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HPLC Separation (hplc + separation)
Selected AbstractsFluorocycloalkylated Fullerenes in the Systems C60/70,C2F4I2EUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 30 2007Anna S. Pimenova Abstract An addition reaction of biradicals thermally generated from C2F4I2 was applied to functionalize fullerenes. This resulted in the formation of a number of C60(C2F4)n and C70(C2F4)m compounds. HPLC separation allowed the determination of the molecular structures of C60(C4F8)2 (two isomers), C60(C4F8)6, and C70(C2F4)2 and revealed that these compounds are formed by different modes of [4,+,2], [4,+,3], and [2,+,2] addition, respectively. A new synthetic approach for the preparation of a series of fluorocycloalkylated derivatives of fullerenes should be further exploited. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Re-evaluation of intramolecular long-range electron transfer between tyrosine and tryptophan in lysozymesFEBS JOURNAL, Issue 17 2003Evidence for the participation of other residues One-electron oxidation of six different c-type lysozymes from hen egg white, turkey egg white, human milk, horse milk, camel stomach and tortoise was studied by gamma- and pulse-radiolysis. In the first step, one tryptophan side chain is oxidized to indolyl free radical, which is produced quantitatively. As shown already, the indolyl radical subsequently oxidizes a tyrosine side chain to the phenoxy radical in an intramolecular reaction. However this reaction is not total and its stoichiometry depends on the protein. Rate constants also vary between proteins, from 120·s,1 to 1000·s,1 at pH 7.0 and room temperature [extremes are hen and turkey egg white (120·s,1) and human milk (1000·s,1)]. In hen and turkey egg white lysozymes we show that another reactive site is the Asn103,Gly104 peptidic bond, which gets broken radiolytically. Tryptic digestion followed by HPLC separation and identification of the peptides was performed for nonirradiated and irradiated hen lysozyme. Fluorescence spectra of the peptides indicate that Trp108 and/or 111 remain oxidized and that Tyr20 and 53 give bityrosine. Tyr23 appears not to be involved in the process. Thus new features of long-range intramolecular electron transfer in proteins appear: it is only partial and other groups are involved which are silent in pulse radiolysis. [source] Synthesis and characterization of tritium labeled 2,3,4,9-tetrahydro-1H -carbazoles as potent DP receptor antagonistsJOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS, Issue 2 2004Carl Berthelette Abstract Tritium labeled 2,3,4,9-tetrahydro-1H -carbazole 1a and 2a were prepared in good yields with a specific activity of 7.0 Ci/mmol (214 GBq/mmol) and 4.2 Ci/mmol (155 GBq/mmol), respectively. Both compounds have been synthesized in high radiochemical purity by catalytic tritium,bromine exchange of the corresponding aryl bromide precursors. The 6-bromocarbazole precursors 7 and 8 were prepared as a mixture by a three step process, involving regioselective bromination of 3c with pyridinium tribromide, oxidation of thioether 4c using m -CPBA and hydrolysis of acylsultamcarbazole 5c. Finally, HPLC separation of the enantiomers afforded the 6-bromo precursors 7 and 8 in high diastereomeric ratio (dr 99% and dr 93% respectively). Copyright © 2004 John Wiley & Sons, Ltd. [source] A novel prohormone processing site in Aplysia californica: the Leu,Leu ruleJOURNAL OF NEUROCHEMISTRY, Issue 6 2002Amanda B. Hummon Abstract Neuropeptides are a complex set of signaling molecules produced through enzymatic cleavages from longer prohormone sequences. The most common cleavage sites in prohormones are basic amino acid residues; however, processing is observed at non-basic sites. Cleavage at Leu,Leu sequences has been observed in three Aplysia californica prohormones. To further investigate this unusual event, native and non-native synthetic peptides containing Leu,Leu residues are incubated with homogenates of Aplysia californica ganglia and the resulting products monitored with MALDI MS. Cleavage near and between Leu,Leu residues is observed in the abdominal and buccal ganglia homogenates, confirming the presence of an unidentified peptidase. In addition, fractions from an HPLC separation of buccal ganglia homogenates also produce cleavages at Leu,Leu residues. Products resulting from cleavage at Leu,Leu sites are observed and are produced in larger amounts in acidic and neutral pH ranges, and cleavage is inhibited by the addition of EDTA, suggesting a metal is required for activity. [source] Complete structure determination of the A chain of mistletoe lectin III from Viscum album L. ssp. albumJOURNAL OF PEPTIDE SCIENCE, Issue 3 2004Roland Wacker Abstract The complete primary structure of the A chain of mistletoe lectin III (ML3A), a type II ribosome-inactivating protein, was determined using proteolytic digests of