectivity for specific groups (e.g., His residues) in peptides (e.g., 3-7) and on protein surfaces (8-13). The number of stationary phases that can be synthesized for efficient chelation of metal ions is unlimited, but the critical consideration is that there must be enough exposure of the metal ion to interact with the proteins, preferably in a biospecific manner. Several examples are presented in Fig. 1. The challenge to produce new immobilized chelating groups, including protein surface metal-binding domains (14,15) is being explored continuously. Table 1 presents a list of published procedures for the synthesis and use of stationary phases with immobilized chelating groups. This is by no means exhaustive, and is intended only to give an idea of the scope and versatility of IMAC. Fig. 1 Schematic illustration of several types of immobilized metal-chelating groups, including, iminodiacetate (IDA), tris(carboxymethyl) ethylenediamine (TED), and the metal-binding peptides (GHHPH)(n)G (where n = 1,2,3, and 5) (14,15). Table 1 Immobilized Chelating Groups and Metal Ions Used for Immobilized Metal Ion Affinity Chromatography Chelating group Suitable metal ions Reference Commercial source Immodiacetate Transitional1,2 Pharmacia LKB Pierce Sigma Boehringer Mannheim TosoHaas 2-Hydroxy-3[N-(2- pyrtdylmethyl) glycme]propyl Transitional3 Not available ?-Alky1 mtrilo triacetic acid Transitional4 Not available Carboxymethylated asparhc acid Ca(II)13 Not available Tris (carboxy- methyl) ethylene Diamme

  4. Concerning the interaction of non-stationary cross-flow vortices in a three-dimensional boundary layer

    NASA Technical Reports Server (NTRS)

    Bassom, Andrew P.; Hall, Philip

    1990-01-01

    Recently there has been much work devoted to considering some of the many and varied interaction mechanisms which may be operative in three-dimensional boundary layer flows. This paper is concerned with resonant triads of crossflow vortices. The effects of interactions upon resonant triads is examined where each member of the triad has the property of being linearly neutrally stable so that the importance of the interplay between modes can be relatively easily assessed. Modes within the boundary layer flow above a rotating disc are investigated because of the similarity between this disc flow and many important practical flows and, secondly, because the selected flow is an exact solution of the Navier-Stokes equations which makes its theoretical analysis especially attractive. It is demonstrated that the desired triads of linearly neutrally stable modes can exist within the chosen boundary layer flow. Evolution equations are obtained to describe the development of the amplitudes of these modes once the interaction mechanism is accounted for. It is found that the coefficients of the interaction terms within the evolution equations are, in general, given by quite intricate expressions although some elementary numerical work shows that the evaluation of these coefficients is practicable. The basis of the work lends itself to generalization to more complicated boundary layers, and effects of detuning or non-parallelism could be provided for within the asymptotic framework.

  5. [Preparation and evaluation of N-acryloyltris (hydroxymethyl) aminomethane-bonded chromatographic stationary phase].

    PubMed

    Cheng, Xiaodong; Feng, Yuqi

    2015-09-01

    The present study described the preparation of N-acryloyltris (hydroxymethyl) aminomethane-bonded silica (NAS) stationary phase based on "thiol-ene" click chemistry. The composition of the surface grafts of NAS stationary phase was determined by elemental analysis and the results demonstrated the successful introduction of the N-acryloyltris (hydroxymethyl) aminomethane groups to the silica surface. Similar elemental composition of three batches of the NAS stationary phases exhibited good reproducibility of the preparation strategy. A set of standard compounds were employed to investigate the retention mechanism of the NAS stationary phase by three different empirical equations. The results indicated the retention of the tested analytes on the NAS stationary phase was based more on a mixed-mechanism rather than a simple partitioning or adsorption process. Eight compounds were selected to study the hydrophobic and hydrophilic properties of the NAS stationary phase in mobile phase with different ACN contents. Due to its hydrophilic triolacrylamide groups and short hydrophobic alkyl chains, the NAS phase was successfully applied in both reversed-phase liquid chromatography (RPLC) mode and hydrophilic interaction liquid chromatography (HILIC) mode. The influence of flow rate on the column efficiency was compared in these two modes. In contrast to RPLC columns, the overall heights equivalent to a theoretical plate (HETP) in HILIC is weakly dependent on the retention of the analyte and the HETP curve is much flatter in RPLC than in HILIC at larger reduced velocities. Furthermore, the separation of alkylbenzenes, nucleosides and nucleobases, water-soluble vitamins was achieved on the new stationary phase, demonstrating the excellent application potential. PMID:26753276

  6. Entropic interaction chromatography: separating proteins on the basis of size using end-grafted polymer brushes.

    PubMed

    Pang, Peter; Koska, Jürgen; Coad, Bryan R; Brooks, Donald E; Haynes, Charles A

    2005-04-01

    Partitioning of a macromolecule into the interfacial volume occupied by a grafted polymer brush decreases the configurational entropy (DeltaSbrush(c)) of the terminally attached linear polymer chains due to a loss of free volume. Self-consistent field theory (SCF) calculations are used to show that DeltaSbrush(c) is a strong function of both the size (MWp) of the partitioning macromolecule and the depth of penetration into the brush volume. We further demonstrate that the strong dependence of DeltaSbrush(c) on MWp provides a novel and powerful platform, which we call entropic interaction chromatography (EIC), for efficiently separating mixtures of proteins on the basis of size. Two EIC columns, differing primarily in polymer grafting density, were prepared by growing a brush of poly(methoxyethyl acrylamide) chains on the surface of a wide-pore (1,000-A pores, 64-microm diameter rigid beads) resin (Toyopearl AF-650M) bearing surface aldehyde groups. Semipreparative 0.1-L columns packed with either EIC resin provide reduced-plate heights of 2 or less for efficient separation of globular protein mixtures over at least three molecular-weight decades. Protein partitioning within these wide-pore EIC columns is shown to be effectively modeled as a thermodynamically controlled process, allowing partition coefficients (K(P)) and elution chromatograms to be accurately predicted using a column model that combines SCF calculation of K(P) values with an equilibrium-dispersion type model of solute transport through the column. This model is used to explore the dependence of column separation efficiency on brush properties, predicting that optimal separation of proteins over a broad MWp range is achieved at low to moderate grafting densities and intermediate chain lengths. PMID:15706591

  7. Hydrophilic interaction liquid chromatography for the separation, purification, and quantification of raffinose family oligosaccharides from Lycopus lucidus Turcz.

    PubMed

    Liang, Tu; Fu, Qing; Li, Fangbing; Zhou, Wei; Xin, Huaxia; Wang, Hui; Jin, Yu; Liang, Xinmiao

    2015-08-01

    A systematic strategy based on hydrophilic interaction liquid chromatography was developed for the separation, purification and quantification of raffinose family oligosaccharides from Lycopus lucidus Turcz. Methods with enough hydrophilicity and selectivity were utilized to resolve the problems encountered in the separation of oligosaccharides such as low retention, low resolution and poor solubility. The raffinose family oligosaccharides in L. lucidus Turcz. were isolated using solid-phase extraction followed by hydrophilic interaction liquid chromatography at semi-preparative scale to obtain standards of stachyose, verbascose and ajugose. Utilizing the obtained oligosaccharides as standards, a quantitative determination method was developed, validated and applied for the content determination of raffinose family oligosaccharides both in the aerial and root parts of L. lucidus Turcz. There were no oligosaccharides in the aerial parts, while in the root parts, the total content was 686.5 mg/g with the average distribution: raffinose 66.5 mg/g, stachyose 289.0 mg/g, verbascose 212.4 mg/g, and ajugose 118.6 mg/g. The result provided the potential of roots of L. lucidus Turcz. as new raffinose family oligosaccharides sources for functional food. Moreover, since the present systematic strategy is efficient, sensitive and robust, separation, purification and quantification of oligosaccharides by hydrophilic interaction liquid chromatography seems to be possible. PMID:26011699

  8. Extracting stationary segments from non-stationary synthetic and cardiac signals

    NASA Astrophysics Data System (ADS)

    Rodríguez, María. G.; Ledezma, Carlos A.; Perpiñán, Gilberto; Wong, Sara; Altuve, Miguel

    2015-01-01

    Physiological signals are commonly the result of complex interactions between systems and organs, these interactions lead to signals that exhibit a non-stationary behaviour. For cardiac signals, non-stationary heart rate variability (HRV) may produce misinterpretations. A previous work proposed to divide a non-stationary signal into stationary segments by looking for changes in the signal's properties related to changes in the mean of the signal. In this paper, we extract stationary segments from non-stationary synthetic and cardiac signals. For synthetic signals with different signal-to-noise ratio levels, we detect the beginning and end of the stationary segments and the result is compared to the known values of the occurrence of these events. For cardiac signals, RR interval (cardiac cycle length) time series, obtained from electrocardiographic records during stress tests for two populations (diabetic patients with cardiovascular autonomic neuropathy and control subjects), were divided into stationary segments. Results on synthetic signals reveal that the non-stationary sequence is divided into more stationary segments than needed. Additionally, due to HRV reduction and exercise intolerance reported on diabetic cardiovascular autonomic neuropathy patients, non-stationary RR interval sequences from these subjects can be divided into longer stationary segments compared to the control group.

  9. Optimization of hydrophobic interaction chromatography using a mathematical model of elution curves of a protein mixture.

    PubMed

    Lienqueo, María Elena; Shene, Carolina; Asenjo, Juan

    2009-01-01

    This paper describes a methodology for optimizing performance of hydrophobic interaction chromatography (HIC) for protein mixtures in which Rate Model simulations and evaluation of cost function are used. The system under study was HIC of a two-protein mixture (alpha-chymotrypsin and alpha-amylase), carried out with different conditions (gradient steepness, salt, concentration, volume of sample, pH, type of matrix, and flow rates). Parameters in the rate model were obtained from different sources. Mass transfer parameters (Reynolds, Peclet, and Biot numbers) were calculated using empirical correlations. Under the experimental conditions Re number was small (0.23) and axial dispersion negligible (PeL>300). Mass transfer was controlled by intraparticle diffusion (Bi>50). The model assumes that equilibrium constant (b) in the Langmuir isotherm was salt concentration (I) dependent [b(I)]. Parameters in the relationship for b(I) were estimated from experimental single protein elution curves and used to simulate protein mixtures. Rate model simulations showed that if protein sample load to the column was below 1 mg, displacement effects were negligible; in other cases protein interactions would limit the proposed mathematical description of HIC. The calibrated Rate Model successfully predicted elution curves of the protein mixture with deviations lower than 6x10(-4) absorbance units. The model was also able to predict that the Butyl Sepharose--NaCl 4 M system allowed to obtain the highest resolution (>1) for the two-protein mixture evaluated. The cost function built for optimizing the performance of HIC considers yield, purity, concentration, and the time needed to accomplish the separation. This function contains two types of parameters that have to be determined. The ones dependent on the HIC system and process conditions were obtained from simulations of elution curves of the two-protein mixture, by the calibrated Rate Model. The other parameters are dependent on

  10. Temperature-induced inversion of the elution order of enantiomers in gas chromatography: N-ethoxycarbonyl propylamides and N-trifluoroacetyl ethyl esters of alpha-amino acids on Chirasil-Val-C11 and Chirasil-Dex stationary phases.

    PubMed

    Levkin, Pavel A; Levkina, Anna; Czesla, Harri; Schurig, Volker

    2007-06-15

    Inversion of the elution order of enantiomers caused by enthalpy-entropy compensation at the isoenantioselective temperature (Tiso) was experimentally observed by gas chromatography on the diamide-type chiral stationary phase (CSP), Chirasil-L-Val-C11, with N-ethoxycarbonyl propylamide (ECPA) derivatives of a number of alpha-amino acids. For the first time, a clear visual representation of the increase of the apparent enantioseparation factor alpha app from 1.00 to 1.08 as the temperature is raised from 120 to 170 degrees C is presented. The increase of alpha app is accompanied by a concomitant reduction of the retention factors of the enantiomers. The Tiso values were in the range from 110 to 130 degrees C depending on the nature of the alpha-amino acid. On the contrary, the Tiso values of the N(O)-trifluoroacetyl ethyl ester derivatives (TFA-Et) of the same alpha-amino acids were approximately 80 degrees higher than that of ECPA derivatives. The comprehensive thermodynamic investigation of the enantioseparation of ECPA and TFA-Et derivatives of valine and alanine using the retention increment method showed that the Delta L,D(DeltaH) difference between the diastereomeric selector-selectand associates was almost the same for ECPA and TFA-Et derivatives despite a much stronger bonded selector-selectand association taking place for the ECPA derivatives. On the other hand, the Delta L,D(DeltaS) values were found to be more negative in the case of ECPA derivatives, resulting in the unusually low values of Tiso. A temperature-dependent inversion of the elution order of enantiomers was also observed on the cyclodextrin-type CSP, Chirasil-Dex, with TFA-Et derivatives of several alpha-amino acids. The Tiso values were in the range from 20 to 170 degrees C depending on the nature of the alpha-amino acid. The results obtained demonstrate the necessity to conduct temperature-dependent studies in order to optimize the enantiomeric separation of single racemates isothermally or

  11. System constants for the bis(cyanopropylsiloxane)-co-methylsilarylene HP-88 and poly(siloxane) Rtx-440 stationary phases.

    PubMed

    Kiridena, Waruna; Patchett, Cheryl C; Koziol, Wladyslaw W; Poole, Colin F

    2005-07-22

    The solvation parameter model is used to characterize the retention properties of the bis(cyanopropylsiloxane)-co-methylsilarylene, HP-88, and poly(siloxane), Rtx-440, stationary phases over the temperature range 60-140 degrees C. HP-88 is among the most cohesive, dipolar/polarizable and hydrogen-bond basic of stationary phases for open-tubular column gas chromatography. It has no hydrogen-bond acidity or capacity for electron lone pair interactions. It exhibits similar selectivity to the poly(cyanopropylsiloxane) stationary phase SP-2340. Rtx-440 is a low-polarity, low-cohesion stationary phase with a moderate capacity for dipolar/polarizable and hydrogen-bond base interactions. It has no hydrogen-bond acidity and possesses weak electron lone pair interactions. It has unique selectivity when compared against a system constants database for 28 common stationary phase compositions. Cluster analysis indicated that the poly(cyanopropylphenyldimethylsiloxane) stationary phase containing 6% cyanopropylphenylsiloxane monomer, DB-1301, the poly(dimethyldiphenylsiloxane) stationary phase containing 20% diphenylsiloxane monomer, Rtx-20, the poly(siloxane) stationary phase of unknown composition, DB-624, and DX-1 [a mixture of poly(dimethylsiloxane) and poly(ethylene glycol) 9:1] are the closest selectivity matches in the database. The selectivity of DB-1301 and Rtx-440 are very similar for solutes with weak hydrogen-bond acidity allowing one stationary phase to be substituted for the other with likely success. For strong hydrogen-bond acids, such as phenols, DB-1301 and Rtx-440 exhibit different selectivity. PMID:16038217

  12. Isolation of fluorescent constituents from soil humic and fulvic acids by hydrophilic interaction chromatography

    NASA Astrophysics Data System (ADS)

    Aoyama, Masakazu

    2014-05-01

    Humic acids (HAs) and fulvic acids (FAs) are the most abundant components of soil organic matter and exhibit fluorescence. Our previous studies using high performance size-exclusion chromatography (HPSEC) and polyacrylamide gel electrophoresis demonstrated that the fluorescence of soil HAs was mainly due to the minor constituents with relatively small molecular sizes. In order to clarify the nature of the fluorescence of soil organic matter, it is necessary to isolate the fluorescent constituents from HAs and FAs. I succeeded in isolating the fluorescent constituents from soil HAs and FAs by using hydrophilic interaction chromatography (HILIC). When HILIC of soil HAs and FAs was carried out under isocratic conditions using a SeQuant ZIC-HILIC column and acetonitrile-water as a mobile phase, the complete separation of fluorescent and non-fluorescent peaks was achieved at the acetonitrile concentration of 90%. Another fluorescent peak was eluted with decreasing concentration of acetonitrile from 90% to 50%. The use of a TSKgel Amide-80 column gave the same results. The best resolution was obtained when HILIC was performed under gradient conditions from 90% to 50% acetonitrile using the ZIC-HILIC and Amide-80 columns linked in series. For both HAs and FAs, a sharp non-fluorescent peak (peak A) followed by a sharp fluorescent peak (peak B) and a broad fluorescent peak (peak C) were eluted under the above optimum operating conditions. The intensity of peak A relative to that of peak B was significantly less in the FAs than in the HAs. The fluorescent peaks (peaks B and C) of the FAs showed considerable UV absorption, whereas those of the HAs did little UV absorption. When the fluorescence emission spectra (excitation at 280 nm) were measured for the fluorescent peaks, two emission peaks were located at 460 and 520 nm for the HAs, while for the FAs, a broad emission peak at 400-450 nm with a small shoulder at around 500 nm was observed. The peaks were collected

  13. Estimation of interaction between oriented immobilized green fluorescent protein and its antibody by high performance affinity chromatography and molecular docking.

    PubMed

    Li, Qian; Wang, Jing; Yang, Lingjian; Gao, Xiaokang; Chen, Hongwei; Zhao, Xinfeng; Bian, Liujiao; Zheng, Xiaohui

    2015-07-01

    Although green fluorescence protein (GFP) and its antibody are widely used to track a protein or a cell in life sciences, the binding behavior between them remains unclear. In this work, diazo coupling method that synthesized a new stationary GFP was oriented immobilized on the surface of macro-porous silica gel by a phase. The stationary phase was utilized to confirm the validation of injection amount-dependent analysis in exploring protein-protein interaction that use GFP antibody as a probe. GFP antibody was proved to have one type of binding site on immobilized GFP. The number of binding site and association constant were calculated to be (6.41 ± 0.76) × 10(-10) M and (1.39 ± 0.12) × 10(9) M(-1). Further analysis by molecular docking showed that the binding of GFP to its antibody is mainly driven by hydrogen bonds and salt bridges. These results indicated that injection amount-dependent analysis is capable of exploring the protein-protein interactions with the advantages of ligand and time saving. It is a valuable methodology for the ligands, which are expensive or difficult to obtain. PMID:25727342

  14. Use of complex configuration interaction calculations and the stationary principle for the description of metastable electronic states of HCl-

    NASA Astrophysics Data System (ADS)

    Honigmann, Michael; Liebermann, Heinz-Peter; Buenker, Robert J.

    2010-07-01

    The complex multireference single- and double-excitation configuration interaction method has been employed to compute potential curves for the anion of the hydrogen chloride molecule. First, conventional CI calculations with real basis functions have been carried out to determine the potential curves of both HCl and its anion over a large range of internuclear distance. It is shown that adding basis functions with very small exponents leads to sharply avoided crossings for the HCl- potentials that greatly complicate the search for resonance states thought to be responsible for features observed in electron collision experiments. By limiting the number of such diffuse-type functions it is possible to describe resonance states at a highly correlated level and still account for their interaction with the continuum in which they are embedded. In the present study of the HCl- anion the complex basis function technique of Moiseyev-Corcoran and McCurdy-Resigno is employed to calculate the energy positions and line-widths of the resonance states. Two states of Σ2+ symmetry are calculated which have potentials that have significantly different shapes than that of the neutral ground state and thus contribute to the cross section for vibrational excitation of the neutral HCl molecule induced by low-energy electron collisions. The lower of these (1 Σ2+) correlates smoothly with the bound anionic ground state at large internuclear distances and is seen to be responsible for the sharp peaks observed in the low-energy region of the spectrum. The upper state (3 Σ2+) has a much larger bond length and is assigned to the broad bands observed with a maximum in the 2.5-3.0 eV range. The present calculations thus stand in contradiction to earlier claims that the above peaks are caused by so-called virtual states without a definite autoionization lifetime.

  15. Large-scale qualitative and quantitative characterization of components in Shenfu injection by integrating hydrophilic interaction chromatography, reversed phase liquid chromatography, and tandem mass spectrometry.

    PubMed

    Song, Yuelin; Zhang, Na; Shi, Shepo; Li, Jun; Zhang, Qian; Zhao, Yunfang; Jiang, Yong; Tu, Pengfei

    2015-08-14

    It is of great importance to clarify in depth the chemical composition, including qualitative and quantitative aspects, of traditional Chinese medicine (TCM) injection that contains a great number of hydrophilic and hydrophobic ingredients to guarantee its safe medication in clinic. Column-switching hydrophilic interaction liquid chromatography-reversed phase liquid chromatography coupled with tandem mass spectrometry (HILIC-RPLC-MS/MS) has been revealed to be advantageous at simultaneous measurement of compounds covering a broad polarity range. Previous studies have profiled the hydrophobic components, mainly aconite alkaloids and ginsenosides, in Shenfu Injection (SFI); however, the hydrophilic substances haven't been taken into account. In the present study, we aim to holistically characterize the hydrophilic constituents and to simultaneously quantitate both hydrophilic and hydrophobic components in SFI. A strategy integrating predefined multiple reaction monitoring, step-wise multiple ion monitoring, and enhanced product ion scans was proposed to universally screen the hydrophilic substances using a hybrid triple quadrupole-linear ion trap mass spectrometer. Structural identification was carried out by comparing with authentic compounds, analyzing MS(2) spectra, and referring to accessible databases (e.g., MassBank, METLIN and HMDB). A total of 157 hydrophilic compounds were detected from SFI, and 154 ones were identified as amino acids, nucleosides, organic acid, carbohydrates, etc. A column-switching HILIC-RPLC-MS/MS system was developed and validated for simultaneously quantitative analysis of 40 primary hydrophilic and hydrophobic ingredients in SFI, including eleven amino acids, nine nucleosides, nine aconite alkaloids, and eleven ginsenosides. Taken together, the findings obtained could provide meaningful information for comprehensively understanding the chemical composition and offer a reliable approach for the quality control of SFI. PMID:26143607

  16. [Determination of melamine and ammeline in eggs and meat using hydrophilic interaction liquid chromatography].

    PubMed

    Li, Yanzhao; Hao, Weiqiang; Wang, Yubo; Chen, Qiang; Li, Jinchun; Sun, Xiaoli

    2012-07-01

    A hydrophilic interaction liquid chromatographic (HILIC) method for the determination of melamine and its degradation product ammeline in eggs and meat has been developed. The separation was carried out on a ZIC-HILIC column with 3 mmol/L NH4H2PO4 (pH 6.9)-acetonitrile (20: 80, v/v) as mobile phase at the flow rate of 0.8 mL/min, and detected at 220 nm. Compared with the reversed-phase liquid chromatography, this method can avoid the use of ion pair reagents and thus simplify the composition of mobile phase. Under the above chromatographic conditions, melamine and ammeline had good peak shapes and moderate retention times. Good separation between these compounds and the substances that were naturally contained in the samples can be achieved. For the sample preparation, the analytes were first extracted with 0.1% phosphoric acid due to the basicity of melamine and ammeline. Then, metaphosphoric acid and acetonitrile were used to remove proteins and saccharides by precipitation. After the filtration and removal of acetonitrile by rotary evaporation under vacuum, the filtrate was cleaned-up by solid-phase extraction (SPE) technique in which a cation exchange column was used. The SPE column was activated by using methanol and 0.1% phosphoric acid. A solution of 5% ammonia methanol was chosen as eluent. The residues obtained from the eluant by evaporating the solvent were resolved in the mobile phase. It was found that there was a good linear relationship between concentration and detector response within the range of 0.4-40 mg/L. The limits of detection were 2 mg/kg for both melamine and ammeline. The average recoveries were between 80% and 105% in the spiked range of 2-10 mg/kg. The relative standard deviations were not more than 10%. The solutions of melamine and ammeline were stable in a month. The established method can be used in practice to determine melamine and ammeline simultaneously in egg and meat samples. PMID:23189668

  17. Mixed-mode ion-exchangers and their comparative chromatographic characterization in reversed-phase and hydrophilic interaction chromatography elution modes.

    PubMed

    Lämmerhofer, Michael; Richter, Martin; Wu, Junyan; Nogueira, Raquel; Bicker, Wolfgang; Lindner, Wolfgang

    2008-08-01

    A set of particulate silica-supported mixed-mode RP/weak anion-exchangers (RP/WAX) (obtained by bonding of N-undecenoylated 3-aminoquinuclidine, 3-aminotropane and 2-dimethylaminoethylamine as well as of N-butenoyl-(2S,4S,5R)-2-aminomethyl-5-[(2-octylthio)ethyl]-quinuclidine to thiol-modified silica) were chromatographically characterized in comparison to selected commercially available columns using two distinct isocratic elution modes, viz. an aqueous-rich RP-type elution mode (with 40% ACN and 60% buffer) as well as an organic solvent-rich hydrophilic interaction chromatography (HILIC)-type elution mode (95 and 90% ACN). The mixed-mode RP/WAX phases showed multimodal applicability, unlike a polar embedded RP material (Synergi Fusion RP), amino phases (Luna NH(2), BioBasic AX) or typical HILIC packings (ZIC-HILIC, TSKGel Amide-80). Principal component analysis (PCA) of the RP test data confirmed that the in-house developed RP/WAX columns as well as the Acclaim Mixed-Mode WAX-1 phase resemble each other in their chromatographic characteristics having slightly lower hydrophobic selectivity (alpha(CH2) of 1.5) than the tested Synergi Fusion RP (alpha(CH2) approximately 1.8). In contrast, a decrease in mixed-mode character due to lowered ion-exchange capacity and concomitantly increased RP-like behavior could be identified for other mixed-mode phases in the order of Obelisc R > Primesep B2 > Uptisphere MM3. PCA on HILIC data revealed that the RP/WAX phases behave dissimilar to TSKGel Amide-80, ZIC-HILIC and polysulfoethyl A under the chosen elution conditions. Hence, they may be regarded as complementary to these commercial stationary phases with applicability profiles for hydrophilic but also hydrophobic solutes. PMID:18693304

  18. Development and validation of a hydrophilic interaction chromatography method coupled with a charged aerosol detector for quantitative analysis of nonchromophoric α-hydroxyamines, organic impurities of metoprolol.

    PubMed

    Xu, Qun; Tan, Shane; Petrova, Katya

    2016-01-25

    The European Pharmacopeia (EP) metoprolol impurities M and N are polar, nonchromophoric α-hydroxyamines, which are poorly retained in a conventional reversed-phase chromatographic system and are invisible for UV detection. Impurities M and N are currently analyzed by TLC methods in the EP as specified impurities and in the United States Pharmacopeia-National Formulary (USP-NF) as unspecified impurities. In order to modernize the USP monographs of metoprolol drug substances and related drug products, a hydrophilic interaction chromatography (HILIC) method coupled with a charged aerosol detector (CAD) was explored for the analysis of the two impurities. A comprehensive column screening that covers a variety of HILIC stationary phases (underivatized silica, amide, diol, amino, zwitterionic, polysuccinimide, cyclodextrin, and mixed-mode) and optimization of HPLC conditions led to the identification of a Halo Penta HILIC column (4.6 × 150 mm, 5 μm) and a mobile phase comprising 85% acetonitrile and 15% ammonium formate buffer (100 mM, pH 3.2). Efficient separations of metoprolol, succinic acid, and EP metoprolol impurities M and N were achieved within a short time frame (<8 min). The HILIC-CAD method was subsequently validated per USP validation guidelines with respect to specificity, robustness, linearity, accuracy, and precision, and could be incorporated into the current USP-NF monographs to replace the outdated TLC methods. Furthermore, the developed method was successfully applied to determine organic impurities in metoprolol drug substance (metoprolol succinate) and drug products (metoprolol tartrate injection and metoprolol succinate extended release tablets). PMID:26580821

  19. An integrated metabolomics workflow for the quantification of sulfur pathway intermediates employing thiol protection with N-ethyl maleimide and hydrophilic interaction liquid chromatography tandem mass spectrometry.

    PubMed

    Ortmayr, Karin; Schwaiger, Michaela; Hann, Stephan; Koellensperger, Gunda

    2015-11-21

    The sulfur metabolic pathway is involved in basic modes of cellular metabolism, including methylation, cell division, respiratory oscillations and stress responses. Hence, the implicated high reactivity of the sulfur pathway intermediates entails challenges for their quantitative analysis. In particular the unwanted oxidation of the thiol group-containing metabolites glutathione, cysteine, homocysteine, γ-glutamyl cysteine and cysteinyl glycine must be prevented in order to obtain accurate snapshots of this important part of cellular metabolism. Suitable analytical methodologies are therefore needed to support studies of drug metabolism and metabolic engineering. In this work, a novel sample preparation strategy targeting thiolic metabolites was established by implementing thiol group protection with N-ethyl maleimide using a cold methanol metabolite extraction procedure. It was shown that N-ethyl maleimide derivatization is compatible with typical metabolite extraction procedures and also allowed for the stabilization of the instable thiolic metabolites in a fully (13)C-labeled yeast cell extract. The stable isotope labeled metabolite analogs could be used for internal standardization to achieve metabolite quantification with high precision. Furthermore, a dedicated hydrophilic interaction liquid chromatography tandem mass spectrometry method for the separation of sulfur metabolic pathway intermediates using a sub-2 μm particle size stationary phase was developed. Coupled with tandem mass spectrometry, the presented methodology proved to be robust, and sensitive (absolute detection limits in the low femtomole range), and allowed for the quantification of cysteine, cysteinyl glycine, cystathionine, cystine, glutamic acid, glutamyl cysteine, reduced glutathione, glutathione disulfide, homocysteine, methionine, S-adenosyl homocysteine and serine in a human ovarian carcinoma cell model. PMID:26451393

  20. Time-resolved Tomo-PIV measurements of the interaction between a stationary held sphere and a turbulent boundary layer.

    NASA Astrophysics Data System (ADS)

    van Hout, Rene; Eisma, Jerke; Overmars, Edwin; Elsinga, Gerrit; Westerweel, Jerry

    2015-11-01

    Time resolved tomographic PIV measurements (acquisition rate 250Hz) were performed in a turbulent boundary layer (TBL) on the side wall of an open channel, water flow facility (cross section 60x60cm, Wx H) , 3.5m downstream of the inlet at a bulk flow velocity of Ub = 0.17m/s (Reb =Ub H / ν = 102x103, δ0 . 99 = 5 . 0 cm, Reθ = 891). The measurement volume was a horizontal slab (6x1.5x6cm3, lx wx h) extending from the side wall, 30cm above the bottom. The Tomo-PIV setup comprised four high-speed ImagerPro HS cameras (2016x2016pixels), a high-speed laser (Nd:YLF, Darwin Duo 80M, Quantronix), optics/prisms and data acquisition/processing software (LaVision, DaVis8.2). A sphere with diameter, D = 6mm (D+ = 51, ``+'' denotes inner wall scaling), was positioned at y = 37.5 and 5.4mm (y+ = 319 and 46) from the wall (measured from the sphere's center). The latter position covers most of the buffer layer while the former is well in the outer layer. Sphere Reynolds numbers based on D and the average streamwise velocity at the sphere's center were 984 (y+ = 319) and 684 (y+ = 46). Results show the interaction between the coherent turbulence structures in the TBL and those generated in the sphere's wake. Total and partial destruction of the log-law layer is observed when the sphere is positioned in the buffer and outer layer, respectively.

  1. An Advanced, Interactive, High-Performance Liquid Chromatography Simulator and Instructor Resources

    ERIC Educational Resources Information Center

    Boswell, Paul G.; Stoll, Dwight R.; Carr, Peter W.; Nagel, Megan L.; Vitha, Mark F.; Mabbott, Gary A.

    2013-01-01

    High-performance liquid chromatography (HPLC) simulation software has long been recognized as an effective educational tool, yet many of the existing HPLC simulators are either too expensive, outdated, or lack many important features necessary to make them widely useful for educational purposes. Here, a free, open-source HPLC simulator is…

  2. Determination of aminoglycoside residues in kidney and honey samples by hydrophilic interaction chromatography-tandem mass spectrometry.

    PubMed

    Kumar, Praveen; Rúbies, Antoni; Companyó, Ramon; Centrich, Francesc

    2012-10-01

    Two methods based on liquid chromatography-tandem mass spectrometry were developed for the determination of ten aminoglycosides (streptomycin, dihydrostreptomycin, spectinomycin, apramycin, paromomycin, kanamycin A, gentamycin C1, gentamycin C2/C2a, gentamycin C1a, and neomycin B) in kidney samples from food-producing animals and in honey samples. The methods involved extraction with an aqueous solution (for the kidney samples) or sample dissolution in water (for the honey samples), solid-phase extraction with a weak cation exchange cartridge and injection of the eluate into a liquid chromatography-tandem mass spectrometry system. A zwitterionic hydrophilic interaction chromatography column was used for separation of aminoglycosides and a triple quadrupole mass analyzer was used for detection. The methods were validated according to Decision 2002/657/EC. The limits of quantitation ranged from 2 to 125 μg/kg in honey and 25 to 264 μg/kg in the kidney samples. Interday precision (RSD%) ranged from 6 to 26% in honey and 2 to 21% in kidney. Trueness, expressed as the percentage of error, ranged from 7 to 20% in honey and 1 to 11% in kidney. PMID:23065931

  3. Rapid hydrophilic interaction chromatography determination of lysine in pharmaceutical preparations with fluorescence detection after postcolumn derivatization with o-phtaldialdehyde.

    PubMed

    Douša, Michal; Břicháč, Jiří; Gibala, Petr; Lehnert, Petr

    2011-04-01

    A rapid procedure for the determination of lysine based on hydrophilic interaction chromatography (HILIC) separation of arginine and lysine with fluorescence detection has been developed. The separation conditions and parameters of lysine postcolumn derivatization with o-phtaldialdehyde (OPA)/2-mercaptoethanol were studied. The various HILIC columns were employed using isocratic elution. Fluorescence detection was performed at excitation and emission wavelength of 345 nm and 450 nm, respectively. An advantage of the reported method is a simple sample pre-treatment and a quick and very sensitive HPLC method. The developed method was successfully applied for analysis of commercial samples of Ibalgin Fast tablets (Zentiva, Czech Republic). PMID:21163603

  4. Potential of hydrophilic interaction chromatography for the analytical characterization of protein biopharmaceuticals.

    PubMed

    Periat, Aurélie; Fekete, Szabolcs; Cusumano, Alessandra; Veuthey, Jean-Luc; Beck, Alain; Lauber, Matthew; Guillarme, Davy

    2016-05-27

    A new stationary phase based on wide-pore hybrid silica bonded with amide ligand has been used to explore the utility of HILIC for the analytical characterization of protein biopharmaceuticals. Various, highly-relevant samples were tested, including different insulins, interferon α-2b and trastuzumab. This work shows that HILIC can be successfully employed for the analysis of therapeutic proteins and mAbs, using mobile phase compositions comprised of between 65 and 80% ACN and 0.1% TFA. In terms of elution order and selectivity, these HILIC separations have proven to be highly orthogonal to RPLC, while the kinetic performance remains comparable. In the case of characterizing trastuzumab, HILIC was uniquely able to resolve several important glycoforms at the middle-up level of analysis (fragments of 25-100kDa). Such a separation of glycoforms has been elusive by other separation mechanisms, such as RPLC and IEX. Besides showing orthogonality to RPLC and improved separations of glycoforms, HILIC offers several additional benefits for biopharmaceutical characterization: i) an inherent compatibility with MS, ii) a reduced requirement for very high mobile phase temperatures that are otherwise needed in RPLC to limit undesirably strong adsorption to the surface of the stationary phase, and iii) the possibility to couple several columns in series to improve resolving power, thanks to comparatively low mobile phase viscosity. PMID:27131959

  5. Multiple protein stationary phases: a review.

    PubMed

    Singh, N S; Habicht, K-L; Dossou, K S S; Shimmo, R; Wainer, I W; Moaddel, R

    2014-10-01

    Cellular membrane affinity chromatography stationary phases have been extensively used to characterize immobilized proteins and provide a direct measurement of multiple binding sites, including orthosteric and allosteric sites. This review will address the utilization of immobilized cellular and tissue fragments to characterize multiple transmembrane proteins co-immobilized onto a stationary phase. This approach will be illustrated by demonstrating that multiple transmembrane proteins were immobilized from cell lines and tissue fragments. In addition, the immobilization of individual compartments/organelles within a cell will be discussed and the changes in the proteins binding/kinetics based on their location. PMID:24780640

  6. Phenylboronate chromatography selectively separates glycoproteins through the manipulation of electrostatic, charge transfer, and cis-diol interactions.

