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  • Development and comparison of chromatographic methods for the analysis of long chain diols and alkenones in biological materials and sediment
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-19
    Marijke W. de Bar, Ellen C. Hopmans, Monique Verweij, Denise J.C. Dorhout, Jaap S. Sinninghe Damsté, Stefan Schouten

    We have compared and assessed the suitability of several chromatographic methods for the analysis of long chain alkenones and long chain diols and the associated paleotemperature proxies (UK’37 and LDI). We evaluated the traditional methods for the analysis of the UK’37 and the LDI, gas chromatography (GC) − flame ionization detection (FID) and GC mass spectrometry (MS) using selected ion monitoring (SIM), respectively, and developed a new method using GC-MS/MS in multiple reaction monitoring mode (MRM) for the analysis of long chain diols as well as a method for automatic silylation of diols using a robot autosampler. Finally, we evaluated liquid chromatography (LC) methods to simultaneously measure the UK’37 and the LDI, using ultra high performance LC (UHPLC) with low (nominal mass) resolution MS in SIM mode, and UHPLC with high resolution MS (HRMS). Detection and quantification limits and reproducibility were assessed by means of serial dilutions of culture extracts.Automated silylation by robot autosampler showed similar reproducibility as off-line silylation while substantially decreasing sample preparation time. The novel MRM method had a slightly lower limit of quantification (LOQ; i.e. 0.3 pg C28 1,13-diol injected on-column) than the traditional method (0.5 pg) and improved reproducibility while allowing more unambiguous identification of LCDs in complex matrices. For diols, UHPLC-MS using SIM had the highest LOQ (i.e. 15 pg) and a comparable reproducibility as GC-MS. UHPLC-HRMS had a LOQ of ca. 1.5 pg, and an improved reproducibility for diol analysis. For alkenone analysis, both UHPLC-HRMS and UHPLC-MS using SIM were 2-3 orders of magnitude more sensitive (LOQ ca. 20 and 2 pg C37:2 alkenone injected on-column, respectively) than GC-FID (LOD ca. 3 ng), with a similar reproducibility of the UK’37 index. Hence, UHPLC-HRMS allows simultaneous analysis of the UK’37 and LDI at an increased sensitivity. In addition, it allows simultaneous measurement of TEX86, a temperature proxy based on the isoprenoid glycerol dialkyl glycerol tetraethers. This reduces the preparation time by excluding the need of derivatization and separation of the ketone (containing the long chain alkenones) and polar fractions (containing the long chain diols and GDGTs). However, synthetic standards are required to fully assess the accuracy of the new methods for determination of the LDI and UK’37.

    更新日期:2017-09-19
  • The 9th International Countercurrent Chromatography Conference held at Dominican University, Chicago, USA, August 1–3, 2016
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-19
    J. Brent Friesen, James B. McAlpine, Shao-Nong Chen, Guido F. Pauli

    The 9th International Countercurrent Chromatography Conference (CCC 2016) was held at Dominican University near Chicago, IL (USA), from August 1st–3rd, 2016. The biennial CCC 20XX conferences provide an opportunity for countercurrent chromatography and centrifugal partition chromatography (CCC/CPC) manufactures, marketers, theorists, and research scientists to gather together socially, learn from each other, and advance countercurrent separation technology. A synopsis of the conference proceedings as well as a series of short reviews of the special edition articles is included in this document. Many productive discussions and collegial conversation at CCC 2016 attested to the liveliness, connectivity, and productivity of the global countercurrent research community and bodes well for the success of the 10th conference at the University of Braunschweig, Germany on August 1–3, 2018.

    更新日期:2017-09-19
  • Development and validation of a high-throughput method for determination of nine fluoroquinolones residues in muscle of different animal species by liquid chromatography coupled to tandem mass spectrometry with low temperature clean up
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-18
    Fabiano Barreto, Cristina B.D. Ribeiro, Rodrigo Barcellos Hoff, Teresa Dalla Costa

    This work describes the development and validation of a quantitative and confirmatory method to determination of nine fluoroquinolones residues in poultry, bovine, swine and fish muscle using LC–MS/MS. Sample preparation was based in a fast solvent extraction with acetonitrile with 1% of formic acid followed by a low-temperature clean up procedure and centrifugation, without further steps. The recoveries ranged between 79% and 115%. The concentration work range was 0–200 μg kg−1. The LOD and LOQ were 5 and 10 μg kg−1, respectively. This high-throughput method was reliable for identification and confirmation of nine relevant compounds in food-producing animal tissues. Validation procedure was performed according the Directive 2002/657/EC criteria and the method showed fitness to purpose in terms of precision and reproducibility as well as for the other performance parameters.

    更新日期:2017-09-19
  • Electrospun nanofibers-based online micro-solid phase extraction for the determination of monohydroxy polycyclic aromatic hydrocarbons in human urine
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-18
    Huifang Zhang, Hui Xu

    In this article, silver nanoparticles (AgNPs) modified nanofibers that represent a novel kind of packing materials for micro-solid phase extraction (μ-SPE) is reported for the first time. The composite material was fabricated through in-situ formation of AgNPs on the polydopamine (PDA) coated polystyrene electrospun fibers (PS@PDA-Ag). The nanofibers displayed desirable hydrophilicity, large surface area, high porosity, good extraction ability and certain capacity to resist matrix interference. An μ-SPE combined with liquid chromatography-mass spectrometry (LC-MS) method was developed for the online determination of three monohydroxy polycyclic aromatic hydrocarbons (OH-PAHs) metabolites in human urine. Under the optimal conditions, good linearity (0.02–5 ng mL−1) was acquired with the correlation coefficients (R2) being larger than 0.9962. Low limits of detection (0.007–0.032 ng mL−1) expressed the satisfactory sensitivity of the method. Moreover, the intraday relative standard deviation values lower than 9.7% and the recoveries among the range of 71-116% were obtained. In general, the online μ-SPE-LC-MS method had the features of simplicity, rapidity, sensitivity and automation, and it was expected to become a promising approach for the online analysis of trace OH-PAHs in complex biological samples.

    更新日期:2017-09-19
  • An easy and fast adenosine 5′-diphosphate quantification procedure based on hydrophilic interaction liquid chromatography-high resolution tandem mass spectrometry for determination of the in vitro adenosine 5′-triphosphatase activity of the human breast cancer resistance protein ABCG2
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-18
    Lea Wagmann, Hans H. Maurer, Markus R. Meyer

    Interactions with the human breast cancer resistance protein (hBCRP) significantly influence the pharmacokinetic properties of a drug and can even lead to drug-drug interactions. As efflux pump from the ABC superfamily, hBCRP utilized energy gained by adenosine 5′-triphosphate (ATP) hydrolysis for the transmembrane movement of its substrates, while adenosine 5′-diphosphate (ADP) and inorganic phosphate were released. The ADP liberation can be used to detect interactions with the hBCRP ATPase. An ADP quantification method based on hydrophilic interaction liquid chromatography (HILIC) coupled to high resolution tandem mass spectrometry (HR-MS/MS) was developed and successfully validated in accordance to the criteria of the guideline on bioanalytical method validation by the European Medicines Agency. ATP and adenosine 5′-monophosphate were qualitatively included to prevent interferences. Furthermore, a setup consisting of six sample sets was evolved that allowed detection of hBCRP substrate or inhibitor properties of the test compound. The hBCRP substrate sulfasalazine and the hBCRP inhibitor orthovanadate were used as controls. To prove the applicability of the procedure, the effect of amprenavir, indinavir, nelfinavir, ritonavir, and saquinavir on the hBCRP ATPase activity was tested. Nelfinavir, ritonavir, and saquinavir were identified as hBCRP ATPase inhibitors and none of the five HIV protease inhibitors turned out to be an hBCRP substrate. These findings were in line with a pervious publication.

    更新日期:2017-09-19
  • Automatic single-step quick, easy, cheap, effective, rugged and safe sample preparation devices for analysis of pesticide residues in foods
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-17
    Jishi Wang, Zeying He, Lu Wang, Yaping Xu, Yi Peng, Xiaowei Liu

    In this research, the manual two-step QuEChERS approach has been streamlined and automated into a one-step method using a cleanup tube fitted within an extraction tube. A novel automatic QuEChERS combination have been developed to simplify the QuEChERS procedures and improve sample preparation efficiency. This combination integrates QuEChERS procedures into a single run via the use of a vortex vibration-centrifuge device and a centrifuge filtration tube. To validate the efficiency of our automatic QuEChERS device, 270 pesticides were analyzed in plant origined foods including celery, tomatoes, leeks, eggplants, grapes, corn, green tea, and soybean oil using this automatic platform. The results were then compared with those obtained using the manual QuEChERS method. Different parameters were validated and compared including recovery, linearity, repeatability and limits of quantification (LOQ). Satisfactory results, comparable to results obtained using the manual QuEChERS method were obtained. The average recoveries ranged between 70% and 120% for most pesticides with associated relative standard deviations (RSDs) < 20% (n = 5) indicating satisfactory accuracy and repeatability. An LOQ of 2 μg/kg was obtained for most pesticides present in celery and corn matrices, and the correlation coefficients (r2) were >0.990 within a linearity range of 2–500 μg/kg. Compared to manual QuEChERS, this novel automatic QuEChERS device and combination could significantly improve the sample preparation efficiency for the multiresidue analysis of pesticides.

    更新日期:2017-09-19
  • Gas chromatographic-mass spectrometric determination of glycosides without prior hydrolysis
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-17
    Valery A. Isidorov, Jolanta Nazaruk

    The article presents for the first time the linear temperature programmed retention indices on a column with stationary phases of 5% phenylpolydimethyl silicone and the mass spectra of trimethylsilyl (TMS) derivatives of 71 glycosides (both commercial preparations and compounds extracted from plant tissues) which were not characterized earlier by these parameters. Converted to their TMS derivatives, the glycosides were thermally stable: they exhibited single peaks on their chromatograms without products of thermal decomposition. Therefore this work demonstrates the suitability of high resolution–high temperature gas chromatography (HR-HT/GC) to analyse different groups of glycosides including compounds with disaccharide moieties without the necessity of their hydrolyses. Since a limited number of commercial and plant-isolated glycosides were available, an attempt was made to assess their retention indices using the known “structure−retention relationships” approach. It was demonstrated that the retention indices of silanised glycosides and their aglycones were characterized by a linear dependence.

    更新日期:2017-09-19
  • Facile and green fabrication of cation exchange membrane adsorber with unprecedented adsorption capacity for protein purification
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-15
    M. Kamran Khan, Jianquan Luo, Rashid Khan, Jinxin Fan, Yinhua Wan

    Fabricating membrane adsorbers with high adsorption capacity and appreciable throughput for the separation and purification of protein products is challenging in biomedical and pharmaceutical industries. Herein, we report the synthesis of a novel membrane adsorber by functionalizing a nylon microfiltration membrane with alginate dialdehyde (ADA) followed by sulphonic addition, without any solvent usage, and its successful application in the purification of lysozyme. Taking advantage of abundant dual cation exchange (CEX) groups on sulphonic-ADA (S-ADA) ligands, this novel S-ADA-nylon membrane adsorber showed an unprecedented static binding capicity of 286 mg/mL for lysozyme adsorption. Meanwhile, the prepared membrane adsorber could be easily regenerated (complete protein elution) under mild conditions and be reused at least for five times. Featured with a unique selectivity, the S-ADA-nylon membrane also captured lysozyme from chicken egg white solution with a high purity (100%) and a high recovery of 98%. The purified lysozyme showed similar specific activity as commercial product. The present work provides a facile, green and low-cost approach for the preparation of high-performance membrane adsorbers, which has a great potential in protein production.