ML3A, HPLC separation of the peptides, Edman degration and MALDI-MS. Based on our results, ML3A consists of 254 amino acid residues, showing a high homology to the A chain of isolectin ML1 with only 24 amino acid residue exchanges. A striking important structural difference compared with ML1A is the lack of the single N-glycosylation site in ML3A due to an amino acid exchange at position 112 (ML1A: N112GS,ML3A: T112GS). The alignment of ML3A with the A chains of ML1, isoabrins, ricin D, Ricinus communis agglutinin and three lectins, identified from the Korean mistletoe Viscum album ssp. coloratum, demonstrates the rigid conservation of all amino acid residues, responsible for the RNA-N-glycosidase activity as reported for ricin D. In addition, the fully determined primary structure of ML3A will give further information about the biological mechanism of mistletoe lectin therapy. Copyright © 2003 European Peptide Society and John Wiley & Sons, Ltd. [source] OILS FROM CENOZOIC RIFT-BASINS IN CENTRAL AND NORTHERN THAILAND: SOURCE AND THERMAL MATURITYJOURNAL OF PETROLEUM GEOLOGY, Issue 1 2007H.I. Petersen Oil is produced from the Suphan Buri, Phitsanulok and Fang Basins onshore central and northern Thailand. Most of the Cenozoic rift-basins onshore Thailand are 2,4 km deep, but the Phitsanulok Basin is the deepest with a basin-fill up to 8 km thick. In this basin, the Sirikit field produces ,18,000,24,000 bbl/day of crude oil. In the Suphan Buri Basin, about 400 bbl/day of crude oil is produced from the U Thong and Sang Kajai fields. Approximately 800 bbl/day of crude oil is produced from the Fang field (Fang Basin), which in reality consists of a number of minor structures including Ban Thi, Pong Nok, San Sai, Nong Yao and Mae Soon. A total of eight oil samples were collected from these structures and from the Sirikit, U Thong and Sang Kajai fields. The oils were subjected to MPLC and HPLC separation and were analysed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS and GC-MS-MS). The U Thong oil was investigated in more detail by separating the oil into a number of fractions suited for the analysis of various specific compounds. The Sirikit oil appears to be the most mature, whereas the Suphan Buri oils and the oil from the San Sai structure (Fang Basin) are the least mature. Apart from the San Sai oil, the other oils in the Fang Basin are of similar maturity. The oils contain small amounts of asphaltenes and the asphaltene-free fractions are completely dominated by saturated hydrocarbons (generally >60%). Long-chain n-alkanes extend to at least C40 and the oils are thus highly waxy. In general the oils were generated from freshwater lacustrine source rocks containing a large proportion of algal material, as indicated by the presence of long-chain n-alkanes, low C3122R/C30 hopane ratios, the presence of 28-Nor-spergulane, T26/T25 (tricyclic triterpanes) ratios of 1.07,1.57 and tetracyclic polyprenoid (TTP) ratios close to 1. Occasional saline conditions may have occurred during deposition of the Sirikit, Ban Thi and Pong Nok source rocks. The Fang Basin oils were sourced from two different kitchens, one feeding the Ban Thi and Pong Nok structures and one feeding the Mae Soon, Nong Yao and San Sai structures. The presence ofcadalene, tetracyclic C24 compounds, oleanane, lupane, bicadinane and trace amounts ofnorpimarane or norisopimarane indicate a contribution from higher land plant organic matter to the oils. The terrestrial organic matter may occur disseminated in the lacustrine facies or concentrated in coal seams associated with the lacustrine mudstones. Thermally immature oil shales (lacustrine mudstones) and coals exposed in numerous basins in central and northern Thailand could upon maturation generate oils with a composition comparable to the investigated oils. [source] Determination of aminoglycoside and macrolide antibiotics in meat by pressurized liquid extraction and LC-ESI-MSJOURNAL OF SEPARATION SCIENCE, JSS, Issue 4-5 2010Houda Berrada Abstract A simple method for the simultaneous determination of dihydrostreptomycin, spectinomycin, spiramycin, streptomycin, tilmicosin, and tylosin in meat has been developed using pressurized liquid extraction and LC-triple quadrupole MS (LC-ESI-MS/MS). The pressurized liquid extraction operational parameters were optimized and no protein precipitating and fat removing steps were required. A gradient HPLC separation was developed with ion-pair mobile phases consisting of aqueous 1,mM heptafluorobutyric acid water and methanol. Protonated molecules were used as precursor ions for CID. Data acquisition under MS/MS was achieved by applying multiple reaction monitoring of three fragment ion transitions to provide a high degree of sensitivity and specificity. Dirithromycin and sisomycin were selected as internal standards. A validation study was conducted for these antibiotics in poultry meat samples. All selected