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Hydrophilic Interaction Chromatography (hydrophilic + interaction_chromatography)
Selected AbstractsSynthesis of Sugar-Based Silica Gels by Copper-Catalysed Azide,Alkyne Cycloaddition via a Single-Step Azido-Activated Silica Intermediate and the Use of the Gels in Hydrophilic Interaction ChromatographyCHEMISTRY - A EUROPEAN JOURNAL, Issue 19 2010Lisa Moni Dr. Abstract Novel sugar-based silica gels were prepared by exploiting the copper-catalysed azide,alkyne cycloaddition (CuAAC) of two different sugar alkynes, namely, ethynyl C -galactoside 1 and propargyl O -lactoside 2, with new single-step azido-activated silica gels. The fully characterised stationary phases were generally used for hydrophilic interaction chromatography (HILIC), with particular application in the stereoselective separation of monosaccharides. Dynamic HILIC (DHILIC) experiments were performed to evaluate the influence of mutarotation on the chromatographic peak shapes of two interconverting sugar anomers. The potential of such materials was shown in the separation of other highly polar compounds, including amino acids and flavonoids. [source] Hydrophilic interaction and reversed-phase ultra-performance liquid chromatography TOF-MS for metabonomic analysis of Zucker rat urineJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2008Helen G. Gika Abstract Hydrophilic interaction chromatography (HILIC) provides a complementary technique to RP methods for the retention of polar analytes for LC-MS-based metabonomic studies. Combining the advantages of both RP and HILIC separations with the efficient and rapid separations obtained using sub-2 ,m particles via the recently introduced ultra-performance LC (UPLC) enables increased coverage of the metabolites present in biological samples to be achieved. Here an HILIC-UPLC-MS method was developed to provide metabolite profiles for urine samples obtained from male Zucker rats. The resulting data were compared with results obtained for the same samples by RP-UPLC-MS and demonstrated the complementary nature of the two separations with both methods enabling discrimination between the different sample types. Interestingly sample type differentiation was based on different markers. [source] Towards a platform for the metabonomic profiling of different strains of Drosophila melanogaster using liquid chromatography,Fourier transform mass spectrometryFEBS JOURNAL, Issue 22 2009Muhammad A. Kamleh A platform based on hydrophilic interaction chromatography in combination with Fourier transform mass spectrometry was developed in order to carry out metabonomics of Drosophila melanogaster strains. The method was able to detect , 230 metabolites, mainly in the positive ion mode, after checking to eliminate false positives caused by isotope peaks, adducts and fragment ions. Two wild-type strains, Canton S and Oregon R, were studied, plus two mutant strains, Maroon Like and Chocolate. In order to observe the differential expression of metabolites, liquid chromatography-mass spectrometry analyses of the different strains were compared using sieve 1.2 software to extract metabolic differences. The output from sieve was searched against a metabolite database using an Excel-based macro written in-house. Metabolic differences were observed between the wild-type strains, and also between both Chocolate and Maroon Like compared with Oregon R. It was established that a metabonomic approach could produce results leading to the generation of new hypotheses. In addition, the structure of a new class of lipid with a histidine head group, found in all of the strains of flies, but lower in Maroon Like, was elucidated. [source] Hydrophilic interaction LC of peptides: Columns comparison and clusteringJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Sylvia Van Dorpe Abstract A wide variety of hydrophilic interaction chromatography (HILIC) stationary phase surface chemistries are currently available. Although their selectivity can be considerably different, column comparison or clustering using peptides is limited. In this study, ten pharmaceutically relevant model peptides are analyzed on seven different HILIC columns (bare silica, amide, poly-hydroxyethyl aspartamide, diol and zwitterionic) for the evaluation of their performance and classification. The responses examined include single and multiple responses: plate number, asymmetry factor, LOD, geometric mean resolution, resolution product, time