    更新日期:2017-09-15
  • Identification and characterization of curcuminoids in turmeric using ultra-high performance liquid chromatography-quadrupole time of flight tandem mass spectrometry
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-14
    Shuailong Jia, Zhifeng Du, Chengwu Song, Shuna Jin, Yang Zhang, Yulin Feng, Chaomei Xiong, Hongliang Jiang

    A three-step strategy was developed for systematic characterization of curcuminoids in turmeric. Based on UHPLC-QTOF-MS/MS analysis, 89 curcuminoids including 16 novel ones were identified in the turmeric samples using this approach. During the identification process, false positive results were excluded by combining the positive and negative ESI-MS/MS analyses. Moreover, the characterization of the keto and enol forms of type A, B and C curcuminoids was first discussed and they were clearly distinguished using negative ESI-MS/MS method with UV spectra analyses. The structures of detected curcuminoids were identified and rationalized in both ion modes. Additionally, the fragmentation behaviors of the 15 types of curcuminoids were clearly illustrated in this work, which will be helpful for detection and identification of corresponding trace curcuminoids in complex turmeric samples using UHPLC-QTOF-MS/MS methods.

    更新日期:2017-09-15
  • LC-HRMS-based Metabolomic Approach for the Detection of CERA’s Effects in Horse Doping Control ☆
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-14
    Céline Joré, Benoît Loup, Patrice Garcia, Anne-Christelle Paris, Marie-Agnès Popot, Michel Audran, Yves Bonnaire, Emmanuelle Varlet-Marie, Ludovic Bailly-Chouriberry

    Erythropoiesis Stimulating Agents (ESAs) were developed for therapeutic purposes to stimulate red blood cell (RBC) production. Consequently, tissue oxygenation is enhanced as athlete's endurance and ESAs misuse now benefits doping. Our hypothesis is that most of ESAs should have similar mechanisms and thus have the same effects on metabolism. Studying the metabolome variations could allow suspecting the use of any ESAs with a single method by targeting their effects. In this objective, a metabolomic study was carried out on 3 thoroughbred horses with a single administration of 4.2 μg/kg of Mircera®, also called Continuous Erythropoiesis Receptor Activator (CERA). Blood and urine samples were collected from D-17 to D+74 and haematological parameters were followed throughout the study as plasmatic CERA concentration (ELISA). Urine and plasma metabolic fingerprints were recorded by Liquid Chromatography coupled to High Resolution Mass Spectrometry (LC-HRMS) in positive and negative mode. After preprocessing steps, normalized data were analyzed by multivariate statistics to build OPLS models. Hemoglobin concentration and hematocrit showed a significant increase after CERA administration unlike reticulocytes. CERA concentration showed a high intensity peak and then a slow decrease until becoming undetectable after D+31. Models built with multivariate statistics allow a discrimination between pre and post-administration plasma and urine samples until 74 days after administration, i.e. 43 days longer than ELISA method. By reducing and studying variables (ions), some potential candidate biomarkers were found.

    更新日期:2017-09-15
  • An automatic on-line 2,2-diphenyl-1-picrylhydrazyl-high performance liquid chromatography method for high-throughput screening of antioxidants from natural products
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-14
    Yanzhen Lu, Nan Wu, Yingtong Fang, Nusrat Shaheen, Yun Wei

    Many natural products are rich in antioxidants which play an important role in preventing or postponing a variety of diseases, such as cardiovascular and inflammatory disease, diabetes as well as breast cancer. In this paper, an automatic on-line 2,2-diphenyl-1-picrylhydrazyl-high performance liquid chromatography (DPPH-HPLC) method was established for antioxidants screening with nine standards including organic acids (4-hydroxyphenylacetic acid, p-coumaric acid, ferulic acid, and benzoic acid), alkaloids (coptisine and berberine), and flavonoids (quercitrin, astragalin, and quercetin). The optimal concentration of DPPH was determined, and six potential antioxidants including 4-hydroxyphenylacetic acid, p-coumaric acid, ferulic acid, quercitrin, astragalin, and quercetin, and three non-antioxidants including benzoic acid, coptisine, and berberine, were successfully screened out and validated by conventional DPPH radical scavenging assay. The established method has been applied to the crude samples of Saccharum officinarum rinds, Coptis chinensis, and Malus pumila leaves, consecutively. Two potential antioxidant compounds from Saccharum officinarum rinds and five potential antioxidant compounds from Malus pumila leaves were rapidly screened out. Then these seven potential antioxidants were purified and identified as p-coumaric acid, ferulic acid, phloridzin, isoquercitrin, quercetin-3-xyloside, quercetin-3-arabinoside, and quercetin-3-rhamnoside using countercurrent chromatography combined with mass spectrometry and their antioxidant activities were further evaluated by conventional DPPH radical scavenging assay. The activity result was in accordance with that of the established method. This established method is cheap and automatic, and could be used as an efficient tool for high-throughput antioxidant screening from various complex natural products.

    更新日期:2017-09-15
  • Influence of mixed electrolytes and ph on adsorption of bovine serum albumin in hydrophobic interaction chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-14
    Eva Hackemann, Hans Hasse

    Using salt mixtures instead of single salts can be beneficial for hydrophobic interaction chromatography (HIC). The effect of electrolytes on the adsorption of proteins, however, depends on the pH. Little is known on that dependence for mixed electrolytes. Therefore, the effect of the pH on protein adsorption from aqueous solutions containing mixed salts is systematically studied in the present work for a model system: the adsorption of bovine serum albumin (BSA) on the mildly hydrophobic resin Toyopearl PPG-600M. The pH is adjusted to 4.0, 4.7 or 7.0 using 25 mM sodium phosphate or sodium citrate buffer. Binary and ternary salt mixtures of sodium chloride, ammonium chloride, sodium sulfate and ammonium sulfate as well as the pure salts are used at overall ionic strengths between 1500 and 4200 mM. The temperature is always 25 °C. The influence of the mixed electrolytes on the adsorption behavior of BSA changes completely with varying pH. Positive as well as negative cooperative effects of the mixed electrolytes are observed. The results are analyzed using a mathematical model which was recently introduced by our group. In that model the influence of the electrolytes is described by a Taylor series expansion in the individual ion molarities. After suitable parametrization using a subset of the data determined in the present work, the model successfully predicts the influence of mixed electrolytes on the protein adsorption. Furthermore, results for BSA from the present study are compared to literature data for lysozyme, which are available for the same adsorbent, temperature and salts. By calculating the ratio of the loading of the adsorbent for both proteins particularly favorable separation conditions can be selected. Hence, a model-based optimization of solvents for protein separation is possible.

    更新日期:2017-09-14
  • Optimized Purification of a Fusion Protein by Reversed-phase High Performance Liquid Chromatography Informed by the Linear Solvent Strength Model
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-13
    Isaac B. Falconer, Colin T. Mant, C. James McKnight, Liliya Vugmeyster, Robert Hodges

    Fusion protein systems are commonly used for expression of small proteins and peptides. An important criterion for a fusion protein system to be useful is the ability to separate the protein of interest from the tag. Additionally, because no protease cleaves fusion proteins with 100% efficiency, the ability to separate the desired peptide from any remaining uncleaved protein is also necessary. This is likely to be the more difficult task as at least a portion of the sequence of the fusion protein is identical to that of the protein of interest. When a high level of purity is required, gradient elution reversed-phase HPLC is frequently used as a final purification step. Shallow gradients are often advantageous for maximizing both the purity and yield of the final product, however, the relationship between relative retention times at shallow gradients and those at steeper gradients typically used for analytical HPLC are not always straightforward. In this work, we report reversed-phase HPLC results for the fusion protein system consisting of the N-terminal domain of ribosomal protein L9 (NTL9) and the 36-residue villin headpiece subdomain (HP36) linked by a recognition sequence for the protease factor Xa. This system represents an excellent example of the difficulties in purification that may arise from this unexpected elution behavior at shallow gradients. Additionally, we report on the sensitivity of this elution behavior to the concentration of the additive trifluoroacetic acid in the mobile phase and present optimized conditions for separating HP36 from the full fusion protein by reversed-phase HPLC using a shallow gradient. Finally, we suggest that these findings are relevant to the purification of other fusion protein systems, for which similar problems may arise, and support this suggestion using insights from the linear solvent strength model of gradient elution liquid chromatography.

    更新日期:2017-09-13
  • Update of on-line coupled liquid chromatography – gas chromatography for the analysis of mineral oil hydrocarbons in foods and cosmetics
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-13
    Maurus Biedermann, Celine Munoz, Koni Grob

    On-line coupled high performance liquid chromatography-gas chromatography-flame ionization detection (HPLC-GC-FID) is the most widely used method for the analysis of mineral oil hydrocarbons in food, food contact materials, tissues and cosmetics. With comprehensive two-dimensional gas chromatography (GCxGC), a tool became available for better establishing the elution sequence of the various types of hydrocarbons from the HPLC column used for isolating the mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH). The performance of a heavily used HPLC column with reduced retention for MOAH was investigated to improve the robustness of the method. Updates are recommended that render the MOSH/MOAH separation less dependent of the state of the HPLC column and more correct in cases of highly refined mineral oil products of high molecular mass. Cyclohexyl cyclohexane (Cycy), used as internal standard, turned out to be eluted slightly after cholestane (Cho); apparently the size exclusion effect predominates the extra retention by ring number on the 60 Å pore size silica gel. Hence, Cycy can be used to determine the end of the MOSH fraction. Long chain alkyl benzenes were eluted earlier than tri-tert. butyl benzene (Tbb). It is proposed to start the MOAH transfer immediately after the MOSH fraction and use a gradient causing breakthrough of dichloromethane (visible in the UV chromatogram) at a time suitable to elute perylene (Per) at the end of the fraction. In this way, a decrease in retention power of the HPLC column can be tolerated without adjustment of the MOAH fraction until some MOAH start being eluted into the MOSH fraction. This critical point can be checked either with di(2-ethylhexyl) benzene (DEHB) as a marker or the HPLC-UV chromatogram. Finally, based on new findings in rats and human tissues, it is recommended to integrate the MOSH and MOAH up to the retention time of the n-alkanes C40.

    更新日期:2017-09-13
  • Anthracenyl polar embedded stationary phases with enhanced aromatic selectivity. Part II: A density functional theory study
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-11
    Mélanie Mignot, Benjamin Schammé, Vincent Tognetti, Laurent Joubert, Pascal Cardinael, Valérie Peulon-Agasse

    New polar embedded aromatic stationary phases (mono- and trifunctional versions) that contain an amide-embedded group coupled with a tricyclic aromatic moiety were developed for chromatographic applications and described in the first paper of this series. These phases offered better separation performance for PAHs than for alkylbenzene homologues, and an enhanced ability to differentiate aromatic planarity to aromatic tridimensional conformation, especially for the trifunctional version and when using methanol instead of acetonitrile. In this second paper, a density functional theory study of the retention process is reported. In particular, it was shown that the selection of the suitable computational protocol allowed for describing rigorously the interactions that could take place, the solvent effects, and the structural changes for the monofunctional and the trifunctional versions. For the first time, the experimental data coupled with these DFT results provided a better understanding of the interaction mechanisms and highlighted the importance of the multimodal character of the designed stationary phases: alkyl spacers for interactions with hydrophobic solutes, amide embedded groups for dipole-dipole and hydrogen-bond interactions, and aromatic terminal groups for π-π interactions.

