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ESI-MS
Terms modified by ESI-MS Selected AbstractsImproved sample preparation method for glycan analysis of glycoproteins by CE-LIF and CE-MSELECTROPHORESIS, Issue 8 2010Zoltan Szabo Abstract CE is a high-resolution separation technique broadly used in the biotechnology industry for carbohydrate analysis. The standard sample preparation protocol for CE analysis of glycans released from glycoproteins generally requires derivatization times of overnight at 37°C, using ,100 fold excess of fluorophore reagent, 8-aminopyrene-1,3,6-trisulfonic-acid, if the sample is unknown, or it is a regulated biotherapeutic product, possibly containing terminal sialic acid(s). In this paper, we report on significant improvements for the standard CE sample preparation method of glycan analysis. By replacing the conventionally used acetic acid catalyst with citric acid, as low as 1:10 glycan to fluorophore molar ratio (versus the typical 1:,100 ratio) maintained the >95% derivatization yield at 55°C with only 50,min reaction time. Terminal sialic acid loss was negligible at 55°C during the derivatization process, and indicating that the kinetics of labeling at 55°C was faster than the loss of sialic acid from the glycan. The reduced relative level of 8-aminopyrene-1,3,6-trisulfonic-acid simplified the removal of excess reagent, important in both CE-LIF (electrokinetic injection bias) and CE-MS (ion suppression). Coupling CE- ESI-MS confirmed that the individual peaks separated by CE corresponded to single glycans and increased the confidence of structural assignment based on glucose unit values. [source] Electrokinetic supercharging-electrospray ionisation-mass spectrometry for separation and on-line preconcentration of hypolipidaemic drugs in water samplesELECTROPHORESIS, Issue 7 2010Mohamed Dawod Abstract Electrokinetic supercharging, a powerful on-line preconcentration technique in CE, was for the first time hyphenated with ESI-MS for the on-line concentration and separation of five hypolipidaemic drugs. The electrophoretic separation was performed in a co-EOF mode using the EOF reversal agent, hexadimethrine bromide, in ammonium bicarbonate electrolyte, pH 9.00. The ionic strength and the amount of methanol in the buffer were optimised in a multivariate manner using artificial neural networks, with the optimal conditions being 60,mM ammonium bicarbonate containing 60% methanol, providing baseline resolution of the five hypolipidaemics within 20,min. Using electrokinetic supercharging, the sensitivity of the method was improved 1000-fold over a conventional injection under field-amplified sample stacking conditions with LODs of 180,ng/L. This is the first report of the separation of hypolipidaemics by CE. The developed method was validated and then applied to the determination of the target drugs in water samples from Hobart city. [source] Online CIEF-ESI-MS in glycerol,water media with a view to hydrophobic protein applicationsELECTROPHORESIS, Issue 23 2009Meriem Mokaddem Abstract A new online coupling of CIEF with ESI-MS has been developed in glycerol,water media. This improved protocol provides: (i) the electric continuity during the whole analysis by a discontinuous filling of the capillary with 60:40 (cm/cm) catholyte/proteins,ampholyte mixture; (ii) the use of an anticonvective medium, i.e. 30:70 glycerol/water, v/v, compatible with MS detection and as an aid to hydrophobic protein solubilization and (iii) the use of unmodified bare fused-silica capillaries, as the glycerol/water medium strongly reduces EOF. Focusing was performed in positive polarity and cathodic mobilization was achieved by both voltage and pressure application. The setup was optimized with respect to analysis time, sensitivity and precision on pI determination. The optimized anolyte and catholyte were composed of 50,mM formic acid/1,mM glutamic acid (pH 2.35) and 100,mM NH3/1,mM lysine (pH 10.6), respectively. The effects of ampholyte concentration, focusing time and ESI parameters were presented for model proteins and discussed. This new integrated protocol should be an easy and effective additional tool in the field of proteome analysis, providing a means for the characterization of a large number of hydrophilic and hydrophobic proteins. [source] On-chip tryptic digest with direct coupling to ESI-MS using magnetic particlesELECTROPHORESIS, Issue 24 2008Anne Le Nel Abstract As a step toward a fully automated front-end microfluidic chip for MS proteomics, we propose a system capable of performing online tryptic digest and ESI-MS, using a replaceable on-chip digestion microcolumn based on self-assembled magnetic particles. [source] Separation of multiphosphorylated peptide isomers by CZEELECTROPHORESIS, Issue 21 2008Marika V. Muetzelburg Abstract A separation of mono, doubly and triply phosphorylated isomers was developed with CZE with an aqueous electrolyte containing 3.9,mol/L formic acid and 30%,v/v trifluoroethanol. Thus a mixture of ten phosphopeptides corresponding to the human tau sequence 226,240 was separated within 70,min. Although peptides with different phosphorylation degrees, i.e. 0,3 phosphate groups, were well separated, some of the phosphopeptide isomers containing one or two phosphate groups were only partially