corrected resolution product, peak capacity and chromatographic response function. Column classification was performed using hierarchical clustering and principal component analysis. Moreover, the overall performance quality of the HILIC columns was compared using a linear desirability function. Hierarchical clustering and principal component analysis showed consistent clusters. The zwitterionic phase was clustered apart from the other HILIC columns and both poly-aspartamide columns were clustered together. In addition, the two bare silica phases represent two different clusters, and thus different selectivities. Overall, the responses showed the best performance for one of the bare silica columns (Alltima-Alltech), followed by the zwitterionic phase (ZIC)-HILIC. Thus, these columns, belonging to different clusters, were found to be the best performing systems in pharmaceutical peptide analysis for the selected peptide set. [source] Influence of stationary phase chemistry and mobile-phase composition on retention, selectivity, and MS response in hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Kenneth J. Fountain Abstract A comprehensive retention and selectivity characterization of several hydrophilic interaction chromatography (HILIC) stationary phases was performed with 28 test probes in order to study the influence of particle type, surface chemistry, and mobile-phase pH on chromatographic retention, selectivity, and MS response. Selectivity differences were compared for columns operated at both low and high pH, while ESI-MS was used to study the effects of mobile-phase pH on signal response. Additionally, acetone was explored as a potential alternative to ACN as the weak HILIC solvent. Moderate differences in selectivity were observed on the same column operated at different pH, mostly due to acidic compounds. In addition, the MS response increased when a high pH mobile phase was used, particularly for analytes that were ionized with negative ESI-MS. Even larger selectivity differences were observed for different stationary phases evaluated with the same mobile phase. Acetone was not a suitable replacement for ACN in routine HILIC separations due to differences in selectivity and MS response. Finally, the data from this study were used to establish guidelines for rapid HILIC method development of polar compounds, which is demonstrated with a mixture of histidine dipeptides and organophosphonate nerve agent metabolites. [source] Retention of arsenic species on zwitterionic stationary phase in hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Dan Xie Abstract Zwitterionic hydrophilic interaction chromatography (ZIC® -HILIC) was used to study the retention of selected organoarsenicals. The retention behavior of nine organic arsenic species on ZIC® -HILIC was investigated to elucidate which is the driving force for their separation, hydrophilic partitioning or adsorption driven by hydrogen bonds with surface H-donor/acceptor groups of the stationary phase. For this, the retention factor of the compounds k was correlated with log PO/W and with the calculated strength of hydrogen bonding of the analytes. By examining aliphatic and phenylic compounds separately, improved correlation was received. This indicates that both phenomena contribute to the separation of these arsenic species on ZIC® -HILIC. The results obtained evidence that considerable electrostatic interactions also occur on ZIC® -HILIC. Retention behavior of arsenic species was investigated by varying the separation conditions, which shows that the composition of the eluent has a strong influence on the retention behavior. It is highly dependent on water/acetonitrile ratio, pH value and salt additives. Dissociation degree and polarity of arsenic species, which are varying with pH, regulate the distribution of arsenic species between stationary and mobile phases in HILIC. Increase in the ammonium acetate concentration leads to shortened or to prolonged retention depending on the structure of the arsenic species. [source] Characterization of sialylated and fucosylated glycopeptides of ,2-glycoprotein I by a combination of HILIC LC and MALDI MS/MSJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Akira Kondo Abstract Characterization of low microgram levels of glycoprotein remains a challenge due to extensive heterogeneity of the conjugated N -glycans at each individual glycosylation site. We present an optimized, sensitive workflow for glycopeptide isolation