compounds could be detected (monitoring ions by multiple reaction monitoring) in meat samples at amounts below the regulatory level of concern. Using the internal standards, pressurized liquid extraction recovery rates were from 70 to 96% (RSD 12,25%). LC-ESI-MS/MS method detection limits of the selected antibiotics were 1,6,,g/kg. Good method reproducibility was found by intra- and inter-day precisions at maximum residue level, yielding the RSDs less than 15 and 16%, respectively. [source] Multiresidue HPLC analysis of ten quinolones in milk after solid phase extraction: Validation according to the European Union Decision 2002/657/ECJOURNAL OF SEPARATION SCIENCE, JSS, Issue 15 2007Eleni A. Christodoulou Abstract A rapid and sensitive analytical method was developed for the residue analysis of ten quinolones (enoxacin (ENO), ofloxacin (OFL), norfloxacin (NOR), ciprofloxacin (CIP), danofloxacin (DAN), enrofloxacin (ENR), sarafloxacin (SAR), oxolinic acid (OXO), nalidixic acid (NAL), and flumequine (FLU)) in cow's milk. The analytes were extracted from milk by a deproteinization step followed by a simple SPE cleanup procedure using LiChrolut RP-18 Merck cartridges. Recoveries varied between 75 and 92%. HPLC separation was performed at 25°C using an ODS-3 PerfectSil® Target (250×4 mm2) 5 ,m analytical column (MZ-Analysentechnik, Germany). The mobile phase consisted of a mixture of TFA 0.1%,CH3CN,CH3OH, delivered by a gradient program at the flow rate of 1.2 mL/min. Elution of the ten analytes and the internal standard (caffeine, 7.5 ng/,L) was completed within 27 min. Column effluent was monitored using a photodiode array detector, set at 275 and 255 nm. The developed method was validated according to the criteria of Commission Decision 2002/657/EC. The LODs of the specific method of quinolones' determination in milk varied between 1.5 and 6.8 ng/,L. [source] HPLC separation of fullerenes on two charge-transfer stationary phasesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6 2006Qiong-Wei Yu Abstract Two charge-transfer stationary phases were prepared by immobilizing p -nitrobenzoic acid and naphthyl acetic acid onto silica. The nitrophenyl moiety and the naphthyl moiety were grafted to silica gel through the spacer of aminoalkyl silanes. The HPLC separation of C60, C70, and higher fullerenes on the new stationary phases was also studied. The influence of mobile phase and column temperature on the separation of C60 and C70 was examined, respectively. The retentions of C60 and C70 on the two stationary phases increased with decreasing toluene content in the mobile phase or with increasing column temperature. Higher fullerenes can be separated well using toluene as the mobile phase on the stationary phase of p -nitrobenzoic acid-bonded silica. [source] Plasma pharmacokinetics of high-dose oral busulfan in children and adults undergoing bone marrow transplantationPEDIATRIC TRANSPLANTATION, Issue 2003Bruce Bostrom Abstract: We have analyzed the plasma pharmacokinetics of busulfan in 272 patients receiving high-dose oral busulfan and intravenous cyclophosphamide in conjunction with allogeneic or autologous bone marrow transplantation. The patients ranged in age from 2 months to 59 yr (mean 10, median 12 yr) and had the following diagnoses: thalassemia or sickle cell anemia (n = 74); leukemia or myelodysplasia (n = 112); inborn errors of metabolism (n = 41) or immunodeficiency (n = 45). Plasma specimens were collected following the first dose for each patient which ranged from 1 to 4 mg/kg (mean ± SD, 1.21 ± 0.41, median 1.15). Busulfan was quantitated using ultraviolet absorbance detection after derivatization and HPLC separation. Pharmacokinetic parameters were derived by modeling the raw data to fit first-order single compartment kinetics. The kinetic parameters showed wide interpatient variability independent of age and diagnosis. There was a statistically significant correlation of age with the following parameters: area under the curve (AUC); maximal concentration; minimum concentration; clearance; volume of distribution and absorption half-time. The coefficients of determination (i.e. correlation coefficient squared) were low ranging from 0.04 to 0.12 implying only a small part (i.e. 4,12%) of the variance was explained by age. Although busulfan pharmacokinetics are age-related most of the variability is not explained by age or diagnosis. [source] High-performance liquid chromatographic separation of natural and synthetic desulphoglucosinolates and their chemical validation by UV, NMR and chemical ionisation-MS methodsPHYTOCHEMICAL ANALYSIS, Issue 4 2001Guy Kiddle Abstract Methods are described for the optimised extraction, desulphation and HPLC separation of desulphoglucosinolates. These methods provide rapid separation, identification and quantitative measurements of glucosinolates extracted from Brassica napus L and related crops, of unusual glucosinolates