separated. The electrolyte system is compatible with both MALDI- and ESI-MS, allowing a direct coupling, and thus could have some interesting applications in proteomics. [source] Separation of cytokinin isomers with a partial filling-micellar electrokinetic chromatography-mass spectrometry approachELECTROPHORESIS, Issue 10 2008Liya Ge Abstract A new method based on partial filling-MEKC (PF-MEKC) directly coupled to ESI-MS was developed for the simultaneous separation and determination of 13 structurally similar cytokinins, including various geometric and positional isomers of cytokinins. On the basis of the resolution of the neighboring isomer peaks, different parameters (i.e., pH and concentration of buffer, surfactant concentrations, length of the injected micellar plug, organic modifier, and applied separation voltage) were optimized to achieve a satisfactory PF-MEKC separation. Under optimum conditions, the separation of 13 cytokinin standards was accomplished within 25,min. MS/MS with multiple reaction monitoring detection was carried out to obtain sufficient selectivity. PF-MEKC-MS/MS allowed for the direct identification and confirmation of the cytokinins present in banana (Musa spp.) pulp sample after extraction and purification. Finally, trans- zeatin riboside (ZR) and trans- zeatin (Z) were unambiguously identified in banana pulp. It is anticipated that the current PF-MEKC-MS method can be applied to analyze cytokinins in a wide range of biological samples. [source] Microfluidic technologies for MALDI-MS in proteomicsELECTROPHORESIS, Issue 18 2006Don L. DeVoe Professor Abstract The field of microfluidics continues to offer great promise as an enabling technology for advanced analytical tools. For biomolecular analysis, there is often a critical need to couple on-chip microfluidic sample manipulation with back-end MS. Though interfacing microfluidics to MS has been most often reported through the use of direct ESI-MS, there are compelling reasons for coupling microfluidics to MALDI-MS as an alternative to ESI-MS for both online and offline analysis. The intent of this review is to provide a summary of recent developments in the integration of microfluidic systems with MALDI-MS, with an emphasis on applications in proteomics. Key points are summarized, followed by a review of relevant technologies and a discussion of outlook for the field. [source] Application of polymeric surfactants in micellar electrokinetic chromatography-electrospray ionization mass spectrometry of benzodiazepines and benzoxazocine chiral drugsELECTROPHORESIS, Issue 5-6 2006Jingguo Hou Abstract Chiral micellar EKC (CMEKC) coupled to ESI-MS using polymeric surfactants as pseudostationary phases is investigated for simultaneous enantioseparation of two benzodiazepines, (±)-oxazepam ((±)-OXA) and (±)-lorazepam ((±)-LOR), and one benzoxazocine, (±)-nefopam ((±)-NEF). First, enantioselectivity and electrospray sensitivity of six chiral polymeric surfactants for all three chiral compounds are compared. Second, using poly(sodium N -undecenoyl- L -leucinate) as pseudostationary phase, the organic modifiers (methanol (MeOH), isopropanol, and ACN) are added into the running buffer to further improve chiral resolution (RS). Next, a CMEKC-ESI-MS method for the simultaneous enantioseparation of two benzodiazepines is further developed by using a dipeptide polymeric surfactant, poly(sodium N -undecenoxy carbonyl- L,L -leucyl-valinate) (poly- L,L -SUCLV). The CMEKC conditions including nebulizer pressure, capillary length, ammonium acetate concentration, pH, poly- L,L -SUCLV concentration, and capillary temperature were optimized to achieve maximum chiral RS and highest sensitivity of MS detection. The spray chamber parameters (drying gas temperature and drying gas flow rate) as well as sheath liquid conditions (MeOH content, pH, flow rate, and ionic strength) were found to significantly influence MS S/N of both (±)-OXA and (±)-LOR. Finally, a comparative study between simultaneous UV and MS detection showed high plate numbers, better chiral RS, and enhanced detectability with CMEKC-MS. However, speed of analysis was faster using CMEKC-UV. [source] Biomarker discovery in rat plasma for estrogen receptor-, actionELECTROPHORESIS, Issue 23 2005Tom G. Holt Dr. Abstract To support in vivo screening efforts for estrogen receptor (ER) subtype selective therapeutic agents, we initiated work to discover surrogate markers (biomarkers) in blood plasma that would change in response to ER subtype-specific action. We used a proteomic approach employing strong anion exchange chromatography (SAX), PAGE, and MS to identify potential plasma markers for selective ER-, action. The methodology was used to compare blood from vehicle-treated rats to blood from rats treated with either 17,-estradiol (an ER-,/ER-, agonist) or compound 1 (17,-ethynyl-[3,2-c]pyrazolo-19-nor-4-androstene-17,-ol, an ER-,-selective agonist). Blood samples were first fractionated by SAX to separate fractions containing dominant common plasma proteins from fractions enriched for less-abundant plasma proteins. 