and characterization that exploits the complementary features of RP (Poros R2) and hydrophilic (zwitter-ionic hydrophilic interaction chromatography) chromatographic resins. The glycopeptide analysis workflow was applied to human ,2-glycoprotein I (,2-GPI, apolipoprotein H), which contains multiple N -glycosylation sites. Conditions for rapid proteolytic digestion of ,2-GPI using low-specificity proteases were optimized to detect ,2-GPI glycopeptides by MS. We demonstrate the importance of ensuring sufficient column capacity of both hydrophobic and hydrophilic stationary phases for optimal glycoprofiling by MS. The enriched glycopeptides were characterized using MALDI quadrupole TOF MS/MS. A total of 23 glycan structures, including sialylated bi- and tri-antennary complex type glycans, were characterized at three N -glycosylation sites, namely Asn-143, Asn-174 and Asn-234, of ,2-GPI. Further exploration of the complementary nature of RP and HILIC stationary phases for glycopeptide isolation prior to MS analysis may eventually enable systematic analysis of complex glycoprotein samples in functional proteomic research and advance our understanding of the biological role of protein glycosylation. [source] Monitoring of mutarotation of monosaccharides by hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010ķ Pazourek Abstract Calibration based on the "single-point calibration method", a simple exponential transformation of the response function of an evaporative light scattering detector was improved and applied to analysis of selected saccharides under hydrophilic interaction chromatography mode (a polar phase LiChrospher100 DIOL, mobile phase acetonitrile/water). The improved approach to the calibration procedure yielded a calibration curve with an excellent linearity (quality coefficient <5%). This quantitative evaluation of chromatograms of D -galactose suggested that not only anomers but even pyranose and furanose forms of the anomers could be resolved , the resulting calculations of abundance of the anomeric form strongly correlated with data from the literature obtained mostly by NMR studies (analogous results were also obtained for D -arabinose, D -glucose, and D -mannose). Because of the rapid separation (retention time less than 10,min), the observed correlation enabled to monitor anomeric conversion (mutarotation) of monosaccharides. [source] Lactic acid quantitation in hand dishwashing liquid using an HILIC-UV methodologyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Mark Storton Abstract Different hydrophilic interaction chromatography (HILIC) columns were screened for lactic acid separation in hand dishwashing liquid products and the influence of mobile phase strength, buffer concentration and column temperature on the retention of lactic acid on a Zorbax NH2 column was investigated. An isocratic HILIC method for the quantitation of lactic acid in hand dishwashing liquid products was developed. The mobile phase consists of 70% methanol and 30% 20,mM sodium phosphate buffer (v/v) at pH 2.5. The HILIC stationary phase is Zorbax NH2, 250×4.6 with a 5,,m particle size. Detection was carried out using a variable wavelength UV-VIS detector at 226,nm. The linear range and percent recovery for lactic acid in the products were 44.68,1206.39,,g/mL and 100.3%, respectively. This paper provides an optimized HILIC methodology for the analysis of an acidic polar analyte (lactic acid) on a basic stationary phase. The proposed method can be used for the routine analysis of lactic acid. [source] Rapid analysis of constituents of Radix Cyathulae using hydrophilic interaction-reverse phase LC-MSJOURNAL OF SEPARATION SCIENCE, JSS, Issue 22 2009Mei-Ting Ren Abstract A hydrophilic interaction chromatography (HILIC) and reverse-phase liquid chromatography (RPLC) coupled with electrospray TOF MS method was developed for the analysis and characterization of constituents in the radix of Cyathula officinalis Kuan. Separation parameters of HILIC such as buffer pH, mobile phase strength, and organic modifier were evaluated. Fructose, glucose, and sucrose were identified by HILIC-ESI/TOF MS. Reverse-phase liquid chromatography-ESI/TOF MS were applied for quick and sensitive identification of major saponins in Cyathula officinalis. In-source collision-induced dissociation has been performed to elucidate the fragmentation pathways of oleanane-, hederagenin-, and gypsogmin-type saponins. Twelve saponins were characterized in this plant for the