    更新日期:2017-09-11
  • Hollow porous molecularly imprinted polymer for highly selective clean-up followed by influential preconcentration of ultra-trace glibenclamide from bio-fluid
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-10
    Abbas Ostovan, Mehrorang Ghaedi, Maryam Arabi, Arash Asfaram

    In the present work, hollow porous molecularly imprinted polymer (HPMIP) was prepared via adopting a sacrificial support approach using glibenclamide (GB) as template, methacrylic acid (MAA) as functional monomer, ethylene glycol dimethacrylate as cross-linker (EGDMA) and mesoporous MCM-48 nanospheres as support. Owing to a short thickness of the foresaid HPMIP, a suitable steric structure was readily available that lead to fast and effective mass transfer of target analyte to sorbent and consequently supply high binding capacity. After ultrasonic-assisted dispersive solid phase extraction of urine sample, the analyte of interest was quantitatively pre-concentrated and determined by high performance liquid chromatography-ultraviolet detection (HPLC-UV). Influence of factors affecting the extraction efficiency such as sonication time, sample pH, sorbent dosage, and volumes of eluent and washing agents as well as their significant interactions were simultaneously optimized by experimental design methodology. Under optimized conditions, present method has linear response over concentration range of 10–3000 μg L−1 in human urine samples with a satisfactory detection limit close to 3.5 μg L−1. The inter-day precision of the current method (coefficient of variation) lower than 5%, while recoveries are more than 87.7%.

    更新日期:2017-09-10
  • Additional band broadening of peptides in the first size-exclusion chromatographic dimension of an automated stop-flow two-dimensional high performance liquid chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-09
    Jucai Xu, Dongxiao Sun-Waterhouse, Chaoying Qiu, Mouming Zhao, Baoguo Sun, Lianzhu Lin, Guowan Su

    The need to improve the peak capacity of liquid chromatography motivates the development of two-dimensional analysis systems. This paper presented a fully automated stop-flow two-dimensional liquid chromatography system with size exclusion chromatography followed by reversed phase liquid chromatography (SEC × RPLC) to efficiently separate peptides. The effects of different stop-flow operational parameters (stop-flow time, peak parking position, number of stop-flow periods and column temperature) on band broadening in the first dimension (1st D) SEC column were quantitatively evaluated by using commercial small proteins and peptides. Results showed that the effects of peak parking position and the number of stop-flow periods on band broadening were relatively small. Unlike stop-flow analysis of large molecules with a long running time, additional band broadening was evidently observed for small molecule analytes due to the relatively high effective diffusion coefficient (Deff). Therefore, shorter analysis time and lower 1st D column temperature were suggested for analyzing small molecules. The stop-flow two-dimensional liquid chromatography (2D-LC) system was further tested on peanut peptides and an evidently improved resolution was observed for both stop-flow heart-cutting and comprehensive 2D-LC analysis (in spite of additional band broadening in SEC). The stop-flow SEC × RPLC, especially heart-cutting analysis with shorter analysis time and higher 1st D resolution for selected fractions, offers a promising approach for efficient analysis of complex samples.

    更新日期:2017-09-10
  • Capillary Electrophoresis–Mass Spectrometry for Direct Structural Identification of Serum N-Glycans
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-09
    Christa M. Snyder, Xiaomei Zhou, Jonathan A. Karty, Bryan R. Fonslow, Milos V. Novotny, Stephen C. Jacobson

    Through direct coupling of capillary electrophoresis (CE) to mass spectrometry (MS) with a sheathless interface, we have identified 77 potential N-glycan structures derived from human serum. We confirmed the presence of N-glycans previously identified by indirect methods, e.g., electrophoretic mobility standards, obtained 31 new N-glycan structures not identified in our prior work, differentiated co-migrating structures, and determined specific linkages on isomers featuring sialic acids. Serum N-glycans were cleaved from proteins, neutralized via methylamidation, and labeled with the fluorescent tag 8-aminopyrene-1,3,6-trisulfonic acid, which renders the glycan fluorescent and provides a −3 charge for electrophoresis and negative-mode MS detection. The neutralization reaction also stabilizes the labile sialic acids. In addition to methylamidation, native charges from sialic acids were neutralized through reaction with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium to amidate α2,6-linked sialic acids in the presence of ammonium chloride and form lactones with α2,3-linked sialic acids. This neutralization effectively labels each type of sialic acid with a unique mass to determine specific linkages on sialylated N-glycans. For both neutralization schemes, we compared the results from microchip electrophoresis and CE.

    更新日期:2017-09-10
  • Evaluation of intensity drift correction strategies using MetaboDrift, a normalization tool for multi-batch metabolomics data
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-09
    Chanisa Thonusin, Heidi B. IglayReger, Tanu Soni, Amy E. Rothberg, Charles F. Burant, Charles R. Evans

    In recent years, mass spectrometry-based metabolomics has increasingly been applied to large-scale epidemiological studies of human subjects. However, the successful use of metabolomics in this context is subject to the challenge of detecting biologically significant effects despite substantial intensity drift that often occurs when data are acquired over a long period or in multiple batches. Numerous computational strategies and software tools have been developed to aid in correcting for intensity drift in metabolomics data, but most of these techniques are implemented using command-line driven software and custom scripts which are not accessible to all end users of metabolomics data. Further, it has not yet become routine practice to assess the quantitative accuracy of drift correction against techniques which enable true absolute quantitation such as isotope dilution mass spectrometry. We developed an Excel-based tool, MetaboDrift, to visually evaluate and correct for intensity drift in a multi-batch liquid chromatography − mass spectrometry (LC-MS) metabolomics dataset. The tool enables drift correction based on either quality control (QC) samples analyzed throughout the batches or using QC-sample independent methods We applied MetaboDrift to an original set of clinical metabolomics data from a mixed-meal tolerance test (MMTT). The performance of the method was evaluated for multiple classes of metabolites by comparison with normalization using isotope-labeled internal standards. QC sample-based intensity drift correction significantly improved correlation with IS-normalized data, and resulted in detection of additional metabolites with significant physiological response to the MMTT. The relative merits of different QC-sample curve fitting strategies are discussed in the context of batch size and drift pattern complexity. Our drift correction tool offers a practical, simplified approach to drift correction and batch combination in large metabolomics studies.

    更新日期:2017-09-10
  • Preparation of a reversed-phase/anion-exchange mixed-mode spherical sorbent by Pickering emulsion polymerization for highly selective solid-phase extraction of acidic pharmaceuticals from wastewater
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-08
    Chaonan Huang, Yun Li, Jiajia Yang, Junyu Peng, Jing Jin, Dhanjai, Jincheng Wang, Jiping Chen

    The present work represents a simple and effective preparation of a novel mixed-mode anion-exchange (MAX) sorbent based on porous poly[2-(diethylamino)ethyl methacrylate-divinylbenzene] (poly(DEAEMA-DVB)) spherical particles synthesized by one-step Pickering emulsion polymerization. The poly(DEAEMA-DVB) particles were quaternized with 1,4-butanediol diglycidyl ether (BDDE) followed by triethylamine (TEA) via epoxy-amine reaction to offer strong anion exchange properties. The synthesized MAX sorbent was characterized by scanning electron microscopy, Fourier-transform infrared spectroscopy, nitrogen adsorption-desorption measurements and elemental analysis. The MAX sorbent possessed regular spherical shape and narrow diameter distribution (15–35 μm), a high IEC of 0.54 meq/g, with carbon and nitrogen contents of 80.3% and 1.62%, respectively. Compared to poly(DEAEMA-DVB), the MAX sorbent exhibited decreased SBET (390.5 vs. 515.3 m2 g−1), pore volume (0.74 vs. 0.85 cm3 g−1) and pore size (16.8 vs. 17.3 nm). Moreover, changes of N content for producing the MAX sorbent reveal a successful two-step quaternization, which can be highly related to such a high IEC. Finally, the MAX sorbent was successfully evaluated for selective isolation and purification of some selected acidic pharmaceuticals (ketoprofen, KEP; naproxen, NAP; and ibuprofen, IBP) from neutral (hydrocortisone, HYC), basic (carbamazepine, CAZ; amitriptyline, AMT) pharmaceuticals and other interferences in water samples using solid phase extraction (SPE). An efficient analytical method based on the MAX-based mixed-mode SPE coupled with HPLC-UV was developed for highly selective extraction and cleanup of acidic KEP, NAP and IBP in spiked wastewater samples. The developed method exhibited good sensitivity (0.009–0.085 μg L−1 limit of detection), satisfactory recoveries (82.1%–105.5%) and repeatabilities (relative standard deviation <7.9%, n = 3).

    更新日期:2017-09-09
  • Enantiomeric Separation of Some Chiral Analytes Using Amylose 3,5-Dimethylphenylcarbamate Covalently Immobilized on Silica by Nano-Liquid Chromatography and Capillary Electrochromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-08
    Giovanni D’Orazio, Chiara Fanali, Marina Karchkhadze, Bezhan Chankvetadze, Salvatore Fanali

    A newexperimental chiral stationary phase containing amylose 3,5-dimethylphenylcarbamate immobilized onto silica gel (i-ADMPC) was packed in 100 μm I.D. fused silica capillary and used for the chiral separation of eight selected flavanone derivatives in polar organic mobile phase by using different miniaturized techniques, such as nano-liquid chromatography (nano-LC), capillary electrochromatography (CEC)and pressure-assisted CEC (pCEC). A comparative study of different elution modes in terms of chromatographic efficiency, analysis time and enantiomeric resolution was carried out. In pCEC mode, the highest chromatography performance wasobtained applying +2.5 kV voltage and inlet pressure 10 bar. Under these conditions, the analysis times were shorter than 20 min, and chromatographic efficiencies were in the range 33,000-49,000 plates/m (first eluted peak).The solvent versatility of packed i- ADMPC capillary column was also investigated. In nano-LC, the CSP was stressed under the use of strong organic solvents such as ethyl acetate, acetone and methyl t-butyl ether (MTBE). After at least 40 working hours and over 50 sample injections, the CSP resulted to be very stable allowing to achieve good repeatability employing again the mobile phase containing polar organic solvent. Enantioresolutions and chromatographic efficiencies decreased by 1.60 and 10%, respectively as the result of using above mentioned strong solvents.