1-D PAGE analysis of fractions depleted of dominant plasma proteins revealed treatment-specific changes in protein profiles. Protein bands that changed reproducibly in response to ER-, action were excised from the gel, separated by capillary LC, and identified by microspray ESI-MS. Using this method, the plasma levels of two proteins, transthyretin and apolipoprotein E, were shown to decrease in response to ER-, agonism. The method lacked the sensitivity to identify the known, 1000-fold less-abundant, estrogenic marker prolactin (PRL). However, using a commercial RIA and immunoblots, we showed that PRL levels increase significantly in response to treatment with the ER-, selective agonist, compound 1. [source] A silica-based monolithic column in capillary HPLC and CEC coupled with ESI-MS or electrospray-atmospheric-pressure laser ionization-MSELECTROPHORESIS, Issue 21 2005Stefan Droste Abstract We describe the successful coupling of CEC and capillary HPLC with the recently developed atmospheric-pressure laser ionization (APLI) method. APLI is suitable for selectively and sensitively ionizing nonpolar aromatic compounds at ambient pressure for subsequent mass-selective detection. The polycyclic aromatic hydrocarbons used as analytes are first separated either by CEC on a silica-based monolithic column or by capillary HPLC. The eluent, along with a sheath flow, is volatilized by microelectrospray and then selectively ionized by excimer laser (KrF*) radiation via two-photon excitation. A QTOF-MS is used as mass-selective detector. This interface combination makes soft ionization of thermally labile nonpolar aromatic analytes possible. [source] Capillary electrophoresis-laser induced fluorescence-electrospray ionization-mass spectrometry: A case studyELECTROPHORESIS, Issue 7-8 2005Carolin Huhn Abstract The simultaneous hyphenation of capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection and electrospray ionization-mass spectrometry (ESI-MS) as a novel combined detection system for CE is presented. ,-Carbolines were chosen as model analytes with a forensic background. Nonaqueous CE as well as conventional CE with an aqueous buffer system are compared concerning efficiency and obtainable detection limits. The distance between the optical detection window and the sprayer tip was minimized by placing the optical cell directly in front of the electrospray interface. Similar separation efficiencies for both detection modes could thus be obtained. No significant peak-broadening induced by the MS interface was observed. The high fluorescence quantum yield and the high proton affinity of the model analytes investigated resulted in limits of detection in the fg (nmol/L) range for both detection methods. The analysis of confiscated ayahuasca samples and ethanolic plant extracts revealed complementary selectivities for LIF and MS detection. Thus, it is possible to improve peak identification of the solutes investigated by the use of these two detection principles. [source] On-line concentration of proteins in pressurized capillary electrochromatography coupled with electrospray ionization-mass spectrometryELECTROPHORESIS, Issue 7-8 2005Zhen Liang Abstract Pressurized capillary electrochromatography (pCEC) and electrospray ionization-mass spectrometry (ESI-MS) have been hyphenated for protein analysis. Taken cytochrome,c, lysozyme, and insulin as samples, the limits of detection (LODs) for absolute concentrations are 10,11,mol (signal-to-noise ratio S/N = 3) with relative standard deviations (RSDs) of retention time and peak area, respectively, of less than 1.7% and 4.8%. In order to improve the detection sensitivity, on-line concentration by field-enhanced sample-stacking effect and chromatographic zone-sharpening effect has been developed, and parameters affecting separation and detection, such as pH and electrolyte concentration in the mobile phase, separation voltage, as well as enrichment voltage and time, have been studied systematically. Under the optimized conditions, the LODs of the three proteins could be decreased up to 100-fold. In addition, the feasibility of such techniques has been further demonstrated by the analysis of modified insulins at a concentration of 20,,g/mL. [source] Characterization of glyco isoforms in plasmaderived human antithrombin by on-line capillary zone electrophoresis-electrospray ionization-quadrupole ion trap-mass spectrometry of the intact glycoproteinsELECTROPHORESIS, Issue 13 2004Uwe M. Demelbauer Abstract The carbohydrate structures of five isoforms of ,-AT and two isoforms of ,-AT were determined by applying capillary zone electrophoresis (CZE) on-line coupled to electrospray ionization-mass spectrometry (ESI-MS) using an ion-trap analyzer. For the AT preparations gained from a plasma pool at least semiquantitative information on the isoform-distributions could be gained. Unlike to the commonly used approaches starting from enzymatically treated glycoproteins, this approach deals with intact proteins. The high accuracy of the molecular mass determination obtained by the ion-trap analyzer allows one to calculate and ascertain the carbohydrate composition assuming no variations in the protein moiety of AT and to exclude or confirm the presence of the potential post-translational or other modifications. Therefore, the direct coupling of CZE with ESI-MS does not only represent a fast alternative technique to two-dimensional electrophoresis (2-DE) but serves as a method which provides structural information