    PubMed

    Carvalho, Rimenys J; Woo, James; Aires-Barros, M Raquel; Cramer, Steven M; Azevedo, Ana M

    2014-10-01

    Phenylboronate chromatography (PBC) has been applied for several years, however details regarding the mechanisms of interactions between the ligand and biomolecules are still scarce. The goal of this work is to investigate the various chemical interactions between proteins and their ligands, using a protein library containing both glycosylated and nonglycosylated proteins. Differences in the adsorption of these proteins over a pH range from 4 to 9 were related to two main properties: charge and presence of glycans. Acidic or neutral proteins were strongly adsorbed below pH 8 although the uncharged trigonal form of phenylboronate (PB) is less susceptible to forming electrostatic and cis-diol interactions with proteins. The glycosylated proteins were only adsorbed above pH 8 when the electrostatic repulsion between the boronate anion and the protein surface was mitigated (at 200 mM NaCl). All basic proteins were highly adsorbed above pH 8 with PB also acting as a cation-exchanger with binding occurring through electrostatic interactions. Batch adsorption performed at acidic conditions in the presence of Lewis base showed that charge-transfer interactions are critical for protein retention. This study demonstrates the multimodal interaction of PBC, which can be a selective tool for separation of different classes of proteins. PMID:25130283

  7. Chromatographic behavior of 12 polar pteridines in hydrophilic interaction chromatography using five different HILIC columns coupled with tandem mass spectrometry.

    PubMed

    Xiong, Xin; Liu, Yanmeng

    2016-04-01

    Retention characteristic of 5 hydrophilic interaction chromatography (HILIC) columns, containing neutral and possibly negatively charged support (silica, diol and amide), cationic phase (triazole) and zwitterionic phase (sulfobetaine), that are commercially available were studied for the separation of a group of 12 polar pteridines. The main factors influencing the retention and selectivity of pteridines for these different HILIC systems have been studied in liquid chromatography-tandem mass spectrometry (LC-MS/MS) conditions: mobile phase composition, buffer type, pH and concentration and the separation mechanism was also investigated. Results of the effects of organic modifier, buffer pH and ion strength indicate that the retention mechanism is a mixed-mode of adsorption and ion exchange, and optimization of HILIC analyses depends on the ionization state of the analytes. For silica, diol, amide and sulfobetaine phases, hydrophilic partitioning mainly contributes to the retention, while electrostatic interactions and hydrogen-bonding should be considered to understand the elution orders for triazole phase. An zwitterionic phase (ZIC-HILIC) provided the stronger retention for all pteridines than other tested columns. PMID:26838435

  8. Profiling the substitution pattern of xyloglucan derivatives by integrated enzymatic hydrolysis, hydrophilic-interaction liquid chromatography and mass spectrometry.

    PubMed

    Liu, Jun; Kisonen, Victor; Willför, Stefan; Xu, Chunlin; Vilaplana, Francisco

    2016-09-01

    Plant polysaccharides constitute arguably the most complex family of biomacromolecules in terms of the stereochemistry and regiochemistry of their intramolecular linkages. The chemical modification of such polysaccharides introduces an additional level of complexity for structural determinations. We have developed an integrated analytical procedure combining selective enzymatic hydrolysis, hydrophilic interaction liquid chromatography (HILIC), and mass spectrometry (MS) to describe the substitution pattern of xyloglucan (XyG) and its chemo-enzymatic derivatives (cationic, anionic, and benzyl aminated). Enzymatic hydrolysis of XyG derivatives by a xyloglucan-specific endoglucanase (XEG) generates oligosaccharides amenable for mass spectrometric identification with distinct structures, based on enzymatic substrate recognition and hydrolytic pattern. Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-ToF-MS) and electrospray ionisation mass spectrometry (ESI-MS) offer qualitative mass profiling of the chemical derivatives. Separation and identification of the complex oligosaccharide profiles released by enzymatic hydrolysis is achieved by hyphenation of hydrophilic interaction liquid chromatography with mass spectrometry (HILIC-ESI-MS). Further fragmentation by tandem mass spectrometry (ESI-MS/MS) in positive mode enables the structural sequencing of modified XyG oligosaccharides and the identification of the substituent position without further derivatisation. This integrated approach can be used to obtain semi-quantitative information of the substitution pattern of hemicellulose derivatives, with fundamental implications for their modification mechanisms and performance. PMID:27524300

  9. Spherical β-cyclodextrin-silica hybrid materials for multifunctional chiral stationary phases.

    PubMed

    Wang, Litao; Dong, Shuqing; Han, Feng; Zhao, Yingwei; Zhang, Xia; Zhang, Xiaoli; Qiu, Hongdeng; Zhao, Liang

    2015-02-27

    Spherical β-CD-silica hybrid materials have been prepared successfully by simple one-pot polymerization, which provide a new strategy to construct new type of HPLC chiral stationary phases. Various β-CD, ethane, triazinyl and 3,5-dimethylphenyl functional groups that can provide multiple interactions were introduced into the pore channels and pore wall framework of mesoporous materials, respectively. The materials towards some chiral, acidic, anilines and phenols compounds showed multiple chromatographic separation functions including racemic resolution, anion exchange and achiral separations with a typical feature of normal/reversed phase chromatography. Multi-tasking including racemic resolution and achiral separations for selected compounds were performed simultaneously on a chiral chromatographic column. The multifunctional character of materials arises from the multiple interactions including hydrophobic interaction, π-π interaction, anion exchange, inclusion interaction and hydrogen bonding interaction. PMID:25637012

  10. Fast HPLC method using ion-pair and hydrophilic interaction liquid chromatography for determination of phenylephrine in pharmaceutical formulations.

    PubMed

    Dousa, Michal; Gibala, Petr

    2010-01-01

    A rapid procedure based on a direct extraction and HPLC determination with fluorescence detection of phenylephrine in pharmaceutical sachets that include a large excess of paracetamol (65 + 1, w/w), ascorbic acid (5 + 1, w/w), and other excipients (aspartame and sucrose) was developed and validated. The final optimized chromatographic method for ion-pair chromatography used an XTerra RP18 column, 3 microm particle size, 50 x 3.0 mm id. The mobile phase consisted of a mixture of acetonitrile and buffer (10 mM sodium octane-1-sulfonate, adjusted with H3PO4 to pH 2.2; 200 + 800, v/v), with a constant flow rate of 0.3 mL/min. The separation was carried out at 30 degrees C, and the injection volume was 3 microL. Fluorescence detection was performed at excitation and emission wavelengths of 275 and 310 nm, respectively. The mobile phase parameters, such as the organic solvent fraction (acetonitrile) in mobile phase as an organic modifier, the concentration of sodium octane-1-sulfonate as a counter-ion, temperature, and pH of mobile phase, were studied. As an alternative to ion-pair chromatography, hydrophilic interaction liquid chromatography (HILIC) was investigated using a Luna HILIC column, 3 microm, 100 x 4.6 mm id. The mobile phase consisted of acetonitrile and buffer (5 mM potassium dihydrogen phosphate, adjusted with H3PO4 to pH 2.5; 750 + 250, v/v) at a flow rate of 0.8 mL/min. The separation was carried out at 25 degrees C, and the injection volume was 5 microL. The proposed method has an advantage of a very simple sample pretreatment, and is much faster than the currently utilized HPLC methods using gradient elution and UV detection. Commercial samples of sachets were successfully analyzed by the proposed HPLC method. PMID:21140654

  11. Interaction between hydroxypropyl methylcellulose and biphasic calcium phosphate after steam sterilisation: capillary gas chromatography studies.

    PubMed

    Bourges, X; Schmitt, M; Amouriq, Y; Daculsi, G; Legeay, G; Weiss, P

    2001-01-01

    The purpose of this study was to check the chemical stability of an injectable bone substitute (IBS) composed of a 50/50 w/w mixture of 2.92% hydroxypropyl methylcellulose (HPMC) solution in deionized water containing biphasic calcium phosphate (BCP) granules (60% hydroxyapatite/40% beta-tricalcium phosphate w/w). After separation of the organic and mineral phases, capillary gas chromatography (GC) was used to study the possible modification of HPMC due to the contact with BCP granules following steam sterilisation and 32 days storage at room temperature. HPMC was extracted from IBS in aqueous medium, and a dialytic method was then used to extract calcium phosphate salts from the HPMC. The percentage of HPMC extracted from BCP was 98.5%+/-0.5%, as measured by UV. GC showed no chemical modifications after steam sterilisation and storage. PMID:11556737

  12. Interaction between hydroxypropyl methylcellulose and biphasic calcium phosphate after steam sterilisation: capillary gas chromatography studies

    PubMed Central

    Bourges, Xavier; Schmitt, Michel; Amouriq, Yves; Daculsi, Guy; Legeay, Gilbert; Weiss, Pierre

    2001-01-01

    The purpose of this study was to check the chemical stability of an injectable bone substitute (IBS) composed of a 50/50 w/w mixture of a 2.92% hydroxypropyl methylcellulose (HPMC) solution in deionised water containing biphasic calcium phosphate (BCP) granules (60% hydroxyapatite/40% β-tricalcium phosphate w/w). After separation of the organic and mineral phases, capillary gas chromatography (GC) was used to study the possible modification of HPMC due to the contact with BCP granules following steam sterilisation and 32 days of storage at room temperature. HPMC was extracted from IBS in aqueous medium, and a dialytic method was then use to extract calcium phosphate salts from HPMC. The percentage of HPMC extracted from BCP was 98.5% ± 0.5% as measured by a UV method. GC showed no chemical modifications after steam sterilisation and storage. PMID:11556737

  13. IDENTIFICATION AND ANALYSIS OF STEREOSELECTIVE DRUG INTERACTIONS WITH LOW DENSITY LIPOPROTEIN BY HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Sobansky, Matthew R.; Hage, David S.

    2012-01-01

    Columns containing immobilized low density lipoprotein (LDL) were prepared for the analysis of drug interactions with this agent by high-performance affinity chromatography (HPAC). R/S-Propranolol was used as a model drug for this study. The LDL columns gave reproducible binding to propranolol over 60 h of continuous use in the presence of pH 7.4, 0.067 M potassium phosphate buffer. Experiments conducted with this type of column through frontal analysis indicated that two types of interactions were occurring between R-propranolol and LDL, while only a single type of interaction was observed between S-propranolol and LDL. The first type of interaction, which was seen for both enantiomers, involved non-saturable binding; this interaction had an overall affinity (nKa) of 1.9 (± 0.1) × 105 M-1 for R-propranolol and 2.7 (± 0.2) × 105 M-1 for S-propranolol at 37 °C. The second type of interaction was observed only for R-propranolol and involved saturable binding that had an association equilibrium constant (Ka) of 5.2 (± 2.3) × 105 M-1 at 37 °C. Similar differences in binding behavior were found for the two enantiomers at 20 °C and 27 °C. This is the first known example of stereoselective binding of drugs by LDL or other lipoproteins. This work also illustrates the ability of HPAC to be used as a tool for characterizing mixed-mode interactions that involve LDL and related binding agents. PMID:22354572

  14. Gradient enhanced-fluidity liquid hydrophilic interaction chromatography of ribonucleic acid nucleosides and nucleotides: A "green" technique.

    PubMed

    Beilke, Michael C; Beres, Martin J; Olesik, Susan V

    2016-03-01

    A "green" hydrophilic interaction liquid chromatography (HILIC) technique for separating the components of mixtures with a broad range of polarities is illustrated using enhanced-fluidity liquid mobile phases. Enhanced-fluidity liquid chromatography (EFLC) involves the addition of liquid CO2 to conventional liquid mobile phases. Decreased mobile phase viscosity and increased analyte diffusivity results when a liquefied gas is dissolved in common liquid mobile phases. The impact of CO2 addition to a methanol:water (MeOH:H2O) mobile phase was studied to optimize HILIC gradient conditions. For the first time a fast separation of 16 ribonucleic acid (RNA) nucleosides/nucleotides was achieved (16min) with greater than 1.3 resolution for all analyte pairs. By using a gradient, the analysis time was reduced by over 100% compared to similar separations conducted under isocratic conditions. The optimal separation using MeOH:H2O:CO2 mobile phases was compared to MeOH:H2O and acetonitrile:water (ACN:H2O) mobile phases. Based on chromatographic performance parameters (efficiency, resolution and speed of analysis) and an assessment of the environmental impact of the mobile phase mixtures, MeOH:H2O:CO2 mixtures are preferred over ACN:H2O or MeOH:H2O mobile phases for the separation of mixtures of RNA nucleosides and nucleotides. PMID:26860052

  15. Influence of binding pH and protein solubility on the dynamic binding capacity in hydrophobic interaction chromatography.

    PubMed

    Baumann, Pascal; Baumgartner, Kai; Hubbuch, Jürgen

    2015-05-29

    Hydrophobic interaction chromatography (HIC) is one of the most frequently used purification methods in biopharmaceutical industry. A major drawback of HIC, however, is the rather low dynamic binding capacity (DBC) obtained when compared to e.g. ion exchange chromatography (IEX). The typical purification procedure for HIC includes binding at neutral pH, independently of the proteins nature and isoelectric point. Most approaches to process intensification are based on resin and salt screenings. In this paper a combination of protein solubility data and varying binding pH leads to a clear enhancement of dynamic binding capacity. This is shown for three proteins of acidic, neutral, and alkaline isoelectric points. High-throughput solubility screenings as well as miniaturized and parallelized breakthrough curves on Media Scout RoboColumns (Atoll, Germany) were conducted at pH 3-10 on a fully automated robotic workstation. The screening results show a correlation between the DBC and the operational pH, the protein's isoelectric point and the overall solubility. Also, an inverse relationship of DBC in HIC and the binding kinetics was observed. By changing the operational pH, the DBC could be increased up to 30% compared to the standard purification procedure performed at neutral pH. As structural changes of the protein are reported during HIC processes, the applied samples and the elution fractions were proven not to be irreversibly unfolded. PMID:25911386

  16. Development of analytical method for catechol compounds in mouse urine using hydrophilic interaction liquid chromatography with fluorescence detection.

    PubMed

    Kanamori, Takahiro; Isokawa, Muneki; Funatsu, Takashi; Tsunoda, Makoto

    2015-03-15

    An analytical method for catecholamines and related compounds using hydrophilic interaction liquid chromatography (HILIC) with native fluorescence detection has been developed. We found that ZIC-cHILIC with phosphorylcholine was suitable for the separation of catechol compounds with good peak shapes among six different HILIC columns (Inertsil SIL, Inertsil Amide, Inertsil Diol, TSKgel NH2-100, ZIC-HILIC, and ZIC-cHILIC). Using ZIC-cHILIC, eight catechol compounds (dopamine, epinephrine, norepinephrine, 3,4-dihydroxyphenylalanine, 3,4-dihydroxyphenylacetic acid, 3,4-dihydroxyphenylglycol, 3,4-dihydroxymandelic acid, and internal standard 3,4-dihydroxybenzylamine) were separated within 15min. The limit of detection at a signal to noise ratio of 3 was 3-28nM. An improved sensitivity was obtained as compared to that of reversed-phase liquid chromatography. This was partly attributed to the increase in the fluorescence intensity of the catechol compounds in the acetonitrile-rich mobile phase. Solid phase extraction using a monolithic silica disk-packed spin column with phenylboronate moieties, which have affinity to catechol compounds, was performed for the selective extraction of catechol compounds from mouse urine. Dopamine, epinephrine, norepinephrine, 3,4-dihydroxyphenylalanine, and 3,4-dihydroxyphenylglycol were successfully quantified in mouse urine. PMID:25682335

  17. Study of photorespiration in marine microalgae through the determination of glycolic acid using hydrophilic interaction liquid chromatography.

    PubMed

    Rigobello-Masini, Marilda; Penteado, José C P; Tiba, Maurício; Masini, Jorge C

    2012-01-01

    Determination of organic acids in intracellular extracts and in the cultivation media of marine microalgae aid investigations about metabolic routes related to assimilation of atmospheric carbon by these organisms, which are known by their role in the carbon dioxide sink. The separation of these acids was investigated by hydrophilic interaction liquid chromatography (HILIC) using isocratic elution with a mobile phase composed of 70:30 v/v acetonitrile/20 mmol/L ammonium acetate buffer (pH 6.8) and detection at 220 nm. HILIC allowed the determinations of glycolic acid, the most important metabolite for the evaluation of the photorespiration process in algae, to be made with better selectivity than that achieved by reversed phase liquid chromatography, but with less detectability. The concentration of glycolic acid was determined in the cultivation media and in intracellular extracts of the algae Tetraselmis gracilis and Phaeodactylum tricornutum submitted to different conditions of aeration: (i) without forced aeration, (ii) aeration with atmospheric air, and (iii) bubbling with N(2). The concentration of glycolic acid had a higher increase as the cultures were aerated with nitrogen, showing higher photorespiratory flux than that occurring in the cultures aerated with atmospheric air. PMID:22128110

  18. Analysis of paralytic shellfish toxins, potential chemical threat agents, in food using hydrophilic interaction liquid chromatography-mass spectrometry.

    PubMed

    Jansson, Daniel; Åstot, Crister

    2015-10-23

    A novel method for determining paralytic shellfish toxin (PST) profiles in food was developed using a combination of silica and strong cation exchange (SCX) solid phase extraction (SPE) coupled to hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS). Besides the risk for natural contamination of seafood and drinking water, PSTs also pose potent threats through intentional contamination of food, due to their high toxicity and the wide distributions of toxin-producing algae. The new preparation method aim to maintain the samples' original toxin profiles by avoiding conditions known to induce interconversion or degradation of the PSTs. The method was evaluated for PST extraction from water, milk, orange juice, apple purée, baby food, and blue mussels (Mytilus edulis). The extracts were found to produce reproducible retention times in HILIC-MS/MS analysis. When an authentic toxic mussel sample was analyzed using the novel method, saxitoxin and gonyautoxin-3 were identified, in agreement with data acquired using the Lawrence pre-column oxidation high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method. Overall recoveries of the PSTs from tested foods by the novel method ranged from 36% to 111%. PMID:26404910

  19. Quantitation of five organophosphorus nerve agent metabolites in serum using hydrophilic interaction liquid chromatography and tandem mass spectrometry

    PubMed Central

    Hamelin, Elizabeth I.; Schulze, Nicholas D.; Shaner, Rebecca L.; Coleman, Rebecca M.; Lawrence, Richard J.; Crow, Brian S.; Jakubowski, E. M.; Johnson, Rudolph C.

    2015-01-01

    Although nerve agent use is prohibited, concerns remain for human exposure to nerve agents during decommissioning, research, and warfare. Exposure can be detected through the analysis of the hydrolysis products in urine as well as blood. An analytical method to detect exposure to five nerve agents, including VX, VR (Russian VX), GB (sarin), GD (soman) and GF (cyclosarin), through the analysis of the hydrolysis products, which are the primary metabolites, in serum has been developed and characterized. This method uses solid phase extraction coupled with high performance liquid chromatography for separation and isotopic dilution tandem mass spectrometry for detection. An uncommon buffer of ammonium fluoride was used to enhance ionization and improve sensitivity when coupled with hydrophilic interaction liquid chromatography resulting in detection limits from 0.3–0.5 ng/mL. The assessment of two quality control samples demonstrated high accuracy (101–105%) and high precision (5–8%) for the detection of these five nerve agent hydrolysis products in serum. PMID:24633507

  20. Quantitation of five organophosphorus nerve agent metabolites in serum using hydrophilic interaction liquid chromatography and tandem mass spectrometry.

    PubMed

    Hamelin, Elizabeth I; Schulze, Nicholas D; Shaner, Rebecca L; Coleman, Rebecca M; Lawrence, Richard J; Crow, Brian S; Jakubowski, E M; Johnson, Rudolph C

    2014-08-01

    Although nerve agent use is prohibited, concerns remain for human exposure to nerve agents during decommissioning, research, and warfare. Exposure can be detected through the analysis of hydrolysis products in urine as well as blood. An analytical method to detect exposure to five nerve agents, including VX, VR (Russian VX), GB (sarin), GD (soman), and GF (cyclosarin), through the analysis of the hydrolysis products, which are the primary metabolites, in serum has been developed and characterized. This method uses solid-phase extraction coupled with high-performance liquid chromatography for separation and isotopic dilution tandem mass spectrometry for detection. An uncommon buffer of ammonium fluoride was used to enhance ionization and improve sensitivity when coupled with hydrophilic interaction liquid chromatography resulting in detection limits from 0.3 to 0.5 ng/mL. The assessment of two quality control samples demonstrated high accuracy (101-105%) and high precision (5-8%) for the detection of these five nerve agent hydrolysis products in serum. PMID:24633507

  1. Determination of nicotine and its metabolites accumulated in fish tissue using hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry.

    PubMed

    Chang, Yun-Wei; Nguyen, Hien P; Chang, Mike; Burket, S Rebekah; Brooks, Bryan W; Schug, Kevin A

    2015-07-01

    The determination of nicotine and its major metabolites (cotinine and anabasine) in fish tissue was performed using liquid chromatography and tandem mass spectrometry. Marine and freshwater fish were purchased from local grocery stores and were prepared based on a quick, easy, cheap, effective, rugged, and safe sample preparation protocol. To determine the highly polar compounds, hydrophilic interaction liquid chromatography was also used. There were modest suppressions on measured nicotine signals (10%) due to the matrix effects from marine fish but no obvious effects on freshwater fish signals. Method validation was incorporated with internal standards and carried out with matrix-matched calibration. The detection limits for nicotine, cotinine, and anabasine were 9.4, 3.0, and 1.5 ng/g in fish, respectively. Precision was quite acceptable returning less than 8% RSD at low, medium, and high concentrations. Acceptable and reproducible extraction recoveries (70-120%) of all three compounds were achieved, except for anabasine at low concentration (61%). The method was then applied to define nicotine bioaccumulation in a fathead minnow model, which resulted in rapid uptake with steady state internal tissue levels, reached within 12 h. This developed method offers a fast, easy, and sensitive way to evaluate nicotine and its metabolite residues in fish tissues. PMID:25953492

  2. PHARMACEUTICAL AND BIOMEDICAL APPLICATIONS OF AFFINITY CHROMATOGRAPHY: RECENT TRENDS AND DEVELOPMENTS

    PubMed Central

    Hage, David S.; Anguizola, Jeanethe A.; Bi, Cong; Li, Rong; Matsuda, Ryan; Papastavros, Efthimia; Pfaunmiller, Erika; Vargas, John; Zheng, Xiwei

    2012-01-01

    Affinity chromatography is a separation technique that has become increasingly important in work with biological samples and pharmaceutical agents. This method is based on the use of a biologically-related agent as a stationary phase to selectively retain analytes or to study biological interactions. This review discusses the basic principles behind affinity chromatography and examines recent developments that have occurred in the use of this method for biomedical and pharmaceutical analysis. Techniques based on traditional affinity supports are discussed, but an emphasis is placed on methods in which affinity columns are used as part of HPLC systems or in combination with other analytical methods. General formats for affinity chromatography that are considered include step elution schemes, weak affinity chromatography, affinity extraction and affinity depletion. Specific separation techniques that are examined include lectin affinity chromatography, boronate affinity chromatography, immunoaffinity chromatography, and immobilized metal ion affinity chromatography. Approaches for the study of biological interactions by affinity chromatography are also presented, such as the measurement of equilibrium constants, rate constants, or competition and displacement effects. In addition, related developments in the use of immobilized enzyme reactors, molecularly imprinted polymers, dye ligands and aptamers are briefly considered. PMID:22305083

  3. Research on the interaction of hydrogen-bond acidic polymer sensitive sensor materials with chemical warfare agents simulants by inverse gas chromatography.

    PubMed

    Yang, Liu; Han, Qiang; Cao, Shuya; Huang, Feng; Qin, Molin; Guo, Chenghai; Ding, Mingyu

    2015-01-01

    Hydrogen-bond acidic polymers are important high affinity materials sensitive to organophosphates in the chemical warfare agent sensor detection process. Interactions between the sensor sensitive materials and chemical warfare agent simulants were studied by inverse gas chromatography. Hydrogen bonded acidic polymers, i.e., BSP3, were prepared for micro-packed columns to examine the interaction. DMMP (a nerve gas simulant) and 2-CEES (a blister agent simulant) were used as probes. Chemical and physical parameters such as heats of absorption and Henry constants of the polymers to DMMP and 2-CEES were determined by inverse gas chromatography. Details concerning absorption performance are also discussed in this paper. PMID:26043177

  4. Research on the Interaction of Hydrogen-Bond Acidic Polymer Sensitive Sensor Materials with Chemical Warfare Agents Simulants by Inverse Gas Chromatography

    PubMed Central

    Yang, Liu; Han, Qiang; Cao, Shuya; Huang, Feng; Qin, Molin; Guo, Chenghai; Ding, Mingyu

    2015-01-01

    Hydrogen-bond acidic polymers are important high affinity materials sensitive to organophosphates in the chemical warfare agent sensor detection process. Interactions between the sensor sensitive materials and chemical warfare agent simulants were studied by inverse gas chromatography. Hydrogen bonded acidic polymers, i.e., BSP3, were prepared for micro-packed columns to examine the interaction. DMMP (a nerve gas simulant) and 2-CEES (a blister agent simulant) were used as probes. Chemical and physical parameters such as heats of absorption and Henry constants of the polymers to DMMP and 2-CEES were determined by inverse gas chromatography. Details concerning absorption performance are also discussed in this paper. PMID:26043177

  5. The second virial coefficient as a predictor of protein aggregation propensity: A self-interaction chromatography study.

    PubMed

    Quigley, A; Williams, D R

    2015-10-01

    The second osmotic virial coefficients (b2) of four proteins - lysozyme, recombinant human lactoferrin, concanavalin A and catalase were measured by self-interaction chromatography (SIC) in solutions of varying salt type, concentration and pH. Protein aggregate sizes based on the initial hydrodynamic radius of the protein solution species present were measured using dynamic light scattering, and the relationship between b2 and protein aggregate size was studied. A linear correlation was established between b2 values and protein aggregate hydrodynamic size for all proteins, and for almost all solution conditions. Aggregate sizes of <∼10nm, indicative of non-aggregated protein systems, were consistently observed to have b2 values >0. The observed b2 trends as a function of solution conditions were very much protein dependent, with notable trends including the existence of attractive interactions (negative b2 values) at low ionic strengths for catalase and concanavalin A, and the highly positive b2 values observed for lactoferrin over a wide range of solution conditions, reflecting lactoferrin's innately high stability. It is concluded that the quantification of protein-protein interactions using SIC based b2 data is a potentially valuable screening tool for predicting protein aggregation propensity. PMID:26259782

  6. Chromatography of carbon nanotubes separated albumin from other serum proteins: a method for direct analysis of their interactions.

    PubMed

    Kuboki, Yoshinori; Koshikawa, Takamitu; Takita, Hiroko; Fujisawa, Ryuichi; Lee, Min-ho; Abe, Shigeaki; Akasaka, Tsukasa; Uo, Motohiro; Watari, Fumio; Sammons, Rachel

    2010-08-01

    Chromatography technology was employed to clarify the mechanism of interaction between multi-wall carbon nanotubes (MWCNT) and proteins. A column (16x100 mm) was packed with purified MWCNT, and various proteins were eluted with phosphate buffered saline (PBS) with and without gradient systems. It was found that albumin in bovine serum was eluted immediately from the column without any adsorption to MWCNT. Conversely, the non-albumin proteins, including a protein of 85 kDa molecular mass and a group of proteins with molecular masses higher than 115 kDa, exhibited considerably high affinity towards MWCNT. A sample of pure bovine serum albumin was also eluted immediately from the column, while lysozyme did not elute as a peak with PBS, but eluted with 0.6 M NaCl. Fundamentally, carbon nanotubes are devoid of any electrical charge. Therefore, other forces including the hydrogen bonds, hydrophilic interactions, and van der Waals forces were most probably responsible for the differential elution behaviors. In conclusion, this chromatographic method provided a simple and direct analysis of the interactions between carbon nanotubes and the various proteins. PMID:20610879

  7. The second virial coefficient as a predictor of protein aggregation propensity: A self-interaction chromatography study

    PubMed Central

    Quigley, A.; Williams, D.R.

    2015-01-01

    The second osmotic virial coefficients (b2) of four proteins – lysozyme, recombinant human lactoferrin, concanavalin A and catalase were measured by self-interaction chromatography (SIC) in solutions of varying salt type, concentration and pH. Protein aggregate sizes based on the initial hydrodynamic radius of the protein solution species present were measured using dynamic light scattering, and the relationship between b2 and protein aggregate size was studied. A linear correlation was established between b2 values and protein aggregate hydrodynamic size for all proteins, and for almost all solution conditions. Aggregate sizes of <∼10 nm, indicative of non-aggregated protein systems, were consistently observed to have b2 values >0. The observed b2 trends as a function of solution conditions were very much protein dependent, with notable trends including the existence of attractive interactions (negative b2 values) at low ionic strengths for catalase and concanavalin A, and the highly positive b2 values observed for lactoferrin over a wide range of solution conditions, reflecting lactoferrin’s innately high stability. It is concluded that the quantification of protein–protein interactions using SIC based b2 data is a potentially valuable screening tool for predicting protein aggregation propensity. PMID:26259782

  8. Escherichia coli produces linoleic acid during late stationary phase.

    PubMed Central

    Rabinowitch, H D; Sklan, D; Chace, D H; Stevens, R D; Fridovich, I

    1993-01-01

    Escherichia coli produces linoleic acid in the late stationary phase. This was the case whether the cultures were grown aerobically or anaerobically on a supplemented glucose-salts medium. The linoleic acid was detected by thin-layer chromatography and was measured as the methyl ester by gas chromatography. The linoleic acid methyl ester was identified by its mass spectrum. Lipids extracted from late-stationary-phase cells generated thiobarbituric acid-reactive carbonyl products when incubated with a free radical initiator. In contrast, extracts from log-phase or early-stationary-phase cells failed to do so, in accordance with the presence of polyunsaturated fatty acid only in the stationary-phase cells. PMID:8366020

  9. Interactions between minimum run time, modifier concentration, and efficiency parameters in a high performance liquid chromatography separation.

    PubMed

    Chester, T L; Stalcup, A M

    2011-01-14

    We modeled and studied the separation of uracil, nicotinamide, resorcinol, theobromine, theophylline, and caffeine on four C-18 columns of different lengths packed with the same stationary phase using water/methanol mobile phase at one temperature. Predictions of retention times and peak widths were compared with experimental results and were found to be sufficiently accurate for performing optimization calculations. With limits set on the required resolution and on maximum values for pressure and flow rate, calculations were performed for numerous virtual column lengths seeking the smallest possible analysis time for each length while allowing methanol concentration and flow rate to vary as required to minimize run time. Predictions were experimentally verified for the column lengths actually available. These calculations revealed the dependence of best-possible analysis time on column length, modifier concentration, flow rate, and pressure for the real system that was modeled, and provided insight into parameter interactions with respect to analysis times meeting the needs and limits specified. We show that when these parameters are considered in concert, rather than individually, conventional guidelines regarding setting their values may not always lead to the optimum. PMID:21130461

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

    PubMed Central

    Murphy, Patrick J. M.

    2014-01-01

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

  11. Enantiomeric separation of amlodipine and its two chiral impurities by nano-liquid chromatography and capillary electrochromatography using a chiral stationary phase based on cellulose tris(4-chloro-3-methylphenylcarbamate).

    PubMed

    Auditore, Roberta; Santagati, Natale A; Aturki, Zeineb; Fanali, Salvatore

    2013-09-01

    In this work, a novel polysaccharide-based chiral stationary phase, cellulose tris(4-chloro-3-methylphenylcarbamate), also called Sepapak 4 has been evaluated for the chiral separation of amlodipine (AML) and its two impurities. AML is a powerful vasodilatator drug used for the treatment of hypertension. Capillary columns of 100 μm id packed with the chiral stationary phase were used for both nano-LC and CEC experiments. The optimization of the mobile phase composed of ACN/water, (90:10, v/v) containing 15 mM ammonium borate pH 10.0 in nano-LC allowed the chiral separation of AML and the two impurities, but not in a single run. With the purpose to obtain the separation of the three pairs of enantiomers simultaneously, CEC analyses were performed in the same conditions achieving better enantioresolution and higher separation efficiencies for each compound. To fully resolve the mixture of six enantiomers, parameters such as buffer pH and concentration sample injection have been then investigated. A mixture of ACN/water (90:10, v/v) containing 5 mM ammonium borate buffer pH 9.0 enabled the complete separation of the three couples of enantiomers in less than 30 min. The optimized CEC method was therefore validated and applied to the analysis of pharmaceutical formulation declared to contain only AML racemate. PMID:23775281

  12. Stationary nonlinear Airy beams

    SciTech Connect

    Lotti, A.; Faccio, D.; Couairon, A.; Papazoglou, D. G.; Panagiotopoulos, P.; Tzortzakis, S.; Abdollahpour, D.

    2011-08-15

    We demonstrate the existence of an additional class of stationary accelerating Airy wave forms that exist in the presence of third-order (Kerr) nonlinearity and nonlinear losses. Numerical simulations and experiments, in agreement with the analytical model, highlight how these stationary solutions sustain the nonlinear evolution of Airy beams. The generic nature of the Airy solution allows extension of these results to other settings, and a variety of applications are suggested.

  13. Quantification of Sunscreen Benzophenone-4 in Hair Shampoos by Hydrophilic Interactions Thin-Layer Chromatography/Densitometry or Derivative UV Spectrophotometry.

    PubMed

    Sobańska, Anna W; Kałębasiak, Katarzyna; Pyzowski, Jarosław; Brzezińska, Elżbieta

    2015-01-01

    Benzophenone-4 (BZ4) was separated from surfactants, dyes, preservatives, and other components of hair shampoos by thin-layer chromatography on silica gel 60 stationary phase, with ethyl acetate-ethanol-water-pH 6 phosphate buffer (15 : 7 : 5 : 1 v/v/v/v) as mobile phase. Densitometry scanning of chromatograms was performed at 285 nm. The densitometric calibration curve for BZ4 was nonlinear (second-degree polynomial), with R > 0.999. The limits of detection and quantification were ca. 0.03 and ca. 0.1 μg spot(-1), respectively. The results obtained by HPTLC-densitometry were compared to those obtained by zero and 2nd derivative UV spectrophotometry. In the case of spectrophotometric methods, calibration curves were linear with R > 0.9998. The chromatographic method was fully validated. PMID:25734022

  14. The accurate measurement of second virial coefficients using self-interaction chromatography: experimental considerations.

    PubMed

    Quigley, A; Heng, J Y Y; Liddell, J M; Williams, D R

    2013-11-01

    Measurement of B22, the second virial coefficient, is an important technique for describing the solution behaviour of proteins, especially as it relates to precipitation, aggregation and crystallisation phenomena. This paper describes the best practise for calculating B22 values from self-interaction chromatograms (SIC) for aqueous protein solutions. Detailed analysis of SIC peak shapes for lysozyme shows that non-Gaussian peaks are commonly encountered for SIC, with typical peak asymmetries of 10%. This asymmetry reflects a non-linear chromatographic retention process, in this case heterogeneity of the protein-protein interactions. Therefore, it is important to use the centre of mass calculations for determining accurate retention volumes and thus B22 values. Empirical peak maximum chromatogram analysis, often reported in the literature, can result in errors of up to 50% in B22 values. A methodology is reported here for determining both the mean and the variance in B22 from SIC experiments, includes a correction for normal longitudinal peak broadening. The variance in B22 due to chemical effects is quantified statistically and is a measure of the heterogeneity of protein-protein interactions in solution. In the case of lysozyme, a wide range of B22 values are measured which can vary significantly from the average B22 values. PMID:23623796

  15. Separation performance of cucurbit[8]uril and its coordination complex with cadmium (II) in capillary gas chromatography.