first time, and four of them were presumed to be new compounds. In addition, one phytoecdysteroid (cyasterone) and one coumarin (6,7-dimethoxycoumarin) were detected at the same time. The present method was capable of rapid characterizing and providing structure information of constituents from herbal drugs. [source] Profiling of polar metabolites in biological extracts using diamond hydride-based aqueous normal phase chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 13 2009Damien L. Callahan Abstract Highly polar metabolites, such as sugars and most amino acids are not retained by conventional RP LC columns. Without sufficient retention low concentration compounds are not detected due ion suppression and structural isomers are not resolved. In contrast, hydrophilic interaction chromatography (HILIC) and aqueous normal phase chromatography (ANP) retain compounds based on their hydrophilicity and therefore provides a means of separating highly polar compounds. Here, an ANP method based on the diamond hydride stationary phase is presented for profiling biological small molecules by LC. A rapid separation system based upon a fast gradient that delivers reproducible chromatography is presented. Approximately 1000 compounds were reproducibly detected in human urine samples and clear differences between these samples were identified. This chromatography was also applied to xylem fluid from soyabean (Glycine max) plants to which 400 compounds were detected. This method greatly increases the metabolite coverage over RP-only metabolite profiling in biological samples. We show that both forms of chromatography are necessary for untargeted comprehensive metabolite profiling and that the diamond hydride stationary phase provides a good option for polar metabolite analysis. [source] Mixed-mode ion-exchangers and their comparative chromatographic characterization in reversed-phase and hydrophilic interaction chromatography elution modesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 14 2008Michael Lämmerhofer Abstract A set of particulate silica-supported mixed-mode RP/weak anion-exchangers (RP/WAX) (obtained by bonding of N -undecenoylated 3-aminoquinuclidine, 3-aminotropane and 2-dimethylaminoethylamine as well as of N -butenoyl-(2S,4S,5R)-2-aminomethyl-5-[(2-octylthio)ethyl]-quinuclidine to thiol-modified silica) were chromatographically characterized in comparison to selected commercially available columns using two distinct isocratic elution modes, viz. an aqueous-rich RP-type elution mode (with 40% ACN and 60% buffer) as well as an organic solvent-rich hydrophilic interaction chromatography (HILIC)-type elution mode (95 and 90% ACN). The mixed-mode RP/WAX phases showed multimodal applicability, unlike a polar embedded RP material (Synergi Fusion RP), amino phases (Luna NH2, BioBasic AX) or typical HILIC packings (ZIC-HILIC, TSKGel Amide-80). Principal component analysis (PCA) of the RP test data confirmed that the in-house developed RP/WAX columns as well as the Acclaim Mixed-Mode WAX-1 phase resemble each other in their chromatographic characteristics having slightly lower hydrophobic selectivity (,CH2 of 1.5) than the tested Synergi Fusion RP (,CH2 ,1.8). In contrast, a decrease in mixed-mode character due to lowered ion-exchange capacity and concomitantly increased RP-like behavior could be identified for other mixed-mode phases in the order of Obelisc R > Primesep B2 > Uptisphere MM3. PCA on HILIC data revealed that the RP/WAX phases behave dissimilar to TSKGel Amide-80, ZIC-HILIC and polysulfoethyl A under the chosen elution conditions. Hence, they may be regarded as complementary to these commercial stationary phases with applicability profiles for hydrophilic but also hydrophobic solutes. [source] Evaluation of mobile phase, ion pairing, and temperature influence on an HILIC-MS/MS method for L -arginine and its dimethylated derivatives detectionJOURNAL OF SEPARATION SCIENCE, JSS, Issue 13 2008Giuseppe Paglia Abstract Asymmetric NG,,NG -dimethylarginine (ADMA) increases in diseases such as renal failure, diabetes mellitus, and hypercholesterolemia. The feasibility and utility of a hydrophilic interaction chromatography (HILIC) method for the separation of free L -arginine (Arg), ADMA, and symmetric NG,,NG, -dimethylarginine (SDMA) on a typical silica column were explored and the impact of some experimental parameters on the chromatographic behavior of these analytes was investigated. The effect of water and TFA content