complementary to that gained from peptide mapping methods. [source] Development of capillary zone electrophoresis-electrospray ionization-mass spectrometry for the determination of lamotrigine in human plasmaELECTROPHORESIS, Issue 13 2004Jack Zheng Abstract A method of coupling capillary zone electrophoresis (CZE) with electrospray ionization-mass spectrometry (ESI-MS) detection has been developed for monitoring an antiepileptic drug, lamotrigine (LTG) in human plasma. The CZE-MS was developed in three stages: (i) CZE separation and ESI-MS detection of LTG and tyramine (TRM, internal standard) were simultaneously optimized by studying the influence of CZE background electrolyte (BGE) pH, BGE ionic strength, and nebulizer pressure of the MS sprayer; (ii) sheath liquid parameters, such as pH, ionic strength, organic modifier content, and flow rate of the sheath liquid, were systematically varied under optimum CZE-MS conditions developed in the first stage; (iii) MS sprayer chamber parameters (drying gas temperature and drying gas flow rate) were varied for the best MS detection of LTG. The developed assay was finally applied for the determination of LTG in plasma samples. The linear range of LTG in plasma sample assay was between 0.1,5.0 ,g/mL with a limit of detection as low as 0.05 ,g/mL and run time less than 6 min. Finally, the concentration-time profile of LTG in human plasma sample was found to correlate well when CZE-ESI-MS was compared to a more established method of high-performance liquid chromatography with ultraviolet detection. [source] Microfluidic device for capillary electrochromatography-mass spectrometryELECTROPHORESIS, Issue 21 2003Iulia M. Lazar Abstract A novel microfabricated device that integrates a monolithic polymeric separation channel, an injector, and an interface for electrospray ionization-mass spectrometry detection (ESI-MS) was devised. Microfluidic propulsion was accomplished using electrically driven fluid flows. The methacrylate-based monolithic separation medium was prepared by photopolymerization and had a positively derivatized surface to ensure electroosmotic flow (EOF) generation for separation of analytes in a capillary electrochromatography (CEC) format. The injector operation was optimized to perform under conditions of nonuniform EOF within the microfluidic channels. The ESI interface allowed hours of stable operation at the flow rates generated by the monolithic column. The dimensions of one processing line were sufficiently small to enable the integration of 4,8 channel multiplexed structures on a single substrate. Standard protein digests were utilized to evaluate the performance of this microfluidic chip. Low- or sub-fmol amounts were injected and detected with this arrangement. [source] High-efficiency peptide analysis on monolithic multimode capillary columns: Pressure-assisted capillary electrochromatography/capillary electrophoresis coupled to UV and electrospray ionization-mass spectrometryELECTROPHORESIS, Issue 21 2003Alexander R. Ivanov Abstract High-efficiency peptide analysis using multimode pressure-assisted capillary electrochromatography/capillary electrophoresis (pCEC/pCE) monolithic polymeric columns and the separation of model peptide mixtures and protein digests by isocratic and gradient elution under an applied electric field with UV and electrospray ionization-mass spectrometry (ESI-MS) detection is demonstrated. Capillary multipurpose columns were prepared in silanized fused-silica capillaries of 50, 75, and 100 ,m inner diameters by thermally induced in situ copolymerization of methacrylic monomers in the presence of n -propanol and formamide as porogens and azobisisobutyronitrile as initiator. N -Ethylbutylamine was used to modify the chromatographic surface of the monolith from neutral to cationic. Monolithic columns were termed as multipurpose or multimode columns because they showed mixed modes of separation mechanisms under different conditions. Anion-exchange separation ability in the liquid chromatography (LC) mode can be determined by the cationic chromatographic surface of the monolith. At acidic pH and high voltage across the column, the monolithic stationary phase provided conditions for predominantly capillary electrophoretic migration of peptides. At basic pH and electric field across the column, enhanced chromatographic retention of peptides on monolithic capillary column made CEC mechanisms of migration responsible for separation. The role of pressure, ionic strength, pH, and organic content of the mobile phase on chromatographic performance was investigated. High efficiencies (exceeding 300,000 plates/m) of the monolithic columns for peptide separations are shown using volatile and nonvolatile, acidic and basic buffers. Good reproducibility and robustness of isocratic and gradient elution pressure-assisted CEC/CE separations were achieved for both UV and ESI-MS detection. Manipulation of the electric field and gradient conditions allowed high-throughput analysis of complex peptide mixtures. A simple design of sheathless electrospray emitter provided effective and robust low dead volume interfacing of monolithic multimode columns with ESI-MS. Gradient elution pressure-assisted mixed-mode separation CE/CEC-ESI-MS mass fingerprinting and data-dependent