    更新日期:2017-09-09
  • High-Resolution 2D-LC Analysis of Key Linker Drug Intermediate Used in Antibody Drug Conjugates (ADCs)
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-08
    C.J. Venkatramani, Shu Rong Huang, Mohammad Al-Sayah, Ila Patel, Larry Wigman

    In this manuscript, the application of high-resolution sampling (HRS) two-dimensional liquid chromatography (2D-LC) in the detailed analysis of key linker drug intermediate is presented. Using HRS, selected regions of the primary column eluent were transferred to a secondary column with fidelity enabling qualitative and quantitative analysis of linker drugs. The primary column purity of linker drug intermediate ranged from 88.9% to 94.5% and the secondary column purity ranged from 99.6% to 99.9%, showing lot-to-lot variability, significant differences between the three lots, and substantiating the synthetic and analytical challenges of ADCs. Over 15 impurities co-eluting with the linker drug intermediate in the primary dimension were resolved in the secondary dimension. The concentrations of most of these impurities were over three orders of magnitude lower than the linker drug. Effective peak focusing and high-speed secondary column analysis resulted in sharp peaks in the secondary dimension, improving the signal-to-noise ratios. − The sensitivity of 2D-LC separation was over five fold better than conventional HPLC separation. The limit of quantitation (LOQ) was less than 0.01%. Many peaks originating from primary dimension were resolved into multiple components in the complementary secondary dimension, demonstrating the complexity of these samples. The 2D-LC was highly reproducible, showing good precision between runs with %RSD of peak areas less than 0.1 for the main component. The absolute difference in the peak areas of impurities less than 0.1% were within +/− 0.01% and for impurities in the range of 0.1% to 0.3%, the absolute difference were +/− 0.02%, which are comparable to 1D-LC. The overall purity of the linker drug intermediate was determined from the product of primary and secondary column purity (HPLC Purity =%peak area of main component in the primary dimension x %peak area of main component in the secondary dimension). Additionally, the 2D-LC separation enables the determination of potential impurities that could impact the downstream process, like ADCs stability, efficacy and patient safety. Peak capacity of this magnitude, sensitivity and reproducibility of 2D-LC for resolving structurally similar impurities co-eluting with the main component has not been demonstrated to date. This application clearly demonstrates the power of 2D-LC in detailed analysis of structurally similar, co-eluting impurities from key linker drug intermediate used in ADCs that is impossible to achieve by conventional 1D-LC.

    更新日期:2017-09-08
  • A useful strategy based on chromatographic data combined with Quality-by-Design approach for food analysis applications. The case study of furanic derivatives in sugarcane honey. ☆
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-08
    Pedro Silva, Catarina L. Silva, Rosa Perestrelo, Fernando M. Nunes, José S. Câmara

    Sugarcane honey (SCH) is one of the Madeira Island products par excellence and it is now popular worldwide. Its sui generis and peculiar sensory properties, explained by a variety of volatile compounds including furanic derivatives (FDs), arise mainly from manufacturing and storage conditions. A simple high-throughput approach based on semi-automatic microextraction by packed sorbent (MEPS) combined with ultra-high performance liquid chromatography (UHPLC) was developed and validated for identification and quantification of target FDs in sugarcane honey. A Quality-by-Design (QbD) approach was used as a powerful strategy to optimize analytical conditions for high throughput analysis of FDs in complex sugar-rich food matrices. The optimum point into MEPS-Method Operable Design: Region (MODR) was obtained with R-CX sorbent, acetonitrile (ACN) as elution solvent, three loading cycles and 500 μL of sample volume. The optimum point into UHPLC-MODR was obtained with a CORTECS column operating at a temperature of 50 °C, ACN as eluent and a flow rate of 125 μL min−1. The robustness was demonstrated by Monte Carlo simulation and capability analysis for estimation of residual errors. The concentration-response relationship for all FDs were described by polynomial function models, being confirmed by Fisher variance (F-test). The % recoveries were in a range of 91.9-112.1%. Good method precision was observed, yielding relative standard deviations (RSDs) less than 4.9% for repeatability and 8.8% for intermediate precision. The limits of quantitation for the analytes ranged from 30.6 to 737.7 μg kg−1. The MEPSR-CX/UHPLCCORTECS-PDA method revealed an effective and potential analytical tool for SCH authenticity control based on target analysis of FDs allowing a strict control and differentiation from other similar or adulterated products.

    更新日期:2017-09-08
  • Rapid Quantitative Detection of Glucose Content in Glucose Injection by Reaction Headspace Gas Chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-08
    Wei-Qi Xie, Yi-Xian Gong, Kong-Xian Yu

    This work investigates an automated method for rapid quantifying the glucose content in glucose injection by reaction headspace gas chromatography (HS-GC). This technique is based on the oxidation reaction of glucose in glucose injection with potassium dichromate under the acidic conditions. The carbon dioxide (CO2) formed from the oxidation reaction can be quantitatively detected by GC. The results show that the conversion of glucose in glucose injection can be achieved under the given conditions (at 85 °C for 20 min), the relative standard deviation (RSD) of this HS-GC technique in the glucose determination was within 2.91%, and the measured glucose contents in glucose injection samples closely match those measured by the reference technique (relative differences <6.45%). The new HS-GC technique is rapid, practical and can be used to the batch detection of the glucose content in glucose injection related applications.

    更新日期:2017-09-08
  • Headspace solid-phase microextraction coupled to comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry for the analysis of aerosol from tobacco heating product
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-07
    Benjamin Savareear, Radoslaw Lizak, Michał Brokl, Chris Wright, Chuan Liu, Jean-Francois Focant

    A method involving headspace solid-phase microextraction (HS-SPME) and comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC × GC-TOFMS) was developed and optimised to elucidate the volatile composition of the particulate phase fraction of aerosol produced by tobacco heating products (THPs). Three SPME fiber types were studied in terms of extraction capacity and precision measurements. Divinylbenzene polydimethylsiloxane appeared as the most efficient coating for these measurements. A central composite design of experiment was utilised for the optimization of the extraction conditions. Qualitative and semi-quantitative analysis of the headspace above THP aerosol condensate was carried out using optimised extraction conditions. Semi-quantitative analyses of detected constituents were performed by assuming that their relative response factors to the closest internal standard (itR) were equal to 1. Using deconvoluted mass spectral data (library similarity and reverse match >750) and linear retention indices(match window of ±15 index units), 205 peaks were assigned to individual compounds, 82 of which (including 43 substances previously reported to be present in tobacco) have not been reported previously in tobacco aerosol. The major volatile fraction of the headspace contained ketones, alcohols, aldehydes, alicyclic hydrocarbons alkenes, and alkanes. The method was further applied to compare the volatiles from the particulate phase of aerosol composition of THP with that of reference cigarette smoke and showed that the THP produced a less complex chemical mixture. This new method showed good efficiency and precision for the peak areas and peak numbers from the volatile fraction of aerosol particulate phase for both THP and reference cigarettes.

    更新日期:2017-09-08
  • Thin layer chromatography in drug discovery process
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-07
    Krzesimir Ciura, Szymon Dziomba, Joanna Nowakowska, Michał J. Markuszewski

    The review is mainly focused on application of thin layer chromatography (TLC) as simple, rapid and inexpensive method for lipophilicity assessment. Among separation techniques, TLC is still one of the most popular for lipophilicity measurement. The principles and methodology of Quantitative Structure Retention Relationship (QSRR) employed to lipophilicity prediction from retention data are presented. Moreover, applications of TLC retention constants in Quantitative Structure Activity Relationship (QSAR) studies were critically overviewed. The paper concerns also bioautography as a TLC method complementary to QSAR studies. In the article, the advantages and limitations of well established and less common planar chromatography modes applied for drug discovery process were discussed.

    更新日期:2017-09-08
  • Towards a chromatographic similarity index to establish localised Quantitative Structure-Retention Relationships for retention prediction. III Combination of Tanimoto similarity index, logP, and retention factor ratio to identify optimal analyte training sets for ion chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-07
    Soo Hyun Park, Paul R. Haddad, Ruth I.J. Amos, Mohammad Talebi, Roman Szucs, Christopher A. Pohl, John W. Dolan

    Retention prediction for unknown compounds based on Quantitative Structure-Retention Relationships (QSRR) can lead to rapid “scoping” method development in chromatography by simplifying the selection of chromatographic parameters. The use of retention factor ratio (or k-ratio) as a chromatographic similarity index can be a potent method to cluster similar compounds into a training set to generate an accurate predictive QSRR model provided that its limitation − that the method is impractical for retention prediction for unknown compounds – is successfully addressed. In this work, we propose a localised QSRR modelling approach with the aim of compensating the critical limitation in the otherwise successful k-ratio filter-based QSRR modelling. The approach is to combine a k-ratio filter with both Tanimoto similarity (TS) and a ΔlogP index (i.e., logP-Dual filter). QSRR models for two retention parameters (a and b) in the linear solvent strength (LSS) model in ion chromatography (IC), logk = a − blog[eluent], were generated for larger organic cations (molecular mass up to 506) on a Thermo Fisher Scientific CS17 column. The application of the developed logP-Dual filter resulted in the production of successful QSRR models for 50 organic cations out of 87 in the dataset. The predicted a- and b-values of the models were then applied to the LSS model to predict the corresponding retention times. External validation showed that QSRR models for a-, b- and tR- values with excellent accuracy and predictability (Qext(F2)2 of 0.96, 0.95, and 0.96, RMSEP of 0.06, 0.02, and 0.38 min) were created successfully, and these models can be employed to speed up the “scoping” phase of method development in IC.

    更新日期:2017-09-08
  • In-line coupling of supported liquid membrane extraction to capillary electrophoresis for simultaneous analysis of basic and acidic drugs in urine
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-07
    Pavla Pantůčková, Pavel Kubáň

    Simultaneous extraction of basic and acidic drugs across thin supported liquid membrane (SLM) and direct injection of the extracted drugs from SLM surface into capillary electrophoresis (CE) were demonstrated. A microextraction device compatible with injection system of commercial CE instrument was filled with 20 μL of sample and 10 μL of acceptor solution, which were interspaced by the SLM impregnated with 5 μL of organic solvent. Extractions of three basic drugs (nortriptyline, haloperidol and loperamide) and two acidic drugs (ketoprofen and naproxen) were achieved at optimized conditions including 1-ethyl-2-nitrobenzene as SLM solvent, natural pH of sample solution, 2.5 mM NaOH acceptor solution and 30 min extraction time. The extracted drugs were directly injected into CE for separation and quantification in a background electrolyte solution consisting of 30 mM ammonium acetate adjusted to pH 4.2 with acetic acid. The entire analytical procedure including drugs extraction, injection, separation and quantification was automated in the CE instrument and the only manual procedures were SLM impregnation and filling the microextraction device with sample and acceptor solutions. The analytical method was suitable for simultaneous determination of basic and acidic drugs in undiluted human urine samples and was used for direct determination of naproxen in urine after oral administration of Nalgesin S tablet. Efficient elimination of sample matrix and selective transfer of basic and acidic drugs were achieved and the hyphenated SLM-CE method was characterized by repeatability of peak areas ranging from 3.7 to 13.4%, linear relationship between peak areas and concentrations (r2 = 0.994–0.999) and limits of detection between 0.05 and 1.5 μg/mL.

    更新日期:2017-09-08
  • Triphenylamine-based hypercrosslinked organic polymer as adsorbent for the extraction of phenylurea herbicides
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Juanjuan Wu, Ruiyang Ma, Lin Hao, Chun Wang, Qiuhua Wu, Zhi Wang

    A hypercrosslinked organic polymer material (named as PPTPA) was prepared by a simple one-step self-polycondensation of triphenylamine. A series of characterization experiments, including N2 adsorption, scanning electron microscopy, X-ray diffraction, Fourier transform infrared spectra, and thermogravimetric analysis, were carried out to evaluate the morphology, structure and other intrinsic properties of the PPTPA. To investigate its adsorption performance, the PPTPA was used as the solid-phase extraction adsorbent for the extraction of phenylurea herbicides from water, milk and tomato juice samples followed by high-performance liquid chromatographic analysis. Under optimum conditions, good linearity for the analytes was observed in the range of 0.05–40.0 ng mL−1, 0.1-40.0 ng mL−1 and 0.5–40.0 ng mL−1 for water, milk and tomato juice samples, respectively. The established method also showed low limits of detection (S/N = 3) in the range of 0.008–0.01 ng mL−1, 0.01–0.03 ng mL−1 and 0.05–0.1 ng mL−1 for water, milk and tomato juice samples, respectively. The possible adsorption mechanism of the PPTPA towards the analytes was investigated, and the results demonstrated that the hydrogen bonding was the main interaction force between the PPTPA and the analytes. It suggests that the PPTPA can serve as a promisingadsorbent for the efficient pre-enrichment of organic compounds with more hydrogen-bonding donor sites.