    PubMed

    Sun, Tao; Ji, Ningning; Qi, Meiling; Tao, Zhu; Fu, Ruonong

    2014-05-23

    Here we report the investigation of using cucurbit[8]uril (CB8) and its coordination complex with cadmium (II) (CB8-Cd) as stationary phases for capillary gas chromatography (GC). The prepared capillary columns of CB8 and CB8-Cd stationary phases achieved column efficiency of 2200plates/m and 1508plates/m, respectively, and showed weak polarity based on the measured McReynolds constants. Their separation performance was investigated by GC separation of mixtures of different types while a commercial column was used for comparison. The CB8 stationary phase achieved high resolution for a wide range of analytes from nonpolar to polar while the CB8-Cd stationary phase exhibited good separation mainly for nonpolar to weak polar analytes. The CB stationary phases differ from the commercial one in terms of retention behaviors and resolving ability due to their different molecular interactions with analytes. Moreover, energy effect on the retention of analytes on CB8 and CB8-Cd stationary phases was examined, showing that retention on CB8 column was determined mainly by enthalpy change for polar analytes and by both enthalpy change and entropy change for weak polar analytes whereas retention on CB8-Cd column was mainly controlled by entropy change. This work demonstrates the great potential of CB8 and CB8-Cd stationary phases as a new type of GC stationary phases in GC analysis. PMID:24745846

  16. Evaluation of the technical variations and the suitability of a hydrophilic interaction liquid chromatography-high resolution mass spectrometry (ZIC-pHILIC-Exactive orbitrap) for clinical urinary metabolomics study.

    PubMed

    Zhang, Tong; Watson, David G

    2016-06-01

    Although many hydrophilic interaction liquid chromatography-high resolution mass spectrometry (HILIC-HRMS) methods have been developed and applied for untargeted metabolite profiling in clinical metabolomics, according to the literature, the suitability of these HILIC-HRMS methods has not been fully evaluated with respect to their performance when they are subjected to statistical analysis. In this study, using a series of human urine samples we investigated the effect of technical variations on multivariate and univariate analysis of the data collected using a previously developed HILIC-HRMS method for untargeted urinary metabolite profiling in clinical metabolomics. The technical variation introduced by sample preparation was more significant than that produced by the HILIC-HRMS method. By using an orthogonal partial least squares (OPLS) model, subtle fold-changes were accurately measured in the urine samples spiked with (13)C and (15)N isotope labelled amino acids at different concentrations. The robustness of this HILIC method was also evaluated by analysing the obtained data from a single urine sample following manipulation of several primary LC parameters. High reproducibility in the chromatographic performance of three ZIC-pHILIC columns with different batch numbers indicated the reliability of the polymer based zwitterionic stationary phase allowing column replacement without compromising the performance of the method. PMID:27107246

  17. Sequential enrichment of singly- and multiply-phosphorylated peptides with zwitterionic hydrophilic interaction chromatography material.

    PubMed

    Sheng, Qianying; Yang, Kaiya; Xue, Xingya; Li, Xiuling; Guo, Zhimou; Shen, Aijin; Ke, Yanxiong; Lan, Minbo; Liang, Xinmiao

    2015-09-25

    An interesting and novel method for the selective and sequential enrichment of singly- and multiply-phosphorylated peptides with a zwitterionic material "Click TE-Cys" is presented. Retention mechanisms between phosphopeptides and Click TE-Cys are systematically investigated by checking the influence of acetonitrile content, pH value, and buffer concentration on the retention of phosphopeptides. Both hydrophilic interaction and electrostatic interaction are involved in retention between phosphopeptides and Click TE-Cys. Based on these results, an optimized method is established for selective enrichment of phosphopeptides using Click TE-Cys. This method not only exhibits high selectivity for phosphopeptides, but also fractionates singly- and multiply-phosphorylated peptides into two fractions. This method was evaluated using relatively complex samples, including peptide mixtures of α-casein and bovine serum albumin (BSA) at a molar ratio of 1:10 and skim milk. This efficient and optimized protocol has great potential for enriching multiply-phosphorylated peptides and could be a valuable tool for specific enrichment of phosphopeptides in phosphoproteome analysis. PMID:26298604

  18. Using Affinity Chromatography to Investigate Novel Protein–Protein Interactions in an Undergraduate Cell and Molecular Biology Lab Course

    PubMed Central

    2009-01-01

    Inquiry-driven lab exercises require students to think carefully about a question, carry out an investigation of that question, and critically analyze the results of their investigation. Here, we describe the implementation and assessment of an inquiry-based laboratory exercise in which students obtain and analyze novel data that contribute to our understanding of macromolecular trafficking between the nucleus and cytoplasm in eukaryotic cells. Although many of the proteins involved in nucleocytoplasmic transport are known, the physical interactions between some of these polypeptides remain uncharacterized. In this cell and molecular biology lab exercise, students investigate novel protein–protein interactions between factors involved in nuclear RNA export. Using recombinant protein expression, protein extraction, affinity chromatography, SDS-polyacrylamide gel electrophoresis, and Western blotting, undergraduates in a sophomore-level lab course identified a previously unreported association between the soluble mRNA transport factor Mex67 and the C-terminal region of the yeast nuclear pore complex protein Nup1. This exercise immersed students in the process of investigative science, from proposing and performing experiments through analyzing data and reporting outcomes. On completion of this investigative lab sequence, students reported enhanced understanding of the scientific process, increased proficiency with cellular and molecular methods and content, greater understanding of data analysis and the importance of appropriate controls, an enhanced ability to communicate science effectively, and an increased enthusiasm for scientific research and for the lab component of the course. The modular nature of this exercise and its focus on asking novel questions about protein–protein interactions make it easily transferable to undergraduate lab courses performed in a wide variety of contexts. PMID:19723816

  19. High performance liquid chromatography analysis of 9-(2',3'-dideoxy-2'beta-fluoro-D-threo-penta furanosyl) adenine and its metabolite in human plasma using solid-phase extraction on a polyfluorinated reversed stationary phase.

    PubMed

    Aboul-Enein, H Y; Abu-Zaid, S

    2001-06-01

    A quick and sensitive reversed-phase HPLC method has been developed for the analysis of 2'-beta -fluoro-2',3'-dideoxy adenosine (F-ddA), the acid-stable anti-AIDS drug, and its metabolite 2'-fluoro-2',3'-dideoxy inosine (F-ddI) in human plasma using polyfluorinated stationary phase column (Fluo fix, 15 cm, 4.0 mm i.d., 5 microm particle size). The mobile phase consisted of ammonium phosphate buffer solution (10 mM) adjusted with phosphoric acid 85% to pH 6.8:dimethyl formamide (97:3, v/v). F-ddA and F-ddI were monitored by UV-visible detector at 258 and 247 nm, respectively. The recoveries of F-ddA and F-ddI from plasma using a C(18) solid-phase extraction cartridge were 99.2% and 99.7%, respectively. PMID:11438969

  20. Modelling of ceramide interactions with porous graphite carbon in non-aqueous liquid chromatography.

    PubMed

    West, C; Cilpa, G; Gaudin, K; Chaminade, P; Lesellier, E

    2005-09-16

    Interactions of solutes on porous graphitic carbon (PGC) with non-aqueous mobile phases are studied by the linear solvation energy relationship (LSER). Studies have been carried out with eight binary mixtures composed of a weak solvent (acetonitrile or methanol) and a strong solvent (tetrahydrofuran, n-butanol, CH2Cl2, 1,1,2-trichloro-2,2,1-trifluoroethane). The systematic analysis of a set of test compounds was performed for each solvent mixture in isocratic mode (50:50). The results were compared to those obtained on PGC with hydro-organic liquids and supercritical fluids. They were then correlated with the observed retention behaviour of lipid compounds, more particularly ceramides. PMID:16130700

  1. Salts employed in hydrophobic interaction chromatography can change protein structure - insights from protein-ligand interaction thermodynamics, circular dichroism spectroscopy and small angle X-ray scattering.

    PubMed

    Komaromy, Andras Z; Kulsing, Chadin; Boysen, Reinhard I; Hearn, Milton T W

    2015-03-01

    Key requirements of protein purification by hydrophobic interaction chromatography (HIC) are preservation of the tertiary/quaternary structure, maintenance of biological function, and separation of the correctly folded protein from its unfolded forms or aggregates. This study examines the relationship between the HIC retention behavior of hen egg white lysozyme (HEWL) in high concentrations of several kosmotropic salts and its conformation, assessed by circular dichroism (CD) spectroscopy. Further, the physicochemical properties of HEWL in the presence of high concentrations of ammonium sulfate, sodium chloride and magnesium chloride were investigated by small angle X-ray scattering (SAXS) at different temperatures. Radii of gyration were extrapolated from Guinier approximations and the indirect transform program GNOM with protein-protein interaction and contrast variation taken into account. A bead model simulation provided information on protein structural changes using ab initio reconstruction with GASBOR. These results correlated to the secondary structure content obtained from CD spectroscopy of HEWL. These changes in SAXS and CD data were consistent with heat capacity ΔCp -values obtained from van't Hoff plot analyses of the retention data. Collectively, these insights enable informed decisions to be made on the choice of chromatographic conditions, leading to improved separation selectivity and opportunities for innovative column-assisted protein refolding methods. PMID:25690783

  2. Interaction of L-glutamate oxidase with triazine dyes: selection of ligands for affinity chromatography.

    PubMed

    Katsos, N E; Labrou, N E; Clonis, Y D

    2004-08-01

    Glutamate oxidase (GOX, EC 1.4.3.11) from Streptomyces catalyses the oxidation of L-glutamate to alpha-ketoglutarate. Its kinetic constants for L-glutamate were measured equal to 2 mM for Km and 85.8 s(-1) for kcat. BLAST search and amino acid sequence alignments revealed low homology to other L-amino acid oxidases (18-38%). Threading methodology, homology modeling and CASTp analysis resulted in certain conclusions concerning the structure of catalytic alpha-subunit and led to the prediction of a binding pocket that provides favorable conditions of accommodating negatively charged aromatic ligands, such as sulphonated triazine dyes. Eleven commercial textile dyes and four biomimetic dyes or minodyes, bearing a ketocarboxylated-structure as their terminal biomimetic moiety, immobilized on cross-linked agarose gel. The resulted mini-library of affinity adsorbents was screened for binding and eluting L-glutamate oxidase activity. All but Cibacron Blue 3GA (CB3GA) affinity adsorbents were able to bind GOX at pH 5.6. One immobilized minodye-ligand, bearing as its terminal biomimetic moiety p-aminobenzyloxanylic acid (BM1), displayed the higher affinity for GOX. Kinetic inhibition studies showed that BM1 inhibits GOX in a non-competitive manner with a Ki of 10.5 microM, indicating that the dye-enzyme interaction does not involve the substrate-binding site. Adsorption equilibrium data, obtained from a batch system with BM1 adsorbent, corresponded well to the Freundlich isotherm with a rate constant k of 2.7 mg(1/2)ml(1/2)/g and Freundlich isotherm exponent n of 1. The interaction of GOX with the BM1 adsorbent was further studied with regards to adsorption and elution conditions. The results obtained were exploited in the development of a facile purification protocol for GOX, which led to 335-fold purification in a single step with high enzyme recovery (95%). The present purification procedure is the most efficient reported so far for L-glutamate oxidase. PMID:15203041

  3. Assessment of estrogen receptor-monoclonal antibody interaction by high performance liquid chromatography

    SciTech Connect

    Brandt, D.W.; Wittliff, J.L.

    1986-05-01

    To define the interrelationships between the various isoforms of the estrogen receptors (ER), a monoclonal antibody-horse radish peroxidase conjugate H222 was used as a probe in conjunction with HPIEC (Synchrom AX-1000) and HPSEC (TSK-3000 SW Toyo Soda) columns. ER from breast tumors was assessed using (16..cap alpha..-/sup 125/I)-iodoestradiol-17..beta.. (3nM) +/-200 fold excess estradiol-17..beta.. and excess H222. When ER was analyzed by HPSEC (size and shape), with 400 mM KCl which caused the dissociation of ER into 4S isoforms, a shift in retension time to higher molecular weight species was seen. The H222 appeared to interact with most isoforms of ER. However, when ER was analyzed by HPIEC (surface charge) with H222, a shift in virtually all of the high salt (150mM) isoform to the flow-through was observed with only 46% shift in elution of the low salt (60-70mM) isoforms. H222 did not alter total ER binding capacity. These data suggest that H222 recognized discrete forms of the ER. Therefore, modification in the receptor may have occurred which masks or removes the antigenic determinant limiting the specificity of H222. These results indicate that H222 may be employed as a tool to elucidate the interrelationships between these ER species.

  4. Spectroscopic characterization of ethyl xanthate oxidation products and analysis by ion interaction chromatography

    PubMed

    Hao; Silvester; Senior

    2000-10-15

    An ion interaction chromatographic separation method, coupled with UV spectroscopic detection, has been developed for the analysis of ethyl xanthate (O-ethyl dithiocarbonate) and its oxidative decomposition products in mineral flotation systems. The effects of the ion-pairing agent (tetrabutylammonium ion), pH modifier (phosphoric acid), and organic modifier (acetonitrile) in the eluant upon the retention characteristics of the ethyl xanthate oxidation products have been determined. The optimized separation procedure has been successfully applied to the analysis of ethyl xanthate and its oxidation products in a nickel-iron sulfide mineral suspension containing a number of other anionic species, including cyanide complexes of nickel and iron, as well as sulfur-oxy anions. The ethyl xanthate oxidation products investigated in this study have been isolated as pure compounds and characterized by UV-visible, FT-IR, and NMR spectroscopies. The UV-visible and FT-IR spectroscopic properties of these species are discussed in terms of the chemical modifications of the thiocarbonate group. PMID:11055697

  5. Simultaneous determination of residues of dipyrone metabolites in goat tissues by hydrophilic interaction liquid chromatography tandem mass spectrometry.

    PubMed

    Zhang, Chong; Zhang, Lifang; Cao, Suqing; Jiang, Zhaoling; Wu, Hao; Yan, Ming; Zhang, Xiaoxiao; Jiang, Shanxiang; Xue, Feiqun

    2016-04-01

    A reliable LC-MS/MS method with high sensitivity was developed and validated for the determination of dipyrone (DIP) metabolites in goat muscle, fat, liver, and kidney samples. Analytes were extracted using acetonitrile mixed with ammonia solution. After dehydration and evaporation to dryness, extracts were purified using an Oasis MAX cartridge. Chromatographic separation was performed on a hydrophilic interaction liquid chromatography column. The analytes were then detected using triple-quadrupole mass spectrometry in positive electrospray ionization and multiple reaction monitoring mode. Calibration plots were constructed using matrix-matched standards and showed good linearity. Limits of quantification for 4-methylaminoantipyrine (MAA), 4-formylaminoantipyrine (FAA), and 4-acetylaminoantipyrone (AAA) ranged from 0.4 μg kg(-1) to 6 μg kg(-1), while those for 4-aminoantipyrone (AA) ranged from 10 μg kg(-1) to 125 μg kg(-1) in all tissues. The developed method was successfully applied in the determination of DIP metabolite residues in actual goat tissues. PMID:26593468

  6. Hydrophilic interaction liquid chromatography-electrospray ionization-tandem mass spectrometry of a complex mixture of native and oxidized phospholipids.

    PubMed

    Losito, I; Facchini, L; Diomede, S; Conte, E; Megli, F M; Cataldi, T R I; Palmisano, F

    2015-11-27

    A mixture of native and oxidized phospholipids (PLs), generated by the soybean lipoxygenase type V-catalyzed partial oxidation of a lipid extract obtained from human platelets, was analyzed by Hydrophilic Interaction Liquid Chromatography-ElectroSpray Ionization-Tandem Mass Spectrometry (HILIC-ESI-MS/MS). The complexity of the resulting mixture was remarkable, considering that the starting lipid extract, containing (as demonstrated in a previous study) about 130 native PLs, was enriched with enzymatically generated hydroperoxylated derivatives and chemically generated hydroxylated forms of PLs bearing polyunsaturated side chains. Nonetheless, the described analytical approach proved to be very powerful; indeed, focusing on phosphatidylcolines (PCs), the most abundant PL class in human platelets, about fifty different native/oxidized species could be identified in a single HILIC-ESI-MS/MS run. Low-energy collision induced dissociation tandem MS (CID-MS/MS) experiments on chromatographically separated species showed single neutral losses of H2O2 and H2O to be typical fragmentation pathways of hydroperoxylated PCs, whereas a single H2O loss was observed for hydroxylated ones. Moreover, diagnostic losses of n-hexanal or n-pentanol were exploited to recognize PCs hydroperoxylated on the last but five carbon atom of a ɷ-6 polyunsaturated side chain. Despite the low resolution of the 3D ion trap mass analyzer used, the described HILIC-ESI-MS/MS approach appears very promising for the identification of oxidized lipids in oxidatively stressed complex biological systems. PMID:26508677

  7. Identification of novel isomeric pectic oligosaccharides using hydrophilic interaction chromatography coupled to traveling-wave ion mobility mass spectrometry.

    PubMed

    Leijdekkers, Antonius G M; Huang, Jie-Hong; Bakx, Edwin J; Gruppen, Harry; Schols, Henk A

    2015-03-01

    Separation and characterization of complex mixtures of pectic oligosaccharides still remains challenging and often requires the use of multiple analytical techniques, especially when isomeric structures are present. In this work, it is demonstrated that the coupling of hydrophilic interaction chromatography (HILIC) to traveling-wave ion mobility mass spectrometry (TWIMMS) enabled the simultaneous separation and characterization of complex mixtures of various isomeric pectic oligosaccharides. Labeling of oligosaccharides with 3-aminoquinoline (3-AQ) improved MS-ionization efficiency of the oligosaccharides and reduced the complexity of the product ion mass spectra, without losing resolution of the HILIC separation. In addition, labeling enabled quantification of oligosaccharides on molar basis using in-line fluorescence detection. Isomeric structures were distinguished using TWIMMS. The 3-AQ-HILIC-TWIMMS method was used to characterize a series of isomeric sugar beet rhamnogalacturonan I derived oligosaccharides carrying a glucuronic acid substituent. Thereby, some novel structural features were identified for the first time: glucuronic acid was attached to O-3 or to O-2 of galacturonic acid residues and a single galacturonic acid residue within an oligomer could contain both an acetyl group and a glucuronic acid substituent. PMID:25647688

  8. A hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS) based metabolomics study on colour stability of ovine meat.

    PubMed

    Subbaraj, Arvind K; Kim, Yuan H Brad; Fraser, Karl; Farouk, Mustafa M

    2016-07-01

    Meat colour is one of the cues available to the consumer to gauge overall meat quality and wholesomeness. Colour stability of meat is determined by several factors both inherent to the animal and post-slaughter conditions, including ageing, storage/packaging and display times. A hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS) based metabolomics study was undertaken to identify and compare polar metabolites between ovine meat samples that were exposed to different durations of ageing, storage conditions, and display times. Primary metabolites comprising amino acids, sugars, nucleotides, nucleosides, organic acids and their breakdown products were mainly identified as discriminating factors. For the first time, boron complexes of sugar and malic acid were also tentatively identified. As expected, most compounds identified were related to myoglobin chemistry, and compounds with antioxidant properties were found in higher levels in colour stable samples. Supplementary studies identifying semi-polar, non-polar and volatile compounds will provide a holistic understanding of the chemical basis of colour stability in ovine meat. PMID:26986230

  9. Identification and separation of saxitoxins using hydrophilic interaction liquid chromatography coupled to traveling wave ion mobility-mass spectrometry.

    PubMed

    Poyer, Salomé; Loutelier-Bourhis, Corinne; Coadou, Gaël; Mondeguer, Florence; Enche, Julien; Bossée, Anne; Hess, Philipp; Afonso, Carlos

    2015-01-01

    The aim of this work was to develop a reliable and efficient analytical method to characterise and differentiate saxitoxin analogues (STX), including sulphated (gonyautoxins, GTX) and non-sulphated analogues. For this purpose, hydrophilic interaction liquid chromatography (HILIC) was used to separate sulphated analogues. We also resorted to ion mobility spectrometry to differentiate the STX analogues because this technique adds a new dimension of separation based on ion gas phase conformation. Positive and negative ionisation modes were used for gonyautoxins while positive ionisation mode was used for non-sulphated analogues. Subsequently, the coupling of these three complementary techniques, HILIC-IM-MS, permitted the separation and identification of STX analogues; isomer differentiation was achieved in HILIC dimension while non-sulphated analogues were separated in the IM-MS dimension. Additional structural characteristics concerning the conformation of STXs could be obtained using IM-MS measurements. Thus, the collision cross sections (CCS) of STXs are reported for the first time in the positive ionisation mode. These experimental CCSs correlated well with the calculated CCS values using the trajectory method. PMID:25601690

  10. Stationary mutation models.

    PubMed

    Simonsson, Ivar; Mostad, Petter

    2016-07-01

    Probability calculations for relationship inference based on DNA tests are often performed with computer packages such as Familias. When mutations are assumed to be a possibility, one may notice a curious and problematic effect of including untested parents: results tend to change slightly. In this paper, we trace this effect back to fundamental model-formulating issues which can only be resolved by using stationary mutation models. We present several methods for obtaining such stationary mutation matrices from original mutation matrices, and evaluate essential properties of these methods. Our conclusion is that typically, stationary mutation models can be obtained, but for many types of markers, it may be impossible to combine specific biologically reasonable requirements for a mutation matrix with the requirement of stationarity. PMID:27231805

  11. Charged particle tunnels from the stationary and non-stationary Kerr-Newman black holes

    NASA Astrophysics Data System (ADS)

    Chen, Deyou; Yang, Shuzheng

    2007-09-01

    Considering the unfixed background space-time and self-gravitational interaction, we view the Hawking radiation of a stationary Kerr-Newman black hole by Hamilton-Jacobi method. Meanwhile, extending this work to non-stationary black holes, we attempt to investigate the Hawking radiation of the non-stationary Kerr-Newman black hole. Both of the results show the tunneling probabilities are related to the change of Bekenstein- Hawking entropy and the radiation spectrums deviate from the purely thermal one, which is in accordance with the known result.

  12. Comprehensive Two-Dimensional Hydrophilic Interaction Chromatography (HILIC) × Reversed-Phase Liquid Chromatography Coupled to High-Resolution Mass Spectrometry (RP-LC-UV-MS) Analysis of Anthocyanins and Derived Pigments in Red Wine.

    PubMed

    Willemse, Chandré M; Stander, Maria A; Vestner, Jochen; Tredoux, Andreas G J; de Villiers, André

    2015-12-15

    Changes in anthocyanin chemistry represent some of the most important transformations involved in red wine aging. However, accurate analysis of the derived pigments, as required to study the evolution of anthocyanins and tannins during aging, is hampered by their extreme structural diversity, low levels, and the fact that many of these compounds have identical mass spectral characteristics. In this context, chromatographic separation is critical. In this contribution, the application of online hydrophilic interaction chromatography (HILIC) × reversed-phase liquid chromatography (RP-LC) separation coupled to high-resolution mass spectrometry (MS) is described for the detailed characterization of anthocyanins and their derived pigments in aged red wine. A systematic approach was followed for the optimization of HILIC × RP-LC separation parameters using a capillary liquid chromatography (LC) system in the first dimension and an ultrahigh-pressure LC system in the second dimension to ensure maximum sensitivity and performance. Ninety four (94) anthocyanin-derived pigments were tentatively identified in one- and six-year-old Pinotage wines using accurate mass and fragmentation information obtained using quadrupole-time-of-flight mass spectrometry (Q-TOF-MS). Online HILIC × RP-LC-MS was found to offer high-resolution separation, because of the combination of two different separation modes, while the structured elution order observed improved the certainty in compound identification. Therefore, this approach shows promise for the detailed elucidation of the chemical alteration of anthocyanins during wine aging. PMID:26554292

  13. Mixed-mode chromatography in pharmaceutical and biopharmaceutical applications.

    PubMed

    Zhang, Kelly; Liu, Xiaodong

    2016-09-01

    Mixed-mode chromatography (MMC) is a fast growing area in recent years, thanks to the new generation of mixed-mode stationary phases and better understanding of multimode interactions. MMC has superior applications in the separation of compounds that are not retained or not well resolved by typical reversed-phase LC methods, especially for polar and charged molecules. Due to the multiple retention modes that a single MMC column can offer, often MMC provides additional dimension to a separation method by adjusting the mobile phase conditions. Mixed-mode media is also an effective way to clean up complex sample matrices for purification purposes or for sensitive detection of trace amounts of analytes. In this article, we discuss mixed-mode stationary phases and separation mechanisms and review recent advances in pharmaceutical and biopharmaceutical applications including the analysis and/or purification of counterions, small molecule drugs, impurities, formulation excipients, peptides and proteins. PMID:27236100

  14. Facile Preparation of Octadecyl Monoliths with Incorporated Carbon Nanotubes and Neutral Monoliths with Coated Carbon Nanotubes Stationary Phases for HPLC of Small and Large Molecules by Hydrophobic and π-π Interactions

    PubMed Central

    Mayadunne, Erandi; Rassi, Ziad El

    2014-01-01

    Two approaches for incorporating carbon nanotubes into monolithic columns for HPLC are described in this report. They pertain to the investigation of carbon nanotubes either (i) as entities to modulate solute retention on monolithic columns bearing well defined retentive ligands or (ii) as entities that constitute the stationary phase responsible for solute retention and separation. Approach (i) involved the incorporation of carbon nanotubes into octadecyl monolithic columns while approach (ii) concerns the preparation and evaluation of an ideal monolithic support and coating it with carbon nanotubes to yield a real “carbon nanotube stationary phase” for the HPLC separation of a wide range of solutes. First, an octadecyl monolithic column based on the in situ polymerization of octadecyl acrylate and trimethylolpropane trimethacrylate was optimized for use in HPLC separations of small and large solutes (e.g., proteins). To further modulate the retention and separation of proteins, small amounts of carbon nanotubes were incorporated into the octadecyl monolith column. In approach (ii), an inert, relatively polar monolith based on the in situ polymerization of glyceryl monomethacrylate (GMM) and ethylene glycol dimethacrylate (EDMA) proved to be the most suitable support for the preparation of “carbon nanotube stationary phase”. This carbon nanotube “coated” monolith proved useful in the HPLC separation of a wide range of small solutes including enantiomers. In approach (ii), a more homogeneous incorporation of carbon nanotubes into the diol monolithic columns (i.e., GMM/EDMA) was achieved when hydroxyl functionalized carbon nanotubes were incorporated into the GMM/EDMA monolithic support. In addition, high power sonication for a short time enhanced further the homogeneity of the monolith incorporated with nanotubes. In all cases, nonpolar and π interactions were responsible for solute retention on the monolith incorporated carbon nanotubes. PMID:25127634

  15. Facile preparation of octadecyl monoliths with incorporated carbon nanotubes and neutral monoliths with coated carbon nanotubes stationary phases for HPLC of small and large molecules by hydrophobic and π-π interactions.

    PubMed

    Mayadunne, Erandi; El Rassi, Ziad

    2014-11-01

    Two approaches for incorporating carbon nanotubes into monolithic columns for HPLC are described in this report. They pertain to the investigation of carbon nanotubes either (i) as entities to modulate solute retention on monolithic columns bearing well defined retentive ligands or (ii) as entities that constitute the stationary phase responsible for solute retention and separation. Approach (i) involved the incorporation of carbon nanotubes into octadecyl monolithic columns while approach (ii) concerns the preparation and evaluation of an ideal monolithic support and coating it with carbon nanotubes to yield a real "carbon nanotube stationary phase" for the HPLC separation of a wide range of solutes. First, an octadecyl monolithic column based on the in situ polymerization of octadecyl acrylate and trimethylolpropane trimethacrylate was optimized for use in HPLC separations of small and large solutes (e.g., proteins). To further modulate the retention and separation of proteins, small amounts of carbon nanotubes were incorporated into the octadecyl monolith column. In approach (ii), an inert, relatively polar monolith based on the in situ polymerization of glyceryl monomethacrylate (GMM) and ethylene glycol dimethacrylate (EDMA) proved to be the most suitable support for the preparation of "carbon nanotube stationary phase". This carbon nanotube "coated" monolith proved useful in the HPLC separation of a wide range of small solutes including enantiomers. In approach (ii), a more homogeneous incorporation of carbon nanotubes into the diol monolithic columns (i.e., GMM/EDMA) was achieved when hydroxyl functionalized carbon nanotubes were incorporated into the GMM/EDMA monolithic support. In addition, high power sonication for a short time enhanced further the homogeneity of the monolith incorporated with nanotubes. In all cases, nonpolar and π interactions were responsible for solute retention on the monolith incorporated carbon nanotubes. PMID:25127634

  16. Evaluation and comparison of the kinetic performance of ultra-high performance liquid chromatography and high-performance liquid chromatography columns in hydrophilic interaction and reversed-phase liquid chromatography conditions.

    PubMed

    Song, Huiying; Adams, Erwin; Desmet, Gert; Cabooter, Deirdre

    2014-11-21

    An intrinsic performance comparison is made of the reduction in analysis time that can be obtained when switching from HPLC to UHPLC column formats in HILIC and reversed-phase conditions. A detailed overview of the packing characteristics of both stationary phase types is given first. It is demonstrated that HILIC columns demonstrate higher external porosity values than their reversed-phase counterparts resulting in lower flow resistance values. Column total porosity values determined from the elution time of a small marker molecule are shown to depend strongly on the composition of the mobile phase. To omit errors that might arise from an over- or underestimation of the column void time, all plate height and kinetic plot data are therefore expressed as a function of the interstitial velocity. Although only a limited number of columns are evaluated in this study, it is shown that the column efficiency of the HILIC columns is lower than that of their reversed-phase counterparts, at least for the compounds evaluated here. Despite this lower efficiency, the kinetic performance of both stationary phase types is similar, due to the much lower viscosity of the mobile phases typically used in HILIC conditions. Finally, it is demonstrated that a similar, yet slightly larger reduction in analysis time can be obtained when switching from HPLC column formats to UHPLC formats in HILIC compared to reversed-phase conditions. PMID:25441074

  17. Applying Chromatography.

    ERIC Educational Resources Information Center

    Klein, Jessie W.; Patev, Paul

    1998-01-01

    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)

  18. Robust analysis of underivatized free amino acids in soil by hydrophilic interaction liquid chromatography coupled with electrospray tandem mass spectrometry.

    PubMed

    Gao, Jiajia; Helmus, Rick; Cerli, Chiara; Jansen, Boris; Wang, Xiang; Kalbitz, Karsten

    2016-06-01

    Amino acids are an important and highly dynamic fraction of organic N in soils and their determination in soil without derivatization is challenging due to the difficulties in separation and detection of trace amounts of these polar analytes. In the present work, we developed an analytical method to quantify 20 free amino acids in aqueous soil extracts without derivatization. The method employed hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) technique combined with a cation exchange solid phase extraction (SPE). Four stable isotope labelled amino acids were used as internal standards to improve the method performance. Good separation of 20 underivatized amino acids was achieved within 12min. The limit of detection (LODs) and limit of quantification (LOQs) were in the range of 13-384ngg(-1) and 43-1267ngg(-1) (dry soil basis), respectively. The results showed that overall recoveries with high precision were obtained for the extracted free amino acids from ten different soils. The overall recoveries of 18 amino acids were similar for the ten soils used, which differed substantially in organic C content and in other properties as soil texture and pH. For most of the amino acids, the average recoveries from soil extracts were between 74% and 117%, with the exception of Met (31%), Pro (52%) and Arg (68%). Variability was within acceptable limits (relative standard deviations were between 4% and 13%), with the exception of Met (relative standard deviation=90%) and Arg (relative standard deviation=53%). Thus the proposed method with high throughout and high analyte specificity shows great promise for consistent analysis of free amino acids extracted from soils and offers new horizons for the analysis of amino acids in terrestrial and aquatic ecosystem. PMID:27157424

  19. Use of quantitative affinity chromatography for characterizing high-affinity interactions: binding of heparin to antithrombin III.

    PubMed

    Hogg, P J; Jackson, C M; Winzor, D J

    1991-02-01

    The versatility of quantitative affinity chromatography (QAC) for evaluating the binding of macromolecular ligands to macromolecular acceptors has been increased substantially as a result of the derivation of the equations which describe the partitioning of acceptor between matrix-bound and soluble forms in terms of total, rather than free, ligand concentrations. In addition to simplifying the performance of the binding experiments, this development makes possible the application of the technique to systems characterized by affinities higher than those previously amenable to investigation by QAC. Addition of an on-line data acquisition system to monitor the concentration of partitioning solute in the liquid phase as a function of time has permitted the adoption of an empirical approach for determining the liquid-phase concentration of acceptor in the system at partition equilibrium, a development which decreases significantly the time required to obtain a complete binding curve by QAC. The application of these new QAC developments is illustrated by the determination of binding constants for the interactions of high-affinity heparin (Mr 20,300) with antithrombin III at three temperatures. Association constants of 8.0 +/- 2.2 x 10(7), 3.4 +/- 0.3 x 10(7), and 1.0 +/- 0.2 x 10(7) M-1 were observed at 15, 25, and 35 degrees C, respectively. The standard enthalpy change of -4.2 +/- 0.6 kcal/mol that is calculated from these data is in good agreement with a reported value obtained from fluorescence quenching measurements. PMID:2035830

  20. Special Report: Affinity Chromatography.

    ERIC Educational Resources Information Center

    Parikh, Indu; Cuatrecasas, Pedro

    1985-01-01

    Describes the nature of affinity chromatography and its use in purifying enzymes, studying cell interactions, exploring hormone receptors, and other areas. The potential the technique may have in treating disease is also considered. (JN)

  1. Separation of intact proteins on γ-ray-induced polymethacrylate monolithic columns: A highly permeable stationary phase with high peak capacity for capillary high-performance liquid chromatography with high-resolution mass spectrometry.

    PubMed

    Simone, Patrizia; Pierri, Giuseppe; Foglia, Patrizia; Gasparrini, Francesca; Mazzoccanti, Giulia; Capriotti, Anna Laura; Ursini, Ornella; Ciogli, Alessia; Laganà, Aldo

    2016-01-01

    Polymethacrylate-based monolithic capillary columns, prepared by γ-radiation-induced polymerization, were used to optimize the experimental conditions (nature of the organic modifiers, the content of trifluoroacetic acid and the column temperature) in the separation of nine standard proteins with different hydrophobicities and a wide range of molecular weights. Because of the excellent permeability of the monolithic columns, an ion-pair reversed-phase capillary liquid chromatography with high-resolution mass spectrometry method has been developed by coupling the column directly to the mass spectrometer without a flow-split and using a standard electrospray interface. Additionally, the high working flow and concomitant high efficiency of these columns allowed us to employ a longer column (up to 50 cm) and achieve a peak capacity value superior to 1000. This work is motivated by the need to develop new materials for high-resolution chromatographic separation that combine chemical stability at elevated temperatures (up to 75°C) and a broad pH range, with a high peak capacity value. The advantage of the γ-ray-induced monolithic column lies in the batch-to-batch reproducibility and long-term high-temperature stability. Their proven high loading capacity, recovery, good selectivity and high permeability, moreover, compared well with that of a commercially available poly(styrene-divinylbenzene) monolithic column, which confirms that such monolithic supports might facilitate analysis in proteomics. PMID:26530449

  2. Evaluation of two sub-2 μm stationary phases, core-shell and totally porous monodisperse, in the second dimension of on-line comprehensive two dimensional liquid chromatography, a case study: separation of milk peptides after expiration date.