pCE/pCEC-ESI-MS/MS analysis of a bovine serum albumin (BSA) tryptic digest in less than 5 min yielding high sequence coverage (73%) demonstrated the potential of the method. [source] 1,3,5-Triazapentadiene Nickel(II) Complexes Derived from a Ketoxime-Mediated Single-Pot Transformation of NitrilesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 16 2010Maximilian N. Kopylovich Abstract A series of cationic (2+) [Ni{HN=C(R)NHC(R)=NH}2](X)2 {R = 4-(Cl)C6H4 (1), 3-(NC)C6H4 (3), 4-(NC)C6H4 (4) and Me (7); X = Cl, (1, 3, 4) or MeCOO,·H2O (7)} and neutral [Ni{HN=C(R)NC(R)=NH}2](solvate) {R = 3-(Cl)-4-py (2), 3-py (5) and 4-py (6); solvate = MeOH and/or H2O; py = pyridyl} N,N -chelating bis(1,3,5-triazapentadiene/ato)nickel(II) [Ni(tap)2]2+/0 complexes has been easily generated by a ketoxime-mediated single-pot reaction of a nickel(II) salt [NiCl2·2H2O or Ni(MeCOO)2·4H2O] with 4-chlorobenzonitrile, isophthalonitrile, terephthalonitrile, acetonitrile, 2-chloro-4-cyanopyridine, 3-cyanopyridine or 4-cyanopyridine, respectively. The obtained compounds have been characterized by IR, 1H and 13C{1H} NMR spectroscopy, FAB-MS(+) or ESI-MS(+), elemental analyses and single-crystal X-ray diffraction [for 7 and solvated mono- {1a·(Me2CO)0.33·(MeOH)0.67} and bis-deprotonated (2b·2Me2CO, 4b·CHCl3, 5b·Me2CO and 6b·MeOH) products, formed upon recrystallization of 1, 2, 4, 5 and 6, respectively]. The crystal structures of all compounds bear similar monomeric Ni(tap)2 units with a nearly square-planar geometry. In addition, the structure of 7 features the formation of infinite 1D zig-zag water,acetate chains {[(H2O)2(MeCOO)2]2,}n, which multiply interact with the [Ni(tap)2]2+ cations to generate a 2D hydrogen-bonded supramolecular assembly. [source] Structural, Spectroscopic, and Proton-Coupled Electron-transfer Behavior of Pyrazolyl-3,5-bis(benzimidazole)-Bridged Homo- and Heterochiral RuIIRuII, OsIIOsII, and OsIIRuII 2,2,-Bipyridine ComplexesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 4 2010Sujoy Baitalik Abstract The homo- and heterobimetallic complexes [(bpy)2MII(H2pzbzim)M,II(bpy)2](ClO4)3·nH2O (1, 3, 5) and their corresponding deprotonated complexes [(bpy)2MII(pzbzim)M,II(bpy)2](ClO4)·nH2O (2, 4, 6) [where MII, M,II = Ru (1, 2) = Os (3, 4); MII = Os and M,II = Ru (3, 5); bpy = 2,2,-bipyridine; H3pzbzim = pyrazole-3,5-bis(benzimidazole)] were synthesized, separated to their heterochiral (a, ,,/,,) and homochiral (b, ,,/,,) diastereoisomers, and characterized by elemental analyses, ESI-MS, and 1H NMR spectroscopy. The X-ray structures of 1a, 3a, and 5a show the involvement of two pyridine rings of two bpy ligands in strong intramolecular nonbonded ,,, interaction. The occurrence of a C,H···, interaction between an aromatic C,H and the ,-cloud of a pyridine ring leads to strong electronic shielding of this proton (1H NMR). In all cases, the two diastereoisomers show practically no differences in their absorption spectra, redox potentials, and pK values. The large shifts in the E1/2 values to less positive potentials and substantial redshifts in the MLCT bands that occur on deprotonation of 1, 3, and 5 are energetically correlated. From the profiles of E1/2(1), (2) vs. pH over the pH range 1,12, the equilibrium constants and standard redox potentials for all the complex species in the metal oxidation states II·II, II·III, and III·III and the bridged ligand in the protonation states H2pzbzim,, Hpzbzim2,, and pzbzim3, have been evaluated. Using these values the bond dissociation free energies for the benzimidazole N,H bonds have been estimated. Spectroelectrochemical studies have been carried out for 1a, 3a, and 5a in the range 400,1100 nm. With stepwise oxidation of the metal centers replacement of MLCT bands by LMCT bands occur gradually with the observation of sharp isosbestic points. In the case of 1a, a band observed at ,max = 910 nm for the RuIIRuIII species has been ascribed to intervalence charge transfer (IVCT) transition. [source] Ferrocene Conjugates Containing Diarginine and Aspartic Acid: Salt Bridge Interactions in WaterEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 29-30 2009Anas Lataifeh Abstract The ferrocene peptide conjugates of diarginine (MeO-Fc-Arg-Arg-NH2) (1) and aspartic acid [Boc-Fca-Asp(OH)-OH] (2) were found to form a stable 1:1 associate in aqueous solution. The molecular recognition was achieved through a combination of multipoint hydrogen bonding (H-bonding) sites and a guanidinium-carboxylate ion pair. The associate stoichiometry was confirmed by using ESI-MS and NMR experiments; the NMR titration curve shows multiple equilibria with stepwise interconversion from 1:2,,,1:1 binding ratios, and the electrochemical behaviour of the ferrocenyl groups (Fc, Fca) confirm the formation of an ion pair. The CD spectra in the peptide region exhibit a characteristic absorption of a more ordered structure, while the ferrocene helical chirality remains intact. The solid-state IR measurements exclude the involvement of the amide backbone in the interaction.