in mobile phase and of column temperature was investigated during the development of a fast and simple HILIC-MS/MS method that might be suitable for the quantification of free Arg, ADMA, and SDMA in plasma for routine analysis. Our results show that a good compromise between efficiency and peak shape with acceptable retention and total chromatographic run time is achieved using an ACN/water (90:10) mobile phase with TFA% as additive ranging from 0.015 to 0.025% and column temperature ranging from 25 to 30°C. [source] Preparation of a sorbitol methacrylate grafted silica as stationary phase for hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2008Jonas Persson Abstract A new highly hydrophilic stationary phase based on graft polymerization of sorbitol methacrylate from the surface of Kromasil silica particles is described. Polymerization was initiated by thermal cleavage of tert -butyl hydroperoxide covalently attached to the silica particle surface. Due to the highly amphiphilic properties of the monomer, an extensive search was needed to find solvent conditions that enabled surface-initiated polymerization. This was finally solved by using a mixture of solvents that only partially dissolved the monomer. The graft polymerization was confirmed by Fourier transform infrared spectroscopy and elemental analysis. The resulting stationary phase was evaluated by HPLC and exhibited a selectivity markedly different from that of commercially available columns and of neat silica. [source] The application of novel 1.7 ,m ethylene bridged hybrid particles for hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2008Eric S. Grumbach Abstract An un-derivatized 1.7 ,m ethylene bridged hybrid (BEH) particle was evaluated for its utility in retaining polar species in hydrophilic interaction chromatography (HILIC), and was compared to a 3 ,m un-derivatized silica material. Retentivity as a function of mobile phase pH, polar modifier and ACN content was examined. Also, the efficiency of the two particle substrates was compared by plotting HETP vs. linear velocity. Improved chemical resistance of the un-derivatized BEH particle was compared to un-derivatized silica at pH 5, demonstrating no performance deterioration over the course of 2000 injections for the BEH particle, while the silica particle deteriorated rapidly after 800 injections. Lastly, ESI-MS sensitivity as a function of particle size and separation mode was demonstrated. A 2.2 to 4.7-times higher ESI-MS response was observed on the 1.7 ,m particle compared to the 3 ,m particle, whereas a 5.6 to 8.8-times higher ESI-MS response was observed using HILIC as when compared to traditional RP chromatography. [source] Stability-indicating assay of sodium cromoglicate in ophthalmic solution using mixed-mode hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2008Mohammed Shahid Ali Abstract A hydrophilic interaction chromatographic (HILIC) procedure for the quantification of Sodium Cromoglicate (SCG) in ophthalmic solution is developed. Mobile phase consists of ACN and buffer, 86:14 v/v. Atlantis HILIC,Si column, 25 cm×4.6 mm, is used as stationary phase. Detection is carried out using a variable wavelength UV-Vis detector at 326 nm. Linearity range and percent recoveries for SCG were 50,400 ,g/mL and 100.44%, respectively. The SCG HILIC-UV assay was validated according to the International Conference on Harmonization guidelines. The method separates two impurities and degradation products resulting from stress environment. Influence of organic solvent, ionic strength and mobile phase pH on the retention of SCG is studied. The paper provides optimization of polar anionic solute (SCG) on unmodified silica by HILIC. Proposed method can be used as a stability-indicating assay for SGC and can be proved to be beneficial in ESI-MS for enhanced sensitivity. [source] Probing genetic algorithms for feature selection in comprehensive metabolic profiling approachRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 8 2008Wei Zou Six different clones of 1-year-old loblolly pine (Pinus taeda L.) seedlings grown under standardized conditions in a green house were used for sample preparation and further analysis. Three independent and complementary analytical techniques for metabolic profiling were applied in the present study: hydrophilic interaction chromatography (HILIC-LC/ESI-MS), reversed-phase liquid chromatography (RP-LC/ESI-MS), and gas chromatography all coupled to mass spectrometry (GC/TOF-MS). Unsupervised