    PubMed

    Sommella, Eduardo; Pepe, Giacomo; Ventre, Giovanni; Pagano, Francesco; Manfra, Michele; Pierri, Giuseppe; Ismail, Omar; Ciogli, Alessia; Campiglia, Pietro

    2015-01-01

    Milk is a rich source of bioactive peptides of great interest for their healthy properties. These peptides are usually encrypted in the sequences of proteins and are released after time dependent proteolysis as very complex hydrolysates. In order to separate and identify the bioactive sequences, we developed an on-line comprehensive two dimensional liquid chromatography approach using the high performance combined with the ultra high performance conditions. A microbore reversed phase (C18 silica, 5 μm) column was employed in first dimension, while, in second dimension, two different UHPLC columns, packed with C18 silica, were tested: a new column based on monodisperse sub-2 μm fully porous particles with high surface area (50 mm × 3.0 mm, 1.9 μm d.p., from Supelco), and a column based on sub-2 μm core-shell particles (50 mm × 3.0 mm, 1.7μm d.p, from Phenomenex(®)). Both set-ups were compared, showing high peak capacity values with respect to a high efficiency monodimensional method, maintaining the same analysis time. Satisfactory selectivity was obtained through the use of different pH between the two dimension, while a very fast continuous shifted gradient in second dimension ensured a good employment of the 2D separation space. PMID:25498555

  3. Parallel, fluorous open-tubular chromatography using microstructured fibers.

    PubMed

    Daley, Adam B; Wright, Ramin D; Oleschuk, Richard D

    2011-04-01

    Although commonly used in gas chromatography, open-tubular columns for liquid chromatography have seen their development hindered by a number of factors both theoretical and practical. Requiring small diameters, great lengths and specialized detection systems to achieve a proper chromatographic response, columns of this sort have largely been ignored despite the highly desirable column performance an optimized system would provide. Here, we introduce the use of microstructured fibers (MSFs) as a platform for the development of multiplexed open-tubular liquid chromatography (OTLC) columns. The multiple, parallel silica channels presented by the MSF act as a promising substrate for an OTLC column, as they have diameters near the ideal range for interactions (1-3 μm), minimize flow-induced backpressure through their many uniform paths, and increase the loading capacity compared to a single capillary channel of similar size. Additionally, with outer diameters comparable to regular fused silica capillaries, MSFs can easily be employed in conventional chromatographic systems, eliminating the need for specialized equipment. Finally, MSF columns of this type can be functionalized using silane coupling techniques to allow the introduction of a wide variety of stationary phase chemistries. While in this report we explore the potential and limitations of fluorine-functionalized MSFs as OTLC columns, other stationary phase materials could easily be substituted by choosing appropriate silanization reagents. Particular attention here will be paid to the physical and performance characteristics of the fabricated columns, as well as avenues for their improvement and implementation. PMID:21435484

  4. Mutually supportive use of stable isotope and gas chromatography techniques to understand ecohydrological interactions in dryland environments

    NASA Astrophysics Data System (ADS)

    Puttock, A.; Brazier, R. E.; Dungait, J. A. J.; Bol, R.; Dixon, E. R.; Macleod, C. J. A.

    2012-04-01

    Many drylands globally are experiencing extensive vegetation change. In the semi-arid Southwestern United States, this change is characterised by the encroachment of woody vegetation into environments previously dominated by grassland (Van Auken. 2009). The transition from grass to woody vegetation results in a change in ecosystem structure and function (Turnbull et al. 2008). Structural change is typically characterised by an increased heterogeneity of soil and vegetation resources, associated with reduced vegetation coverage and an increased vulnerability to soil erosion and the potential loss of key nutrients to adjacent fluvial systems. This project uses an ecohydrological approach, monitoring natural rainfall-runoff events and resulting water and sediment fluxes over six bounded plots with different vegetation coverage at the Sevilleta National Wildlife Refuge, New Mexico, USA. The experiment takes advantage of a shift in the photosynthetic pathway of dominant vegetation from C3 piñon-juniper (Pinus edulis-Juniperus monosperma) mixed stand through a C4 pure-grass (Bouteloua eriopoda) to C3 shrub (Larrea tridentate). This allows for the utilisation of natural abundance tracing techniques, specifically stable 13C isotope and gas chromatography lipid biomarker analyses. Results collected during the 2010 and 2011 monsoon seasons will be presented, using biogeochemical signatures, to trace and partition fluvial soil organic matter and carbon fluxes during runoff generating rainfall events. Results show that biogeochemical signatures specific to individual plant species can be used to define the provenance of carbon, quantifying whether more Pinus edulis-Juniperus monosperma derived carbon is mobilised from the upland plots, or whether more Larrea tridentata carbon is lost when compared to bouteloa eripoda losses in the lowlands. Results also show that biogeochemical signatures vary with event characteristics, raising the possibility of using these tracing

  5. HYDROCARBON POLLUTANTS FROM STATIONARY SOURCES

    EPA Science Inventory

    The report gives results of a study of hydrocarbon pollutants from stationary sources. Early in the study, readily available information was assembled on stationary sources of hydrocarbon emissions and effluents. Information was also obtained on process descriptions, operating pa...

  6. Relaminarization under stationary vortices

    NASA Astrophysics Data System (ADS)

    Breidenthal, Robert

    2005-11-01

    Flow visualization reveals that a turbulent boundary layer is relaminarized when stationary streamwise vortices are introduced. Following a suggestion of Balle, the vortices are stabilized by large streamwise ``Karman'' grooves in a wavy wall. In a water tunnel, upstream vortex generators place a large streamwise vortex in the middle of each groove, at the stationary point where Prandtl's vortex force vanishes. According to a theory by Cotel, the wall fluxes of a turbulent boundary layer should decline to laminar values under such ``persistent'' vortices. The observed relaminarization is consistent with this theory and with previous measurements of heat transfer by Touel and Balle. However, the structure of the transverse flow resembles the cats-eye pattern of a temporal shear layer rather than the anticipated von Karman wake. The cats-eye pattern corresponds to the forced shear layers of Oster-Wygnanski and Roberts, who found that the Reynolds stresses and mixing rate also decline to laminar values.

  7. Size-exclusion capillary electrochromatographic separation of polysaccharides using polymeric stationary phases.

    PubMed

    Mistry, Kavita; Krull, Ira; Grinberg, Nelu

    2003-06-01

    We report the successful size-based separations of large, neutral polysaccharides using capillary electrochromatography (CEC). As the polysaccharides possessed little chromophore for photometric detection, two separate approaches were taken. In the first approach, indirect detection was combined with size-exclusion chromatography using a sulfonated polystyrene/divinylbenzene stationary phase. The separations were performed using a 300 A pore size stationary phase under aqueous conditions. Non-size based interactions were minimal using this material, resulting in an effective calibration range of molecular masses 180 to 112 000 g.mol(-1) for pullulans. In the second approach, the polysaccharides were derivatized with phenylisocyanate and were subsequently separated on columns made using a combination of high capacity ion-exchanger and a neutral polystyrene/divinylbenzene material of various pore sizes. The sulfonated ion-exchange phase provided the electroosmotic flow, while the mixed pore size material provided the extended calibration range. The linear range for this primarily nonaqueous system using tetrahydrofuran was determined to be from molecular masses 738 to 404 000 g.mol(-1) of the original, untagged pullulan. This approach overcame the limited solubility issue associated with analysis of some polysaccharides. Analysis of pullulan and amylose samples by CEC correlated well with results obtained by conventional high-performance liquid chromatography (HPLC). The size-exclusion electrochromatographic separations provide an alternative mode for determining the relative molecular weights of polysaccharides with reduced sample and solvent consumption, as well as analysis times. PMID:12783452

  8. Global characterization of neutral saccharides in crude and processed Radix Rehmanniae by hydrophilic interaction liquid chromatography tandem electrospray ionization time-of-flight mass spectrometry.

    PubMed

    Liu, Zhirui; Lou, Ziyang; Ding, Xuan; Li, Xiang; Qi, Yunpeng; Zhu, Zhenyu; Chai, Yifeng

    2013-12-01

    The combined use of the state-of-the-art hybrid mass spectrometer (MS) together with high efficient liquid chromatography may prove to be a useful tool for neutral saccharide analysis. In the present work, we used hydrophilic interaction liquid chromatography (HILIC) to separate selected saccharides (fructose, glucose, galactose, sucrose, melibiose, raffinose, manninotriose, stachyose and verbascose). Influences of the column type, additive type, temperature, pH and other separation factors on analyte retention were evaluated. The new method did not require reduction, derivatization or postcolumn addition of reagents, which are commonly used in conventional saccharide analysis. Our results demonstrate the potential of HILIC-MS for sensitive and robust determination of saccharides in crude and processed Radix Rehmanniae, and may promote new perspectives in the research of other medicinal herbs. PMID:23871031

  9. Separation and analysis of phenolic acids from Salvia miltiorrhiza and its related preparations by off-line two-dimensional hydrophilic interaction chromatography×reversed-phase liquid chromatography coupled with ion trap time-of-flight mass spectrometry.

    PubMed

    Sun, Wanyang; Tong, Ling; Miao, Jingzhuo; Huang, Jingyi; Li, Dongxiang; Li, Yunfei; Xiao, Hongting; Sun, Henry; Bi, Kaishun

    2016-01-29

    Salvia miltiorrhiza (SM) is one of the most widely used Traditional Chinese Medicine. Active constituents of SM mainly contain hydrophilic phenolic acids (PAs) and lipophilic tanshinones. However, due to the existing of multiple ester bonds and unsaturated bonds in the structures, PAs have numerous chemical conversion products. Many of them are so low-abundant that hard to be separated using conventional methods. In this study, an off-line two-dimensional liquid chromatography (2D-LC) method was developed to separate PAs in SM and its related preparations. In the first dimension, samples were fractionated by hydrophilic interaction chromatography (HILIC) (Acchrom×Amide, 4.6×250mm, 5μm) mainly based on the hydrogen bonding effects. The fractions were then separated on reversed-phase liquid chromatography (RP-LC) (Acquity HSS T3, 2.1×50mm, 1.7μm) according to hydrophobicity. For the selective identification of PAs, diode array detector (DAD) and electrospray ionization tandem ion trap time-of-flight mass spectrometry (ESI-IT-TOF-MS) were employed. Practical and effective peak capacities of all the samples were greater than 2046 and 1130, respectively, with the orthogonalities ranged from 69.7% to 92.8%, which indicated the high efficiency and versatility of this method. By utilizing the data post-processing techniques, including mass defect filter, neutral loss filter and product ion filter, a total of 265 compounds comprising 196 potentially new PAs were tentatively characterized. Twelve kinds of derivatives, mainly including glycosylated compounds, O-alkylated compounds, condensed compounds and hydrolyzed compounds, constituted the novelty of the newly identified PAs. The HILIC×RP-LC/TOF-MS system expanded our understanding on PAs of S. miltiorrhiza and its related preparations, which could also benefit the separation and characterization of polar constituents in complicated herbal extracts. PMID:26792448

  10. Development of a hydrophilic interaction liquid chromatography-mass spectrometry method for detection and quantification of urea thermal decomposition by-products in emission from diesel engine employing selective catalytic reduction technology.

    PubMed

    Yassine, Mahmoud M; Dabek-Zlotorzynska, Ewa; Celo, Valbona

    2012-03-16

    The use of urea based selective catalytic reduction (SCR) technology for the reduction of NOx from the exhaust of diesel-powered vehicles has the potential to emit at least six thermal decomposition by-products, ammonia, and unreacted urea from the tailpipe. These compounds may include: biuret, dicyandiamine, cyanuric acid, ammelide, ammeline and melamine. In the present study, a simple, sensitive and reliable hydrophilic interaction liquid chromatography (HILIC)-electrospray ionization (ESI)/mass spectrometry (MS) method without complex sample pre-treatment was developed for identification and determination of urea decomposition by-products in diesel exhaust. Gradient separation was performed on a SeQuant ZIC-HILIC column with a highly polar zwitterionic stationary phase, and using a mobile phase consisting of acetonitrile (eluent A) and 15 mM ammonium formate (pH 6; eluent B). Detection and quantification were performed using a quadrupole ESI/MS operated simultaneously in negative and positive mode. With 10 μL injection volume, LODs for all target analytes were in the range of 0.2-3 μg/L. The method showed a good inter-day precision of retention time (RSD<0.5%) and peak area (RSD<3%). Satisfactory extraction recoveries from spiked blanks ranged between 96 and 98%. Analyses of samples collected during transient chassis dynamometer tests of a bus engine equipped with a diesel particulate filter (DPF) and urea based SCR technology showed the presence of five target analytes with cyanuric acid and ammelide the most abundant compounds in the exhaust. PMID:22318005

  11. Affinity chromatography: a historical perspective.

    PubMed

    Hage, David S; Matsuda, Ryan

    2015-01-01

    Affinity chromatography is one of the most selective and versatile forms of liquid chromatography for the separation or analysis of chemicals in complex mixtures. This method makes use of a biologically related agent as the stationary phase, which provides an affinity column with the ability to bind selectively and reversibly to a given target in a sample. This review examines the early work in this method and various developments that have lead to the current status of this technique. The general principles of affinity chromatography are briefly described as part of this discussion. Past and recent efforts in the generation of new binding agents, supports, and immobilization methods for this method are considered. Various applications of affinity chromatography are also summarized, as well as the influence this field has played in the creation of other affinity-based separation or analysis methods. PMID:25749941

  12. Ceramic stationary gas turbine

    SciTech Connect

    Roode, M. van

    1995-10-01

    The performance of current industrial gas turbines is limited by the temperature and strength capabilities of the metallic structural materials in the engine hot section. Because of their superior high-temperature strength and durability, ceramics can be used as structural materials for hot section components (blades, nozzles, combustor liners) in innovative designs at increased turbine firing temperatures. The benefits include the ability to increase the turbine inlet temperature (TIT) to about 1200{degrees}C ({approx}2200{degrees}F) or more with uncooled ceramics. It has been projected that fully optimized stationary gas turbines would have a {approx}20 percent gain in thermal efficiency and {approx}40 percent gain in output power in simple cycle compared to all metal-engines with air-cooled components. Annual fuel savings in cogeneration in the U.S. would be on the order of 0.2 Quad by 2010. Emissions reductions to under 10 ppmv NO{sub x} are also forecast. This paper describes the progress on a three-phase, 6-year program sponsored by the U.S. Department of Energy, Office of Industrial Technologies, to achieve significant performance improvements and emissions reductions in stationary gas turbines by replacing metallic hot section components with ceramic parts. Progress is being reported for the period September 1, 1994, through September 30, 1995.

  13. Ceramic stationary gas turbine

    SciTech Connect

    Roode, M. van

    1995-12-31

    The performance of current industrial gas turbines is limited by the temperature and strength capabilities of the metallic structural materials in the engine hot section. Because of their superior high-temperature strength and durability, ceramics can be used as structural materials for hot section components (blades, nozzles, combustor liners) in innovative designs at increased turbine firing temperatures. The benefits include the ability to increase the turbine inlet temperature (TIT) to about 1200{degrees}C ({approx}2200{degrees}F) or more with uncooled ceramics. It has been projected that fully optimized stationary gas turbines would have a {approx}20 percent gain in thermal efficiency and {approx}40 percent gain in output power in simple cycle compared to all metal-engines with air-cooled components. Annual fuel savings in cogeneration in the U.S. would be on the order of 0.2 Quad by 2010. Emissions reductions to under 10 ppmv NO{sub x} are also forecast. This paper describes the progress on a three-phase, 6-year program sponsored by the U.S. Department of Energy, Office of Industrial Technologies, to achieve significant performance improvements and emissions reductions in stationary gas turbines by replacing metallic hot section components with ceramic parts. Progress is being reported for the period September 1, 1994, through September 30, 1995.

  14. Combined effects of potassium chloride and ethanol as mobile phase modulators on hydrophobic interaction and reversed-phase chromatography of three insulin variants.

    PubMed

    Johansson, Karolina; Frederiksen, Søren S; Degerman, Marcus; Breil, Martin P; Mollerup, Jørgen M; Nilsson, Bernt

    2015-02-13

    The two main chromatographic modes based on hydrophobicity, hydrophobic interaction chromatography (HIC) and reversed-phase chromatography (RPC), are widely used for both analytical and preparative chromatography of proteins in the pharmaceutical industry. Despite the extensive application of these separation methods, and the vast amount of studies performed on HIC and RPC over the decades, the underlying phenomena remain elusive. As part of a systematic study of the influence of mobile phase modulators in hydrophobicity-based chromatography, we have investigated the effects of both KCl and ethanol on the retention of three insulin variants on two HIC adsorbents and two RPC adsorbents. The focus was on the linear adsorption range, separating the modulator effects from the capacity effects, but some complementary experiments at higher load were included to further investigate observed phenomena. The results show that the modulators have the same effect on the two RPC adsorbents in the linear range, indicating that the modulator concentration only affects the activity of the solute in the mobile phase, and not that of the solute-ligand complex, or that of the ligand. Unfortunately, the HIC adsorbents did not show the same behavior. However, the insulin variants displayed a strong tendency toward self-association on both HIC adsorbents; on one in particular. Since this causes peak fronting, the retention is affected, and this could probably explain the lack of congruity. This conclusion was supported by the results from the non-linear range experiments which were indicative of double-layer adsorption on the HIC adsorbents, while the RPC adsorbents gave the anticipated increased tailing at higher load. PMID:25595534

  15. Triangular Helical Column for Centrifugal Countercurrent Chromatography.

    PubMed

    Ito, Yoichiro; Yu, Henry

    2009-01-01

    Effective column space and stationary phase retention have been improved by changing the configuration of the helical column originally used for toroidal coil countercurrent chromatography. The use of an equilateral triangular core for the helix column doubles effective column space and retains the stationary phase over 40% of the total column capacity without increasing the column pressure. The present results suggest that the stationary phase retention and the peak resolution will be further improved using new column designs fabricated by a new technology called "laser sintering for rapid prototyping." PMID:20046940

  16. Protein separation and enrichment by counter-current chromatography using reverse micelle solvent systems.

    PubMed

    Shen, Ching-Wei; Yu, Tiing

    2007-06-01

    A protein mixture consisting of myoglobin, cytochrome c, and lysozyme was separated by high-speed counter-current chromatography using a two-phase aqueous/reverse micelle-containing organic solvent system. About 50% stationary phase retention ratio was obtained in most chromatographic experiments. Separations were manipulated mainly by pH gradients that controlled the electrostatic interactions between the protein molecules and reverse micelles. Separations were further improved by incorporating an ionic strength gradient along with the pH gradient. Control of ionic strength in the aqueous solution helped fine-tune protein partitioning between the stationary and mobile phases. Although non-specific protein interactions affected baseline resolution, recovery of cytochrome c and lysozyme reached 90% and 82%. Furthermore, concentration or enrichment of these two proteins was achieved from a large-volume sample load. This technique can potentially be employed in the recovery and enrichment of proteins from large-volume aqueous solutions. PMID:17289061

  17. Stationary engineering handbook

    SciTech Connect

    Petrocelly, K.L.

    1989-01-01

    Years ago, the only qualifications you needed to become to become an operating engineer were the ability to shovel large chunks of coal through small furnace doors and the fortitude to sweat profusely for hours without fainting. As a consequence of technological evolution, the engineer's coal shovels have been replaced with computers and now perspiration is more the result of job stress than exposure to high temperatures. The domain of the operator has been extended far beyond the smoke-filled caverns that once encased him, out into the physical plant, and his responsibilities have been expanded accordingly. Unlike his less sophisticated predecessor, today's technician must be well versed in all aspects of the operation. The field of power plant operations has become a full-fledged profession and its principals are called Stationary Engineers. This book addresses the areas of responsibility and the education and skills needed for successful operation of building services equipment.

  18. Simultaneous Enrichment of Plasma Soluble and Extracellular Vesicular Glycoproteins Using Prolonged Ultracentrifugation-Electrostatic Repulsion-hydrophilic Interaction Chromatography (PUC-ERLIC) Approach*

    PubMed Central

    Sok Hwee Cheow, Esther; Hwan Sim, Kae; de Kleijn, Dominique; Neng Lee, Chuen; Sorokin, Vitaly; Sze, Siu Kwan

    2015-01-01

    Plasma glycoproteins and extracellular vesicles represent excellent sources of disease biomarkers, but laboratory detection of these circulating structures are limited by their relatively low abundance in complex biological fluids. Although intensive research has led to the development of effective methods for the enrichment and isolation of either plasma glycoproteins or extracellular vesicles from clinical materials, at present it is not possible to enrich both structures simultaneously from individual patient sample, a method that affords the identification of biomarker combinations from both entities for the prediction of clinical outcomes will be clinically useful. We have therefore developed an enrichment method for use in mass spectrometry-based proteomic profiling that couples prolonged ultracentrifugation with electrostatic repulsion-hydrophilic interaction chromatography, to facilitate the recovery of both glycoproteins and extracellular vesicles from nondepleted human plasma. Following prolonged ultracentrifugation, plasma glycoproteins and extracellular vesicles were concentrated as a yellow suspension, and simultaneous analyses of low abundant secretory and vesicular glycoproteins was achieved in a single LC-MS/MS run. Using this systematic prolonged ultracentrifugation-electrostatic repulsion-hydrophilic interaction chromatography approach, we identified a total of 127 plasma glycoproteins at a high level of confidence (FDR ≤ 1%), including 48 glycoproteins with concentrations ranging from pg to ng/ml. The novel enrichment method we report should facilitate future human plasma-based proteome and glycoproteome that will identify novel biomarkers, or combinations of secreted and vesicle-derived biomarkers, that can be used to predict clinical outcomes in human patients. PMID:25862729

  19. Affinity Chromatography.

    ERIC Educational Resources Information Center

    Gray, Gary R.

    1980-01-01

    Presents selected recent advances in immobilization chemistry which have important connections to affinity chromatography. Discusses ligand immobilization and support modification. Cites 51 references. (CS)

  20. A hydrophilic interaction liquid chromatography electrospray tandem mass spectrometry method for the simultaneous determination of γ-hydroxybutyrate and its precursors in forensic whole blood.

    PubMed

    Sørensen, Lambert K; Hasselstrøm, Jørgen B

    2012-10-10

    A liquid-chromatography-tandem-mass-spectrometry method using pneumatically assisted electrospray ionisation (LC-ESI-MS/MS) was developed for the simultaneous determination of γ-hydroxybutyric acid (GHB), γ-butyrolactone (GBL) and 1,4-butanediol (1,4-BD) in human ante-mortem and post-mortem whole blood. The blood proteins were precipitated using a mixture of methanol and acetonitrile, and the extract was cleaned-up by passage through a polymeric strong cation exchange sorbent. Separation of the analytes and their structural isomers was obtained using a column with a zwitterionic stationary phase. Matrix-matched calibrants, combined with isotope dilution, were used for quantitative analysis. GHB was determined in both positive and negative ion modes. The relative intra-laboratory reproducibility standard deviations were better than 10% and 6% for blood samples at concentrations of 2 mg/L and 20-150 mg/L, respectively. The mean true extraction recoveries were 80% for GHB and greater than 90% for GBL and 1,4-BD at concentration levels of 20-50 mg/L. The limits of detection were approximately 0.5 mg/L for GHB and GBL, and 0.02 mg/L for 1,4-BD in ante-mortem blood. The corresponding lower limits of quantification were less than 1 mg/L for GHB and GBL, and less than 0.1 mg/L for 1,4-BD. GBL was unstable in whole blood freshly preserved with a sodium fluoride oxalate mixture, but the stability could be improved significantly by preservation with a sodium fluoride citrate EDTA mixture. PMID:22917943

  1. Stationary Engineering Laboratory Manual--2.

    ERIC Educational Resources Information Center

    Steingress, Frederick M.; Frost, Harold J.

    The Stationary Engineering Laboratory Manual 2 was designed for vocational/technical high school students who have received instruction in the basics of stationary engineering. It was developed for students who will be operating a live plant and who will be responsible for supplying steam for heating, cooking, and baking. Each lesson in the manual…

  2. Stationary Engineering. Science Manual--2.

    ERIC Educational Resources Information Center

    Frost, Harold J.; Steingress, Frederick M.

    This second-year student manual contains 140 brief related science lessons applying science and math to trade activities in the field of stationary engineering. The lessons are organized into 16 units: (1) Introduction to Stationary Engineering, (2) Engineering Fundamentals, (3) Steam Boilers, (4) Boiler Fittings, (5) Boilerroom System, (6)…

  3. Enabling Microliquid Chromatography by Microbead Packing of Microchannels

    NASA Technical Reports Server (NTRS)

    Balvin, Manuel; Zheng, Yun

    2013-01-01

    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

  4. Enabling Microliquid Chromatography by Microbead Packing of Microchannels

    NASA Technical Reports Server (NTRS)

    Balvin, Manuel; Zheng, Yun

    2014-01-01

    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

  5. High perfomance liquid chromatography in pharmaceutical analyses.

    PubMed

    Nikolin, Branko; Imamović, Belma; Medanhodzić-Vuk, Saira; Sober, Miroslav

    2004-05-01

    complex electrostatic hydrophobic and steric interactions exist between the solute and both stationary and mobile phases. These enable the effective separation of samples of different nature. The main advantages of the use of a micelar solution in reversed-phase liquid chromatography are the solvent and the lower cost and toxicity, the biodegradability of the solvent and the easy dissolution of analytical samples, that enables the determination of drugs in physiological fluids without the need for previous separation of the proteins present in the samples. Using tetrabutylammonium phosphate as a competing base in the investigation of sulphonamides and heptanes sulfonate as ion pairing reagent. Ion pairing reagent is term used to describe enhanced retention as the result of the addition to the mobile phase of a large ion opposite charge to the molecular ions to be separated. For molecular cations alkyl sulphates or sulfonates are generally utilised. PMID:15629016

  6. Recent advances in cell membrane chromatography for traditional Chinese medicines analysis.

    PubMed

    Hou, Xiaofang; Wang, Sicen; Zhang, Tao; Ma, Jing; Zhang, Jie; Zhang, Yanmin; Lu, Wen; He, Huaizhen; He, Langchong

    2014-12-01

    Traditional Chinese medicines (TCMs) have been used for preventative care for thousands of years. The active components are the basis for the pharmacodynamics of TCMs, and they can be an important source of lead compounds. As a bioaffinity chromatography technique, cell membrane chromatography (CMC) has been developed for almost 20 years since 1996. It has been proven to be a useful method for studying drug-receptor interactions and screening active components from medicinal herbs. In our review in 2007 (Drug Discov. Ther., 1 (2007) 104-107), the preparation, identification, evaluation, and preliminary applications of CMC stationary phases were presented. In this article, we briefly review some of the latest progress and applications about CMC including instrument development, research on drug-receptor interactions, screening active components from TCMs, and quality control of TCMs. PMID:24972757

  7. Gas Chromatography.

    ERIC Educational Resources Information Center

    Karasek, Francis W.; And Others

    1984-01-01

    This review covers fundamental developments in gas chromatography during 1982 and 1983. Literature is considered under these headings: columns; liguid phases; solid supports; sorption processes and solvents; open tubular column gas chromatography; instrumentation; high-resolution columns and applications; other techniques; qualitative and…

  8. Identification of the AntiListerial Constituents in Partially Purified Column Chromatography Fractions of Garcinia kola Seeds and Their Interactions with Standard Antibiotics

    PubMed Central

    Penduka, D.; Buwa, L.; Mayekiso, B.; Basson, A. K.; Okoh, A. I.

    2014-01-01

    Partially purified fractions of the n-hexane extract of Garcinia kola seeds were obtained through column chromatography and their constituents were identified through the use of gas chromatography coupled to mass spectrometry (GC-MS). Three fractions were obtained by elution with benzene as the mobile phase and silica gel 60 as the stationery phase and these were named Benz1, Benz2, and Benz3 in the order of their elution. The antiListerial activities of these fractions were assessed through MIC determination and only Benz2 and Benz3 were found to be active with MIC's ranging from 0.625 to 2.5 mg/mL. The results of the GC-MS analysis showed Benz2 to have 9 compounds whilst Benz3 had 7 compounds, with the major compounds in both fractions being 9,19-Cyclolanost-24-en-3-ol, (3.β.) and 9,19-Cyclolanostan-3-ol,24-methylene-, (3.β.). The Benz2 fraction was found to have mainly indifferent interactions with ampicillin and penicillin G whilst mainly additive interactions were observed with ciprofloxacin. The Benz3 fraction's interactions were found to be 50% synergistic with penicillin G and 25% synergistic with ciprofloxacin and ampicillin. A commercially available 9,19-Cyclolanost-24-en-3-ol, (3.β.) was found not to exhibit any antiListerial activities at maximum test concentrations of 5 mg/mL, suggesting that the compound could be acting in synergy with the other compounds in the eluted fractions of Garcinia kola seeds. PMID:24527056

  9. Identification of the AntiListerial Constituents in Partially Purified Column Chromatography Fractions of Garcinia kola Seeds and Their Interactions with Standard Antibiotics.

    PubMed

    Penduka, D; Buwa, L; Mayekiso, B; Basson, A K; Okoh, A I

    2014-01-01

    Partially purified fractions of the n-hexane extract of Garcinia kola seeds were obtained through column chromatography and their constituents were identified through the use of gas chromatography coupled to mass spectrometry (GC-MS). Three fractions were obtained by elution with benzene as the mobile phase and silica gel 60 as the stationery phase and these were named Benz1, Benz2, and Benz3 in the order of their elution. The antiListerial activities of these fractions were assessed through MIC determination and only Benz2 and Benz3 were found to be active with MIC's ranging from 0.625 to 2.5 mg/mL. The results of the GC-MS analysis showed Benz2 to have 9 compounds whilst Benz3 had 7 compounds, with the major compounds in both fractions being 9,19-Cyclolanost-24-en-3-ol, (3. β .) and 9,19-Cyclolanostan-3-ol,24-methylene-, (3. β .). The Benz2 fraction was found to have mainly indifferent interactions with ampicillin and penicillin G whilst mainly additive interactions were observed with ciprofloxacin. The Benz3 fraction's interactions were found to be 50% synergistic with penicillin G and 25% synergistic with ciprofloxacin and ampicillin. A commercially available 9,19-Cyclolanost-24-en-3-ol, (3. β .) was found not to exhibit any antiListerial activities at maximum test concentrations of 5 mg/mL, suggesting that the compound could be acting in synergy with the other compounds in the eluted fractions of Garcinia kola seeds. PMID:24527056

  10. Mechanistic insights on chaotropic interactions of liophilic ions with basic pharmaceuticals in polar ionic mode liquid chromatography.

    PubMed

    Sanganyado, Edmond; Lu, Zhijiang; Gan, Jay

    2014-11-14

    We report for the first time the effect of liophilic mobile phase additives on the mechanism of chiral recognition of basic chiral pharmaceutical on a vancomycin based chiral stationary phase (CSP). Using methanol as the mobile phase and 0.005% formic acid as pH modifier, we evaluated the effect of different concentrations of three types of liophilic anions, formate (HCOO(-)), nitrate (NO3(-)), and acetate (CH3COO(-)), on enantioresolution (Rs), enantioselectivity (α) and retention factor (k) of enantiomers of fluoxetine and atenolol. The effect of liophilic ion types on k followed the Hofmeister series: CH3COO(-)>HCOO(-)>NO3(-). Increasing concentration from 4 to 20mM resulted in decreases in Rs and k in accordance to hydrophobicity of the liophilic anion. The effect of temperature or mobile phase composition on enantioseparation was determined at 13-40°C. For both analytes, standard changes in enthalpy (ΔH°) and entropy (ΔS°) calculated using van't Hoff plots (lnk against 1/T) to varied from -4.99 to -0.63 kJ/mol and -11.82 to 9.47 J/mol, respectively. The van't Hoff plots showed elution order of the enantiomers of each analyte did not reverse in the temperature range studied. Chiral recognition of the enantiomers of atenolol and fluoxetine in the presence of liophilic ions was enthalpy driven. PMID:25311487

  11. Mixed-mode anion-cation exchange/hydrophilic interaction liquid chromatography-electrospray mass spectrometry as an alternative to reversed phase for small molecule drug discovery

    PubMed

    Strege; Stevenson; Lawrence

    2000-10-01

    Within pharmaceutical drug discovery, significant needs currently exist for the analysis and purification of structurally diverse samples prior to or immediately following high-throughput screening. These processes are required to facilitate rapid and accurate biological profiling, structural determination, and resupply of new drug candidates. Reversed-phase high-performance liquid chromatography (RP-HPLC) coupled with electrospray ionization mass spectrometry (ESI-MS) for both analytical and preparative applications has become the small molecule separation/detection tool of choice for meeting many of these needs. However, the separation selectivity provided by RP-HPLC has been limited to the hydrophobicity-based resolution of relatively nonpolar sample components, and for high-throughput drug discovery applications, no sufficient alternative procedures have been identified. In this investigation, a mixed-mode anion-cation exchange/hydrophilic interaction chromatography (ACE-HILIC) method has been developed to provide both direct compatibility with ESI-MS and evaporative light-scattering detection (ELSD) and separation selectivity highly orthogonal to RP-HPLC. The technique employed silica-based small-pore weak ion exchange resins eluted with a combined aqueous and pH gradient. A diverse set of dipeptide probes was employed for the elucidation of the relative contributions of three retention mechanisms. ACE-HILIC-ESI-MS-ELSD should prove useful for the analysis and purification of compounds from both biological (e.g., natural products) and synthetic (e.g., combinatorial chemistry) sources of molecular diversity. PMID:11028621

  12. Analytical characterization of a monoclonal antibody therapeutic reveals a three-light chain species that is efficiently removed using hydrophobic interaction chromatography

    PubMed Central

    Wollacott, Rachel B; Casaz, Paul L; Morin, Trevor J; Zhu, H Lily; Anderson, Roger S; Babcock, Gregory J; Que, John; Thomas Jr, William D; Ozturk, Sadettin S

    2013-01-01

    Size exclusion high performance liquid chromatography analysis of a human monoclonal antibody (mAb) showed the presence of a new species that eluted with a retention time between the dimeric and monomeric species of the antibody. Extensive characterization of this species, referred to as “shoulder,” indicated that it was a mAb containing an extra light chain and had a molecular weight of approximately 175 kDa. The extra light chain was found to be non-covalently associated with the Fab portion of the protein. The relative amount of shoulder (typically 1−3% of the total mAb present) varied with the Chinese hamster ovary cell line producing the mAb and was not influenced by the growth conditions. Our three-step mAb purification platform using protein A, anion exchange, and cation exchange process steps was successful at removing dimer and higher and lower molecular weight species, but not the shoulder impurity. It was found that hydrophobic interaction chromatography could be used in place of cation exchange to exploit the subtle differences in hydrophobicity between monomer and shoulder. We developed an antibody polishing process using Butyl Sepharose HP resin that is capable of removing the majority of high and low molecular weight impurities yielding 99% pure mAb monomer, virtually devoid of the shoulder species, with a step recovery of about 80%. PMID:23995619

  13. Analysis of Mycosporine-Like Amino Acids in Selected Algae and Cyanobacteria by Hydrophilic Interaction Liquid Chromatography and a Novel MAA from the Red Alga Catenella repens

    PubMed Central

    Hartmann, Anja; Becker, Kathrin; Karsten, Ulf; Remias, Daniel; Ganzera, Markus

    2015-01-01

    Mycosporine-like amino acids (MAAs), a group of small secondary metabolites found in algae, cyanobacteria, lichens and fungi, have become ecologically and pharmacologically relevant because of their pronounced UV-absorbing and photo-protective potential. Their analytical characterization is generally achieved by reversed phase HPLC and the compounds are often quantified based on molar extinction coefficients. As an alternative approach, in our study a fully validated hydrophilic interaction liquid chromatography (HILIC) method is presented. It enables the precise quantification of several analytes with adequate retention times in a single run, and can be coupled directly to MS. Excellent linear correlation coefficients (R2 > 0.9991) were obtained, with limit of detection (LOD) values ranging from 0.16 to 0.43 µg/mL. Furthermore, the assay was found to be accurate (recovery rates from 89.8% to 104.1%) and precise (intra-day precision: 5.6%, inter-day precision ≤6.6%). Several algae were assayed for their content of known MAAs like porphyra-334, shinorine, and palythine. Liquid chromatography-mass spectrometry (LC-MS) data indicated a novel compound in some of them, which could be isolated from the marine species Catenella repens and structurally elucidated by nuclear magnetic resonance spectroscopy (NMR) as (E)-3-hydroxy-2-((5-hydroxy-5-(hydroxymethyl)-2-methoxy-3-((2-sulfoethyl)amino)cyclohex-2-en-1-ylidene)amino) propanoic acid, a novel MAA called catenelline. PMID:26473886

  14. Characterization and quantification of histidine degradation in therapeutic protein formulations by size exclusion-hydrophilic interaction two dimensional-liquid chromatography with stable-isotope labeling mass spectrometry.