    更新日期:2017-09-07
  • Preparation of a magnetic porous organic polymer for the efficient extraction of phenylurea herbicides
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Juntao Wang, Menghua Li, Caina Jiao, Yuhong Song, Chun Wang, Qiuhua Wu, Zhi Wang

    A magnetic porphyrin-based porous organic polymer (M-PPOP) with good porosity, high surface area and strong magnetism was prepared and employed as the adsorbent for the magnetic solid phase extraction of the phenylurea herbicides (metoxuron, monolinuron, chlorotoluron, and buturon) from bottled grape juice and tomato samples prior to their determination by high performance liquid chromatography. Under the optimized conditions, the developed method exhibited a good linear range, low limits of detection, low relative standard deviations (<6.8%), good method recoveries between 80.8% and 117%, and high enrichment factors (51–106). For a better elucidation of the adsorption of the M-PPOP towards the analytes, the extraction performance of the M-PPOP for different types of organic compounds including polycyclic aromatic hydrocarbons, phenylurea insecticides, phenylurea herbicides and phenols was studied. The results indicated that the π- stacking, hydrogen-bonding and hydrophobic interactions between the M-PPOP and the analytes played a main role for the adsorption. The results revealed that the M-PPOP material had a great potential for the enrichment of more organic pollutants from real samples.

    更新日期:2017-09-07
  • Counter-current Chromatography with Off-line Detection by Ultra High Performance Liquid Chromatography/High Resolution Mass Spectrometry in the Study of the Phenolic Profile of Lippia origanoides
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Suzana Guimaraes Leitão, Gilda Guimarães Leitão, Douglas K.T. Vicco, João Paulo Barreto Pereira, Gustavo de Moraes Simão, Danilo R. Oliveira, Rita Celano, Luca Campone, Anna Lisa Piccinelli, Luca Rastrelli

    Lippia origanoides (Verbenaceae) is an important Brazilian medicinal plant, also used for culinary purposes. Most chemical studies with this plant have been focused on its volatile composition. In this work, we combined High-Speed Counter-current Chromatography (HSCCC) and High Performance Liquid Chromatography coupled to Ultra Violet detection and High Resolution Mass Spectrometry (HPLC-UV-HRMSn) methodologies to access the non-volatile chemical composition of L. origanoides. The crude ethanol extract of L. origanoides (LOEF) was first analyzed by HPLC-UV-HRMSn and allowed the identification of 7 major compounds. Among them, eriodictyol, naringenin and pinocembrin, were determined and are phytochemical markers of this plant. However, owing to the complexity of this plant matrix, LOEF was fractionated by HSCCC (hexane-ethanol-water, 4:3:1) as a tool for preparative pre-purification, affording a flavonoid-rich fraction. A column screening with the chromatographic stationary phases ZIC-HILIC, monolithic and particulate RP18 was performed. The best column separation was achieved with a Purospher STAR RP18e, which was used for HPLC-DAD-HRMSn studies. By this approach 12 compounds were further identified in addition to the major ones identified in the raw extract. Two of them, 6,8-di-C-hexosyl-luteolin and 6,8-di-C-glucosyl-apigenin, are being reported for the first time in the family Verbenaceae. This work shows the integration of HSCCC as a preparative tool for the fractionation and purification of natural products from a complex plant extract with other analytical techniques, with the purpose of showing each technique’s potential.

    更新日期:2017-09-07
  • Determination of Molar Masses of Macromolecules by SEC-Light Scattering not Requiring Knowledge of Refractive Index Increments
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Simona Lavric, Jasmin Preis, Christine Rosenauer, Wolfgang Radke

    A new approach for the calibration of SEC-light scattering (SEC-LS) setups is proposed, which requires solely the molar mass of a reference polymer. Neither the specific refractive index increment of the calibrant nor of the analyte is required. Comparison of the molar masses derived in different solvents for a large number of chemically different polymers shows that the new approach yields the same molar masses as if molar masses were derived using dn/dc to calibrate the light scattering setup. The approach therefore allows easier determination of molar masses by SEC-LS.

    更新日期:2017-09-07
  • Hollow mesoporous carbon spheres-based fiber coating for solid-phase microextraction of polycyclic aromatic hydrocarbons
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Xingru Hu, Chao Liu, Jiansheng Li, Rui Luo, Hui Jiang, Xiuyun Sun, Jinyou Shen, Weiqing Han, Lianjun Wang

    In this study, a novel hollow mesoporous carbon spheres-based fiber (HMCSs-F) was fabricated to immobilize HMCSs onto a stainless steel wire for solid-phase microextraction (SPME). Characterization results showed that the HMCSs-F possessed a large specific surface area, high porosity and uniform pore size. To demonstrate the extraction performance, a series of polycyclic aromatic hydrocarbons (PAHs) was chosen as target analytes. The experimental parameters including extraction and desorption conditions were optimized. Compared to commercial fibers, the HMCSs-F exhibited better extraction efficiency for PAHs. More interestingly, a good extraction selectivity for PAHs from the complex matrix was observed in these HMCSs-F. The enhanced SPME performance was attributed to the unique pore structure and special surface properties of the HMCSs. Furthermore, under the optimum conditions, the limits of detection (LODs) for the HMCSs-F were in the range of 0.20 to 1.15 ng L−1 with a corresponding relative standard deviation that was below 8.6%. The method was successfully applied for the analysis of PAHs in actual environmental water samples with recoveries ranging from 85.9% to 112.2%. These results imply that the novel HMCSs-F have potential application in environmental water analysis.

    更新日期:2017-09-07
  • New perspectives in high efficient and ultrafast chiral liquid chromatography through zwitterionic teicoplanin-based 2-micron superficially porous particles
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    Omar H. Ismail, Michela Antonelli, Alessia Ciogli, Claudio Villani, Alberto Cavazzini, Martina Catani, Simona Felletti, David S. Bell, Francesco Gasparrini

    With the aim of pushing forward the limits of high efficient and ultrafast chiral liquid chromatography, a new Chiral Stationary Phase (CSP) has been prepared by covalently bonding the teicoplanin selector on 2.0 μm Superficially Porous Particles (SPPs). An already validated bonding protocol, which permits to achieve teicoplanin-based CSPs exhibiting zwitterionic behaviour, has been employed to prepare not only the 2.0 μm version of the CSP but also two other analogous CSPs based, respectively, on 2.7 μm SPPs and 1.9 μm Fully Porous Particles (FPPs). The kinetic performance of these CSPs has been compared through the analysis of both van Deemter curves and kinetic plots by employing in-house packed columns of 4.6 mm internal diameter and different lengths (20, 50 and 100 mm). In particular on the columns packed with 2.0 μm SPPs, extremely large efficiencies were observed for both achiral ( > 310,000 theoretical plates/meter, N/m; hr: 1.61) and chiral compounds (>290,000 N/m; hr: 1.72) in HILIC conditions. Thanks to their efficiency and enantioselectivity, these CSPs were successfully employed in ultrafast chiral separations. As an example, the enantiomers of haloxyfop were baseline resolved in about 3 seconds, with a resolution higher than 2.0, (flow rate: 8 mL/min) on a 2 cm long column packed with the 2.0 μm chiral SPPs.

    更新日期:2017-09-06
  • Ion-pair in-tube solid phase microextraction for the simultaneous determination of phthalates and their degradation products in atmospheric particulate matter ☆
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-06
    M. Fernández-Amado, M.C. Prieto-Blanco, P. López-Mahía, S. Muniategui-Lorenzo, D. Prada-Rodríguez

    An in-tube solid phase microextraction, coupled with high-performance liquid chromatography with diode array detection (IT-SPME-HPLC-DAD) method,has been developed for the simultaneous determination of 13 diesters(from dimethyl to dioctylphthalateplus diisobutyl, benzylbutyl, 2-ethylhexyl,diisononyl, diisodecylphthalate)and 2 monoesters of phthalic acid (mono-butyl and mono-(2-ethylhexyl) phthalate) in particulate matter (PM10). Triethylamine at pH = 3 was used as an ion-pair reagent with a double function, of regulating the chromatographic retention of the monoesters and the most hydrophilic diesters on a monolithic silica column, and of improving their extraction on a porous polymer with divinylbenzene-4-vinylpyridine capillary. The chromatographic separation was achieved in 13 minutes. A previous ultrasound-assisted extraction from PM10filters was also optimized using methanol as solvent. The method detection limits were 0.09-0.52 ng m−3, the inter-day precision at concentration of 20 ng mL−1 was between 4.2% and 12.7% (n = 15), and the average recovery was 87.3%. The average absolute IT-SPME recovery was 26.2% and the linear range reached up to 109 ng m−3 for most analytes. The method was applied to PM10 samples from different environments collected in Galicia (Spain).DiBP was the major phthalate, followed by its isomer DnBP in urban sites and by DEP in the suburban area. In all samples, DEHP quantified correlates with the isomers of dibutylphthalate. Total PAE concentration was between 14.5 and 245.5 ng m−3. To the best of our knowledge, this is the first time that a method allows the simultaneous determination of 13 phthalates and their degradation products in particulate matter.

    更新日期:2017-09-06
  • Matrix-effect free quantitative Liquid Chromatography Mass Spectrometry analysis in complex matrices using nanoflow LC with integrated emitter tip and high dilution factors ☆
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-05
    David Moreno-González, Jaime Alcántara-Durán, Bienvenida Gilbert-López, Juan F. García-Reyes, Antonio Molina-Díaz

    Matrix effects are probably the Achilles heel of most quantitative liquid chromatography mass spectrometry (LC-MS) methods based on electrospray ionization. This work reports the evaluation of matrix effects in challenging matrices such as food extracts, human urine or wastewater at different dilution factors using nanoflow liquid chromatography-high resolution mass spectrometry (LC-MS). For this purpose, a suite of representative low-molecular weight compounds such as pesticides, drugs of abuse, sport drugs or environmental contaminants were selected. The approach is based on the use of reversed-phase C18 nano columns furnished with an integrated emitter tip. The nanoflow LC system was combined with full-scan high resolution mass spectrometry using a HRMS (orbitrap) instrument operated at a resolution of 70000. The sensitivity achieved with this configuration enables the implementation of high dilution factors (eg. 1:20, 1:50 or beyond). When combining nanoflow LC-MS analysis with such high dilution factors (eg. 1:50), signal suppression was negligible in most cases, so that matrix-matched standards may eventually be skipped, simplifying laboratory workflows by using external calibration in demanding applications such as drug analysis in urine, environmental contaminants in wastewater or pesticide testing in food, thus, eliminating the need for standard addition, matrix-matched calibration or isotopically-labelled standards.