methods, such as principle component analysis (PCA) and clustering, and supervised methods, such as classification, were used for data mining. Genetic algorithms (GA), a multivariate approach, was probed for selection of the smallest subsets of potentially discriminative classifiers. From more than 2000 peaks found in total, small subsets were selected by GA as highly potential classifiers allowing discrimination among six investigated genotypes. Annotated GC/TOF-MS data allowed the generation of a small subset of identified metabolites. LC/ESI-MS data and small subsets require further annotation. The present study demonstrated that combination of comprehensive metabolic profiling and advanced data mining techniques provides a powerful metabolomic approach for biomarker discovery among small molecules. Utilizing GA for feature selection allowed the generation of small subsets of potent classifiers. Copyright © 2008 John Wiley & Sons, Ltd. [source] On-line desalting and determination of morphine, morphine-3-glucuronide and morphine-6-glucuronide in microdialysis and plasma samples using column switching and liquid chromatography/tandem mass spectrometryRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 15 2005Jörgen Bengtsson A sensitive and reproducible method for the determination of morphine and the metabolites morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) was developed. The method was validated for perfusion fluid used in microdialysis as well as for sheep and human plasma. A C18 guard column was used to desalt the samples before analytical separation on a ZIC HILIC (hydrophilic interaction chromatography) column and detection with tandem mass spectrometry (MS/MS). The mobile phases were 0.05% trifluoroacetic acid (TFA) for desalting and acetonitrile/5,mM ammonium acetate (70:30) for separation. Microdialysis samples (5,µL) were directly injected onto the system. The lower limits of quantification (LLOQ) for morphine, M3G and M6G were 0.50, 0.22 and 0.55,ng/mL, respectively, and the method was linear from LLOQ to 200,ng/mL. For plasma, a volume of 100,µL was precipitated with acetonitrile containing internal standards (deuterated morphine and metabolites). The supernatant was evaporated and reconstituted in 0.05% TFA before the desalting process. The LLOQs for sheep plasma were 2.0 and 3.1,ng/mL and the ranges were 2.0,2000 and 3.1,3100,ng/mL for morphine and M3G, respectively. For human plasma, the LLOQs were 0.78, 1.49 and 0.53,ng/mL and the ranges were 0.78,500, 1.49,1000 and 0.53,500,ng/mL for morphine, M3G and M6G, respectively. Copyright © 2005 John Wiley & Sons, Ltd. [source] Simple and rapid determination of 1-deoxynojirimycin in mulberry leavesBIOFACTORS, Issue 1-4 2004Toshiyuki Kimura Abstract A simple and rapid method for determining 1-deoxynojirimycin (DNJ), a potent glucosidase inihibitor present in mulberry leaves (Morus alba and Morus bombysis), by high performance liquid chromatography coupled to an evaporative light scattering detector (ELSD) has been developed. DNJ was separated from extract of mulberry leaves on TSK gel Amide-80 column, which is a representative column for hydrophilic interaction chromatography. During post column detection, DNJ was detected by ELSD and concurrently identified by mass spectrometry. The detection limit was 100 ng. This method is sufficiently sensitive for determining DNJ in mulberry leaves and other related products. [source] Synthesis of Sugar-Based Silica Gels by Copper-Catalysed Azide,Alkyne Cycloaddition via a Single-Step Azido-Activated Silica Intermediate and the Use of the Gels in Hydrophilic Interaction ChromatographyCHEMISTRY - A EUROPEAN JOURNAL, Issue 19 2010Lisa Moni Dr. Abstract Novel sugar-based silica gels were prepared by exploiting the copper-catalysed azide,alkyne cycloaddition (CuAAC) of two different sugar alkynes, namely, ethynyl C -galactoside 1 and propargyl O -lactoside 2, with new single-step azido-activated silica gels. The fully characterised stationary phases were generally used for hydrophilic interaction chromatography (HILIC), with particular application in the stereoselective separation of monosaccharides. Dynamic HILIC (DHILIC) experiments were performed to evaluate the influence of mutarotation on the chromatographic peak shapes of two interconverting sugar anomers. The potential of such materials was shown in the separation of other highly polar compounds, including amino acids and flavonoids. [source] | ||||||||||