    PubMed

    Wang, Chunlei; Chen, Sike; Brailsford, John A; Yamniuk, Aaron P; Tymiak, Adrienne A; Zhang, Yingru

    2015-12-24

    Two dimensional liquid chromatography (2D-LC) coupling size exclusion (SEC) and hydrophilic interaction chromatography (HILIC) is demonstrated as a useful tool to study polar excipients, such as histidine and its degradant, in protein formulation samples. The SEC-HILIC setup successfully removed interferences from complex sample matrices and enabled accurate mass measurement of the histidine degradation product, which was then determined to be trans-urocanic acid. Because the SEC effluent is a strong solvent for the second dimension HILIC, experimental parameters needed to be carefully chosen, i.e., small transferring loop, fast gradient at high flow rates for the second dimension gradient, in order to mitigate the solvent mismatch and to ensure good peak shapes for HILIC separations. In addition, the generation of trans-urocanic acid was quantified by single heart-cutting SEC-HILIC 2D-LC combined with stable-isotope labeling mass spectrometry. Compared with existing 2D quantification methods, the proposed approach is fast, insensitive to solvent mismatch between dimensions, and tolerant of small retention time shifts in the first dimension. Finally, the first dimension diode array detector was found to be a potential degradation source for photolabile analytes such as trans-urocanic acid. PMID:26674608

  15. Determination of therapeutic γ-aminobutyric acid analogs in forensic whole blood by hydrophilic interaction liquid chromatography-electrospray tandem mass spectrometry.

    PubMed

    Sørensen, Lambert K; Hasselstrøm, Jørgen B

    2014-05-01

    Vigabatrin, pregabalin, gabapentin and baclofen are γ-aminobutyric acid analogs that are used in the treatment of epileptic seizures (vigabatrin, pregabalin and gabapentin) and spasticity (baclofen). The intake of these drugs may induce adverse reactions and impair the ability of an individual to drive a vehicle. There have also been reports of cases of intoxication and fatalities from overdoses. For rapid and accurate quantification of these drugs in forensic cases, an ultraperformance liquid chromatography tandem mass spectrometry method using pneumatically assisted electrospray ionization has been developed. The technique has been validated on both ante- and postmortem human whole blood. The protein in the blood samples was removed by the addition of a mixture of methanol and acetonitrile, and the extract was ultrafiltered and diluted with acetonitrile. The separation was performed by hydrophilic interaction liquid chromatography. Calibration of the system was achieved through use of matrix-matched calibrants combined with isotope dilution. The lower limits of quantification were 0.02-0.04 mg/L, and the relative intra-laboratory reproducibility standard deviations were <4 and 8% at concentrations of 10 and 1 mg/L, respectively. The mean true recoveries were >89%. The trueness expressed as the relative bias of the test results was within ±7% at concentrations of 1-40 mg/L for vigabatrin, pregabalin and gabapentin and of 0.1-4 mg/L for baclofen. PMID:24523295

  16. Rapid determination of cocamidopropyl betaine impurities in cosmetic products by core-shell hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Wang, Perry G; Zhou, Wanlong

    2016-08-26

    Cocamidopropyl betaine (CAPB) is a common surfactant widely used in personal care products. Dimethylaminopropylamine (DMAPA) and lauramidopropyldimethylamine (LAPDMA) are two chemicals present as impurities in CAPB and have been reported as skin sensitizers. A rapid and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method, using a core shell hydrophilic interaction liquid chromatography (HILIC) column, has been developed to quantify DMAPA and LAPDMA in cosmetic products. Corresponding stable isotopically labeled analogues of the above native compounds were used as internal standards to compensate for matrix effect and for loss of recovery. Each sample was first screened to determine whether the sample needed to be diluted to minimize matrix effects as well as to fit the calibration range. The concept of matrix effect factor (MEF) was introduced to quantitatively evaluate each sample with a unique matrix using the internal standards. Recoveries at three spiking levels of low, medium, and high concentrations ranged from 98.4 to 112% with RSDs less than 5%. This method has been validated and is the first UHPLC-MS/MS method, which uses core shell HILIC column and stable isotopically labeled internal standards to simultaneously determine these two CAPB impurities in cosmetic products. PMID:27473511

  17. Analysis of Mycosporine-Like Amino Acids in Selected Algae and Cyanobacteria by Hydrophilic Interaction Liquid Chromatography and a Novel MAA from the Red Alga Catenella repens.

    PubMed

    Hartmann, Anja; Becker, Kathrin; Karsten, Ulf; Remias, Daniel; Ganzera, Markus

    2015-10-01

    Mycosporine-like amino acids (MAAs), a group of small secondary metabolites found in algae, cyanobacteria, lichens and fungi, have become ecologically and pharmacologically relevant because of their pronounced UV-absorbing and photo-protective potential. Their analytical characterization is generally achieved by reversed phase HPLC and the compounds are often quantified based on molar extinction coefficients. As an alternative approach, in our study a fully validated hydrophilic interaction liquid chromatography (HILIC) method is presented. It enables the precise quantification of several analytes with adequate retention times in a single run, and can be coupled directly to MS. Excellent linear correlation coefficients (R² > 0.9991) were obtained, with limit of detection (LOD) values ranging from 0.16 to 0.43 µg/mL. Furthermore, the assay was found to be accurate (recovery rates from 89.8% to 104.1%) and precise (intra-day precision: 5.6%, inter-day precision ≤6.6%). Several algae were assayed for their content of known MAAs like porphyra-334, shinorine, and palythine. Liquid chromatography-mass spectrometry (LC-MS) data indicated a novel compound in some of them, which could be isolated from the marine species Catenella repens and structurally elucidated by nuclear magnetic resonance spectroscopy (NMR) as (E)-3-hydroxy-2-((5-hydroxy-5-(hydroxymethyl)-2-methoxy-3-((2-sulfoethyl)amino)cyclohex-2-en-1-ylidene)amino) propanoic acid, a novel MAA called catenelline. PMID:26473886

  18. Poissonian steady states: From stationary densities to stationary intensities

    NASA Astrophysics Data System (ADS)

    Eliazar, Iddo

    2012-10-01

    Markov dynamics are the most elemental and omnipresent form of stochastic dynamics in the sciences, with applications ranging from physics to chemistry, from biology to evolution, and from economics to finance. Markov dynamics can be either stationary or nonstationary. Stationary Markov dynamics represent statistical steady states and are quantified by stationary densities. In this paper, we generalize the notion of steady state to the case of general Markov dynamics. Considering an ensemble of independent motions governed by common Markov dynamics, we establish that the entire ensemble attains Poissonian steady states which are quantified by stationary Poissonian intensities and which hold valid also in the case of nonstationary Markov dynamics. The methodology is applied to a host of Markov dynamics, including Brownian motion, birth-death processes, random walks, geometric random walks, renewal processes, growth-collapse dynamics, decay-surge dynamics, Ito diffusions, and Langevin dynamics.

  19. An HPLC chromatographic framework to analyze the β-cyclodextrin/solute complexation mechanism using a carbon nanotube stationary phase.

    PubMed

    Aljhni, Rania; Andre, Claire; Lethier, Lydie; Guillaume, Yves Claude

    2015-11-01

    A carbon nanotube (CNT) stationary phase was used for the first time to study the β-cyclodextrin (β-CD) solute complexation mechanism using high performance liquid chromatography (HPLC). For this, the β-CD was added at various concentrations in the mobile phase and the effect of column temperature was studied on both the retention of a series of aniline and benzoic acid derivatives with the CNT stationary phase and their complexation mechanism with β-CD. A decrease in the solute retention factor was observed for all the studied molecules without change in the retention order. The apparent formation constant KF of the inclusion complex β-CD/solute was determined at various temperatures. Our results showed that the interaction of β-CD with both the mobile phase and the stationary phase interfered in the complex formation. The enthalpy and entropy of the complex formation (ΔHF and ΔSF) between the solute molecule and CD were determined using a thermodynamic approach. Negative enthalpies and entropies indicated that the inclusion process of the studied molecule in the CD cavity was enthalpically driven and that the hydrogen bonds between carboxylic or aniline groups and the functional groups on the β-CD rim play an important role in the complex formation. PMID:26452814

  20. Determination of nucleosides and nucleotides in baby foods by hydrophilic interaction chromatography coupled to tandem mass spectrometry in the presence of hydrophilic ion-pairing reagents.

    PubMed

    Mateos-Vivas, María; Rodríguez-Gonzalo, Encarnación; Domínguez-Álvarez, Javier; García-Gómez, Diego; Carabias-Martínez, Rita

    2016-11-15

    In this work we propose a rapid and efficient method for the joint determination of nucleosides and nucleotides in dairy and non-dairy baby foods based on hydrophilic interaction chromatography coupled to tandem mass spectrometry in the presence of diethylammonium (DEA) as a hydrophilic ion-pairing reagent (IP-HILIC-MS/MS). Sample treatment of the baby food included dilution with water and centrifugal ultrafiltration (CUF) with an additional washing step that notably improved the global performance of the process. Later dilution of the extract with acetonitrile allowed adequate separation in the HILIC system. With the proposed treatment, we obtained extraction recoveries higher than 80% and, additionally, no matrix effects were observed. The CUF-IP-HILIC-MS/MS method was validated according to the 2002/657/EC decision and was used for the quantification of nucleotides and nucleosides in sixteen samples of commercial baby foods. PMID:27283702

  1. Quantitation of Cotinine and its Metabolites in Rat Plasma and Brain Tissue by Hydrophilic Interaction Chromatography Tandem Mass Spectrometry (HILIC-MS/MS)

    PubMed Central

    Li, Pei; Beck, Wayne D.; Callahan, Patrick M.; Terry, Alvin V.; Bartlett, Michael G.

    2014-01-01

    In this work, we developed a sensitive method to quantify cotinine (COT), norcotinine (NCOT), trans-3′-hydroxycotinine (OHCOT) and cotinine-N-oxide (COTNO) in rat plasma and brain tissue, using solid phase extraction (SPE), hydrophilic interaction liquid chromatography (HILIC) and tandem mass spectrometry (MS/MS). The linear range was 1–100 ng/ml for each analyte in rat plasma and brain homogenate (3–300 ng/g brain tissue). The method was validated with precision within 15% relative standard deviation (RSD) and accuracy within 15% relative error (RE). Stable isotope-labeled internal standards (IS) were used for all the analytes to achieve good reproducibility, minimizing the influence of recovery and matrix effects. This method can be used in future studies to simultaneously determine the concentrations of COT and three major metabolites in rat plasma and brain tissue. PMID:23022114

  2. Quantification of structurally related aliphatic amino alcohols in l-valinol by hydrophilic interaction liquid chromatography separation combined with postcolumn derivatization and fluorescence detection.

    PubMed

    Douša, Michal; Stach, Jan; Gibala, Petr; Lemr, Karel

    2016-03-01

    The amino alcohols in l-valinol were effectively separated and quantified using hydrophilic interaction chromatography with fluorescence detection. The influence of the mobile phase (salt type, buffer concentration, and pH) on retention was studied. A column TSKgel amide and mobile phase consisting of 10 mM acetate buffer pH 4.0 and acetonitrile (20:80, v/v) provided well-separated symmetric peaks of analytes. Fluorescence detection was performed using postcolumn derivatization with o-phtaldialdehyde/2-mercaptoethanol at an excitation and emission wavelength of 345 and 450 nm, respectively. Simple sample pretreatment and very high sensitivity represent the main advantages of the developed method. After validation, the method was successfully applied to the analysis of commercial samples of l-valinol. PMID:26697727

  3. Higher order stationary subspace analysis

    NASA Astrophysics Data System (ADS)

    Panknin, Danny; von Bünau, Paul; Kawanabe, Motoaki; Meinecke, Frank C.; Müller, Klaus-Robert

    2016-03-01

    Non-stationarity in data is an ubiquitous problem in signal processing. The recent stationary subspace analysis procedure (SSA) has enabled to decompose such data into a stationary subspace and a non-stationary part respectively. Algorithmically only weak non- stationarities could be tackled by SSA. The present paper takes the conceptual step generalizing from the use of first and second moments as in SSA to higher order moments, thus defining the proposed higher order stationary subspace analysis procedure (HOSSA). The paper derives the novel procedure and shows simulations. An obvious trade-off between the necessity of estimating higher moments and the accuracy and robustness with which they can be estimated is observed. In an ideal setting of plenty of data where higher moment information is dominating our novel approach can win against standard SSA. However, with limited data, even though higher moments actually dominate the underlying data, still SSA may arrive on par.

  4. Stationary Fuel Cell Evaluation (Presentation)

    SciTech Connect

    Kurtz, J.; Wipke, K.; Sprik, S.; Ramsden, T.; Ainscough, C.

    2012-05-01

    This powerpoint presentation discusses its objectives: real world operation data from the field and state-of-the-art lab; collection; analysis for independent technology validation; collaboration with industry and end users operating stationary fuel cell systems and reporting on technology status, progress and technical challenges. The approach and accomplishments are: A quarterly data analysis and publication of first technical stationary fuel cell composite data products (data through June 2012).

  5. Gas Chromatography.

    ERIC Educational Resources Information Center

    Cram, Stuart P.; And Others

    1980-01-01

    Selects fundamental developments in theory, methodology, and instrumentation in gas chromatography (GC). A special section reviews GC in the People's Republic of China. Over 1,000 references are cited. (CS)

  6. Remediation of undesirable secondary interactions encountered in hydrophilic interaction chromatography during development of a quantitative LC-MS/MS assay for a dipeptidyl peptidase IV (DPP-IV) inhibitor in monkey serum.

    PubMed

    Kadar, Eugene P; Wujcik, Chad E

    2009-02-15

    PF-00734200 (3,3-Difluoropyrrolidin-1-yl)-((2S,4S)-4-(4-(pyrimidin-2-yl) piperazin-1-yl)pyrrolidin-2-yl)methanone) is an inhibitor of dipeptidyl peptidase IV (DPP-IV) for the treatment of diabetic complications and other disorders. A sensitive and selective LC-MS/MS assay capable of quantifying PF-00734200 in monkey serum was required to support regulated safety studies. Due to the polar nature of this compound and for ease of sample processing, hydrophilic interaction chromatography (HILIC) was identified as an ideal assay technique. During the initial phase of method development significant peak tailing was observed. The effects of polar organic modifier percentage, buffer concentration, column particle size, and flow rate were assessed to determine the final optimal conditions. PF-00734200 demonstrated a strong dependence on buffer concentration with respect to height equivalent to a theoretical plate (HETP), capacity factor (k'), and tailing factor (T). Improvements in chromatography were observed with increasing buffer concentration due to reduction of electrostatic secondary interactions with ionized silanols. A plot of logk' versus percentage organic modifier at an elevated buffer concentration, produced a linear fit with a correlation coefficient of 0.996, indicating that the primary chromatographic retention mechanism was partitioning. A LC-MS/MS assay was successfully developed and validated for GLP bioanalysis of PF-00734200 in monkey serum utilizing the optimized HILIC conditions. Additionally, carryover was effectively minimized through fortification of ethylene glycol to the sample extract. PMID:19162567

  7. Peptide Orientation Affects Selectivity in Ion-Exchange Chromatography

    SciTech Connect

    Alpert, Andrew J.; Petritis, Konstantinos; Kangas, Lars J.; Smith, Richard D.; Mechtler, Karl; Mitulovic, Goran; Mohammed, Shabaz; Heck, Albert J.

    2010-06-15

    Here we demonstrate that separation of proteolytic peptides, having the same net charge and one basic residue, is affected by their specific orientation toward the stationary phase in ion-exchange chromatography. In electrostatic repulsion-hydrophilic interaction chromatography (ERLIC) with an anion-exchange material, the C-terminus of the peptides is, on average, oriented toward the stationary phase. In cation exchange, the average peptide orientation is the opposite. Data with synthetic peptides, serving as orientation probes, indicate that in tryptic/Lys-C peptides the C-terminal carboxyl group appears to be in a zwitterionic bond with the side chain of the C-terminal Lys/Arg residue. In effect, the side chain is then less basic than the N-terminus, accounting for the specific orientation of tryptic and Lys-C peptides. Analyses of larger sets of peptides, generated from lysates by either Lys-N, Lys-C, or trypsin, reveal that specific peptide orientation affects the ability of harged side chains, such as phosphate residues, to influence retention. Phosphorylated residues that are remote in the sequence from the binding site affect retention less than those that are closer. When a peptide contains multiple charged sites, then orientation is observed to be less rigid and retention tends to be governed by the peptide’s net charge rather than its sequence. These general observations could be of value in confirming a peptide’s identification and, in particular, phosphosite assignments in proteomics analyses. More generally, orientation accounts for the ability of chromatography to separate peptides of the same compositionbut different sequence.

  8. Peptide Orientation Affects Selectivity in Ion-Exchange Chromatography

    SciTech Connect

    Alpert, Andrew J.; Petritis, Konstantinos; Kangas, Lars J.; Smith, R. D.; Mechtler, Karl; Mitulovic, Goran; Mohammed, Shabaz; Heck, Albert J.

    2010-06-15

    Here we demonstrate that separation of proteolytic peptides, having the same net charge and one basic residue, is affected by their specific orientation toward the stationary phase in ion-exchange chromatography. In electrostatic repulsion-hydrophilic interaction chromatography (ERLIC) with an anion-exchange material, the C-terminus of the peptides is, on average, oriented toward the stationary phase. In cation exchange, the average peptide orientation is the opposite. Data with synthetic peptides, serving as orientation probes, indicate that in tryptic/ Lys-C peptides the C-terminal carboxyl group appears to be in a zwitterionic bond with the side chain of the C-terminal Lys/Arg residue. In effect, the side chain is then less basic than the N-terminus, accounting for the specific orientation of tryptic and Lys-C peptides. Analyses of larger sets of peptides, generated from lysates by either Lys-N, Lys-C, or trypsin, reveal that specific peptide orientation affects the ability of charged side chains, such as phosphate residues, to influence retention. Phosphorylated residues that are remote in the sequence from the binding site affect retention less than those that are closer. When a peptide contains multiple charged sites, then orientation is observed to be less rigid and retention tends to be governed by the peptide’s net charge rather than its sequence. These general observations could be of value in confirming a peptide’s identification and, in particular, phosphosite assignments in proteomics analyses. More generally, orientation accounts for the ability of chromatography to separate peptides of the same composition but different sequence.

  9. Peptide Orientation Affects Selectivity in Ion-Exchange Chromatography

    PubMed Central

    2010-01-01

    Here we demonstrate that separation of proteolytic peptides, having the same net charge and one basic residue, is affected by their specific orientation toward the stationary phase in ion-exchange chromatography. In electrostatic repulsion−hydrophilic interaction chromatography (ERLIC) with an anion-exchange material, the C-terminus of the peptides is, on average, oriented toward the stationary phase. In cation exchange, the average peptide orientation is the opposite. Data with synthetic peptides, serving as orientation probes, indicate that in tryptic/Lys-C peptides the C-terminal carboxyl group appears to be in a zwitterionic bond with the side chain of the C-terminal Lys/Arg residue. In effect, the side chain is then less basic than the N-terminus, accounting for the specific orientation of tryptic and Lys-C peptides. Analyses of larger sets of peptides, generated from lysates by either Lys-N, Lys-C, or trypsin, reveal that specific peptide orientation affects the ability of charged side chains, such as phosphate residues, to influence retention. Phosphorylated residues that are remote in the sequence from the binding site affect retention less than those that are closer. When a peptide contains multiple charged sites, then orientation is observed to be less rigid and retention tends to be governed by the peptide’s net charge rather than its sequence. These general observations could be of value in confirming a peptide’s identification and, in particular, phosphosite assignments in proteomics analyses. More generally, orientation accounts for the ability of chromatography to separate peptides of the same composition but different sequence. PMID:20481592

  10. Exploring Enantiospecific Ligand-Protein Interactions Using Cellular Membrane Affinity Chromatography: Chiral Recognition as a Dynamic Process

    PubMed Central

    Jozwiak, Krzysztof; Moaddel, Ruin; Ravichandran, Sarangan; Plazinska, Anita; Kozak, Joanna; Patel, Sharvil; Yamaguchi, Rika; Wainer, Irving

    2008-01-01

    The chiral recognition mechanisms responsible for the enantioselective binding on the α3β4 nicotinic acetyl choline receptor (α3β4 nAChR) and human organic cation transporter 1 (hOCT1) have been reviewed. The results indicate that chiral recognition on the α3β4 nAChR is a process involving initial tethering of dextromethorphan and levomethorphan at hydrophobic pockets within the central lumen followed by hydrogen bonding interactions favoring dextromethorphan. The second step is the defining enantioselective step. Studies with the hOCT1 indentified four binding sites within the transporter that participated in chiral recognition. Each of the enantiomers of the compounds used in the study interacted with three of these sites, while (R)-verapamil interacted with all four. Chiral recognition arose from the conformational adjustments required to produce optimum interactions. With respect to the prevailing interaction-based models, the data suggest that chiral recognition is a dynamic process and that the static point-based models should be amended to reflect this. PMID:18723411

  11. Decay of stationary light pulses in ultracold atoms

    SciTech Connect

    Wu Jinhui; Artoni, M.; La Rocca, G. C.

    2010-03-15

    We develop a general scheme for studying the optical response of ultracold atoms driven into a regime of standing-wave electromagnetically induced transparency. We rely on full numerical solutions of the Maxwell-Liouville equations without invoking secular and adiabatic approximations and arbitrary initial state assumptions. These approximations and assumptions can conceal, e.g., significant loss and diffusion responsible for the decay of stationary light pulses in cold atomic samples. The complex decay dynamics of a stationary light pulse is here analyzed in terms of higher-order spin and optical coherences that arise from nonlinear interactions of the stationary light pulse with the two counterpropagating components of a standing-wave driving field. Specific results for stationary light pulses in cold {sup 87}Rb atoms have been discussed for temperature regimes where the residual Doppler broadening is negligible.

  12. Temperature-responsive molecular recognition chromatography using phenylalanine and tryptophan derived polymer modified silica beads.

    PubMed

    Hiruta, Yuki; Kanazashi, Ryosuke; Ayano, Eri; Okano, Teruo; Kanazawa, Hideko

    2016-02-01

    Temperature-responsive polymers incorporating molecular-recognition sites were developed as stationary phases for high-performance liquid chromatography (HPLC). The grafted stationary phases consisted of functional copolymers composed of N-isopropylacrylamide (NIPAAm) and N-acryloyl aromatic amino acid methyl esters, i.e., phenylalanine and tryptophan methyl esters (Phe-OMe and Trp-OMe). Three novel temperature-responsive polymers, P(NIPAAm-co-Phe-OMe5), P(NIPAAm-co-Phe-OMe10), and P(NIPAAm-co-Trp-OMe5), were synthesized. These copolymers exhibited a reversible hydrophilic/hydrophobic phase transition at their lower critical solution temperatures (LCSTs). The polymers were grafted onto aminopropyl silica using an activated ester-amine coupling method, and were packed into a stainless steel column, which was connected to an HPLC system. Temperature-responsive chromatography was conducted using water as the sole mobile phase. More hydrophobic analytes were retained longer, and the retention times of aromatic steroids and aromatic amino acids were dramatically increased. This indicated that π-π interactions occurred between the phenyl or indole moieties of phenylalanine or tryptophan, respectively, and the aromatic compounds. Furthermore, the retention times of compounds with hydrogen bond acceptors were higher with P(NIPAAm-co-Trp-OMe5), which contained indole as a hydrogen bond donor, than with P(NIPAAm-co-Phe-OMe5). This indicated that hydrogen bonding occurred between the stationary phase and the analytes. These results indicate that hydrophobic, π-π, and hydrogen bonding interactions all affected the separation mode of the temperature-responsive chromatography, and led to selective separation with molecular recognition. Both temperature-response and molecular recognition characteristics are present in the proposed separation system that utilizes a temperature-responsive polymer bearing aromatic amino acid derivatives. PMID:26646169

  13. The use of linear expressions of solute boiling point versus retention to indicate special interactions with the molecular rings of modified cyclodextrin phases in gas chromatography

    PubMed

    Betts

    2000-08-01

    The boiling points (degrees C, 1 x 10) of diverse C10 polar solutes from volatile oils are set against their relative retention times versus n-undecane to calculate linear equations for 12 commercial modified cyclodextrin (CD) capillary phases. Ten data points are considered for each CD, then solutes are rejected until 5 or more remain that give an expression with a correlation coefficient of at least 0.990 and a standard deviation of less than 5.5. Three phases give almost perfect correlation, and 3 other CDs have difficulty complying. Solutes involved in the equations (most frequently cuminal, linalol, and carvone) are presumed to have a 'standard' polar transient interaction with the molecular rings of the CDs concerned. Several remaining solutes (mostly citral, fenchone, and menthol) exhibit extra retention over the calculated standard (up to 772%), which is believed to indicate a firm 'host' CD or 'guest' solute molecular fit in some cases. Other solutes show less retention than calculated (mostly citronellal, citronellol, estragole, and pulegone). This suggests rejection by the CD, which behaves merely as a conventional stationary phase to them. The intercept constant in the equation for each phase is suggested to be a numerical relative polarity indicator. These b values indicate that 3 hydroxypropyl CDs show the most polarity with values from 28 to 43; and CDs that are fully substituted with inert groups fall in the range of 15 to 20. PMID:10955511

  14. Facile "one-pot" synthesis of poly(methacrylic acid)-based hybrid monolith via thiol-ene click reaction for hydrophilic interaction chromatography.

    PubMed

    Lv, Xumei; Tan, Wangming; Chen, Ye; Chen, Yingzhuang; Ma, Ming; Chen, Bo; Yao, Shouzhuo

    2016-07-01

    A novel sol-gel "one-pot" approach in tandem with a radical-mediated thiol-ene reaction for the synthesis of a methacrylic acid-based hybrid monolith was developed. The polymerization monomers, tetramethoxysilane (TMOS) and 3-mercaptopropyl trimethoxysilane (MPTS), were hydrolyzed in high-concentration methacrylic acid solution that also served as a hydrophilic functional monomer. The resulting solution was then mixed with initiator (2, 2'-azobis (2-methylpropionamide) dihydrochloride) and porogen (urea, polyethylene glycol 20,000) in a capillary column and polymerized in water bath. The column had a uniform porous structure and a good permeability. The evaluation of the monolith was performed by separation of small molecules including nucleosides, phenols, amides, bases and Triton X-100. The calibration curves for uridine, inosine, adenosine and cytidine were determined. All the calibration curves exhibited good linear regressions (R(2)≥0.995) within the test ranges of 0.5-40μg/mL for four nucleosides. Additionaliy, atypical hydrophilic mechanism was proved by elution order from low to high according to polarity retention time increased with increases in the content of the organic solvent in the mobile phase. Further studies indicated that hydrogen bond and electrostatic interactions existed between the polar analytes and the stationary phase. This was the mechanism of retention. The excellent separation of the BSA digest showed good hydrophility of the column and indicated the potential in separation of complex biological samples. PMID:27264742

  15. Continuous stationary phase gradients for planar chromatographic media.

    PubMed

    Kannan, Balamurali; Marin, Michael A; Shrestha, Kushal; Higgins, Daniel A; Collinson, Maryanne M

    2011-12-30

    A simple, elegant method for the formation of a continuous stationary phase gradient for use in chromatographic separations is described. Its applicability to separation science is demonstrated using thin-layer chromatography as a test case. Gradient stationary phases were formed on activated High Performance Thin-Layer Chromatography (HP-TLC) plates using a newly developed methodology termed "controlled rate infusion". Specifically, the SiOH groups on the activated HP-TLC plates were reacted with 3-aminopropyltriethoxysilane (APTEOS) in a time dependent fashion by using a programmable syringe pump to control the rate of APTEOS infusion into the deposition reservoir. The shape (profile) of the gradient was controlled by the rate of infusion and imaged by taking advantage of the concentration-dependent color formation reaction between amine groups and ninhydrin. The advantages of such gradients in optimizing the retention and separation of various components in different mixtures were illustrated using mixtures of (1) four weak acids and bases and (2) three widely used over-the-counter drugs. The separation of the individual components on the gradient stationary phase was clearly improved relative to those on either traditional normal-phase TLC plates or uniformly amine-modified TLC plates. Precise control over component retention and separation was also demonstrated by strategically modifying the steepness of the gradient. PMID:22119610

  16. Analysis of multi-site drug-protein interactions by high-performance affinity chromatography: Binding by glimepiride to normal or glycated human serum albumin.

    PubMed

    Matsuda, Ryan; Li, Zhao; Zheng, Xiwei; Hage, David S

    2015-08-21

    High-performance affinity chromatography (HPAC) was used in a variety of formats to examine multi-site interactions between glimepiride, a third-generation sulfonylurea drug, and normal or in vitro glycated forms of the transport protein human serum albumin (HSA). Frontal analysis revealed that glimepiride interacts with normal HSA and glycated HSA at a group of high affinity sites (association equilibrium constant, or Ka, 9.2-11.8×10(5)M(-1) at pH 7.4 and 37°C) and a group of lower affinity regions (Ka, 5.9-16×10(3)M(-1)). Zonal elution competition studies were designed and carried out in both normal- and reversed-role formats to investigate the binding by this drug at specific sites. These experiments indicated that glimepiride was interacting at both Sudlow sites I and II. Allosteric effects were also noted with R-warfarin at Sudlow site I and with tamoxifen at the tamoxifen site on HSA. The binding at Sudlow site I had a 2.1- to 2.3-fold increase in affinity in going from normal HSA to the glycated samples of HSA. There was no significant change in the affinity for glimepiride at Sudlow site II in going from normal HSA to a moderately glycated sample of HSA, but a slight decrease in affinity was seen in going to a more highly glycated HSA sample. These results demonstrated how various HPAC-based methods can be used to profile and characterize multi-site binding by a drug such as glimepiride to a protein and its modified forms. The information obtained from this study should be useful in providing a better understanding of how drug-protein binding may be affected by glycation and of how separation and analysis methods based on HPAC can be employed to study systems with complex interactions or that involve modified proteins. PMID:26189669

  17. Analysis of Multi-Site Drug-Protein Interactions by High-Performance Affinity Chromatography: Binding by Glimepiride to Normal or Glycated Human Serum Albumin

    PubMed Central

    Matsuda, Ryan; Li, Zhao; Zheng, Xiwei; Hage, David S.

    2015-01-01

    High-performance affinity chromatography (HPAC) was used in a variety of formats to examine multi-site interactions between glimepiride, a third-generation sulfonylurea drug, and normal or in vitro glycated forms of the transport protein human serum albumin (HSA). Frontal analysis revealed that glimepiride interacts with normal HSA and glycated HSA at a group of high affinity sites (association equilibrium constant, or Ka, 9.2–11.8 × 105 M−1 at pH 7.4 and 37°C) and a group of lower affinity regions (Ka, 5.9–16.2 × 103 M−1). Zonal elution competition studies were designed and carried out in both normal- and reversed-role formats to investigate the binding by this drug at specific sites. These experiments indicated that glimepiride was interacting at both Sudlow sites I and II. Allosteric effects were also noted with R-warfarin at Sudlow site I and with tamoxifen at the tamoxifen site on HSA. The binding at Sudlow site I had a 2.1- to 2.3-fold increase in affinity in going from normal HSA to the glycated samples of HSA. There was no significant change in the affinity for glimepiride at Sudlow site II in going from normal HSA to a moderately glycated sample of HSA, but a slight decrease in affinity was seen in going to a more highly glycated HSA sample. These results demonstrated how various HPAC-based methods can be used to profile and characterize multi-site binding by a drug such as glimepiride to a protein and its modified forms. The information obtained from this study should be useful in providing a better understanding of how drug-protein binding may be affected by glycation and of how separation and analysis methods based on HPAC can be employed to study systems with complex interactions or that involve modified proteins. PMID:26189669

  18. Differential chemical derivatization integrated with chromatographic separation for analysis of isomeric sialylated N-glycans: a nano-hydrophilic interaction liquid chromatography-MS platform.

    PubMed

    Tousi, Fateme; Bones, Jonathan; Hancock, William S; Hincapie, Marina

    2013-09-01

    MS analysis of sialylated glycans is challenging due to their low ionization efficiency in positive ion mode as well as the possibility of in-source fragmentation. Chemical derivatization strategies have been developed to address this issue focused on removal of the labile acidic proton prior to MS analysis. Highly sialylated negatively charged glycans also exhibit high retention and unsatisfactory separation efficiency when analyzed by hydrophilic interaction liquid chromatography (HILIC) due to their high polarity. Here, we combined linkage specific derivatization of sialic acids by reaction with the condensation reagent 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM) in methanol with nanoscale liquid chromatographic separation prior to accurate mass Orbitrap MS analysis. Coupling DMT-MM charge neutralization of sialic acids with nano-HILIC-Orbitrap-MS not only allows for linkage specific characterization of sialylated glycans directly from the precursor mass but also improves the preceding HILIC separation by increasing the hydrophobicity and altering the selectivity of the oligosaccharide analytes. We focused on the trisialylated N-glycan fraction from haptoglobin and human plasma, enriched using weak anion exchange chromatography, as this trisialylated fraction has been linked with cancer associated changes in the serum N-glycome. The developed methodology was applied to investigate whether structural alterations in this oligosaccharide pool, enriched from the sera of pathological stage and sex matched patients bearing lung, breast, ovarian, pancreatic, or gastric cancer, demonstrate any degree of cancer specificity or whether changes in expression levels are purely cancer associated. The results of this pilot study indicate limited degrees of cancer specificity, particularly for pancreatic cancer, based on alterations in the relative abundance of specific trisialylated isomers. PMID:23901877

  19. Optimized Analytical Procedures for the Untargeted Metabolomic Profiling of Human Urine and Plasma by Combining Hydrophilic Interaction (HILIC) and Reverse-Phase Liquid Chromatography (RPLC)-Mass Spectrometry.

    PubMed

    Contrepois, Kévin; Jiang, Lihua; Snyder, Michael

    2015-06-01

    Profiling of body fluids is crucial for monitoring and discovering metabolic markers of health and disease and for providing insights into human physiology. Since human urine and plasma each contain an extreme diversity of metabolites, a single liquid chromatographic system when coupled to mass spectrometry (MS) is not sufficient to achieve reasonable metabolome coverage. Hydrophilic interaction liquid chromatography (HILIC) offers complementary information to reverse-phase liquid chromatography (RPLC) by retaining polar metabolites. With the objective of finding the optimal combined chromatographic solution to profile urine and plasma, we systematically investigated the performance of five HILIC columns with different chemistries operated at three different pH (acidic, neutral, basic) and five C18-silica RPLC columns. The zwitterionic column ZIC-HILIC operated at neutral pH provided optimal performance on a large set of hydrophilic metabolites. The RPLC columns Hypersil GOLD and Zorbax SB aq were proven to be best suited for the metabolic profiling of urine and plasma, respectively. Importantly, the optimized HILIC-MS method showed excellent intrabatch peak area reproducibility (CV < 12%) and good long-term interbatch (40 days) peak area reproducibility (CV < 22%) that were similar to those of RPLC-MS procedures. Finally, combining the optimal HILIC- and RPLC-MS approaches greatly expanded metabolome coverage with 44% and 108% new metabolic features detected compared with RPLC-MS alone for urine and plasma, respectively. The proposed combined LC-MS approaches improve the comprehensiveness of global metabolic profiling of body fluids and thus are valuable for monitoring and discovering metabolic changes associated with health and disease in clinical research studies. PMID:25787789

  20. Hydrophilic interaction liquid chromatography-tandem mass spectrometry quantitative method for the cellular analysis of varying structures of gemini surfactants designed as nanomaterial drug carriers.