    更新日期:2017-09-06
  • Construction of a new hydrazone-linked chiral covalent organic framework–silica composite as the stationary phase for high performance liquid chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-05
    Kai Zhang, Song-Liang Cai, Yi-Lun Yan, Zi-Hao He, Hui-Mei Lin, Xiao-Ling Huang, Sheng-Run Zheng, Jun Fan, Wei-Guang Zhang

    Covalent organic frameworks (COFs), as an emerging class of crystalline porous organic polymers, have great potential for applications in chromatographic separation owning to their fascinating crystalline structures and outstanding properties. However, development of COF materials as novel stationary phases in high performance liquid chromatography (HPLC) is just in its infancy. Herein, we report the design and construction of a new hydrazone-linked chiral COF, termed BtaMth COF, from a chiral hydrazide building block (Mth) and present a one-pot synthetic method for the fabrication of BtaMth@SiO2 composite for HPLC separation of isomers. The as-synthesized BtaMth chiral COF displays good crystallinity, high porosity, as well as excellent chemical stability. Meanwhile, the fabricated HPLC column by using BtaMth@SiO2 composite as the new stationary phase exhibits high resolution performances for the separation of positional isomers including nitrotoluene and nitrochlorobenzene, as well as cis-trans isomers including beta-cypermethrin and metconazole. Additionally, some effects such as the composition of the mobile phase and column temperature for HPLC separations on the BtaMth@SiO2 packed column also have been studied in detail. The successful applications indicate the great potentials of hydrazone-linked chiral COF–silica composite as novel stationary phase for the efficient HPLC separation.

    更新日期:2017-09-06
  • The Development of a Monolith-Based PURIFICATION Process for Orthopoxvirus vaccinia virus Lister strain
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-05
    David Vincent, Petra Kramberger, Rosana Hudej, Aleš Štrancar, Yaohe Wang, Yuhong Zhou, Ajoy Velayudhan

    The purification of large viruses remains an important field of research and development. The development of efficient purification trains is limited by limited analytical methods, as well as by the complexity of large viruses, as well as the high variability in starting material from cell culture. Vaccinia virus holds great potential as an oncolytic and immunotherapeutic vaccine against a broad spectrum of cancers. In this work, monolith-based capture and polishing chromatographic steps for vaccinia virus Lister strain has been developed. Virus produced in CV-1 cells was harvested and passed through a 0.8 μm pre-filter before loading onto CIEX, AIEX and HIC CIM monoliths. Without the need for nuclease treatment, up to 99% of the total DNA loaded can be removed from the vaccinia feed stream by the CIM OH monolith, which also reduces the total protein concentration in the product pool to LLOQ levels, and achieves infectious virus recoveries of 90%. Binding capacities of greater than 1 × 109 pfu of vaccinia per mL of matrix were obtained on both CIM SO3 and CIM OH monoliths. Multiple orthogonal analytical methods have been used to develop process knowledge and understanding.

    更新日期:2017-09-06
  • The use of dissolvable layered double hydroxide components in an in situ solid-phase extraction for chromatographic determination of tetracyclines in water and milk samples
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-04
    Nattaphorn Phiroonsoontorn, Sira Sansuk, Yanawath Santaladchaiyakit, Supalax Srijaranai

    This research presents a simple and green in situ solid phase extraction (is-SPE) combined with high-performance liquid chromatography (HPLC) for the simultaneous analysis of tetracyclines (TCs) including tetracycline, oxytetracycline, and chlortetracycline. In is-SPE, TCs were efficiently extracted through the precipitation formation of dissolvable layered double hydroxides (LDHs) by mixing the LDH components such as magnesium and aluminum ions (both in metal chloride salts) thoroughly in an alkaline sample solution. After the centrifugation, the precipitate was completely dissolved with trifluoroacetic acid to release the enriched TCs, and then analyzed by HPLC. Under optimized conditions, this method gave good enrichment factors (EFs) of 41-93 with low limits of detection (LODs) of 0.7–6 μg/L and limits of quantitation (LOQs) of 3–15 μg/L. Also, the proposed method was successfully applied for the determination of TCs in water and milk samples with the recoveries ranging from 81.7-108.1% for water and 55.7-88.7% for milk.

    更新日期:2017-09-04
  • Novel acylhydrazone bond dynamic covalent polymer gel monolithic column online coupling to high-performance liquid chromatography for analysis of sulfonamides and fluorescent whitening agents in food
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-04
    Chengjiang Zhang, Xialin Luo, Tianfu Wei, Yufei Hu, Gongke Li, Zhuomin Zhang

    A new dynamic covalent polymer (DCP) gel was well designed and constructed based on imine chemistry. Polycondensation of 4,4′-biphenyldicarboxaldehyde and 1,3,5-benzenetricarbohydrazide via Schiff-base reaction resulted in an acylhydrazone bond gel (AB-gel) DCP. AB-gel DCP had three-dimensional network of interconnected nanoparticles with hierarchically porous structure. AB-gel DCP was successfully fabricated as a monolithic column by an in-situ chemical bonding method for online enrichment and separation purpose with excellent permeability. AB-gel DCP based monolithic column showed remarkable adsorption affinity towards target analytes including sulfonamides (SAs) and fluorescent whitening agents (FWAs) due to its strong π-π affinity, hydrophobic effect and hydrogen bonding interaction. Then, AB-gel DCP based monolithic column was applied for online separation and analysis of trace SAs and FWAs in food samples coupled with high-performance liquid chromatography (HPLC). Sulfathiazole (ST) and sulfadimidine (SM2) in one positive weever sample were actually found and determined with concentrations of 273.8 and 286.3 μg/kg, respectively. 2,5-Bis(5-tert-butyl-2-benzoxazolyl) thiophene (FWA184) was actually quantified in one tea infusion sample with the concentration of 268.5 ng/L. The spiked experiments suggested the good recoveries in range of 74.5–110% for SAs in weever and shrimp samples with relative standard deviations (RSDs) less than 9.7% and in range of 74.0-113% for FWAs in milk and tea infusion samples with RSDs less than 9.0%. AB-gel DCP monolithic column was proved to be a promising sample preparation medium for online separation and analysis of trace analytes in food samples with complex matrices.

    更新日期:2017-09-04
  • Synthesis and evaluation of pseudopeptide chiral stationary phases for enantioselective resolution
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-01
    Huifang Shen, Ganhong Du, Keyuan Liu, Long Ye, Shoulei Xie, Liming Jiang

    Poly(2-oxazoline)s are regarded as bioinspired polymers due to their structural relation to polypeptides. In this work, a new kind of poly(2-oxazoline)s containing dipeptide segments in the side chains was synthesized through a bottom-up protocol, which involves ring-opening copolymerization of 2-(N-Boc-L-2-pyrrolidinyl)-2-oxazoline (PyOXBoc) with 2-(3-butenyl)-2-oxazoline (BuOX) followed by deprotection and amide coupling with N-protected L-proline. The resulting vinyl-functionalized polymers were subsequently immobilized onto mercaptopropylated silica bead matrices by means of thio-click chemistry and their potential as the chiral stationary phase (CSP) for high-performance liquid chromatography was preliminarily evaluated with a series of structurally different racemates. The results showed that this class of pseudopeptide CSPs is particularly adapted to the enantiomeric separation of 1,1'-bi-2-naphthol and acyloin compounds (such as benzoin) under normal-phase conditions. Moreover, an increase in the length of polymer main chains is beneficial to the enhancement of both enantioselectivity and resolution ability. The chiral discrimination of analytes by the polymeric selectors stems primarily from hydrogen bonding and π-π interactions as well as steric hindrance.

    更新日期:2017-09-04
  • Characterization and Purification of Anthocyanins from Black Peanut (Arachis hypogaea L.) Skin by Combined Column Chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-01
    Zhenlei Zhao, Min Wu, Yali Zhan, Kanghua Zhan, Xiulian Chang, Hongshun Yang, Zhanming Li

    Black peanut skins as a byproduct from peanut industry contain abundant anthocyanins, evaluated as 8.61 ± 0.27 mg/g dry black peanut skins, are currently poorly exploited. In this work, four anthocyanins and three major flavonols were detected and identified by HPLC-PDA-ESI-MS/MS from the acidified water extract of black peanut skins of Arachis hypogaea L. After preliminary removal of flavonols by ethyl acetate (EtOAc), further purification of the anthocyanins was conducted using a combination of Amberlite XAD-7HP and ODS-AQ-HG column chromatography methods Two most abundant monomeric anthocyanins cyanidin-3-O-sophoroside (5.77 ± 0.42 mg) and cyanidin-3-O-sambubioside (4.10 ± 0.17 mg) were eventually obtained from 2 g dry black peanut skins, and their purities were determined by HPLC-PDA as 97.29% and 98.28% at the yields of 87.47% and 64.27% on the basis of their total amount in the crude extracts, respectively. These sequential treatments can be easily adapted to large-scale fractionation of pure anthocyanin monomers.

    更新日期:2017-09-04
  • Quantitation of lanosterol in the vitreous humor of rabbits after ocular administration of lanosterol/thermogel formulation by ultra high performance liquid chromatography–tandem mass spectrometry with the electrospray ionization mode
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-01
    Lei Lv, Dan Li, Hui Wang, Chengjian Li, Xian Qian, Heng-tao Dong, Jianguo Sun, Liang Zhao

    Cataracts are the most common cause of blindness worldwide affecting tens of millions of people. Here, we report a simple, rapid, sensitive and specific method by ultra performance liquid chromatography–tandem mass spectrometry with the electrospray ionization mode (UPLC-ESI-MS/MS) for quantitation of lanosterol, a possible effective drug for cataracts, in the vitreous humor of rabbits after ocular administration. The injected lanosterol was prepared by dispersing lanosterol molecules into the poly-(DL-lactic acid-co-glycolic acid)–poly(ethylene glycol)–poly-(DL-lactic acid-co-glycolic acid) (PLGA–PEG–PLGA) thermogel solution. The analyte and internal standard (IS, panaxadiol) were extracted by the simple protein precipitation with methanol. The chromatographic separation used an Agilent RRHD SB-C18 column with a methanol mobile phase containing 50 mM of ammonium acetate aqueous solution (with 0.1% formic acid) (95:5, v/v). The protonated analyte was quantitated in positive ionization by multiple reaction monitoring (MRM) with a mass spectrometer. The mass transitions m/z 443.5 → 235 and m/z 461 → 127 were used to measure the analyte and IS, respectively. The assay exhibited a linear dynamic range of 1–1250 ng mL−1 for lanosterol in vitreous samples. The lower limit of quantitation (LLOQ) was 1 ng mL−1 with a relative standard deviation (RSD) of less than 15%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 5 min per sample offered a throughput of more than 200 samples per day. This validated method was used to analyze vitreous samples of New Zealand white rabbits for pharmacokinetic studies. The results provided useful information on pharmacological action mechanism of lanosterol and were meaningful for cataract treatment among the elderly population.

    更新日期:2017-09-04
  • Quantification of Piperazine in Chicken and Pig Tissues by Gas Chromatography-Electron Ionization Tandem Mass Spectrometry Employing Pre-Column Derivatization with Acetic Anhydride
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-01
    Bo Wang, Maoda Pang, Xing Xie, Kaizhou Xie, Yangyang Zhang, Lulu Cui, Xia Zhao, Yajuan Wang, Huiqiang Shi, Yawen Guo, Ran Wang, Genxi Zhang, Guojun Dai, Jinyu Wang

    This paper describes a novel method that combines acetic anhydride derivatization with gas chromatography-electron ionization tandem mass spectrometry (GC-EI/MS/MS) for the sensitive and selective determination of piperazine in chicken and pig tissues. Samples were extracted using an accelerated solvent extraction (ASE) apparatus, purified by solid-phase extraction (SPE) and derivatized with acetic anhydride. This optimized method was validated according to the requirements defined by the European Union and the Food and Drug Administration. At the limit of quantification (LOQ) spiked levels of 50.0, 100.0, 500.0, 1000.0 and 2000.0 μg/kg, the average recoveries of piperazine in chicken and pig tissues were 77.46 ∼ 96.26%, with relative standard deviations (RSDs) of 1.55 ∼ 6.64%. The intra-day RSDs were 1.39 ∼ 5.92%, and the inter-day RSDs were 2.24 ∼ 8.39%. The limits of detection (LODs) and the LOQs were 1.4 ∼ 1.6 μg/kg and 4.8 ∼ 5.2 μg/kg, respectively. The decision limits (CCα) were 102.02 ∼ 105.17 μg/kg, and the detection capabilities (CCβ) were 104.03 ∼ 109.09 μg/kg. Finally, the new approach was verified for the quantitative determination of piperazine in 30 commercial chicken and pig tissues from local supermarkets.