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009) [source] (4-Acyl-5-pyrazolonato)titanium Derivatives: Oligomerization, Hydrolysis, Voltammetry, and DFT StudyEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 17 2003Francesco Caruso Abstract Twenty 4-acyl-5-pyrazolonato (Q) titanium derivatives of varied nuclearity have been synthesized from Ti(OR)4 or TiCl4 and characterized with spectroscopic methods (IR, NMR, ESI-MS). While Ti,(,-diketonato) cleavage is not seen in isolated solids, Ti,O(alkoxy) (or Ti,Cl) bonds cleave upon hydrolysis, leading to several structural forms, including oligomers. Ionic Q species with no Ti, i.e., obtained after Ti,Q cleavage, are seen for some Ti,Q derivatives by ESI-MS, which also indicates a varied nuclearity for a given species, e.g., the isolated polynuclear [Q2Ti-,-O]n has several "n" values. Mononuclear Ti complexes are obtained under rigorous anhydrous conditions. The cis structures of the mononuclear species (QT)2Ti(OCH3)2, QT = 3-methyl-4-(neopentylcarbonyl)-1-phenylpyrazol-5-onato have been analyzed with DFT methods. A trans influence is a major driving force that accounts for several sets of Ti,O bonds. One of the cis stereoisomers is 56 kcal/mol higher in energy than the other two. In contrast, all (QT)2TiCl2cis isomers show similar energies. Voltammetry of the mononuclear species (QT)2Ti(OnPr)2 and the antitumor tetranuclear compound [(QB)2Ti-,-O]4, (QB = 4-benzoyl-3-methyl-1-phenylpyrazol-5-onato) indicate that the TiIV is less prone to reduction to TiIII in the latter (Epc for the TiIV/TiIII couple is ,1.71 V and ,1.46 V versus Fc+/Fc, respectively). Potential antitumor compounds having a Ti/Q ratio of 1:1 do not disproportionate, unlike the equivalent acetylacetonato derivatives, and are water-soluble. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003) [source] Reduction of [(C5Me5)2Mo2O5] and [(C5Me5)2Mo2O4] in Methanol/Water/Trifluoroacetate Solutions Investigated by Combined On-Line Electrochemistry/Electrospray-Ionization Mass SpectrometryEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 12 2003Jenny Gun Abstract Complexes [Cp*2Mo2O5] (Cp* = ,5 -C5Me5) and [Cp*2Mo2O4] were investigated by combined on-line electrochemical (EC) reduction and electrospray-ionization mass spectrometry (ESI-MS) techniques in a trifluoroacetic acid buffered water/methanol solution. The reduction products at the larger negative potentials are identical for both compounds. The studies reveal the existence of a wide range of previously unknown di- and trinuclear MoV, MoIV, MoIII, and mixed-valence complexes that were identified on the basis of their masses and characteristic isotope patterns. The structures of the initial compounds and the product of electroreduction with m/z = 713,729 were supported by in situ MSn experiments that allowed the elucidation of the fragmentation pathway for the collision-induced dissociation. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003) [source] Aspergillus nidulans,-galactosidase of glycoside hydrolase family 36 catalyses the formation of ,-galacto-oligosaccharides by transglycosylationFEBS JOURNAL, Issue 17 2010Hiroyuki Nakai The ,-galactosidase from Aspergillus nidulans (AglC) belongs to a phylogenetic cluster containing eukaryotic ,-galactosidases and ,-galacto-oligosaccharide synthases of glycoside hydrolase family 36 (GH36). The recombinant AglC, produced in high yield (0.65 g·L,1 culture) as His-tag fusion in Escherichia coli, catalysed efficient transglycosylation with ,-(1,6) regioselectivity from 40 mm 4-nitrophenol ,- d -galactopyranoside, melibiose or raffinose, resulting in a 37,74% yield of 4-nitrophenol ,- d -Galp -(1,6)- d -Galp, ,- d -Galp -(1,6)-,- d -Galp -(1,6)- d -Glcp and ,- d -Galp -(1,6)-,- d -Galp -(1,6)- d -Glcp -(,1,,2)- d -Fruf (stachyose), respectively. Furthermore, among 10 monosaccharide acceptor candidates (400 mm) and the donor 4-nitrophenol ,- d -galactopyranoside (40 mm), ,-(1,6) linked galactodisaccharides were also obtained with galactose, glucose and mannose in high yields of 39,58%. AglC did not transglycosylate monosaccharides without the 6-hydroxymethyl group, i.e. xylose, l -arabinose, l -fucose and l -rhamnose, or with axial 3-OH, i.e. gulose, allose, altrose and l -rhamnose. Structural modelling using Thermotoga maritima GH36 ,-galactosidase as the template and superimposition of melibiose from the complex with human GH27 ,-galactosidase supported that recognition at subsite +1 in AglC presumably requires a hydrogen bond between 3-OH and Trp358 and a hydrophobic environment around the C-6 hydroxymethyl group. In addition, successful transglycosylation of eight of 10 disaccharides (400 mm), except xylobiose and arabinobiose, indicated broad specificity for interaction with the +2 subsite. AglC thus transferred ,-galactosyl to 6-OH of the terminal residue in the ,-linked melibiose, maltose, trehalose, sucrose and turanose in 6,46% yield and the ,-linked lactose, lactulose and cellobiose in 28,38% yield. The product structures were identified using NMR and ESI-MS and five of the 13 identified products were novel, i.e. ,- d -Galp -(1,6)- d -Manp; ,- d -Galp -(1,6)-,- d -Glcp -(1,4)- d -Glcp; ,- d -Galp -(1,6)-,- d -Galp -(1,4)- d -Fruf; ,- d -Galp -(1,6)- d -Glcp -(,1,,1)- d -Glcp; and ,- d -Galp -(1,6)-,- d -Glcp -(1,3)- d -Fruf. [source] Native and subunit molecular mass and quarternary structure of the hemoglobin from the primitive branchiopod crustacean Triops cancriformisFEBS JOURNAL, Issue 17 2006Morgane Rousselot Many branchiopod crustaceans are endowed with