    PubMed

    Donkuru, McDonald; Michel, Deborah; Awad, Hanan; Katselis, George; El-Aneed, Anas

    2016-05-13

    Diquaternary gemini surfactants have successfully been used to form lipid-based nanoparticles that are able to compact, protect, and deliver genetic materials into cells. However, what happens to the gemini surfactants after they have released their therapeutic cargo is unknown. Such knowledge is critical to assess the quality, safety, and efficacy of gemini surfactant nanoparticles. We have developed a simple and rapid liquid chromatography electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method for the quantitative determination of various structures of gemini surfactants in cells. Hydrophilic interaction liquid chromatography (HILIC) was employed allowing for a short simple isocratic run of only 4min. The lower limit of detection (LLOD) was 3ng/mL. The method was valid to 18 structures of gemini surfactants belonging to two different structural families. A full method validation was performed for two lead compounds according to USFDA guidelines. The HILIC-MS/MS method was compatible with the physicochemical properties of gemini surfactants that bear a permanent positive charge with both hydrophilic and hydrophobic elements within their molecular structure. In addition, an effective liquid-liquid extraction method (98% recovery) was employed surpassing previously used extraction methods. The analysis of nanoparticle-treated cells showed an initial rise in the analyte intracellular concentration followed by a maximum and a somewhat more gradual decrease of the intracellular concentration. The observed intracellular depletion of the gemini surfactants may be attributable to their bio-transformation into metabolites and exocytosis from the host cells. Obtained cellular data showed a pattern that grants additional investigations, evaluating metabolite formation and assessing the subcellular distribution of tested compounds. PMID:27086283

  1. Single column comprehensive analysis of pharmaceutical preparations using dual-injection mixed-mode (ion-exchange and reversed-phase) and hydrophilic interaction liquid chromatography.

    PubMed

    Kazarian, Artaches A; Taylor, Mark R; Haddad, Paul R; Nesterenko, Pavel N; Paull, Brett

    2013-12-01

    The comprehensive separation and detection of hydrophobic and hydrophilic active pharmaceutical ingredients (APIs), their counter-ions (organic, inorganic) and excipients, using a single mixed-mode chromatographic column, and a dual injection approach is presented. Using a mixed-mode Thermo Fisher Acclaim Trinity P1 column, APIs, their counter-ions and possible degradants were first separated using a combination of anion-exchange, cation-exchange and hydrophobic interactions, using a mobile phase consisting of a dual organic modifier/salt concentration gradient. A complementary method was also developed using the same column for the separation of hydrophilic bulk excipients, using hydrophilic interaction liquid chromatography (HILIC) under high organic solvent mobile phase conditions. These two methods were then combined within a single gradient run using dual sample injection, with the first injection at the start of the applied gradient (mixed-mode retention of solutes), followed by a second sample injection at the end of the gradient (HILIC retention of solutes). Detection using both ultraviolet absorbance and refractive index enabled the sensitive detection of APIs and UV-absorbing counter-ions, together with quantitative determination of bulk excipients. The developed approach was applied successfully to the analysis of a dry powder inhalers (Flixotide(®), Spiriva(®)), enabling comprehensive quantification of all APIs and excipients in the sample. PMID:24001905

  2. ANALYSIS OF DRUG INTERACTIONS WITH MODIFIED PROTEINS BY HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY: BINDING OF GLIBENCLAMIDE TO NORMAL AND GLYCATED HUMAN SERUM ALBUMIN

    PubMed Central

    Matsuda, Ryan; Anguizola, Jeanethe; Joseph, K.S.; Hage, David S.

    2012-01-01

    High-performance affinity chromatography (HPAC) was used to examine the changes in binding that occur for the sulfonylurea drug glibenclamide with human serum albumin (HSA) at various stages of glycation for HSA. Frontal analysis on columns containing normal HSA or glycated HSA indicated glibenclamide was interacting through both high affinity sites (association equilibrium constant, Ka, 1.4–1.9 × 106 M−1 at pH 7.4 and 37°C) and lower affinity sites (Ka, 4.4–7.2 × 104 M−1). Competition studies were used to examine the effect of glycation at specific binding sites of HSA. An increase in affinity of 1.7- to 1.9-fold was seen at Sudlow site I with moderate to high levels of glycation. An even larger increase of 4.3- to 6.0-fold in affinity was noted at Sudlow site II for all of the tested samples of glycated HSA. A slight decrease in affinity may have occurred at the digitoxin site, but this change was not significant for any individual glycated HSA sample. These results illustrate how HPAC can be used as tool for examining the interactions of relatively non-polar drugs like glibenclamide with modified proteins and should lead to a more complete understanding of how glycation can alter the binding of drugs in blood. PMID:23092871

  3. Quantification of 1-(13) C-L-methionine in rat serum with hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry and its application in pharmacokinetic study.

    PubMed

    Xu, Ying; Huang, Xiao; Nie, Xiuli; Yang, Li; Yan, Weili; Wang, Zhengtao; Wang, Changhong; Hu, Zhibi

    2011-09-01

    A rapid, selective and sensitive hydrophilic interaction liquid chromatography (HILIC) coupled with tandem mass spectrometry (MS/MS) method was developed to determine 1-(13) C-l-methionine in rat serum. Proteins in serum were precipitated using acetonitrile and the supernatant was separated after centrifugation. 1-(13) C-l-phenylalnine was used as the internal standard. HILIC-tandem mass spectrometry analysis was performed on a hydrophilic interaction silica column (TSK-GEL AMIDE-80) using a linear gradient elution system, acetonitrile-5 mm ammonium acetate containing 0.1% formic acid and multiple reaction monitoring mode for 1-(13) C-l-methionine and 1-(13) C-l-phenylalnine. The assay was validated with a linear range between 10 and 150 ng mL(-1) (r ≥ 0.99) and a lower limit of quantification of 10 ng mL(-1) , calculated with weighted (1/x(2) ) least squares linear regression. The RSD of intra-day precision was smaller than 3.6% and the inter-day RSD less than 6.5%, while the average recovery was 100.48% with an RSD of accuracy within 2.9%, determined from quality control samples. The HILIC-MS/MS method was fully validated and successfully applied to the in vivo pharmacokinetic study of stable-isotope 1-(13) C-l-methionine in rats. PMID:21287581

  4. Stationary surgical smoke evacuation systems.

    PubMed

    2001-03-01

    Two types of systems are available for evacuating the surgical smoke created by electrosurgery and laser surgery: portable and stationary surgical smoke evacuation systems. While portable systems dominate the market today, stationary systems are an alternative worth considering--even though they are still in their infancy, with fewer than 90 systems installed to date. Stationary systems represent a major commitment on the part of the healthcare facility. Several system components must be installed as part of the physical plant (for instance, within the walls), making the system a permanent fixture in the surgical suite. Installation of these systems is often carried out during building construction or major renovation--although the systems can be cost-effective even if no renovations are planned. For this Evaluation, we tested three stationary systems. All three are adequate to capture surgical smoke and evacuate it from the operating room. These systems are easy to use, are quietter than their portable counterparts, and require minimal user maintenance. They represent an excellent option for most hospitals actively evacuating surgical smoke. In this article, we discuss the factors to consider when selecting from among these systems. We also offer guidance on choosing between stationary systems and portable ones. PMID:11321758

  5. Specific recognition of supercoiled plasmid DNA by affinity chromatography using a synthetic aromatic ligand.

    PubMed

    Caramelo-Nunes, Catarina; Tomaz, Cândida T

    2015-01-01

    Liquid chromatography is the method of choice for the purification of plasmid DNA (pDNA), since it is simple, robust, versatile, and highly reproducible. The most important features of a chromatographic procedure are the use of suitable stationary phases and ligands. As conventional purification protocols are being replaced by more sophisticated and selective procedures, the focus changes toward designing and selecting ligands of high affinity and specificity. In fact, the chemical composition of the chromatographic supports determines the interactions established with the target molecules, allowing their preferential retention over the undesirable ones. Here it is described the selective recognition and purification of supercoiled pDNA by affinity chromatography, using an intercalative molecule (3,8-diamino-6-phenylphenanthridine) as ligand. PMID:25749945

  6. HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY AND THE ANALYSIS OF DRUG INTERACTIONS WITH MODIFIED PROTEINS: BINDING OF GLICLAZIDE WITH GLYCATED HUMAN SERUM ALBUMIN

    PubMed Central

    Matsuda, Ryan; Anguizola, Jeanethe; Joseph, K.S.; Hage, David S.

    2011-01-01

    This study used high-performance affinity chromatography (HPAC) to examine the binding of gliclazide (i.e., a sulfonylurea drug used to treat diabetes) with the protein human serum albumin (HSA) at various stages of modification due to glycation. Frontal analysis conducted with small HPAC columns was first used to estimate the number of binding sites and association equilibrium constants (Ka) for gliclazide with normal HSA and glycated HSA. Both normal and glycated HSA interacted with gliclazide according to a two-site model, with a class of high affinity sites (average Ka, 7.1-10 × 104 M−1) and a group of lower affinity sites (average Ka, 5.7-8.9 × 103 M−1) at pH 7.4 and 37°C. Competition experiments indicated that Sudlow sites I and II of HSA were both involved in these interactions, with the Ka values for gliclazide at these sites being 1.9 × 104 M−1 and 6.0 × 104 M−1, respectively, for normal HSA. Two samples of glycated HSA had similar affinities to normal HSA for gliclazide at Sudlow site I, but one sample had a 1.9-fold increase in affinity at this site. All three glycated HSA samples differed from normal HSA in their affinity for gliclazide at Sudlow site II. This work illustrated how HPAC can be used to examine both the overall binding of a drug with normal or modified proteins and the site-specific changes that can occur in these interactions as a result of protein modification. PMID:21922305

  7. Mixed-mode chromatography with zwitterionic phosphopeptidomimetic selectors from Ugi multicomponent reaction.

    PubMed

    Gargano, Andrea F G; Leek, Tomas; Lindner, Wolfgang; Lämmerhofer, Michael

    2013-11-22

    In the present contribution a novel Ugi multicomponent reaction (MCR) was used to generate zwitterionic chromatographic selectors with capability for application in mixed-mode chromatography featuring complementary selectivities in reversed-phase (RP) and hydrophilic interaction liquid chromatography (HILIC). Aminophosphonate zwitterionic chromatographic ligands were synthesized adopting a one pot microwave assisted three-component UGI-MCR synthesis and, after purification, were immobilized by thiol-ene click chemistry on silica beads. Chromatographic characteristics of these stationary phases were evaluated by variation of experimental conditions for a set of diverse analytes with neutral, acidic, basic and zwitterionic character revealing the presence of multimodal retention capabilities (i.e. tunable retention increments from ion exchange, hydrophobic and hydrophilic interaction were observed). To further investigate and classify the retention properties of the novel stationary phases we performed a comparative chromatographic study with commercially available mixed mode, HILIC and RP columns. The resultant chromatographic data were analyzed by principal component analysis (PCA). PCA revealed that the new reversed-phase/zwitterionic ion-exchangers (RP/ZWIX) are complementary to common RP, HILIC and mixed-mode phases on the market and could be a promising alternative in impurity profiling and 2D-HPLC concepts. Moreover, the adopted synthetic approach offers the capability to generate chemical diversity simply by the variation of the starting aldehyde, aminophosphonic acid and/or isonitrile components. This unique characteristics offer great possibility for the design of novel selectors for mixed mode chromatography like RP/ZWIX, HILIC, affinity and chiral chromatography. PMID:23932032

  8. Gas Chromatography

    NASA Astrophysics Data System (ADS)

    Qian, Michael C.; Peterson, Devin G.; Reineccius, Gary A.

    The first publication on gas chromatography (GC) was in 1952 (1), while the first commercial instruments were manufactured in 1956. James and Martin (1) separated fatty acids by GC, collected the column effluent, and titrated the individual fatty acids for quantitation. GC has advanced greatly since that early work and is now considered to be a mature field that is approaching theoretical limitations.

  9. Ion Chromatography.

    ERIC Educational Resources Information Center

    Mulik, James D.; Sawicki, Eugene

    1979-01-01

    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)

  10. The hydrophilicity vs. ion interaction selectivity plot revisited: The effect of mobile phase pH and buffer concentration on hydrophilic interaction liquid chromatography selectivity behavior.

    PubMed

    Iverson, Chad D; Gu, Xinyun; Lucy, Charles A

    2016-08-01

    This work systematically investigates the selectivity changes on many HILIC phases from w(w)pH 3.7-6.8, at 5 and 25mM buffer concentrations. Hydrophilicity (kcytosine/kuracil) vs. ion interaction (kBTMA/kuracil) selectivity plots developed by Ibrahim et al. (J. Chromatogr. A 1260 (2012) 126-131) are used to investigate the effect of mobile phase changes on the selectivity of 18 HILIC columns from various classes. "Selectivity change plots" focus on the change in hydrophilicity and ion interaction that the columns exhibit upon changing mobile phase conditions. In general, the selectivity behavior of most HILIC columns is dominated by silanol activity. Minimal changes in selectivity are observed upon changing pH between w(w)pH 5 and 6.8. However, a reduction in ionic interaction is observed when the buffer concentration is increased at w(w)pH≥5.0 due to ionic shielding. Reduction of the w(w)pH to<5.0 results in decreasing cation exchange activity due to silanol protonation. Under all eluent conditions, the majority of phases show little change in their hydrophilicity. PMID:27388658

  11. Are Eddy Covariance series stationary?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Spectral analysis via a discrete Fourier transform is used often to examine eddy covariance series for cycles (eddies) of interest. Generally the analysis is performed on hourly or half-hourly data sets collected at 10 or 20 Hz. Each original series is often assumed to be stationary. Also automated ...

  12. Stationary measure in the multiverse

    SciTech Connect

    Linde, Andrei; Vanchurin, Vitaly; Winitzki, Sergei E-mail: vitaly@cosmos2.phy.tufts.edu

    2009-01-15

    We study the recently proposed ''stationary measure'' in the context of the string landscape scenario. We show that it suffers neither from the ''Boltzmann brain'' problem nor from the ''youngness'' paradox that makes some other measures predict a high CMB temperature at present. We also demonstrate a good performance of this measure in predicting the results of local experiments, such as proton decay.

  13. Quantification of L-ergothioneine in whole blood by hydrophilic interaction ultra-performance liquid chromatography and UV-detection.

    PubMed

    Sotgia, Salvatore; Zinellu, Angelo; Pintus, Gianfranco; Pinna, Gerard Aime; Deiana, Luca; Carru, Ciriaco

    2013-03-01

    A new hydrophilic interaction ultra-performance LC method was established for the whole blood measurement of L-ergothioneine. Chromatographic separation was achieved in a fairly short time, less than 4 min, on a 100 × 2.1 mm Acquity UPLC BEH HILIC 1.7 μm column with a mobile phase consisting of a mixture of 100 mmol/L ammonium acetate/ACN/water (5:85:10, v/v/v) that flowed isocratically at 0.250 mL/min. The LOD and the limit of quantification were 3.85 and 11.67 μmol/L, respectively. The method exhibited linearity in a concentration range of 15.63-1000 μmol/L (R(2) > 0.999). Mean recovery was 96.34% whereas intraassay and interassay precision were 1.52 and 1.82% RSD, respectively. On the whole, the developed method is simple, fast, precise, accurate, and sensitive and may be useful for routine analyses. PMID:23418129

  14. Determination of glutathione and cysteine in yeasts by hydrophilic interaction liquid chromatography followed by on-line postcolumn derivatization.

    PubMed

    Karakosta, Theano D; Tzanavaras, Paraskevas D; Themelis, Demetrius G

    2013-06-01

    In the present study, we report a new method for the determination of two primary thiols, cysteine (CYS) and glutathione (GSH), by hydrophilic interaction LC. The polar analytes are separated isocratically using a mobile phase consisting of 65% acetonitrile/35% ammonium acetate (15 mmol/L, pH 2.0) and are detected at 285 nm following on-line postcolumn derivatization by the thiol-selective reagent methyl propiolate. The main figures of merit included linearity in the range of 5-200 μmol/L and an LOD 0.6 μmol/L for both compounds. The absence of matrix effect allowed the determination of CYS and GSH in various yeast samples. GSH was present in most of the samples at levels ranging between 0.9 and 3.1 mg/g, whereas CYS was determined in only one sample at a significantly lower concentration. In terms of accuracy, the percent recoveries ranged between 91.2 and 105.6% for GSH, and 91.6 and 106.9% for CYS. PMID:23559570

  15. KINETIC STUDIES OF DRUG-PROTEIN INTERACTIONS BY USING PEAK PROFILING AND HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY: EXAMINATION OF MULTI-SITE INTERACTIONS OF DRUGS WITH HUMAN SERUM ALBUMIN COLUMNS

    PubMed Central

    Tong, Zenghan; Schiel, John E.; Papastavros, Efthimia; Ohnmacht, Corey M.; Smith, Quentin R.; Hage, David S.

    2010-01-01

    Carbamazepine and imipramine are drugs that have significant binding to human serum albumin (HSA), the most abundant serum protein in blood and a common transport protein for many drugs in the body. Information on the kinetics of these drug interactions with HSA would be valuable in understanding the pharmacokinetic behavior of these drugs and could provide data that might lead to the creation of improved assays for these analytes in biological samples. In this report, an approach based on peak profiling was used with high-performance affinity chromatography to measure the dissociation rate constants for carbamazepine and imipramine with HSA. This approach compared the elution profiles for each drug and a non-retained species on an HSA column and control column over a board range of flow rates. Various approaches for the corrections of non-specific binding between these drugs and the support were considered and compared in this process. Dissociation rate constants of 1.7 (± 0.2) s-1 and 0.67 (± 0.04) s-1 at pH 7.4 and 37 °C were estimated by this approach for HSA in its interactions with carbamazepine and imipramine, respectively. These results gave good agreement with rate constants that have determined by other methods or for similar solute interactions with HSA. The approach described in this report for kinetic studies is not limited to these particular drugs or HSA but can also be extended to other drugs and proteins. PMID:21067755

  16. Impact of immobilized polysaccharide chiral stationary phases on enantiomeric separations.

    PubMed

    Ali, Imran; Aboul-Enein, Hassan Y

    2006-04-01

    Immobilized polysaccharide-based chiral stationary phases (CSPs) are gaining importance in the resolution of racemic compounds due to their stable nature on working with normal solvents and those prohibited for use with coated phases (tetrahydrofuran, chloroform, dichloromethane, acetone, 1,4-dioxane, ethyl acetate, and certain other ethers). This review discusses the use of immobilized polysaccharide CSPs in the chiral resolution of various racemates by liquid chromatography. The discussion includes immobilization methodologies, enantioselectivities, efficiencies, and a comparison of chiral recognition capabilities of coated vs. immobilized CSPs. Some applications of immobilized CSPs to the chiral resolution of racemic compounds are also presented. PMID:16830488

  17. Recombinant protein purification using gradient-assisted simulated moving bed hydrophobic interaction chromatography. Part I: selection of chromatographic system and estimation of adsorption isotherms.

    PubMed

    Palani, Sivakumar; Gueorguieva, Ludmila; Rinas, Ursula; Seidel-Morgenstern, Andreas; Jayaraman, Guhan

    2011-09-16

    The design of gradient simulated moving bed (SMB) chromatographic processes requires an appropriate selection of the chromatographic system followed by the determination of adsorption isotherm parameters in the relevant range of mobile phase conditions. The determination of these parameters can be quite difficult for recombinant target proteins present in complex protein mixtures. The first part of this work includes the estimation of adsorption isotherm parameters for streptokinase and a lumped impurity fraction present in an Escherichia coli cell lysate for a hydrophobic interaction chromatography (HIC) matrix. Perturbation experiments were carried out using a Butyl Sepharose matrix with purified recombinant protein on buffer equilibrated columns as well as with crude cell lysate saturated columns. The Henry constants estimated for streptokinase were found to exhibit in a wide range a linear dependence on the salt concentration in the mobile phase. These parameters were applied in subsequent investigations to design a simulated moving bed (SMB) process capable to purify in a continuous manner recombinant streptokinase from the E. coli cell lysate. PMID:21816402

  18. Rapid determination of endogenous cytokinins in plant samples by combination of magnetic solid phase extraction with hydrophilic interaction chromatography-tandem mass spectrometry.

    PubMed

    Liu, Zhao; Cai, Bao-Dong; Feng, Yu-Qi

    2012-04-01

    A 2-acrylamido-2-methyl-1-propanesulfonic acid-co-ethylene glycol dimethacrylate (Fe₃O₄/SiO₂/P(AMPS-co-EGDMA)) copolymer was prepared and used as a magnetic solid phase extraction (MSPE) medium for recovery of endogenous cytokinins (CKs) from plant extracts. This magnetic porous polymer was characterized by electron microscopy, nitrogen sorption experiments, elemental analysis and Fourier-transformed infrared spectroscopy. It was demonstrated to have high extraction capacity toward CKs in plants due to its specificity, surface area and porous structure. Coupled with hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-MS/MS), a rapid, simple, and effective MSPE-HILIC-MS/MS analytical method for the quantitative analysis of endogenous CKs in Oryza sativa (O. sativa) roots was successfully established. Good linearities were obtained for all CKs investigated with correlation coefficients (R²>0.9975. The results showed that LODs (S/N=3) were ranged from 0.18 to 3.65 pg mL⁻¹. Reproducibility of the method was obtained with intra-day and inter-day relative standard deviations (RSDs) less than 16.1% and the recoveries in plant samples ranged from 72.8% to 115.5%. Finally, the MSPE-HILIC-MS/MS method was applied to several plant samples, and the amounts of endogenous CKs in O. sativa roots, leaves and Arabidopsis thaliana (A. thaliana) were successfully determined. PMID:22401906

  19. Affinity chromatography using 2' fluoro-substituted RNAs for detection of RNA-protein interactions in RNase-rich or RNase-treated extracts.

    PubMed

    Hovhannisyan, Ruben; Carstens, Russ

    2009-02-01

    Use of RNA affinity chromatography is commonly used to identify RNA binding proteins that interact with specific RNA cis-elements that function in post-transcriptional gene regulation. These purifications can be complicated by residual RNase activity in cellular extracts that can degrade the RNAs on these affinity columns. Furthermore, some proteins may associate indirectly with the column as a component of multi-protein complexes that are "tethered" through the binding of cellular RNAs. We present a protocol for an RNA affinity procedure that can be used in conjunction with RNase-rich or RNase-treated extracts by using RNAs synthesized with 2' fluoro-substituted cytidine triphosphate (CTP) and uridine triphosphate (UTP). The resulting RNAs are shown to be RNase A-resistant and capable of direct coupling to adipic acid dihydrazide agarose beads. Using an RNA cis-element previously shown to bind hnRNP M, we demonstrated that the substituted RNAs preserve binding capability by a common class of RNA binding proteins. Our results provide a method that may be used more generally for RNA affinity purification or as a validation step to verify more direct binding of a given RNA binding protein to a target RNA. PMID:19317654

  20. Hydrophilic interaction liquid chromatography-tandem mass spectrometry methylphosponic and alkyl methylphosphonic acids determination in environmental samples after pre-column derivatization with p-bromophenacyl bromide.

    PubMed

    Baygildiev, T M; Rodin, I A; Stavrianidi, A N; Braun, A V; Lebedev, A T; Rybalchenko, I V; Shpigun, O A

    2016-04-15

    Once exposed to the environment organophosphate nerve agents readily degrade by rapid hydrolysis to the corresponding alkyl methylphosphonic acids which do not exist in nature. These alkyl methylphosphonic acids are finally slowly hydrolyzed to methylphosphonic acid. Methylphosphonic acid is the most stable hydrolysis product of organophosphate nerve agents, persisting in environment for a long time. A highly sensitive method of methylphosphonic acid and alkyl methylphosphonic acids detection in dust and ground mixed samples has been developed and validated. The fact that alkyl methylphosphonic acids unlike methylphosphonic acid did not react with p-bromophenacyl bromide under chosen conditions was discovered. This allowed simultaneous chromatographic separation and mass spectrometric detection of derivatized methylphosphonic acid and underivatized alkyl methylphosphonic acids using HILIC-MS/MS method. Very simple sample pretreatment with high recoveries for each analyte was developed. Methylphosphonic acid pre-column derivate and alkyl methylphosphonic acids were detected using tandem mass spectrometry with electrospray ionization after hydrophilic interaction liquid chromatography separation. The developed approach allows achieving ultra-low detection limits: 200 pg mL(-1) for methylphosphonic acid, 70 pg mL(-1) for ethyl methylphosphonic acid, 8 pg mL(-1) for i-propyl methylphosphonic acid, 8 pg mL(-1) for i-butyl methylphosphonic acid, 5 pg mL(-1) for pinacolyl methylphosphonic acid in the extracts of dust and ground mixed samples. This approach was successfully applied to the dust and ground mixed samples from decommissioned plant for the production of chemical weapons. PMID:26965649

  1. Analysis of 76 veterinary pharmaceuticals from 13 classes including aminoglycosides in bovine muscle by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Dasenaki, Marilena E; Michali, Christina S; Thomaidis, Nikolaos S

    2016-06-24

    A multiresidue/multiclass method for the simultaneous determination of 76 veterinary drugs and pharmaceuticals in bovine muscle tissue has been developed and validated according to the requirements of European Commission Decision 2002/657/EC. The analytes belong in 13 different classes, including aminoglycoside antibiotics, whose different physicochemical properties (extremely polar character) render their simultaneous determination with other veterinary drugs quite problematic. The method combines a two-step extraction procedure (extraction with acetonitrile followed by an acidic aqueous buffer extraction) with hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) determination, allowing confirmation and quantification in a single chromatographic run. Further cleanup with solid phase extraction was performed using polymeric SPE cartridges. A thorough ionization study of aminoglycosides was performed in order to increase their sensitivity and significant differences in the abundance of the precursor ions of the analytes were revealed, depending on the composition of the mobile phase tested. Further gradient elution optimization and injection solvent optimization were performed for all target analytes.The method was validated according to the European Commission Decision 2002/657. Quantitative analysis was performed by means of standard addition calibration. Recoveries varied from 37.4% (bromhexine) to 106% (kanamycin) in the lowest validation level and 82% of the compounds showed recovery >70%. Detection capability (CCβ) varied from 2.4 (salinomycin) to 1302 (apramycin) μgkg(-1). PMID:27215463

  2. Analytical high-performance affinity chromatography: evaluation by studies of neurophysin self-association and neurophysin-peptide hormone interaction using glass matrices

    SciTech Connect

    Swaisgood, H.E.; Chaiken, I.M.

    1986-07-01

    Bovine neurophysin II (BNP II) was covalently immobilized on both nonporous and porous (200-nm pore diameter) glass beads and incorporated in a high-performance liquid chromatograph to evaluate analytical high-performance affinity chromatography as a microscale method for characterizing biomolecular interactions. The self-association of neurophysin and its binding of the peptide hormone vasopressin were characterized by using a single chromatograhic column containing immobilized neurophysin predominantly in the monomer form. Both (/sup 3/H)(Arg/sup 8/)vasopressin (AVP) and /sup 125/I-BNP II were rapidly eluted (<25 min). The relatively symmetrical elution peaks obtained allowed calculation of both equilibrium dissociation constants and kinetic dissociation rate constants. In contrast to the agreement of chromatographic equilibrium binding constants with those measured in solution, the dissociation rate, k..sqrt../sub 3/, determined from the variance of the affinity chromatographic elution profile with nonporous beads, was several orders of magnitude smaller than the solution counterpart. This latter difference may reflect the probability of rebinding to contiguous sites immobilized on a surface, a feature which would be related to that for contiguous sites on a membrane.

  3. Semi-automated identification of N-Glycopeptides by hydrophilic interaction chromatography, nano-reverse-phase LC-MS/MS, and glycan database search.

    PubMed

    Pompach, Petr; Chandler, Kevin B; Lan, Renny; Edwards, Nathan; Goldman, Radoslav

    2012-03-01

    Glycoproteins fulfill many indispensable biological functions, and changes in protein glycosylation have been observed in various diseases. Improved analytical methods are needed to allow a complete characterization of this complex and common post-translational modification. In this study, we present a workflow for the analysis of the microheterogeneity of N-glycoproteins that couples hydrophilic interaction and nanoreverse-phase C18 chromatography to tandem QTOF mass spectrometric analysis. A glycan database search program, GlycoPeptideSearch, was developed to match N-glycopeptide MS/MS spectra with the glycopeptides comprised of a glycan drawn from the GlycomeDB glycan structure database and a peptide from a user-specified set of potentially glycosylated peptides. Application of the workflow to human haptoglobin and hemopexin, two microheterogeneous N-glycoproteins, identified a total of 57 distinct site-specific glycoforms in the case of haptoglobin and 14 site-specific glycoforms of hemopexin. Using glycan oxonium ions and peptide-characteristic glycopeptide fragment ions and by collapsing topologically redundant glycans, the search software was able to make unique N-glycopeptide assignments for 51% of assigned spectra, with the remaining assignments primarily representing isobaric topological rearrangements. The optimized workflow, coupled with GlycoPeptideSearch, is expected to make high-throughput semiautomated glycopeptide identification feasible for a wide range of users. PMID:22239659

  4. Rapid Quantification of Four Anthocyanins in Red Grape Wine by Hydrophilic Interaction Liquid Chromatography/Triple Quadrupole Linear Ion Trap Mass Spectrometry.

    PubMed

    Sun, Yongming; Xia, Biqi; Chen, Xiangzhun; Duanmu, Chuansong; Li, Denghao; Han, Chao

    2015-01-01

    The identification and quantification of four anthocyanins (cyanidin-3-O-glucoside, peonidin-3-O-glucoside, delphinidin-3-O-glucoside, and malvidin-3-O-glucoside) in red grape wine were carried out by hydrophilic interaction liquid chromatography/triple quadrupole linear ion trap MS (HILIC/QTrap-MS/MS). Samples were diluted directly and separated on a Merck ZIC HILIC column with 20 mM ammonium acetate solution-acetonitrile mobile phase. Quantitative data acquisition was carried out in the multiple reaction monitoring mode. Additional identification and confirmation of target compounds were performed using the enhanced product ion mode of the linear ion trap. The LOQs were in the range 0.05-1.0 ng/mL. The average recoveries were in the range 94.6 to 104.5%. The HILIC/QTrap-MS/MS platform offers the best sensitivity and specificity for characterization and quantitative determination of the four anthocyanins in red grape wines and fulfills the quality criteria for routine laboratory application. PMID:26651575

  5. Direct injection human plasma analysis for the quantification of antihypertensive drugs for therapeutic drug monitoring using hydrophilic interaction liquid chromatography/electrospray ionization mass spectrometry.

    PubMed

    Meimaroglou, Stefanos; Vonaparti, Ariadni; Migias, George; Gennimata, Dimitra; Poulou, Sofia; Panderi, Irene

    2015-11-01

    The concept of personalized medicine is related to the development of new sensitive, precise and accurate analytical methods for therapeutic drug monitoring. In this article a rapid, sensitive and specific method was developed for the quantification of aliskiren, losartan, valsartan and hydrochlorothiazide in human plasma. Sample preparation was performed by protein precipitation with acetonitrile followed by filtration. All analytes and the internal standard (tiamulin) were separated by hydrophilic interaction liquid chromatography using an X-Bridge-HILIC analytical column (150.0×2.1mm i.d., particle size 3.5μm) under isocratic elution. The mobile phase was composed of a 10% 5mM ammonium formate water solution pH 4.5, adjusted with formic acid, in acetonitrile and pumped at a flow rate of 0.25mLmin(-1). The assay was linear over the concentration range of 5-500ngmL(-1) for all the analytes. Intermediate precision was less than 5.2% over the tested concentration ranges. The method is the first reported application of HILIC in the analysis antihypertensives in human plasma. With a small sample size (50μL human plasma) and a run time less than 6.0min for each sample the method can be used to support a wide range of clinical studies and therapeutic drug monitoring. PMID:26432361

  6. Testing the capability of a polynomial-modified gaussian model in the description and simulation of chromatographic peaks of amlodipine and its impurity in ion-interaction chromatography.

    PubMed

    Colović, Jelena; Vemić, Ana; Kostić, Nađa; Malenović, Anđelija; Medenica, Mirjana

    2014-07-01

    In this paper, the capability of a polynomial-modified Gaussian model to relate the peak shape of basic analytes, amlodipine, and its impurity A, with the change of chromatographic conditions was tested. For the accurate simulation of real chromatographic peaks the authors proposed the three-step procedure based on indirect modeling of peak width at 10% of peak height (W0.1), individual values of left-half width (A) and right-half width (B), number of theoretical plates (N), and tailing factor (Tf). The values of retention factors corresponding to the peak beginning (k(B)), peak apex (k(A)), peak ending (k(E)), and peak heights (H0) of the analytes were directly modeled. Then, the investigated experimental domain was divided to acquire a grid of appropriate density, which allowed the subsequent calculation of W0.1, A, B, N, and Tf. On the basis of the predicted results for Tf and N, as well as the defined criteria for the simulation the following conditions were selected: 33% acetonitrile/67% aqueous phase (55 mM perchloric acid, pH 2.2) at 40°C column temperature. Perfect agreement between predicted and experimental values was obtained confirming the ability of polynomial modified Gaussian model and three-step procedure to successfully simulate the real chromatograms in ion-interaction chromatography. PMID:24798430

  7. Determination of polar pesticides in olive oil and olives by hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry and high resolution mass spectrometry.

    PubMed

    Nortes-Méndez, Rocío; Robles-Molina, José; López-Blanco, Rafael; Vass, Andrea; Molina-Díaz, Antonio; Garcia-Reyes, Juan F

    2016-09-01

    This article reports the development of two HPLC-MS methods for the determination of polar pesticides in olive oil and olive samples by hydrophilic interaction liquid chromatography (HILIC) separation followed by mass spectrometry detection with tandem mass spectrometry using a triple quadrupole instrument operated in multiple reaction monitoring mode (HILIC-MS/MS) or electrospray time-of-flight mass spectrometry (HILIC-TOFMS). The selected polar pesticides included in the study were: amitrol, cyromazine, diquat, paraquat, mepiquat, trimethylsulfonium (trimesium, glyphosate counterion) and fosetyl aluminium. The simple sample treatment procedure was based on liquid partitioning with methanol. The performance of the sample extraction was evaluated in terms of recovery rates and matrix effects in both olive oil and olives matrices. The results obtained for olive oil were satisfactory while, due to the high complexity of olives, poor recovery rates were obtained for the extraction of diquat, paraquat and amitrol, although with a reasonable precision enabling its use in routine analysis. Similarly, matrix effects were minor in the case of olive oil (ca. 20% suppression average), while significantly higher suppression was observed for olives (30-50% suppression average). The studied approaches were found to be useful for the determination of the pesticides studied in olive oil and olives with limits of quantitation below 5µgkg(-1) in most cases when tandem mass spectrometry was used, thus being in compliance with MRLs set by current EU regulation. PMID:27343599

  8. Study of retention and peak shape in hydrophilic interaction chromatography over a wide pH range.

    PubMed

    McCalley, David V

    2015-09-11

    Retention factor and column efficiency measurements were made for 14 test compounds comprising acids, bases and neutrals on two pairs of amide and bare silica HILIC columns, each pair obtained from a different manufacturer. The columns were tested with up to 6 different mobile phases with acetonitrile-water containing formic (FA), trifluoroacetic (TFA), heptafluorobutyric acids (HFBA) and ammonium salt buffers at w(w)pH 3, 6 and 9. Measurements of mobile phase pH in water (w(w)pH) and in the aqueous-organic mixture (w(s)pH) were performed, and calculations of ionic strength made, in order to aid interpretation of the chromatographic results. Stronger acids like TFA produced very different selectivity compared with ammonium formate buffers at similar aqueous pH. On a given column using TFA as additive, the retention of strongly acidic solutes was considerably increased relative to that of bases. Some bases even showed exclusion on both amide, and on a hybrid silica column. Conversely, in ammonium formate buffers of similar aqueous pH, bases had increased retention compared with acids, particularly on the bare silica columns. This result can be attributed to the higher pH of the salt buffers when measured in the aqueous-organic phase and interaction with negatively charged silanols. It is possible that the silica surface becomes positively charged at the low pH of TFA, leading to anion exchange properties that become competitive with the cation exchange properties normally attributed to silanol dissociation, although other explanations of these results are possible. Very marked selectivity differences were obtained by use of TFA in the mobile phase. Useful selectivity differences may also be obtained with salt buffers at different pH if the use of TFA is not desired due to its relatively unfavourable properties in mass spectrometry. PMID:26275863

  9. ANALYSIS OF PERFLUORINATED CARBOXYLIC ACIDS IN SOILS II: OPTIMIZATION OF CHROMATOGRAPHY AND EXTRACTION

    EPA Science Inventory

    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...