    更新日期:2017-09-04
  • The reciprocal iso-inhibition volume concept: A procedure for the evaluation in effect-directed analysis with thin-layer chromatography - using the thin-layer chromatography-luminescent bacteria assay as an example
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-09-01
    Wolfgang Schulz, Stefan C. Weiss, Walter H. Weber, Rudi Winzenbacher

    In effect-directed analysis (EDA) with high-performance thin-layer chromatography (HPTLC), the effect is often detected using images. Thus, an approach to create inhibition chromatograms from these images was developed using the example of the HPTLC bioluminescence inhibition test. A comparison between the cuvette test and the HPTLC test shows that the test on the plate is significantly more sensitive. To describe the strength of the effect, the EC50 value is determined from the dose-response relationship. However, the inhibiting compounds are generally unknown and thus their concentrations are also unknown. Therefore, instead of the concentration, the known application volumes are used. This enables the calculation of the application volume necessary to achieve 50% inhibition. Since the volume is inversely proportional to the concentration, the reciprocal value of the calculated volume is indicated and is referred to as the reciprocal iso-inhibition volume (RIV). Using this RIV-concept, it is now possible to compare inhibition bands within and between plates. The entire evaluation is described by the means of two samples from a contaminated site using the bioluminescence inhibition.

    更新日期:2017-09-04
  • Studies of Drug Interactions with Alpha1-Acid Glycoprotein by Using On-Line Immunoextraction and High-Performance Affinity Chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-31
    Cong Bi, Ryan Matsuda, Chenhua Zhang, Zitha Isingizwe, William Clarke, David S. Hage

    A method that combined on-line immunoextraction with high-performance affinity chromatography was developed to examine the binding of drugs with α1-acid glycoprotein (AGP). Affinity microcolumns containing immobilized polyclonal anti-AGP antibodies were developed that had a capture efficiency of up to 98.4% for AGP and a binding capacity of 0.72 nmol AGP when using a 20 mm × 2.1 mm i.d. microcolumn. These microcolumns were employed in various formats to examine the binding of drugs to normal AGP and AGP that had been adsorbed from serum samples for patients with systemic lupus erythematosus (SLE). Drugs that were screened in zonal elution experiments for their overall binding to these types of AGP included chlorpromazine, disopyramide, imipramine, propranolol, and warfarin. Most of these drugs showed an increase in their binding to the AGP from SLE serum when compared to normal AGP (i.e., an increase of 13-76%); however, disopyramide gave a 21-25% decrease in retention when the same AGP samples were compared. Frontal analysis was used to further evaluate the binding of disopyramide and imipramine to these forms of AGP. Both drugs gave a good fit to a model that involved a combination of saturable and non-saturable interactions with AGP. Changes in the non-saturable interactions accounted for most of variations seen in the binding of disopyramide and imipramine with the AGP samples. The methods used in this study could be adapted for use in personalized medicine and the study of other proteins or drugs using aqueous mixtures or clinical samples.

    更新日期:2017-09-04
  • Capillary moving-boundary isotachophoresis with electrospray ionization mass-spectrometric detection and hydrogen ion used as essential terminator: Methodology for sensitive analysis of hydroxyderivatives of s-triazine herbicides in waters
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Zdena Malá, Petr Gebauer

    Capillary isotachophoresis (ITP) is an electrophoretic technique offering high sensitivity due to permanent stacking of the migrating analytes. Its combination with electrospray-ionization mass-spectrometric (ESI-MS) detection is limited by the narrow spectrum of ESI-compatible components but can be compensated by experienced system architecture. This work describes a methodology for sensitive analysis of hydroxyderivatives of s-triazine herbicides, based on implementation of the concepts of moving-boundary isotachophoresis and of H+ as essential terminating component into cationic ITP with ESI-MS detection. Theoretical description of such kind of system is given and equations for zone-related boundary mobilities are derived, resulting in a much more general definition of the effective mobility of the terminating H+ zone than used so far. Explicit equations allowing direct calculation for selected simple systems are derived. The presented theory allows prediction of stacking properties of particular systems and easy selection of suitable electrolyte setups. A simple ESI-compatible system composed of acetic acid and ammonium with H+ and ammonium as a mixed terminator was selected for the analysis of 2-hydroxyatrazine and 2-hydroxyterbutylazine, degradation products of s-triazine herbicides. The proposed method was tested with direct injection without any sample pretreatment and provided excellent linearity and high sensitivity with limits of detection below 100 ng/L (0.5 nM). Example analyses of unspiked and spiked drinking and river water are shown.

    更新日期:2017-08-31
  • Rapid capillary electrophoresis approach for the quantification of ewe milk adulteration with cow milk
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Francesca Trimboli, Valeria Maria Morittu, Caterina Cicino, Camillo Palmieri, Domenico Britti

    The substitution of ewe milk with more economic cow milk is a common fraud. Here we present a capillary electrophoresis method for the quantification of ewe milk in ovine/bovine milk mixtures, which allows for the rapid and inexpensive recognition of ewe milk adulteration with cow milk. We utilized a routine CE method for human blood and urine proteins analysis, which fulfilled the separation of skimmed milk proteins in alkaline buffer. Under this condition, ovine and bovine milk exhibited a recognizable and distinct CE protein profiles, with a specific ewe peak showing a reproducible migration zone in ovine/bovine mixtures. Based on ewe specific CE peak, we developed a method for ewe milk quantification in ovine/bovine skimmed milk mixtures, which showed good linearity, precision and accuracy, and a minimum amount of detectable fraudulent cow milk equal to 5%.

    更新日期:2017-08-31
  • Assessment of the removal of side nanoparticulated populations generated during one-pot synthesis by asymmetric flow field-flow fractionation coupled to elemental mass spectrometry
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Diego Bouzas-Ramos, Marta García-Cortes, Alfredo Sanz-Medel, Jorge Ruiz Encinar, José M. Costa-Fernández

    Coupling of asymmetric flow field-flow fractionation (AF4) to an on-line elemental detection (inductively coupled plasma-mass spectrometry, ICP-MS) has been recently proposed as a powerful diagnostic tool for characterization of the bioconjugation of CdSe/ZnS core-shell Quantum Dots (QDs) to antibodies. Such approach has been used herein to demonstrate that cap exchange of the native hydrophobic shell of core/shell QDs with the bidentate dihydrolipoic acid ligands directly removes completely the eventual side nanoparticulated populations generated during simple one-pot synthesis, which can ruin the subsequent final bioapplication. The critical assessment of the chemical and physical purity of the surface-modified QDs achieved allows to explain the transmission electron microscopy findings obtained for the different nanoparticle surface modification assayed.

    更新日期:2017-08-31
  • Analysis of underivatised low volatility compounds by comprehensive two dimensional gas chromatography with a short primary column
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Fábio Junior Moreira Novaes, Chadin Kulsing, Humberto Ribeiro Bizzo, Francisco Radler de Aquino Neto, Claudia Moraes de Rezende, Philip John Marriott

    Comprehensive two dimensional gas chromatography (GC×GC) approaches with cryogenic modulation were developed for the qualitative analysis of selected low volatility compounds in raw coffee bean extracts, without derivatisation. The approaches employed short first (1D) and second (2D) dimension columns, specifically a 1D 65% phenyl methyl siloxane column (11 m) and a 2D 5% phenyl methyl siloxane (1 m), which allowed elution of high molar mass compounds (e.g. > 600 Da). Solutes included hydrocarbons, fatty acids, diterpenes, tocopherols, sterols, diterpene esters, and di- and triacylglycerides. An oven temperature program up to 350 °C was employed. The effects of experimental conditions were investigated, revealing that the GC × GC results strongly depended on the cryogenic trap T, and oven T program. An appropriate condition was selected and further applied for group type analysis of low volatility compounds in green Arabica coffee beans. Retention indices were compiled for 1D analysis and were similar for the composite column data in GC × GC. The elution of some compounds was confirmed by use of authentic standards. The approach allowed direct analysis of coffee extract in ethyl acetate solution, with improved analyte peak capacity (approximately 200 compounds were detected) without prior fractionation or pre-treatment of the sample. This avoided potential hydrolysis of high molar mass conjugate esters as well as degradation of thermally labile compounds such as the derivatives of the diterpenes cafestol and kahweol.

    更新日期:2017-08-31
  • An automated and self-cleaning nano liquid chromatography mass spectrometry platform featuring an open tubular multi-hole crystal fiber solid phase extraction column and an open tubular separation column
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Meire Ribeiro da Silva, Ole Kristian Brandtzaeg, Tore Vehus, Fernando Mauro Lanças, Steven Ray Wilson, Elsa Lundanes

    An open tubular (OT) sample preparation/separation platform was developed. A multi-channel polymer layer open tubular (mPLOT) solid phase extraction (SPE) column was prepared by wall-coating the 126 channels (8 μm inner diameter (ID) each) of a crystal fiber capillary with an organic polymer, namely poly(styrene-co-octadecene-co-divinylbenzene) (PS-OD-DVB). The mPLOT SPE was coupled on-line with a 10 μm × 2 m poly(styrene-co-divinylbenzene) (PS-DVB) OT liquid chromatography column with nanospray mass spectrometry (OTLC-MS). Compared to using monolithic/particle-packed SPEs, mPLOT-SPE-OTLC allowed both fast loading and sufficient refocusing on the OT analytical column of small model compounds (sulfonamides ≈ 300 Da). Using automatic filtration/filter back-flushing (AFFL) plumbing, the mPLOT SPE column gave a constant and low back-pressure ≈35 bar at 0.5 μL/min. Surprisingly large sample volumes (10 μL) were possible to be injected using a 12 cm mPLOT.

    更新日期:2017-08-31
  • A multiple-dimension liquid chromatography coupled with mass spectrometry data strategy for the rapid discovery and identification of unknown compounds from a Chinese herbal formula (Er-xian decoction)
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Caihong Wang, Jinlan Zhang, Caisheng Wu, Zhe Wang

    It is very important to rapidly discover and identify the multiple components of traditional Chinese medicine (TCM) formula. High performance liquid chromatography with high resolution tandem mass spectrometry (HPLC single bond HRMS/MS) has been widely used to analyze TCM formula and contains multiple-dimension data including retention time (RT), high resolution mass (HRMS), multiple-stage mass spectrometric (MSn), and isotope intensity distribution (IID) data. So it is very necessary to exploit a useful strategy to utilize multiple-dimension data to rapidly probe structural information and identify chemical compounds. In this study, a new strategy to initiatively use the multiple-dimension LC-MS data has been developed to discover and identify unknown compounds of TCM in many styles. The strategy guarantees the fast discovery of candidate structural information and provides efficient structure clues for identification. The strategy contains four steps in sequence: (1) to discover potential compounds and obtain sub-structure information by the mass spectral tree similarity filter (MTSF) technique, based on HRMS and MSn data; (2) to classify potential compounds into known chemical classes by discriminant analysis (DA) on the basis of RT and HRMS data; (3) to hit the candidate structural information of compounds by intersection sub-structure between MTSF and DA (M,D-INSS); (4) to annotate and confirm candidate structures by IID data. This strategy allowed for the high exclusion efficiency (greater than 41%) of irrelevant ions in er-xian decoction (EXD) while providing accurate structural information of 553 potential compounds and identifying 66 candidates, therefore accelerating and simplifying the discovery and identification of unknown compounds in TCM formula.