extracellular, high-molecular-weight hemoglobins whose exact structural characteristics have remained a matter of conjecture. By using a broad spectrum of techniques, we provide precise and coherent information on the hemoglobin of one of the phylogenetically ,oldest' extant branchiopods, the tadpole shrimp Triops cancriformis. The hemoglobin dissociated under reducing conditions into two subunits, designated TcHbA and TcHbB, with masses of 35 775 ± 4 and 36 055 ± 4 Da, respectively, determined by ESI-MS. Nonreducing conditions showed only two disulfide-bridged dimers, a homodimer of TcHbA, designated D1 (71 548 ± 5 Da), and the heterodimer D2 (71 828 ± 5 Da). Carbamidomethylation of free SH groups revealed the presence of three cysteines per subunit and indicated one intrasubunit and one intersubunit disulfide bridge. Ultracentrifugation and light-scattering experiments under nondenaturating conditions yielded mass estimates that suggested an uneven number of 17 subunits forming the native hemoglobin. This unrealistic number resulted from the presence of two size classes (16-mer and 18-mer), which were recognized by native PAGE and Ferguson plot analysis. ESI-MS revealed three hemoglobin isoforms with masses of 588.1 kDa, 662.0 kDa, and 665.0 kDa. The 16-mer and the smaller 18-mer species are supposed to be composed of TcHbA only, given the dominance of this subunit type in SDS/PAGE. Transmission electron microscopy of negatively stained specimens showed a population of compact molecules with geometrical extensions of 14, 16 and 9 nm. The proposed stoichiometric model of quarternary structure provides the missing link to achieve a mechanistic understanding of the structure,function relationships among the multimeric arthropodan hemoglobins. [source] Structural characterization of a novel branching pattern in the lipopolysaccharide from nontypeable Haemophilus influenzaeFEBS JOURNAL, Issue 14 2003Martin Månsson Structural analysis of the lipopolysaccharide (LPS) from nontypeable Haemophilus influenzae strain 981 has been achieved using NMR spectroscopy and ESI-MS on O -deacylated LPS and core oligosaccharide (OS) material as well as by ESI-MSn on permethylated dephosphorylated OS. A heterogeneous glycoform population was identified, resulting from the variable length of the OS branches attached to the glucose residue in the common structural element of H. influenzae LPS, l -,- d -Hepp -(1,2)-[PEtn,6]- l -,- d -Hepp -(1,3)-[,- d -Glcxp-(1,4)]- l -,- d -Hepp -(1,5)-[PPEtn,4]-,-Kdop -(2,6)-Lipid A. Notably, the O-6 position of the ,- d -Glcp residue was either substituted by PCho or the disaccharide branch ,- d -Galp -(1,4)- d -,- d -Hepp, while the O-4 position was substituted by the globotetraose unit, ,- d -GalpNAc-(1,3)-,- d -Galp -(1,4)-,- d -Galp -(1,4)-,- d -Glcp, or sequentially truncated versions thereof. This is the first time a branching sugar residue has been reported in the outer-core region of H. influenzae LPS. Additionally, a PEtn group was identified at O-3 of the distal heptose residue in the inner-core. [source] Structural analysis of the lipopolysaccharide from nontypeable Haemophilus influenzae strain 1003FEBS JOURNAL, Issue 3 2002Martin Månsson Structural analysis of the lipopolysaccharide (LPS) of nontypeable Haemophilus influenzae strain 1003 has been achieved by the application of high-field NMR techniques, ESI-MS, capillary electrophoresis coupled to ESI-MS, composition and linkage analyses on O-deacylated LPS,and core oligosaccharide material. It was found that,the LPS contains the common structural element of,H. influenzae, l -,- d -Hepp -(1,2)-[PEtn,6]- l -,- d -Hepp -(1,3)-[,- d -Glcp -(1,4)]- l -,- d -Hepp -(1,5)-[PP Etn,4]-,-Kdop -(2,6)-Lipid A, in which the ,- d -Glcp residue is substituted by phosphocholine at O-6 and an acetyl group at O-4. A second acetyl group is located at O-3 of the distal heptose residue (HepIII). HepIII is chain elongated at O-2 by either a ,- d -Glcp residue (major), lactose or sialyllactose (minor, i.e. ,-Neu5Ac-(2,3)-,- d -Galp -(1,4)-,- d -Glcp), where a third minor acetylation site was identified at the glucose residue. Disialylated species were also detected. In addition, a minor substitution of ester-linked glycine at HepIII and Kdo was observed. [source] Phosphorylation and oligomerization states of native pig brain HSP90 studied by mass spectrometryFEBS JOURNAL, Issue 8 2001Cyrille Garnier HSP90 is one of the most abundant proteins in the cytosol of eukaryotic cells. HSP90 forms transient or stable complexes with several key proteins involved in signal transduction including protooncogenic protein kinases and nuclear receptors, it interacts with cellular structural elements such as actin-microfilament, tubulin-microtubule and intermediate filaments, and also exhibits conventional chaperone functions. This protein exists in two isoforms ,-HSP90 and ,-HSP90, and it forms dimers which are crucial species for its biological activity. PAGE, ESI-MS and MALDI-MS were used to study HSP90 purified from pig brain. The two protein isoforms were clearly distinguished by ESI-MS, the , isoform being ,,six times more abundant than the , isoform. ESI-MS in combination with ,,phosphatase treatment provided direct evidence of the existence of four phosphorylated forms of native pig brain ,-HSP90, with the diphosphorylated form being the most abundant. For the , isoform, the di-phosphorylated was also the most abundant. MALDI mass spectra of HSP90 samples after chemical cross-linking showed a high percentage of ,,, homodimers. In addition, evidence for the existence of higher HSP90 oligomers was obtained. [source] Iridoid Glucosides from Eremostachys moluccelloidesBungeHELVETICA CHIMICA ACTA, Issue 8 2007hsan Çal Abstract From the aerial parts of Eremostachys moluccelloidesBunge, a new iridoid glucoside, lamalbidic acid (7), was isolated as its choline salt 7a together with six known iridoid glucosides, 5-deoxysesamoside (1), 6, -hydroxy-7-epiloganin (2), lamalbide (3), shanzhiside methyl ester (4), sesamoside (5), and 5-deoxypulchelloside I (6). The structures of 7a and 7 (obtained from 7a) were elucidated by spectroscopic (UV, IR, 1D- and 2D-NMR, and ESI-MS) methods. [source] Substitution- and Elimination-Free Phosphorylation of Functionalized Alcohols Catalyzed by Oxidomolybdenum TetrachlorideADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 1 2010Cheng-Yuan Liu Abstract Among 14 oxidometallic species examined for catalytic phosphorylation of the tested alcohols, oxidomolybdenum tetrachloride (MoOCl4) was found to be the most efficient with a negligible background reaction mediated by triethylamine (Et3N). The new catalytic protocol can be applied to the chemoselective phosphorylations of primary, secondary and tertiary alcohols as well as the substitution-free phosphorylations of allylic, propargylic, and benzylic alcohols. Functionalized alcohols bearing acetonide, tetrahydropyranyl ether, tert -butyldimethylsilyl ether, or ester group are also amenable to the new catalytic protocol. The most difficult scenarios involve substitution-free phosphorylations of 1-phenylethanol and 1-(2-naphthyl)ethanol which can be effected in 95 and 90% yields, respectively. ESI-MS, IR, 1H, and 31P,NMR spectroscopic analyses of the reaction progress suggest the intermediacy of an alkoxyoxidomolybdenum trichloride-triethylamine adduct such as [(RO)Mo(O)Cl3 -Et3N] to be responsible for the catalytic turnover. [source] Candida antarctica Lipase B (CAL-B)-Catalyzed Carbon-Sulfur Bond Addition and Controllable Selectivity in Organic MediaADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 13 2008Feng-Wen Lou Abstract A novel enzymatic, promiscuous protocol for Candida antarctica lipase B (CAL-B)-catalyzed carbon-sulfur bond addition is described. Some control experiments have been designed to demonstrate the catalytic specificity of CAL-B. Selectivity between anti-Markovnikov addition and Markovnikov addition was achieved in different organic media. A series of thioether-containing ester functional groups was synthesized under the catalysis of CAL-B at 50,°C. All the products were characterized by spectroscopic methods (IR, NMR, ESI-MS). [source] CHRACTERIZATION AND 1,1-DIPHENYL-2-PICRYLHYDRAZYL RADICAL SCAVENGING ACTIVITY OF METHANOL AND SUPERCRITICAL CARBON DIOXIDE EXTRACTS FROM LEAVES OF ADINANDRA NITIDAJOURNAL OF FOOD BIOCHEMISTRY, Issue 4 2008BENGUO LIU ABSTRACT Leaves of Adinandra nitida are consumed in southern China as health tea (Shiyacha) and as herbal medicine. In this study, the methanol and supercritical fluid extracts from leaves of A. nitida were obtained by traditional solvent extraction and supercritical carbon dioxide extraction, respectively. Both the extracts showed high 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity. By using ultraviolet-visible spectrometry (UV), infrared spectrometry (IR), nuclear magnetic resonance, electrospray ionization mass spectrometry (ESI-MS), high-performance liquid chromatography-ESI/MS, the main bioactive constituents in the methanol extract (ME) were identified as camellianin A, camellianin B, apigenin. By analysis of gas chromatography-mass spectrometry, a total of 16 compounds accounting for 98.79% of the supercritical fluid extract (SFE) were identified as ,-sitosterol, vitamin E, ,-tocopherol and so on. These compounds found in ME and SFE could contribute to the DPPH radical scavenging performance of the extracts in this study. PRACTICAL APPLICATION Adinandra nitida is a kind of particular wild plant in South China. Few reports have been published about it in the world. In this study, the methanol and supercritical fluid extracts from leaves of A. nitida were respectively obtained by two kinds of industrially significant methods, traditional solvent extraction and supercritical carbon dioxide extraction. By using ultraviolet-visible spectrometry (UV), infrared spectrometry (IR), nuclear magnetic resonance, electrospray ionization mass spectrometry (ESI-MS), high-performance liquid chromatography-ESI/MS, gas chromatography-MS, the main bioactive constituents in the two extracts were identified as flavonoids and plant sterols. Both the extracts showed high 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity and this activity of the flavonoid-rich methanol extract was 10 times more than that of butylated hydroxytoluene. These results showed that leaves of A. nitida is a new kind of natural antioxidant-rich, flavonoid-rich plant source with great commercial interest in the food and phytopharmaceutical market. 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