  10. [Determination of the interaction kinetics between meloxicam and β-cyclodextrin using the quantitative high-performance affinity chromatography coupled with mass spectrometry].

    PubMed

    Wang, Cai-fen; Li, Zhuo; Wang, Xiao-bo; Li, Hai-yan; Zhang, Ji-wen; Sun, Li-xin

    2015-09-01

    The association rate constant and dissociation rate constant are important parameters of the drug-cyclodextrin supermolecule systems, which determine the dissociation of drugs from the complex and the further in vivo absorption of drugs. However, the current studies of drug-cyclodextrin interactions mostly focus on the thermodynamic parameter of equilibrium constants (K). In this paper, a method based on quantitative high performance affinity chromatography coupled with mass spectrometry was developed to determine the apparent dissociation rate constant (k(off,app)) of drug-cyclodextrin supermolecule systems. This method was employed to measure the k(off,app) of meloxicam and acetaminophen. Firstly, chromatographic peaks of drugs and non-retained solute (uracil) on β-cyclodextrin column at different flow rates were acquired, and the retention time and variance values were obtained via the fitting the peaks. Then, the plate heights of drugs (H(R)) and uracil (H(M,C)) were calculated. The plate height of theoretical non-retained solute (H(M,T)) was calculated based on the differences of diffusion coefficient and the stagnant mobile phase mass transfer between drugs and uracil. Finally, the k(off,app) was calculated from the slope of the regression equation between (H(R)-H(M,T)) and uk/(1+k)2, (0.13 ± 0.00) s(-1) and (4.83 ± 0.10) s(-1) for meloxicam and acetaminophen (control drug), respectively. In addition, the apparent association rate constant (k(on,app)) was also calculated through the product of K (12.53 L x mol(-1)) and k(off,app). In summary, it has been proved that the method established in our study was simple, efficiently fast and reproducible for investigation on the kinetics of drug-cyclodextrin interactions. PMID:26757555

  11. Spin projection chromatography

    NASA Astrophysics Data System (ADS)

    Danieli, E. P.; Pastawski, H. M.; Levstein, P. R.

    2004-01-01

    We formulate the many-body spin dynamics at high temperature within the non-equilibrium Keldysh formalism. For the simplest XY interaction, analytical expressions in terms of the one particle solutions are obtained for linear and ring configurations. For small rings of even spin number, the group velocities of excitations depend on the parity of the total spin projection. This should enable a dynamical filtering of spin projections with a given parity i.e., a spin projection chromatography.

  12. Influence of carboxylic ion-pairing reagents on retention of peptides in thin-layer chromatography systems with C18 silica-based adsorbents.

    PubMed

    Gwarda, Radosław Ł; Aletańska-Kozak, Monika; Klimek-Turek, Anna; Ziajko-Jankowska, Agnieszka; Matosiuk, Dariusz; Dzido, Tadeusz H

    2016-04-01

    One of the main problems related to chromatography of peptides concerns adverse interactions of their strong basic groups with free silanol groups of the silica based stationary phase. Influence of type and concentration of ion-pairing regents on peptide retention in reversed-phase high-performance liquid chromatography (RP-HPLC) systems has been discussed before. Here we present influence of these mobile phase additives on retention of some peptide standards in high-performance thin-layer chromatography (HPTLC) systems with C18 silica-based adsorbents. We prove, that due to different characteristic of adsorbents used in both techniques (RP HPLC and HPTLC), influence of ion-pairing reagents on retention of basic and/or amphoteric compounds also may be quite different. C18 silica-based HPTLC adsorbents provide more complex mechanism of retention and should be rather considered as mixed-mode adsorbents. PMID:26944833

  13. Lipid Vesicle-mediated Affinity Chromatography using Magnetic Activated Cell Sorting (LIMACS): a Novel Method to Analyze Protein-lipid Interaction

    PubMed Central

    Bieberich, Erhard

    2011-01-01

    The analysis of lipid protein interaction is difficult because lipids are embedded in cell membranes and therefore, inaccessible to most purification procedures. As an alternative, lipids can be coated on flat surfaces as used for lipid ELISA and Plasmon resonance spectroscopy. However, surface coating lipids do not form microdomain structures, which may be important for the lipid binding properties. Further, these methods do not allow for the purification of larger amounts of proteins binding to their target lipids. To overcome these limitations of testing lipid protein interaction and to purify lipid binding proteins we developed a novel method termed lipid vesicle-mediated affinity chromatography using magnetic-activated cell sorting (LIMACS). In this method, lipid vesicles are prepared with the target lipid and phosphatidylserine as the anchor lipid for Annexin V MACS. Phosphatidylserine is a ubiquitous cell membrane phospholipid that shows high affinity to the protein Annexin V. Using magnetic beads conjugated to Annexin V the phosphatidylserine-containing lipid vesicles will bind to the magnetic beads. When the lipid vesicles are incubated with a cell lysate the protein binding to the target lipid will also be bound to the beads and can be co-purified using MACS. This method can also be used to test if recombinant proteins reconstitute a protein complex binding to the target lipid. We have used this method to show the interaction of atypical PKC (aPKC) with the sphingolipid ceramide and to co-purify prostate apoptosis response 4 (PAR-4), a protein binding to ceramide-associated aPKC. We have also used this method for the reconstitution of a ceramide-associated complex of recombinant aPKC with the cell polarity-related proteins Par6 and Cdc42. Since lipid vesicles can be prepared with a variety of sphingo- or phospholipids, LIMACS offers a versatile test for lipid-protein interaction in a lipid environment that resembles closely that of the cell membrane

  14. Hydrophilic interaction chromatography combined with dispersive liquid-liquid microextraction as a preconcentration tool for the simultaneous determination of the panel of underivatized neurotransmitters in human urine samples.

    PubMed

    Konieczna, Lucyna; Roszkowska, Anna; Niedźwiecki, Maciej; Bączek, Tomasz

    2016-01-29

    A simple and sensitive method using dispersive liquid-liquid microextraction (DLLME) followed by liquid chromatography coupled to mass spectrometry (LC-MS) with a hydrophilic interaction chromatography (HILIC) column was developed for the simultaneous determination of 13 compounds of different polarities, comprising monoamine neurotransmitters (dopamine, norepinephrine, epinephrine and serotonin) along with their respective precursors and metabolites, in human urine samples. The microextraction procedure was based on the fast injection of a mixture of ethanol (disperser solvent) and dichloromethane (extraction solvent) into a human urine sample, forming a cloudy solution in the Eppendorf tube. After centrifugation, the sedimented phase was collected and subsequently analyzed by LC-HILIC-MS in about 12min without a derivatization step. The separation was performed on an XBridge Amide™ BEH column 3.0×100mm, 3.5mm and the mobile phase consisted of phase A: 10mM ammonium formate buffer in water pH 3.0 and phase B: 10 mM ammonium formate buffer in acetonitrile, under gradient program elution. Tyrosine, tryptophan, 5-hydroxytryptophan, dopamine, epinephrine, norepinephrine, serotonin, 3-methoxytyramine, 5-hydroxyindole-3-acetic acid, 3,4-dihydroxy-l-phenylalanine and norvaline (internal standard) were detected in the positive ionization mode. While vanillylmandelic acid, homovanillic acid, 3,4-dihydroxyphenylacetic acid and 3,4-dihydroxybenzylamine (internal standard) were detected in the negative ionization mode. Parameters influencing DLLME and LC-HILIC-MS were investigated. Under the optimum conditions, the proposed method exhibited a low detection limit (5-10ngmL(-1)), and good linearity with R between 0.9991 and 0.9998. The recoveries in human urine samples were 99.0%±3.6%. for the 13 studied biogenic amines with intra- and inter-day RSDs of 0.24-9.55% and 0.31-10.0%, respectively. The developed DLLME-LC-MS method could be successfully applied for the

  15. Non-planar microfabricated gas chromatography column

    DOEpatents

    Lewis, Patrick R.; Wheeler, David R.

    2007-09-25

    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.

  16. Simultaneous high-throughput determination of interaction kinetics for drugs and cyclodextrins by high performance affinity chromatography with mass spectrometry detection.

    PubMed

    Wang, Caifen; Wang, Xiaobo; Xu, Xiaonan; Liu, Botao; Xu, Xu; Sun, Lixin; Li, Haiyan; Zhang, Jiwen

    2016-02-25

    The individual determination of the apparent dissociation rate constant (kd,app) using high performance affinity chromatography (HPAC) is a tedious process requiring numerous separate tests and massive data fitting, unable to provide the apparent association rate constant (ka) and equilibrium binding constant (Ka). In this study, a HPAC with mass spectrometry detection (HPAC-MS/MS) was employed to determine the drug-cyclodextrin (CD) interaction kinetics with low sample loading quantity (<10 ng per injection for single compound) and high-throughput yield as twenty drugs determined in one injection. The kd,app measured by HPAC-MS/MS approach were 0.89 ± 0.07, 4.34 ± 0.01, 1.48 ± 0.01 and 7.77 ± 0.04 s(-1) for ketoprofen, trimethoprim, indapamide and acetaminophen, with kd,app for acetaminophen consistent with that from the HPAC method with UV detector in our previous studies. For twenty drugs with diverse structures and chemical properties, good correlationship was found between kd,app measured by single compound analysis method and high-throughput HPAC-MS/MS approach, with the correlation coefficient of 0.987 and the significance F less than 0.001. Comprehensive quantification of ka,app, kd,app and Ka values was further performed based on the measurement of kd,app by peak profiling method and Ka by the peak fitting method. And the investigation of the drug-CD interaction kinetics under different conditions indicated that the column temperature and mobile phase composition significantly affected the determination of ka,app, kd,app and Ka while also dependent on the acidity and basicity of drugs. In summary, the high-throughput HPAC-MS/MS approach has been demonstrated high efficiency in determination of the drug-CD primary interaction kinetic parameter, especially, kd,app, being proven as a novel tool in screening the right CD for the solubilization of the right drug. PMID:26851087

  17. New trends in fast liquid chromatography for food and environmental analysis.

    PubMed

    Núñez, Oscar; Gallart-Ayala, Héctor; Martins, Claudia P B; Lucci, Paolo

    2012-03-01

    There is an increasing need for applications in food and environmental areas able to cope with a large number of analytes in very complex matrices. The new analytical procedures demand sensitivity, robustness and high resolution within an acceptable analysis time. The purpose of this review is to describe new trends based on fast liquid chromatography applied to the food and environmental analysis. It includes different column technologies, such as monolithic, sub-2 μm, porous shell, as well as different stationary phases such as reversed phase (C8 and C18), hydrophilic interaction liquid chromatography (HILIC) and fluorinated columns. Additionally, recent sample extraction and clean-up methodologies applied to reduce sample manipulation and total analysis time in food and environmental analysis--QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe), on line solid phase extraction coupled to ultrahigh pressure liquid chromatography (on line SPE-UHPLC), turbulent flow chromatography (TFC) and molecularly imprinted polymers (MIPs), were also addressed. The advantages and drawbacks of these methodologies applied to the fast and sensitive analyses of food and environmental samples are going to be discussed. PMID:22153282

  18. Forced-flow planar chromatography in the rear view mirror.

    PubMed

    Kalász, Huba

    2015-03-01

    Mobile phase progress in planar stationary phase can be evoked by either external or internal forces. An internal force is capillarity, while gravity, electric field, a pump and centrifugal forces belong to external forces. Overpressured layer chromatography gives a widely used special chapter of forced-flow planar chromatography, a special bridge between high-performance liquid column chromatography and thin-layer chromatography (TLC). A simple and special rule characterizes the progress of mobile phase. Optimal efficiency is composed by the doubled effect of flow resulting from the pump-forced mobile phase (convex profile of laminar flow) and capillary forces on the dry stationary phase (concave laminar flow). This review describes the most important aspects of forced-flow TLC, including how the set-ups are developed and also the progress of detection methods used. PMID:25681205

  19. Nucleophilic substitution in preparation and surface modification of hypercrosslinked stationary phases.

    PubMed

    Janků, Simona; Škeříková, Veronika; Urban, Jiří

    2015-04-01

    Four linear diaminoalkanes (1,2-diaminoethane, 1,4-diaminobutane, 1,6-diaminohexane, and 1,8-diaminooctane) have been used to hypercrosslink poly(styrene-co-vinylbenzyl chloride-co-divinylbenzene) monolithic stationary phases by nucleophilic substitution reaction. The column efficiency of polymer monoliths improved with longer diaminoalkane with 1,8-diaminoctane providing the highest efficiency. The concentration of 1,8-diaminoctane, together with hypercrosslinking time and temperature has been optimized. To improve the permeability of prepared columns, the hypercrosslinking modification has been combined with an early termination of polymerization reaction and decrease in polymerization temperature. The optimal column has been prepared by a polymerization reaction for 2h at 65°C and hypercrosslinked in the presence of 3% 1,8-diaminooctane for 2h at 95°C. The repeatability study of the presented protocol provided relative standard deviation for nine columns prepared independently out of three individual polymerization mixtures in between 2.0-12.0% for retention factors and 1.5-6.5% for plate heights, respectively. Further, we have modified residual chloromethyl groups with 2-aminoethanesulfonic acid (taurine) to prepare monolithic columns suitable for separation of small polar molecules in hydrophilic interaction chromatography. The highest retention of polar thiourea showed the column modified at 70°C for 20 h. Taurine-modified hypercrosslinked column showed the minimum of van Deemter curve of 20 μm. The prepared column provided dual-retention mechanism, including hydrophilic interaction and reversed-phase liquid chromatography that can be controlled by the composition of the mobile phase. The prepared column has been successfully used for an isocratic separation of low-molecular phenolic acids. PMID:25728663

  20. High efficiency stationary hydrogen storage

    SciTech Connect

    Hynek, S.; Fuller, W.; Truslow, S.

    1995-09-01

    Stationary storage of hydrogen permits one to make hydrogen now and use it later. With stationary hydrogen storage, one can use excess electrical generation capacity to power an electrolyzer, and store the resultant hydrogen for later use or transshipment. One can also use stationary hydrogen as a buffer at fueling stations to accommodate non-steady fueling demand, thus permitting the hydrogen supply system (e.g., methane reformer or electrolyzer) to be sized to meet the average, rather than the peak, demand. We at ADL designed, built, and tested a stationary hydrogen storage device that thermally couples a high-temperature metal hydride to a phase change material (PCM). The PCM captures and stores the heat of the hydriding reaction as its own heat of fusion (that is, it melts), and subsequently returns that heat of fusion (by freezing) to facilitate the dehydriding reaction. A key component of this stationary hydrogen storage device is the metal hydride itself. We used nickel-coated magnesium powder (NCMP) - magnesium particles coated with a thin layer of nickel by means of chemical vapor deposition (CVD). Magnesium hydride can store a higher weight fraction of hydrogen than any other practical metal hydride, and it is less expensive than any other metal hydride. We designed and constructed an experimental NCM/PCM reactor out of 310 stainless steel in the form of a shell-and-tube heat exchanger, with the tube side packed with NCMP and the shell side filled with a eutectic mixture of NaCL, KCl, and MgCl{sub 2}. Our experimental results indicate that with proper attention to limiting thermal losses, our overall efficiency will exceed 90% (DOE goal: >75%) and our overall system cost will be only 33% (DOE goal: <50%) of the value of the delivered hydrogen. It appears that NCMP can be used to purify hydrogen streams and store hydrogen at the same time. These prospects make the NCMP/PCM reactor an attractive component in a reformer-based hydrogen fueling station.

  1. High-throughput wide dynamic range procedure for the simultaneous quantification of nicotine and cotinine in multiple biological matrices using hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Pérez-Ortuño, Raúl; Martínez-Sánchez, Jose M; Fernández, Esteve; Pascual, José A

    2015-11-01

    A straightforward, high-throughput method was developed and fully validated for the simultaneous determination of the specific tobacco biomarkers nicotine and its main metabolite cotinine in a wide dynamic range and supporting the most common human biological matrices (urine, oral fluid and hair). Sample preparation was performed inside the very HPLC injection vials by pipetting 0.5 mL of the liquid samples, deuterated internal standards in alkaline solution and dichloromethane as extraction solvent. Solid samples (i.e. around 10 mg hair) were first submitted to alkaline digestion in the HPLC vials and processed accordingly. The organic phase (reached through the upper aqueous layer) was directly injected without further treatment. Instrumental analysis was performed using hydrophilic interaction (HILIC) ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Total chromatographic time was 2 min. The method covers a wide dynamic range making it fit-for-purpose for the analysis of samples covering entire populations, irrespective of the level of exposure or tobacco use. Calibration curves (r (2) > 0.995) covered the range 1-2000 ng/mL (or 0.05-100 ng/mg hair) for nicotine and 0.1-2000 ng/mL (or 0.005-100 ng/mg hair) for cotinine. Within-run and between-run precision and accuracy were typically below 10 %, and always below 20 % at the lower limit of quantification. The method was successfully applied to the analysis of samples from different projects involving multiple matrices. PMID:26342312

  2. Selective quantitation of the neurotoxin BMAA by use of hydrophilic-interaction liquid chromatography-differential mobility spectrometry-tandem mass spectrometry (HILIC-DMS-MS/MS).

    PubMed

    Beach, Daniel G; Kerrin, Elliott S; Quilliam, Michael A

    2015-11-01

    The neurotoxin β-N-methylamino-L-alanine (BMAA) has been reported in cyanobacteria and shellfish, raising concerns about widespread human exposure. However, inconsistent results for BMAA analysis have led to controversy. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is the most appropriate method for analysis of BMAA, but the risk of interference from isomers, other sample components, and the electrospray background is still present. We have investigated differential mobility spectrometry (DMS) as an ion filter to improve selectivity in the hydrophilic interaction liquid chromatographic (HILIC)-MS/MS determination of BMAA. We obtained standards for two BMAA isomers not previously analyzed by HILIC-MS, β-amino-N-methylalanine and 3,4-diaminobutanoic acid, and the typically used 2,4-diaminobutanoic acid and N-(2-aminoethyl)glycine. DMS separation of BMAA from these isomers was achieved and optimized conditions were used to develop a sensitive and highly selective multidimensional HILIC-DMS-MS/MS method. This work revealed current technical limitations of DMS for trace quantitation, and practical solutions were implemented. Accurate control of low levels of DMS carrier gas modifier was essential, but required external metering. The linearity of our optimized method was excellent from 0.01 to 6 μmol L(-1). The instrumental LOD was 0.4 pg BMAA injected on-column and the estimated method LOD was 20 ng g(-1) dry weight for BMAA in sample matrix. The method was used to analyze cycad plant tissue, a cyanobacterial reference material, and mussel tissues, by use of isotope-dilution quantitation with deuterated BMAA. This confirmed the presence of BMAA and several of its isomers in cycad and mussel tissues, including commercially available mussel tissue reference materials certified for other biotoxins. Graphical Abstract Differential Mobility Spectrometry is used to increases the selectivity of BMAA analysis by HILIC-MS/MS. PMID:26396078

  3. Salting-out assisted extraction method coupled with hydrophilic interaction liquid chromatography for determination of selected β-blockers and their metabolites in human urine.

    PubMed

    Magiera, Sylwia; Kolanowska, Anna; Baranowski, Jacek

    2016-06-01

    In this study, a new analytical method was developed and validated for the simultaneous analysis of β-blockers (metoprolol, propranolol, carvedilol) and their metabolites (5'-hydroxycarvedilol, O-desmethylcarvedilol, α-hydroxymetoprolol, O-desmethylmetoprolol, 5-hydroxypropranolol) in human urine. A salting-out assisted liquid-liquid extraction (SALLE) procedure was used for sample preparation. Several parameters affecting the extraction efficiency and method sensitivity including the type and volume of the extraction solvent, the type and quantity of the inorganic salt, extraction time and sample pH were investigated. Hydrophilic interaction liquid chromatography-ultraviolet detection (HILIC-UV) was used for the determination of all analytes. During method development, the effects of mobile phase components (type, pH, concentration of salt, organic modifier type and content, flow rate, column temperature) on the retention and separation of β-blockers and metabolites on the five different HILIC columns were examined. The method was linear for concentrations ranging from 0.1 to 8.0μg/mL, with determination coefficients higher than 0.993 for all analytes. The limits of quantification were in the range from 0.1 to 0.2μg/mL. Intra- and inter-day precision ranged from 0.1 to 8.9%, and accuracy was within±13% interval for all analytes. Under the optimized conditions, extraction efficiency was greater than 83.4% for determined compounds. The validated method was then applied to the measurement of β-blockers and their metabolites in human urine samples. PMID:27085018

  4. Structural characterization and profiling of lyso-phospholipids in fresh and in thermally stressed mussels by hydrophilic interaction liquid chromatography-electrospray ionization-Fourier transform mass spectrometry.

    PubMed

    Facchini, Laura; Losito, Ilario; Cianci, Costantina; Cataldi, Tommaso R I; Palmisano, Francesco

    2016-07-01

    The separation efficiency of hydrophilic interaction liquid chromatography and the high resolution/accuracy of electrospray ionization-Fourier transform MS were successfully applied to the detailed characterization of lyso-phosphatidylcholines (LPCs) and lyso-phosphatidylethanolamines (LPEs) contained in the lipid extracts of Mytilus galloprovincialis (Mediterranean mussel). As a result, 57 LPCs, including regio- and positional isomers, and 45 LPEs, including acyl and plasma(e)nyl species, were identified. Four lyso-phosphonocholines were also identified among mussel Lyso-Phospholipids. To the best of our knowledge this represents the first characterization, at a molecular level, ever reported for LPEs in mussels. No significant variation was observed in the composition of both LPCs and LPEs when mussels were refrigerated at +4°C for up to 48 h, i.e. under conditions usually employed for seafood transportation and storage. Treatments mimicking more severe thermal stresses, namely eight day-refrigeration at + 4°C, two week-freezing at -15°C and 6 h-storage at 25°C, resulted in a significant increase in the molar abundance of LPCs and LPEs (expressed with respect to that of their precursors, PCs and PEs, respectively) and was accompanied by the death of all or part of the molluscs. These results were interpreted invoking the generation of lyso-phospholipids, mediated by endogenous phospholipases, as an intermediate process toward the partial replacement of side chains in phospholipids, perhaps functional to a better adaptation of mussels to adverse temperature conditions. Interestingly, the relative abundances of specific compounds belonging to the LPC and LPE classes were found to follow the seasonal variations of sea temperature. PMID:26860242

  5. Determination of glycine in biofluid by hydrophilic interaction chromatography coupled with tandem mass spectrometry and its application to the quantification of glycine released by embryonal carcinoma stem cells.

    PubMed

    Tang, Ya-Bin; Teng, Lin; Sun, Fan; Wang, Xiao-Lin; Peng, Liang; Cui, Yong-Yao; Hu, Jin-Jia; Luan, Xin; Zhu, Liang; Chen, Hong-Zhuan

    2012-09-15

    Because glycine plays a prominent role in living creatures, an accurate and precise quantitative analysis method for the compound is needed. Herein, a new approach to analyze glycine by hydrophilic interaction chromatography (HILIC) coupled with electrospray ionization tandem mass spectrometry (ESI-MS/MS) was developed. This method avoids the use of derivatization and/or ion-pairing reagents. N-methyl-D-aspartate (NMDA) is used as the internal standard (IS). The mobile phase for the isocratic elution consisted of 10 mM ammonium formate in acetonitrile-water (70:30, v/v, adjusted to pH 2.8 with formic acid), and a flow rate of 250 μL/min was used. Two microliters of sample was injected for analysis. The signal was monitored in the positive multiple reaction monitoring (MRM) mode. The total run time was 5 min. The dynamic range was 40-2000 ng/mL for glycine in the biological matrix. The LLOQ (lower limit of quantification) of this method was 40 ng/mL (80 pg on column). The validated method was applied to determine the dynamic release of glycine from P19 embryonal carcinoma stem cells (ECSCs). Glycine spontaneously released from the ECSCs into the intercellular space gradually increased from 331.02±60.36 ng/mL at 2 min in the beginning to 963.52±283.80 ng/mL at 60 min and 948.27±235.09 ng/mL at 120 min, finally reaching a plateau, indicating that ECSCs consecutively release glycine until achieving equilibration between the release and the reuptake of the compound; on the contrary, the negative control NIH/3T3 embryonic fibroblast cells did not release glycine. This finding will help to improve our understanding of the novel effects of neurotransmitters, including glycine, on non-neural systems. PMID:22906796

  6. Comparison of hydrophilic interaction and reversed phase liquid chromatography coupled with tandem mass spectrometry for the determination of eight artificial sweeteners and common steviol glycosides in popular beverages.

    PubMed

    Kubica, Paweł; Namieśnik, Jacek; Wasik, Andrzej

    2016-08-01

    Hydrophilic interaction liquid chromatography (HILIC) coupled with tandem mass spectrometry (MS/MS) was used to separate artificial and natural sweeteners approved for use in European Union (EU). Among three tested HILIC columns (BlueOrchid PAL-HILIC, Ascentis Express Si and Acclaim™ Trinity™ P2) the last one was selected for the development of HILIC method due to the best results obtained with it. Early eluting and coeluting compounds in HILIC (acesulfame-K, saccharin, cyclamate, sucralose and aspartame) were successfully separated by the HILIC-based approach for the first time. The developed HILIC method allows for determination of all high potency sweeteners in one analytical run. The calibration curves for all analytes had good linearity within the tested ranges. The limits of detection and quantitation were in the range 0.81-3.30ng/mL and 2.32-9.89ng/mL, respectively. The obtained recoveries used for trueness and precision estimation were from 98.6% to 106.2% with standard deviation less than 4.1%. Sample preparation was reduced to a necessary minimum and contained only proper dilution and centrifugation. More than twenty samples of beverages were analyzed with the developed HILIC method. Finally, the chromatographic parameters of peaks (reduced retention time, width at baseline, width at 50% of peak height, tailing factor and efficiency) obtained in HILIC mode and in RPLC mode were compared. Developed HILIC method along with RPLC method can be applied for rapid evaluation of sweeteners' content, quality and safety control. PMID:26782293

  7. Determination of vanadium as 4-(2-pyridylazo)resorcinol-hydrogen peroxide ternary complexes by ion-interaction reversed-phase liquid chromatography.

    PubMed

    Vachirapatama, Narumol; Dicinoski, Greg W; Townsend, Ashley T; Haddad, Paul R

    2002-05-17

    The separation and determination of the vanadium(V) ternary complex formed with 4-(2-pyridylazo)resorcinol (PAR) and hydrogen peroxide using ion-interaction reversed-phase high-performance liquid chromatography on a C18 column has been investigated. The optimal mobile phase was a methanol-water solution (32:68, v/v) containing 3 mM tetrabutylammonium bromide, 5 mM acetic acid and 5 mM citrate buffer at pH 7, with absorbance detection at 540 nm. The stoichiometry of the ternary complex of vanadium at pH 6 in 10 mM acetate buffer using the mole ratio and Job's method by HPLC indicated that the mole ratio of V(V):PAR:H2O2 was 1:1:1. The optimal conditions for precolumn formation of the ternary complex were 10 mM acetate, 7 mM H2O2, 0.3 mM PAR, and pH 6. The method gave relative standard deviations of retention time, peak area and peak height for the ternary complex of 0.187, 0.45 and 0.57%, respectively. The detection limit (at a signal-to-noise ratio of 3) for V(V) was 0.09 ng/ml in the digested sample using a 100-microl injection loop (or 0.09 microg/g in the solid fertiliser sample). The method was applied to the analysis of fertilisers (phosphate rocks and nitrogen, phosphorus and potassium fertiliser). The results for vanadium obtained by the HPLC method agreed well with those from magnetic sector inductively coupled plasma MS analysis. PMID:12108654

  8. Determination of salivary cotinine through solid phase extraction using a bead-injection lab-on-valve approach hyphenated to hydrophilic interaction liquid chromatography.

    PubMed

    Ramdzan, Adlin N; Barreiros, Luísa; Almeida, M Inês G S; Kolev, Spas D; Segundo, Marcela A

    2016-01-15

    Cotinine, the first metabolite of nicotine, is often used as a biomarker in the monitoring of environmental tobacco smoke (ETS) exposure due to its long half-life. This paper reports on the development of an at-line automatic micro-solid phase extraction (μSPE) method for the determination of salivary cotinine followed by its analysis via hydrophilic interaction liquid chromatography (HILIC). The SPE methodology is based on the bead injection (BI) concept in a mesofluidic lab-on-valve (LOV) flow system to automatically perform all SPE steps. Three commercially available reversed-phase sorbents were tested, namely, Oasis HLB, Lichrolut EN and Focus, and the spherically shaped sorbents (i.e., Oasis HLB and Focus) provided better packing within the SPE column and hence higher column efficiency. An HILIC column was chosen based on its potential for achieving higher sensitivity and better retention of polar compounds such as cotinine. The method uses an isocratic program with acetonitrile:100mM ammonium acetate buffer, pH 5.8 in 95:5 v/v ratio as the mobile phase at a flow rate of 1.0 mL min(-1). Using this approach, the linear calibration range was from 10 to 1000 ng which corresponded to 5-500 μg L(-1). The corresponding μSPE-BI-LOV system was proven to be reliable in the handing and analysis of viscous biological samples such as saliva, achieving a sampling rate of 6h(-1) and a limit of detection and quantification of 1.5 and 3μgL(-1), respectively. PMID:26747690

  9. Mildronate (Meldonium) in professional sports - monitoring doping control urine samples using hydrophilic interaction liquid chromatography - high resolution/high accuracy mass spectrometry.

    PubMed

    Görgens, Christian; Guddat, Sven; Dib, Josef; Geyer, Hans; Schänzer, Wilhelm; Thevis, Mario

    2015-01-01

    To date, substances such as Mildronate (Meldonium) are not on the radar of anti-doping laboratories as the compound is not explicitly classified as prohibited. However, the anti-ischemic drug Mildronate demonstrates an increase in endurance performance of athletes, improved rehabilitation after exercise, protection against stress, and enhanced activations of central nervous system (CNS) functions. In the present study, the existing evidence of Mildronate's usage in sport, which is arguably not (exclusively) based on medicinal reasons, is corroborated by unequivocal analytical data allowing the estimation of the prevalence and extent of misuse in professional sports. Such data are vital to support decision-making processes, particularly regarding the ban on drugs in sport. Due to the growing body of evidence (black market products and athlete statements) concerning its misuse in sport, adequate test methods for the reliable identification of Mildronate are required, especially since the substance has been added to the 2015 World Anti-Doping Agency (WADA) monitoring program. In the present study, two approaches were established using an in-house synthesized labelled internal standard (Mildronate-D3 ). One aimed at the implementation of the analyte into routine doping control screening methods to enable its monitoring at the lowest possible additional workload for the laboratory, and another that is appropriate for the peculiar specifics of the analyte, allowing the unequivocal confirmation of findings using hydrophilic interaction liquid chromatography-high resolution/high accuracy mass spectrometry (HILIC-HRMS). Here, according to applicable regulations in sports drug testing, a full qualitative validation was conducted. The assay demonstrated good specificity, robustness (rRT=0.3%), precision (intra-day: 7.0-8.4%; inter-day: 9.9-12.9%), excellent linearity (R>0.99) and an adequate lower limit of detection (<10 ng/mL). PMID:25847280

  10. Two complementary liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods to study the excretion and metabolic interaction of edaravone and taurine in rats.

    PubMed

    Tang, Dao-quan; Zheng, Xiao-xiao; Li, Yin-jie; Bian, Ting-ting; Yu, Yan-yan; Du, Qian; Yang, Dong-zhi; Jiang, Shui-shi

    2014-11-01

    In this study, two independent and complementary liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods were respectively developed and validated for the determination of edaravone or taurine in rat urine, feces and bile after intravenous administration, using 3-methyl-l-p-tolyl-5-pyrazolone and sulfanilic acid as the internal standards (IS). Edaravone was separated on an Agilent Eclipse Plus C18 column (100×2.1 mm, 3.5 μm) using methanol and water (containing 5 mM ammonium formate and 0.02% formic acid) as mobile phase, while taurine was performed on a Waters Atlantis HILIC Silica column (150×2.1 mm, 3 μm) using acetonitrile and water (containing 5mM ammonium formate and 0.2% formic acid) as mobile phase. The mass analysis was performed in a Triple Quadrupole mass spectrometer via multiple reaction monitoring (MRM) with negative ionization mode. The optimized mass transition ion pairs (m/z) for quantification were 173.1→92.2 and 187.2→106.0 for edaravone and its IS, 124.1→80.0 and 172.0→80.0 for taurine and its IS, respectively. The validated methods have been successfully applied to the excretion and metabolism interaction study of edaravone and taurine in rats after independent intravenous administration and co-administration with a single dose. The results demonstrated that there were no significant alternations on the metabolism and cumulative excretion rate of edaravone and taurine, implying that the proposed combination therapy was pharmacologically viable. PMID:25222743

  11. Rapid determination of chlormequat in meat by dispersive solid-phase extraction and hydrophilic interaction liquid chromatography (HILIC)-electrospray tandem mass spectrometry.

    PubMed

    Li, Chunmei; Jin, Fen; Yu, Zhiyong; Qi, Yamei; Shi, Xiaomei; Wang, Miao; Shao, Hua; Jin, Maojun; Wang, Jing; Yang, Mingqi

    2012-07-11

    A rapid method for analyzing trace levels of chlormequat (CQ) in meat samples by hydrophilic interaction liquid chromatography (HILIC)-electrospray tandem mass spectrometry was developed. The samples were extracted with acetonitrile, followed by a rapid cleanup through a dispersive solid-phase extraction (DSPE) technique with octadecyl (C18) DSPE sorbents. The chromatographic separation was achieved within 6 min using a HILIC column with 10 mM ammonium acetate and 0.1% (v/v) formic acid in water/acetonitrile (v/v, 40:60) as the mobile phase. Quantification was performed using a matrix-matched calibration curve, which was linear in the range of the 0.05-100 μg/L. The limit of detection (LOD) was estimated at 0.03 μg/kg for CQ on the basis of a peak to peak signal noise (S/N = 3). The limit of quantification (LOQ) was 0.1 μg/kg on the basis of the lowest spiked concentration with suitable precision and accuracy. The average recovery of CQ in spiked meat samples was 86.4-94.7% at 2, 20, and 200 μg/kg. Finally, this method was applied to determine CQ in the livestock and poultry meats purchased from markets in Beijing in 2011. CQ was detected in all 12 samples, and the concentration was 0.4-636.0 μg/kg. Concentrations in a chicken sample (636.0 μg/kg) and a goat meat sample (486.0 μg/kg) were found to be 15.9 and 2.43 times the corresponding Codex maximum residue limits, respectively. PMID:22686367

  12. Less common applications of monoliths III. Gas chromatography

    PubMed Central

    Svec, Frantisek; Kurganov, Alexander A.

    2008-01-01

    Porous polymer monoliths emerged about two decades ago. Despite this short time, they are finding applications in a variety of fields. In addition to the most common and certainly best known use of this new category of porous media as stationary phases in liquid chromatography, monolithic materials also found their applications in other areas. This review article focuses on monoliths in capillaries designed for separations in gas chromatography. PMID:17645884

  13. Landau superfluids as nonequilibrium stationary states

    SciTech Connect

    Wreszinski, Walter F.