    更新日期:2017-08-31
  • Comprehensive two dimensional liquid chromatography as analytical strategy for pharmaceutical analysis
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-30
    Marion Iguiniz, Florent Rouvière, Estelle Corbel, Nicolas Roques, Sabine Heinisch

    Comprehensive on-line two-dimensional liquid chromatography (LCxLC) is expected to generate impressive peak capacities, which makes it a method of choice for the analysis of complex samples such as pharmaceuticals. A comparative study of different sets of chromatographic conditions including stationary phase, pH additive and organic modifier was carried out with two real pharmaceutical samples in order to find out the best analytical conditions for implementation of one or several generic on-line LCxLC separations. Our choice was based on the evaluation of both degree of orthogonality and practical sample peak capacity under linear gradient conditions. The potential of 190 combinations of chromatographic systems was compared. A set of 3 RPLCxRPLC configurations was found to be very attractive for both samples and in good agreement with the findings of a previous study carried out with 17 model compounds, thereby supporting the idea of using generic LCxLC conditions in the pharmaceutical area. The three selected 2D-systems were implemented for the on-line RPLCxRPLC-UV/MS analysis of two pharmaceutical samples. It was shown, for each sample, that these 2D-systems were able to generate an effective peak capacity close to 1000 in less than 50 min. For each sample, baseline separation was obtained for every known compound and furthermore a large number of unknown impurities could also be separated and identified. Finally, in the proposed conditions, the total number of compounds detected was significantly improved from one RPLC separation to one RPLCxRPLC separation. Only a small additional gain was observed by performing a second RPLCxRPLC separation or even a third one.

    更新日期:2017-08-31
  • Dispersive micro solid phase extraction (DMSPE) using polymer anion exchange (PAX) as the sorbent followed by UPLC–MS/MS for the rapid determination of four bisphenols in commercial edible oils
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-28
    Yanping Xian, Yuluan Wu, Hao Dong, Xindong Guo, Bin Wang, Li Wang

    The present work presents a novel and rapid analytical method for the simultaneous analysis of bisphenol A (BPA), bisphenol B (BPB), bisphenol F (BPF) and bisphenol S (BPS) in edible oil based on dispersive micro solid phase extraction (DMSPE) for the first time followed by isotope dilution-ultra high performance liquid chromatography tandem mass spectrometry (UPLC–MS/MS). The edible oil sample was dispersed by n-hexane and extracted with ammoniated methanol-water solution. Then the target analytes were dispersedly absorbed using the polymer anion exchange (PAX) as the sorbent and eluted by acidic methanol. After that, four bisphenols were separated on a C18 column by gradient elution with methanol and 0.05% ammonium hydroxide in water as mobile phase, detected by MS/MS under multiple reactions monitoring (MRM) mode and quantified by internal standard method. The PAX amounts, adsorption time, concentrations of formic acid in the elution solvent and volume of elution solvent for the DMSPE technique were optimized. The limit of detection and quantitation (LOD and LOQ), matrix effect, recovery and precision of the developed method were investigated. Results indicated that BPS and the rest three bisphenols displayed excellent linearity in the concentration ranges of 0.1–50 μg/L and 0.5–250 μg/L, respectively, with correlation coefficients (R2) all larger than 0.998. Achieved MLODs (S/N = 3) varied between 0.1–0.4 μg/kg for all bisphenols. The mean recoveries at three spiked levels in edible oil were in the range of 87.3–108%. Intra-day precision (n = 6) and inter-day precision (n = 5) were <9% and <11%, respectively. This method is of rapid-and-simple pretreatment, accurate and sensitive, and suitable for the simultaneous determination of bisphenols in edible oil.

    更新日期:2017-08-29
  • Application of dispersive solid phase extraction based on a surfactant-coated titanium-based nanomagnetic sorbent for preconcentration of bisphenol A in water samples
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Hamid Reza Sobhi, Mahnaz Ghambarian, Mohammad Behbahani, Ali Esrafili

    Herein, a new extraction method employing a surfactant-coated titanium-based nanomagnetic sorbent for the effective extraction of bisphenol A (BPA) from various water samples was developed. Initially, the titanium-based nanomagnetic particles (Fe3O4/SiO2/TiO2 NPs) were successfully synthesized and subsequently characterized by Transmission Electron Microscopy and Fourier-transform infrared spectrometry. Two cationic surfactants were then incorporated into the particles to form a new sorbent for enhancing the extraction of BPA through micelle formation. Once the analyte was extracted, it was desorbed from the sorbent and quantified by high performance liquid chromatography with ultra violet detection (HPLC-UV). Various factors affecting the extraction and desorption of the analyte were investigated in detail and the optimum conditions established. Under these established conditions, the calibration curve was linear over the concentration range of 1–500 ng/mL. The limit of detection was determined to be 0.5 ng/mL based on a signal-to-noise ratio (S/N) = 3. To test the extraction efficiency, the method was applied to various real water samples that were spiked. The average recoveries obtained from the spiked samples ranged between 92 − 105% with relative standard deviations of 3.2 − 7.8%. Finally, the approach was determined to be effective for BPA environmental analysis.

    更新日期:2017-08-28
  • Nano reversed phase versus nano hydrophilic interaction liquid chromatography on a chip in the analysis of hemopexin glycopeptides
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Petr Kozlik, Miloslav Sanda, Radoslav Goldman

    Analysis of the glycosylation of proteins is a challenge that requires orthogonal methods to achieve separation of the diverse glycoforms. A combination of reversed phase chromatography with tandem mass spectrometry (RP-LC-MS/MS) is one of the most powerful tools for glycopeptide analysis. In this work, we developed and compared RP-LC and hydrophilic interaction liquid chromatography (HILIC) in nanoscale on a chip combined with MS/MS in order to separate glycoforms of two peptides obtained from the tryptic digest of hemopexin. We observed reduction of the retention time with decreasing polarity of glycans attached to the same peptide backbone in HILIC. The opposite effect was observed for RP-LC. The presence of sialic acids prolonged the retention of glycopeptides in both chromatographic modes. The nanoHILIC method provided higher selectivity based on the composition of glycan, compared to nanoRP-LC but a lower sensitivity. The nanoHILIC method was able to partially separate linkage isomers of fucose (core and outer arm) on bi-antennary glycoform of SWPAVGDCSSALR glycopeptide, which is beneficial in the elucidation of the structure of the fucosylated glycoforms.

    更新日期:2017-08-28
  • Accurate Prediction of Retention in Hydrophilic Interaction Chromatography (HILIC) by Back Calculation of High Pressure Liquid Chromatography (HPLC) Gradient Profiles
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Nu Wang, Paul G. Boswell

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

    更新日期:2017-08-28
  • Active modulation in neat carbon dioxide packed column comprehensive two-dimensional supercritical fluid chromatography
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Orjen Petkovic, Pierre Guibal, Patrick Sassiat, Jérôme Vial, Didier Thiébaut

    After demonstrating in a first paper the feasibility of SFCxSFC without decompression of the mobile phase, a modified interface has been developed in order to perform active modulation between the two SFC dimensions. In this paper, it is shown that the new interface enabled independent control of modulation parameters in SFCxSFC and performed a band compression effect of solutes between the two SFC dimensions. The effectiveness of this new modulation process was studied using a Design of Experiments. The SFCxSFC prototype was applied to the analysis of a real oil sample to demonstrate the benefits of the active modulator; in comparison to our previous results obtained without active modulation, better separation was obtained with the new interface owing to the peak compression occurring in the modulator.

    更新日期:2017-08-28
  • Development and comprehensive comparison of two on-line capillary electrophoretic methods for β-secretase inhibitor screening ☆
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Roman Řemínek, Lucie Slezáčková, Jan Schejbal, Zdeněk Glatz

    Alzheimer’s disease is the most common cause of dementia, afflicting over 34 million patients worldwide. Since β-secretase is a rate-limiting enzyme of the production of neurotoxic β-amyloid peptide oligomers abnormally accumulated in the affected brain tissue, its specific inhibition appears to be a promising approach to slowing down or even stopping the progression of the disease. Hence two on-line capillary electrophoretic methods for studies of β-secretase activity based on the principles of transverse diffusion of laminar flow profiles and electrophoretically mediated microanalysis were developed, both using a simple unlabeled peptide substrate and UV detection. The optimized procedures were thoroughly validated and applied for determining the enzyme’s kinetic parameters and the inhibition characteristics of two potent probe inhibitors. The resulting values were found to be comparable to literature data obtained with other analytical techniques. The suitability of the employed methodologies for different experimental designs is discussed on the basis of a statistical evaluation of the experimental data. The presented methods constitute a miniaturized and fully automated tool, which should be suitable for kinetic and inhibition studies of β-secretase as a target for Alzheimer’s disease drug discovery in the early stages of the development of a new drug.

    更新日期:2017-08-28
  • Determination of 3-hydroxypropylmercapturic acid in urine by three column-switching high-performance liquid chromatography with electrochemical detection using a diamond electrode
    J. Chromatogr. A (IF 3.981) Pub Date : 2017-08-26
    Kyohei Higashi, Mana Shibasaki, Kyoshiro Kuni, Takeshi Uemura, Masaaki Waragai, Kenichi Uemura, Kazuei Igarashi, Toshihiko Toida

    A three column-switching high-performance liquid chromatography (HPLC) using an electrochemical detector (ECD) equipped with a diamond electrode was established to determine 3-hydroxypropylmercapturic acid (3-HPMA) in urine. An extracted urine sample was consecutively fractionated using a strong anion-exchange column (first column) and a C8 column (second column) via a switching valve before application on an Octa Decyl Silyl (ODS) column (third column), followed by ECD analysis. The% recovery of 3-HPMA standard throughout the three-column process and limit of detection (LOD) were 94 ± 1% and 0.1 pmol, respectively. A solid phase extraction step is required for the sensitive analysis of 3-HPMA in urine by column-switching HPLC-ECD despite a decreased% recovery (55%) of urine sample spiked with 100 pmol of 3-HPMA. To test the utility of our column-switching HPLC-ECD method, 3-HPMA levels of 27 urine samples were determined, and the correlation between HPLC-ECD and LC-Electrospray ionization (ESI)-MS/MS method was examined. As a result, the median values of μmol 3-HPMA/g Creatinine (Cre) in urine obtained by column-switching HPLC-ECD and LC–MS/MS were 2.19 ± 2.12 μmol/g Cre and 2.13 ± 3.38 μmol/g Cre, respectively, and the calibration curve (y = 1.5171x − 1.007) exhibited good linearity within a defined range (r2 = 0.907). These results indicate that the combination of column-switching HPLC and ECD is a powerful tool for the specific, reliable detection of 3-HPMA in urine.

    更新日期:2017-08-28
Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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