    2015-01-15

    We define a superfluid state to be a nonequilibrium stationary state (NESS), which, at zero temperature, satisfies certain metastability conditions, which physically express that there should be a sufficiently small energy-momentum transfer between the particles of the fluid and the surroundings (e.g., pipe). It is shown that two models, the Girardeau model and the Huang-Yang-Luttinger (HYL) model, describe superfluids in this sense and, moreover, that, in the case of the HYL model, the metastability condition is directly related to Nozières’ conjecture that, due to the repulsive interaction, the condensate does not suffer fragmentation into two (or more) parts, thereby assuring its quantum coherence. The models are rigorous examples of NESS in which the system is not finite, but rather a many-body system.

  14. Separation and identification of oligomeric phenylethoxysiloxanols by liquid chromatography-electrospray ionization mass spectrometry.

    PubMed

    Jiang, Yuting; Wan, Qian-Hong

    2015-05-15

    Liquid chromatography-electrospray ionization mass spectrometry has been applied to qualitative analysis of oligomeric phenylethoxysiloxanols, a class of organosilanols as active intermediates to polyhedral silsesquioxanes. The phenylethoxysiloxanol samples were prepared by controlled acid-catalyzed hydrolysis and condensation of phenyltriethoxysilane at various molar equivalents of water (r1) and characterized by standard spectroscopic techniques. Using a gradient binary water-methanol mobile phase, these reaction products were resolved on octadecylsiloxane silica stationary phase and subsequently identified by online electrospray ionization mass spectrometric detection. Results show that the reaction products are composed of a multitude of linear and monocyclic siloxanol oligomers with various numbers of silicon atoms and hydroxyl groups, depending upon the reaction conditions used. With the r1 value increasing from 0.5 to 2.0, the chain lengths of the oligomers increase slightly but the numbers of hydroxyl groups increase considerably, accompanying by structural evolution from chains to rings. Characterization of the retention behavior of these oligomers indicates that hydrophobic interactions of phenyl and ethoxy groups with the stationary phase are responsible for their retention in reversed-phase liquid chromatography. PMID:25843423

  15. Gas Chromatography

    NASA Astrophysics Data System (ADS)

    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.

  16. Impact of injection solvents on supercritical fluid chromatography.

    PubMed

    Abrahamsson, Victor; Sandahl, Margareta

    2013-09-01

    Even though there has been a rapid development in instrumentation and applications of supercritical fluid chromatography (SFC), relatively little is known about retention mechanisms compared to high-performance liquid chromatography (HPLC). Much effort has been made to characterize the influence of injection solvents on chromatographic efficiency in HPLC, however has been left rather uninvestigated in the domain of SFC. In this study properties of different injection solvents have been studied and correlated with properties of seven various analytes on three different columns, a C18, a 2-ethylpyridine and a bare-silica column. Aided by calculations of correlation coefficients and principal component analysis (PCA), the physical properties of injection solvents and the interactions between injection solvent, solute and stationary phase were investigated. The findings of this work shows that interactions capable of masking accessible silanol groups on a C18 column are of importance in order to maximize the plate number. While solvents with dipolar and hydrogen bond interaction properties are associated negatively with chromatographic efficiency using polar columns. Properties such as molar density, vapor pressure and boiling point were related to sharper peaks, mostly likely because of solubility issues of the injection solvent into the methanol-modified carbon dioxide. However, no additional solubility due to hydrogen interactions between the injection solvent and the carbon dioxide in SFC was observed. Surface tension and viscosity was not particularly associated with a decrease in plate numbers. By increasing the injection volume a stronger correlation between solubility related properties and plate numbers were obtained. Additional experiments showed that the resistance in solubility became an issue when performing partial-loop injection where additional washing solvent entered the system, thus providing broadened peaks. PMID:23899383

  17. Fractionation of different PEGylated forms of a protein by chromatography using environment-responsive membranes.

    PubMed

    Yu, Deqiang; Shang, Xiaojiao; Ghosh, Raja

    2010-08-27

    PEGylation of therapeutic proteins can enhance their efficacy as biopharmaceuticals through increased stability and hydrophilicity, and decreased immunogenicity. A site-specific PEGylated protein (e.g. mono-PEGylated at N-terminus) is frequently desirable as a product. However, multiple-PEGylated forms are frequently produced as byproducts. In this paper we discuss the fractionation of the different PEGylated forms of a protein by hydrophobic interaction chromatography using a stack of hydrophilized PVDF membrane, which has been shown to be environment responsive, as stationary phase. With the model protein examined in this study (i.e. lysozyme), the apparent hydrophobicity in the presence of a lyotropic salt increased with the degree of PEGylation. Based on this, unmodified lysozyme and its mono-, di- and tri-PEGylated forms could each be resolved into separate chromatographic peaks. Such fractionation was not feasible using conventional hydrophobic interaction chromatography using a butyl column. The use of membrane chromatography also ensured that the fractionation was fast and hence suitable for analytical applications such as product purity determination and monitoring of the extent of PEGylation reactions. PMID:20638664

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

    SciTech Connect

    Dumont, P.J.

    1996-04-23

    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.

  19. Development, validation and application of a hydrophilic interaction liquid chromatography-evaporative light scattering detection based method for process control of hydrolysis of xylans obtained from different agricultural wastes.

    PubMed

    Li, Fangbing; Wang, Hui; Xin, Huaxia; Cai, Jianfeng; Fu, Qing; Jin, Yu

    2016-12-01

    Purified standards of xylooligosaccharides (XOSs) (DP2-6) were first prepared from a mixture of XOSs using solid phase extraction (SPE), followed by semi-preparative liquid chromatography both under hydrophilic interaction liquid chromatography (HILIC) modes. Then, an accurate quantitative analysis method based on hydrophilic interaction liquid chromatography-evaporative light scattering detection (HILIC-ELSD) was developed and validated for simultaneous determination of xylose (X1), xylobiose (X2), xylotriose (X3), xylotetraose (X4), xylopentaose (X5), and xylohexaose (X6). This developed HILIC-ELSD method was applied to the comparison of different hydrolysis methods for xylans and assessment of XOSs contents from different agricultural wastes. The result indicated that enzymatic hydrolysis was preferable with fewer by-products and high XOSs yield. The XOSs yield (48.40%) from sugarcane bagasse xylan was the highest, showing conversions of 11.21g X2, 12.75g X3, 4.54g X4, 13.31g X5, and 6.78g X6 from 100g xylan. PMID:27374519

  20. Method developments approaches in supercritical fluid chromatography applied to the analysis of cosmetics.

    PubMed

    Lesellier, E; Mith, D; Dubrulle, I

    2015-12-01

    Analyses of complex samples of cosmetics, such as creams or lotions, are generally achieved by HPLC. These analyses are often multistep gradients, due to the presence of compounds with a large range of polarity. For instance, the bioactive compounds may be polar, while the matrix contains lipid components that are rather non-polar, thus cosmetic formulations are usually oil-water emulsions. Supercritical fluid chromatography (SFC) uses mobile phases composed of carbon dioxide and organic co-solvents, allowing for good solubility of both the active compounds and the matrix excipients. Moreover, the classical and well-known properties of these mobile phases yield fast analyses and ensure rapid method development. However, due to the large number of stationary phases available for SFC and to the varied additional parameters acting both on retention and separation factors (co-solvent nature and percentage, temperature, backpressure, flow rate, column dimensions and particle size), a simplified approach can be followed to ensure a fast method development. First, suited stationary phases should be carefully selected for an initial screening, and then the other operating parameters can be limited to the co-solvent nature and percentage, maintaining the oven temperature and back-pressure constant. To describe simple method development guidelines in SFC, three sample applications are discussed in this paper: UV-filters (sunscreens) in sunscreen cream, glyceryl caprylate in eye liner and caffeine in eye serum. Firstly, five stationary phases (ACQUITY UPC(2)) are screened with isocratic elution conditions (10% methanol in carbon dioxide). Complementary of the stationary phases is assessed based on our spider diagram classification which compares a large number of stationary phases based on five molecular interactions. Secondly, the one or two best stationary phases are retained for further optimization of mobile phase composition, with isocratic elution conditions or, when

  1. A novel method for the simultaneous analysis of seven biothiols in rice (Oryza sativa L.) using hydrophilic interaction chromatography coupled with electrospray tandem mass spectrometry.

    PubMed

    Cao, Zhao-Yun; Sun, Li-Hua; Mou, Ren-Xiang; Zhou, Rong; Zhu, Zhi-Wei; Chen, Ming-Xue

    2015-01-22

    Analysis of biothiols is still problematic, due to their high polarity, oxidation sensitivity and time-consuming sample preparation. In this paper, a direct, rapid and sensitive method was developed for simultaneous quantification of unbound cysteine (Cys), glutathione (GSH) and phytochelatins (PCs) in rice leaf, stem and root samples by hydrophilic interaction chromatography coupled with electrospray tandem mass spectrometry (HILIC-MS/MS). Homogenized samples were extracted with water containing 50mM dithiothreitol, without derivatization and further clean-up, and the extracts were injected directly onto an Xbridge Amide-HILIC column (3.5μm, 150mm×2.1mm i.d.). The best chromatographic separation and MS sensitivity was achieved using a linear gradient elution with 10mM aqueous ammonium formate and acetonitrile as the mobile phase. In MS/MS mode the detection limit (S/N≥3) of seven biothiols was 3-105nM. Good linearities were observed (r>0.995) with linear dynamic range at least over three orders of magnitude. Recoveries for most analytes were within the range of 77-128%, with relative standard deviations less than 18.2%. The intra-day precision (n=7) was 6.1-11.7%, and the inter-day precision over 15 d (n=15) was 8.5-16.3% for all biothiols. The optimized HILIC-MS/MS method was applied to study the influence of different cadmium (Cd) concentrations (0, 1 and 50μM) on contents of Cys, GSH and PC2-6 in rice tissue. With increasing Cd concentrations in nutrient solutions, contents of PC2-4 in rice roots increased but contents of Cys and GSH decreased. Contents of PC2-4 in both rice leafs and stems increased markedly at high dose Cd (50μM) treatment compared with controls, compared with low Cd concentrations (1μM). However, both PC5 and PC6 were not detected throughout the stress experiment. PMID:25436484

  2. Hydrophobic and electrostatic interaction chromatography for estimating changes in cell surface charge of Escherichia coli cells treated with pulsed electric fields.

    PubMed

    Ukuku, Dike O; Yuk, Hyun-Gyun; Zhang, Howard

    2011-10-01

    Pulsed electric field (PEF) treatments, a nonthermal process, have been reported to injure and inactivate bacteria in liquid foods. However, the effect of this treatment on bacterial cell surface charge and hydrophobicity has not been investigated. Apple juice (pH 3.8) purchased from a wholesale distributor was inoculated with cocktail of Escherichia coli O157:H7 at 7.4 log CFU/mL, processed with a PEF at a field strength of 18.4 kV/cm and 32.2 kV/cm at 25°C, 35°C, and 45°C with a treatment time of 160 μs and a flow rate of 120 mL/min. Bacterial cell surface charge and hydrophobicity of untreated and PEF-treated E. coli O157:H7 were determined immediately and after storage at 5°C and 23°C using hydrophobic and electrostatic interaction chromatography. Similarly, the populations surviving the PEF treatments including injured cells were determined by plating 0.1 mL of the sample on sorbitol MacConkey agar and tryptic soy agar (TSA) plates. The surviving populations of E. coli cells after PEF treatment varied depending on field strength and treatment temperature used. Percent injury in the surviving populations was high immediately after PEF treatment and varied among treatment temperatures. Cell surface charge of E. coli bacteria before PEF treatment averaged 32.10±8.12. PEF treatments at 25°C, 35°C, and 45°C reduced the above surface charge to 26.34±1.24, 14.24±3.30, and 6.72±2.82, respectively. Similarly, the surface hydrophobicity of untreated E. coli cells at 0.194±0.034 was increased to an average of 0.268±0.022, 0.320±0.124, and 0.586±0.123 after PEF treatments at 25°C, 35°C, and 45°C, respectively. The results of this study indicate that PEF treatment affects the outer cell envelope of E. coli bacteria as evidenced by the changes in surface hydrophobicity and cell surface charge leading to injury and subsequent inactivation of the cells. PMID:21668373

  3. [Progresses in screening active compounds from herbal medicine by affinity chromatography].

    PubMed

    Feng, Ying-shu; Tong, Shan-shan; Xu, Xi-ming; Yu, Jiang-nan

    2015-03-01

    Affinity chromatography is a chromatographic method for separating molecules using the binding characteristics of the stationary phase with potential drug molecules. This method can be performed as a high throughput screening method and a chromatographic separation method to screen a variety of active drugs. This paper summarizes the history of affinity chromatography, screening technology of affinity chromatography, and application of affinity chromatography in screening bio-active compounds in herbal medicines, and then discusses its application prospects, in order to broaden applications of the affinity chromatography in drug screening. PMID:26226740

  4. Thoracic injury metrics with side airbag: Stationary and dynamic occupants

    PubMed Central

    Hallman, Jason J; Yoganandan, N; Pintar, Frank A

    2010-01-01

    Objective Injury risk from side airbag deployment has been assessed using stationary out-of-position occupant test protocols. However, stationary conditions may not always represent real world environments. Therefore, the objective of the present study was to evaluate the effects of torso side airbag deployment on close-proximity occupants, comparing a stationary test protocol with dynamic sled conditions. Methods Chest compression and viscous metrics were quantified from sled tests utilizing post-mortem human specimens and computational simulations with three boundary conditions: rigid wall, ideal airbag interaction, and close-proximity airbag deployment. PMHS metrics were quantified from chestband contour reconstructions. The parametric effect of ΔV on close-proximity occupant was examined with the computational model. Results PMHS injuries suggested close-proximity occupants may sustain visceral trauma, which was not observed in occupants subjected to rigid wall or ideal airbag boundary conditions. Peak injury metrics were also elevated with close-proximity occupant relative to other boundary conditions. The computational model indicated decreasing influence of airbag on compression metrics with increasing ΔV. Airbag influence on viscous metric was greatest with close-proximity occupant at ΔV = 7.0 m/s, at which the response magnitude was greater than linear summation of metrics resulting from rigid impact and stationary close-proximity interaction. Conclusions These results suggest that stationary close-proximity occupants may not represent the only scenario of side airbag deployment harmful to the thoracoabdominal region. The sensitivity of the viscous metric and implications for visceral trauma are also discussed. PMID:20730691

  5. Fluctuations in Stationary Nonequilibrium States of Irreversible Processes

    SciTech Connect

    Bertini, L.; De Sole, A.; Gabrielli, D.; Jona-Lasinio, G.; Landim, C.

    2001-07-23

    We formulate a dynamical fluctuation theory for stationary nonequilibrium states (SNS) which covers situations in a nonlinear hydrodynamic regime and is verified explicitly in stochastic models of interacting particles. In our theory a crucial role is played by the time reversed dynamics. Our results include the modification of the Onsager-Machlup theory in the SNS, a general Hamilton-Jacobi equation for the macroscopic entropy and a nonequilibrium, nonlinear fluctuation dissipation relation valid for a wide class of systems.

  6. The spectral analysis of cyclo-non-stationary signals

    NASA Astrophysics Data System (ADS)

    Abboud, D.; Baudin, S.; Antoni, J.; Rémond, D.; Eltabach, M.; Sauvage, O.

    2016-06-01

    Condition monitoring of rotating machines in speed-varying conditions remains a challenging task and an active field of research. Specifically, the produced vibrations belong to a particular class of non-stationary signals called cyclo-non-stationary: although highly non-stationary, they contain hidden periodicities related to the shaft angle; the phenomenon of long term modulations is what makes them different from cyclostationary signals which are encountered under constant speed regimes. In this paper, it is shown that the optimal way of describing cyclo-non-stationary signals is jointly in the time and the angular domains. While the first domain describes the waveform characteristics related to the system dynamics, the second one reveals existing periodicities linked to the system kinematics. Therefore, a specific class of signals - coined angle-time cyclostationary is considered, expressing the angle-time interaction. Accordingly, the related spectral representations, the order-frequency spectral correlation and coherence functions are proposed and their efficiency is demonstrated on two industrial cases.

  7. Preparation of cyclodextrin chiral stationary phases by organic soluble catalytic 'click' chemistry.

    PubMed

    Wang, Yong; Chen, Hui; Xiao, Yin; Ng, Cheong Hengq; Oh, Ting Shan; Tan, Timothy Thatt Yang; Ng, Siu Choon

    2011-07-01

    We describe an effective and simple protocol that uses click chemistry to attach native β-cyclodextrin (β-CD) to silica particles, resulting in a chiral stationary phase (CCNCSP) that can be used for the enantioseparation of chiral drugs by high-performance liquid chromatography (HPLC). Starting from β-CD, the CCNCSP is prepared in several steps: (i) reaction of β-CD with 1-(p-toluenesulfonyl)-imidazole to afford mono-6-toluenesulfonyl-β-CD; (ii) azidolysis of mono-6-toluenesulfonyl-β-CD in dimethylformamide to give mono-6-azido-β-CD (N(3)-CD); (iii) reaction of cuprous iodide with triphenylphosphine to form an organic soluble catalyst CuI(PPh(3)); (iv) preparation of alkynyl-modified silica particles; and (v) click chemistry immobilization of N(3)-CD onto alkynyl-modified silica to afford the desired chiral stationary phase. Synthesis of the stationary phase and column packing takes ∼1 week. PMID:21720308

  8. Instrumentation: Ion Chromatography.

    ERIC Educational Resources Information Center

    Fritz, James S.

    1987-01-01

    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. Method for making a non-extractable stationary phase of polymer within a capillary column

    DOEpatents

    Springston, Stephen R.

    1990-01-01

    A method for coating interior capillary column surfaces, or packing material of a packed column, used for gas chromatography, with a stationary polymer phase that is cross-linked by exposing it to a low-temperature plasma that is uniformly distributed over the column or packing material for a predetermined period of time to effect the desired degree of cross-linking of the coating.

  10. Method for making a non-extractable stationary phase of polymer within a capillary column

    DOEpatents

    Springston, S.R.

    1990-10-30

    A method is described for coating interior capillary column surfaces, or packing material of a packed column, used for gas chromatography, with a stationary polymer phase that is cross-linked by exposing it to a low-temperature plasma that is uniformly distributed over the column or packing material for a predetermined period of time to effect the desired degree of cross-linking of the coating. 7 figs.

  11. Dielectrokinetic chromatography devices

    DOEpatents

    Chirica, Gabriela S; Fiechtner, Gregory J; Singh, Anup K

    2014-12-16

    Disclosed herein are methods and devices for dielectrokinetic chromatography. As disclosed, the devices comprise microchannels having at least one perturber which produces a non-uniformity in a field spanning the width of the microchannel. The interaction of the field non-uniformity with a perturber produces a secondary flow which competes with a primary flow. By decreasing the size of the perturber the secondary flow becomes significant for particles/analytes in the nanometer-size range. Depending on the nature of a particle/analyte present in the fluid and its interaction with the primary flow and the secondary flow, the analyte may be retained or redirected. The composition of the primary flow can be varied to affect the magnitude of primary and/or secondary flows on the particles/analytes and thereby separate and concentrate it from other particles/analytes.

  12. Isolation and purification of blood group antigens using immuno-affinity chromatography on short monolithic columns.

    PubMed

    Mönster, Andrea; Hiller, Oliver; Grüger, Daniela; Blasczyk, Rainer; Kasper, Cornelia

    2011-02-01

    Monolithic columns have gained increasing attention as stationary phases for the separation of biomolecules and biopharmaceuticals. In the present work the performance of monolithic convective interaction media (CIM(®)) chromatography for the purification of blood group antigens was established. The proteins employed in this study are derived from blood group antigens Knops, JMH and Scianna, equipped both with a His-tag and with a V5-tag by which they can be purified. In a first step a monoclonal antibody directed against the V5-tag was immobilized on a CIM(®) Disk with epoxy chemistry. After this, the immobilized CIM(®) Disk was used in immuno-affinity chromatography to purify the three blood group antigens from cell culture supernatant. Up-scaling of the applied technology was carried out using CIM(®) Tubes. In comparison to conventional affinity chromatography, blood group antigens were also purified via His-tag using a HiTrap(®) metal-affinity column. The two purifications have been compared regarding purity, yield and purification speed. Using the monolithic support, it was possible to isolate the blood group antigens with a higher flow rate than using the conventional bed-packed column. PMID:21194702

  13. Space plasma physics: I - Stationary processes

    NASA Technical Reports Server (NTRS)

    Hasegawa, Akira; Sato, Tetsuya

    1989-01-01

    The physics of stationary processes in space plasmas is examined theoretically in an introduction intended for graduate students. The approach involves the extensive use of numerical simulations. Chapters are devoted to fundamental principles, small-amplitude waves, and the stationary solar plasma system; typical measurement data and simulation results are presented graphically.

  14. Stationary rotating strings as relativistic particle mechanics

    SciTech Connect

    Ogawa, Kouji; Ishihara, Hideki; Saito, Shinya; Kozaki, Hiroshi; Nakano, Hiroyuki

    2008-07-15

    Stationary rotating strings can be viewed as geodesic motions in appropriate metrics in two-dimensional space. We obtain all solutions describing stationary rotating strings in flat spacetime as an application. These rotating strings have infinite length with various wiggly shapes. Averaged value of the string energy, the angular momentum, and the linear momentum along the string are discussed.

  15. Stationary Engineering Laboratory--2. Teacher's Guide.

    ERIC Educational Resources Information Center

    Steingress, Frederick M.; Frost, Harold J.

    The Stationary Engineering Laboratory Manual 2 Teacher's Guide was designed as an aid to the instructors of vocational-technical high school students who have received instruction in the basics of stationary engineering. The course of study was developed for students who will be operating a live plant and who will be responsible for supplying…

  16. Extraction of stationary components in biosignal discrimination.

    PubMed

    Martinez-Vargas, J D; Cardenas-Pena, D; Castellanos-Dominguez, G

    2012-01-01

    Biosignal recordings are widely used in the medical environment to support the evaluation and the diagnosis of pathologies. Nevertheless, the main difficulty lies in the non-stationary behavior of the biosignals, difficulting the obtention of patterns characterizing the changes in physiological or pathological states. Thus, the obtention of the stationary and non-stationary components of a biosignal is still an open issue. This work proposes a methodology to detect time-homogeneities based on time-frequency analysis with aim to extract the non-stationary behavior of the biosignal. Results show an increase in the stationarity and in the distance between classes of the reconstructions from the enhanced time-frequency representations. The stationary components extracted with the proposed approach can be used to solve biosignal classification problems. PMID:23365817

  17. Stationary Plasma Thruster Plume Emissions

    NASA Technical Reports Server (NTRS)

    Manzella, David H.

    1994-01-01

    The emission spectrum from a xenon plasma produced by a Stationary Plasma Thruster provided by the Ballistic Missile Defense Organization (BMDO) was measured. Approximately 270 individual Xe I, Xe II, and XE III transitions were identified. A total of 250 mW of radiated optical emission was estimated from measurements taken at the thruster exit plane. There was no evidence of erosion products in the emission signature. Ingestion and ionization of background gas at elevated background pressure was detected. The distribution of excited states could be described by temperatures ranging from fractions of 1 eV to 4 eV with a high degree of uncertainty due to the nonequilibrium nature of this plasma. The plasma was over 95 percent ionized at the thruster exit plane. Between 10 and 20 percent of the ions were doubly charged. Two modes of operation were identified. The intensity of plasma emission increased by a factor of two during operation in an oscillatory mode. The transfer between the two modes of operation was likely related to unidentified phenomena occurring on a time scale of minutes.

  18. Stationary Plasma Thruster Plume Characteristics

    NASA Technical Reports Server (NTRS)

    Myers, Roger M.; Manzella, David H.

    1994-01-01

    Stationary Plasma Thrusters (SPT's) are being investigated for application to a variety of near-term missions. This paper presents the results of a preliminary study of the thruster plume characteristics which are needed to assess spacecraft integration requirements. Langmuir probes, planar probes, Faraday cups, and a retarding potential analyzer were used to measure plume properties. For the design operating voltage of 300 V the centerline electron density was found to decrease from approximately 1.8 x 10 exp 17 cubic meters at a distance of 0.3 m to 1.8 X 10 exp 14 cubic meters at a distance of 4 m from the thruster. The electron temperature over the same region was between 1.7 and 3.5 eV. Ion current density measurements showed that the plume was sharply peaked, dropping by a factor of 2.6 within 22 degrees of centerline. The ion energy 4 m from the thruster and 15 degrees off-centerline was approximately 270 V. The thruster cathode flow rate and facility pressure were found to strongly affect the plume properties. In addition to the plume measurements, the data from the various probe types were used to assess the impact of probe design criteria

  19. Instability of stationary liquid sheets.

    PubMed

    Ardekani, A M; Joseph, D D

    2009-03-31

    The rupture of a 3D stationary free liquid film under the competing effects of surface tension and van der Waals forces is studied as a linearized stability problem in a purely irrotational analysis utilizing the dissipation method. The results of the foregoing analysis are compared with a 2D long-wave approximation that has given rise to an extensive literature on the rupture problem. The irrotational and long-wave approximations are here compared with the exact 2D solution. The exact solution and the two approximate theories give the same results for infinitely long waves. The problem considered depends on two dimensionless parameters, the Hamaker number and the Ohnesorge number. The Hamaker number is a dimensionless number defined as a measure of the ratio of van der Waals forces to surface tension. The exact solution and the two approximate solutions differ by < 1% when the Hamaker number is small for all values of the Ohnesorge number. When the Ohhnesorge number is close to one, as in the case of water films separated by distance 100 A, the long-wave approximation overestimates and the potential flow approximation underestimates the exact solution by similar small amounts. The high accuracy of the dissipation method shows that the effects of vorticity are small for small to moderate Hamaker numbers. PMID:19279213

  20. Sampling and analysis information aids for stationary source personnel

    SciTech Connect

    Jackson, M.D.; Johnson, L.D.

    1994-12-31

    The Environmental Protection Agency, in developing and evaluating samples and analysis methodology for stationary sources, has compiled information on availability and applicability of sampling and analytical methods. Information has also been summarized on the applicability of the gas chromatography/mass spectrometry as the analytical method. All of this information is accessible in three documents: ``Stationary Source Sampling and Analysis Directory, Version 2`` (SSSADIR), ``Handbook of GC/MS Data and Information for Selected Clean Air Act Amendments Compounds`` (Handbook), and ``Literature Review of CAAA Compounds`` (LitRev). The SSSADIR has information on which sampling and analytical methods to use for organic compounds listed in Title 3 of the Clean Air Act Amendments (CAAA) of 1990, as well as Appendices 8 and 9 of RCRA compounds, and the status of method evaluation for these analytes. The Handbook provides information on the mass spectra of selected CAAA analytes, primary quantitation ions, relative retention times and compatibility of the organic compounds in solution. The LitRev provides information on CAAA compounds for which EPA has no potential methods available but provides suggestions on ways to develop methods.

  1. Large eddy simulation of longitudinal stationary vortices

    NASA Astrophysics Data System (ADS)

    Sreedhar, Madhu; Ragab, Saad

    1994-07-01

    The response of longitudinal stationary vortices when subjected to random perturbations is investigated using temporal large-eddy simulation. Simulations are obtained for high Reynolds numbers and at a low subsonic Mach number. The subgrid-scale stress tensor is modeled using the dynamic eddy-viscosity model. The generation of large-scale structures due to centrifugal instability and their subsequent breakdown to turbulence is studied. The following events are observed. Initially, ring-shaped structures appear around the vortex core. These structures are counter-rotating vortices similar to the donut-shaped structures observed in a Taylor-Couette flow between rotating cylinders. These structures subsequently interact with the vortex core resulting in a rapid decay of the vortex. The turbulent kinetic energy increases rapidly until saturation, and then a period of slow decay prevails. During the period of maximum turbulent kinetic energy, the normalized mean circulation profile exhibits a logarithmic region, in agreement with the universal inner profile of Hoffman and Joubert [J. Fluid Mech. 16, 395 (1963)].

  2. Quantitative structure retention relationship studies for predicting relative retention times of chlorinated phenols on gas chromatography.

    PubMed

    Li, Shi-Yin; Sun, Cheng; Wang, Yu; Xu, Shi-Fen; Yao, Shu-Chun; Wang, Lian-Sheng

    2002-07-01

    A new method of quantitative structure-retention relationship (QSRR) studies was reported for predicting gas chromatography (GC) relative retention times (RRTs) of chlorinated phenols (CPs) using a DB-5 column. Chemical descriptors were calculated from the molecular structure of CPs and related to their gas chromatographic RRTs by using multiple linear regression analysis. The proposed model had a multiple square correlation coefficient R2 = 0.970, standard error SE = 0.0472, and significant level P = 0.0000. The QSRR model also reveals that the gas chromatographic relative retention times of CPs are associated with physicochemical property interactions with the stationary phase, and influenced by the number of chlorine and oxygen in the CP mOlecules. PMID:12211996

  3. Investigation of retention mechanism of resorcinarene based cavitands by linear and nonlinear chromatography.

    PubMed

    Bartó, Endre; Prauda, Ibolya; Kilár, Ferenc; Kiss, Ibolya; Felinger, Attila

    2016-07-22

    Cavitands are cavity-shaped cyclic oligomers and they can create host-guest interactions with various analytes, therefore they have applications in supramolecular chemistry, nanoscale reactions, chromatographic separations, drug encapsulation and delivery, biochemistry. The investigation of the chromatographic behavior of large molecules, such as resorcinarenes and cavitands is meager up to now. To understand the retention of resorcinarenes and cavitands in liquid chromatography, we studied their retention mechanism by the thermodynamic parameters calculated from the van't Hoff equation and by generation of an adsorption isotherm, which can describe the adsorption of the solute on the stationary phase surface. We compared the thermodynamics of the retention for cyclic oligomers in acetonitrile:water and methanol:water mobile phases. Furthermore, we determined the equilibrium adsorption isotherm by inverse method and we made an error analysis of the estimation obtained with the inverse method to ascertain the validity of the obtained isotherm parameters over a broader concentration range. PMID:27317005

  4. Polymeric Cryogel-Based Boronate Affinity Chromatography for Separation of Ribonucleic Acid from Bacterial Extracts.

    PubMed

    Shakya, Akhilesh Kumar; Srivastava, Akshay; Kumar, Ashok

    2015-01-01

    Three-dimensional monolithic columns are preferred stationary phase in column chromatography. Conventional columns based on silica or particles are efficient in bioseparation though associated with limitations of nonspecific interaction and uneven porosity that causes high mass transfer resistance for the movement of big molecules. Cryogels as a monolith column have shown promising application in bioseparation. Cryogels column can be synthesized in the form of a monolith at sub-zero temperature through gelation of pre-synthesized polymers or polymerization of monomers. Cryogels are macroporous and mechanically stable materials. They have open interconnected micron-sized pores with a wide range of porosity (10-200 μm). Current protocol demonstrated the ability of poly(hydroxymethyl methacrylate)-co-vinylphenyl boronic acid p(HEMA-co-VPBA) cryogel matrix for selective separation of RNA from the bacterial crude extract. PMID:26623972

  5. Calix[4]pyrroles: highly selective stationary phases for gas chromatographic separations.

    PubMed

    Fan, Jing; Wang, Zhenzhong; Li, Qian; Qi, Meiling; Shao, Shijun; Fu, Ruonong

    2014-10-01

    Calix[4]pyrroles offer a great potential as stationary phases for gas chromatography (GC) due to their unique structures and physicochemical properties. Herein we present the first report of using two calix[4]pyrroles, namely meso-tetra-cyclohexylcalix[4]pyrrole (THCP) and meso-octamethylcalix[4]pyrrole (OMCP). These stationary phases were statically coated onto capillary columns and investigated in terms of column efficiency, polarity, separation performance, thermal stability and repeatability. The columns achieved column efficiencies of 2200-3000plates/m and exhibited nonpolar nature with an average polarity of 67 for THCP and 64 for OMCP, respectively. THCP stationary phase shows high selectivity for analytes of different polarity and exhibits nice peak shapes, especially for aldehydes, alcohols and anilines that are prone to severe peak tailing in GC analysis. Interestingly, THCP stationary phase possesses superior resolving ability for aniline and benzenediol positional isomers while OMCP shows preferential selectivity for nonpolar analytes such as hexane isomers. Moreover, calix[4]pyrrole columns also have good thermal stability up to 260°C and repeatability with a relative standard deviation (RSD%) of less than 0.10% for run-to-run and less than 5.2% for column-to-column. This work demonstrates the unique separation performance of calix[4]pyrroles and their promising future as a new class of GC stationary phases. PMID:25173993

  6. Genetics Home Reference: autosomal dominant congenital stationary night blindness

    MedlinePlus

    ... stationary night blindness autosomal dominant congenital stationary night blindness Enable Javascript to view the expand/collapse boxes. ... Close All Description Autosomal dominant congenital stationary night blindness is a disorder of the retina , which is ...

  7. Evaluating the robustness of the enantioselective stationary phases on the Rosetta mission against space vacuum vaporization

    NASA Astrophysics Data System (ADS)

    Meierhenrich, Uwe J.; Cason, Julie R. L.; Szopa, Cyril; Sternberg, Robert; Raulin, François; Thiemann, Wolfram H.-P.; Goesmann, Fred

    2013-12-01

    The European Space Agency's Rosetta mission was launched in March 2004 in order to reach comet 67P/Churyumov-Gerasimenko by August 2014. The Cometary Sampling and Composition experiment (COSAC) onboard the Rosetta mission's lander "Philae" has been designed for the cometary in situ detection and quantification of organic molecules using gas chromatography coupled to mass spectrometry (GC-MS). The GC unit of COSAC is equipped with eight capillary columns that will each provide a specific stationary phase for molecular separation. Three of these stationary phases will be used to chromatographically resolve enantiomers, as they are composed of liquid polymers of polydimethylsiloxane (PDMS) to which chiral valine or cyclodextrin units are attached. Throughout the ten years of Rosetta's journey through space to reach comet 67P, these liquid stationary phases have been exposed to space vacuum, as the capillary columns within the COSAC unit were not sealed or filled with carrier gas. Long term exposures to space vacuum can cause damage to such liquid stationary phases as key monomers, volatiles, and chiral selectors can be vaporized and lost in transit. We have therefore exposed identical spare units of COSAC's chiral stationary phases over eight years to vacuum conditions mimicking those experienced in space and we have now investigated their resolution capabilities towards different enantiomers both before and after exposure to space vacuum environments. We have observed that enantiomeric resolution capabilities of these chiral liquid enantioselective stationary phases has not been affected by exposure to space vacuum conditions. Thus we conclude that the three chiral stationary phases of the COSAC experiment onboard the Rosetta mission lander "Philae" can be considered to have maintained their resolution capacities throughout their journey prior to cometary landing in November 2014.

  8. Separation of stationary and non-stationary sources with a generalized eigenvalue problem.

    PubMed

    Hara, Satoshi; Kawahara, Yoshinobu; Washio, Takashi; von Bünau, Paul; Tokunaga, Terumasa; Yumoto, Kiyohumi

    2012-09-01

    Non-stationary effects are ubiquitous in real world data. In many settings, the observed signals are a mixture of underlying stationary and non-stationary sources that cannot be measured directly. For example, in EEG analysis, electrodes on the scalp record the activity from several sources located inside the brain, which one could only measure invasively. Discerning stationary and non-stationary contributions is an important step towards uncovering the mechanisms of the data generating system. To that end, in Stationary Subspace Analysis (SSA), the observed signal is modeled as a linear superposition of stationary and non-stationary sources, where the aim is to separate the two groups in the mixture. In this paper, we propose the first SSA algorithm that has a closed form solution. The novel method, Analytic SSA (ASSA), is more than 100 times faster than the state-of-the-art, numerically stable, and guaranteed to be optimal when the covariance between stationary and non-stationary sources is time-constant. In numerical simulations on wide range of settings, we show that our method yields superior results, even for signals with time-varying group-wise covariance. In an application to geophysical data analysis, ASSA extracts meaningful components that shed new light on the Pi 2 pulsations of the geomagnetic field. PMID:22551683

  9. Hydrophilic interaction chromatographic analysis of anthocyanins.

    PubMed

    Willemse, Chandré M; Stander, Maria A; de Villiers, André

    2013-12-01

    Hydrophilic interaction chromatography (HILIC) provides an alternative separation mode for the analysis of phenolic co