found in crucifer weed species, and also of synthetic alkylglucosinolates. The desulphoglucosinolates used in these studies were either chemically synthesised (at least one example from each major structural class), or purified from various plant sources. Validation of the identities of the desulphoglucosinolates was by comparison of retention times with standards, and by UV, 1H- and 13C-NMR and chemical ionisation MS analysis. A list of useful species, and the specific tissues, from which high concentrations of standards can be extracted is included. Copyright © 2001 John Wiley & Sons, Ltd. [source] Detailed proteome analysis of growing cells of the planctomycete Rhodopirellula baltica SH1TPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 8 2008Cao Xuan Hieu Abstract Rhodopirellula baltica SH1T, which was isolated from the water column of the Kieler Bight, a bay in the southwestern Baltic Sea, is a marine aerobic, heterotrophic representative of the ubiquitous bacterial phylum Planctomycetes. We analyzed the R. baltica proteome by applying different preanalytical protein as well as peptide separation techniques (1-D and 2-DE, HPLC separation) prior to MS. That way, we could identify a total of 1115 nonredundant proteins from the intracellular proteome and from different cell wall protein fractions. With the contribution of 709 novel proteins resulting from this study, the current comprehensive R. baltica proteomic dataset consists of 1267 unique proteins (accounting for 17.3% of the total putative protein-coding ORFs), including 261 proteins with a predicted signal peptide. The identified proteins were functionally categorized using Clusters of Orthologous Groups (COGs), and their potential cellular locations were predicted by bioinformatic tools. A unique protein family that contains several YTV domains and is rich in cysteine and proline was found to be a component of the R. baltica proteinaceous cell wall. Based on this comprehensive proteome analysis a global schema of the major metabolic pathways of growing R. baltica cells was deduced. [source] Role of allatostatin-like factors from the brain of Tenebrio molitor females,ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 4 2009O. Wasielewski Abstract The effect of brain extract from females of freshly emerged Tenebrio molitor on ovary, oocyte development, total protein content of hemolymph, and ovary was studied in 4-day-old adult mealworm females. Injections of extracts of 2-brain equivalents into intact (unligatured) Tenebrio females did not affect ovarian and oocyte development. Injections of ligated females, however, with 2-brain equivalents on day 1 and 2 after adult emergence strongly inhibited ovarian growth and oocyte development. At day 4, ligated and injected females did not develop their ovaries and pre-vitellogenic oocytes were not found. The changes in ovarian development correlated with an increase in the concentration of soluble proteins in the hemolymph as compared with the saline-injected controls. Additionally, a strong reduction of total protein content in ovarian tissue was observed. Reverse phase HPLC separation of a methanolic brain extract of T. molitor females showed that fraction 5 has a similar retention time to synthetic cockroach allatostatin. Fraction 5 was eluted at 12.88,min, which was closest to the internal standard Dippu-AST I, which eluted at 12.77,min. An ELISA of fraction 5 from the methanolic brain extract using antibodies against allatostatins Grybi-AST A1 and Grybi-AST B1 from cricket Gryllus bimaculatus showed that fraction 5 cross-reacted with Grybi-AST A1 antibodies. The cross-reactivity was similar to the synthetic allatostatin from D. punctata, which was used as a positive control. These observations demonstrate a possible role for allatostatin-like brain factor(s) in regulating the reproductive cycle of Tenebrio molitor. © 2009 Wiley Periodicals, Inc. [source] Enantioresolution of dl -penicillamineBIOMEDICAL CHROMATOGRAPHY, Issue 1 2010Ravi Bhushan Abstract Penicillamine (PenA) is a nonproteinogenic amino acid containing a thiol group. The three functional groups in penicillamine undergo characteristic chemical reactions and differ in their ability to participate in various chemical and biochemical reactions. d -penicillamine is more active pharmacologically, while the l -isomer occurs ,naturally'. This review deals with the enantioresolution of PenA both by direct and indirect methods using liquid chromatography. HPLC separation of its diastereomers prepared with different chiral derivatizing reagents (on reversed-phase columns) and separation of the derivatives prepared with achiral reagents (on chiral columns or via ligand exchange mode) has been discussed. Separation of enantiomers tagged with achiral reagent (to add a chromophore for ehanced detection) when there is no diastereomer formation has been considered separately. In addition, application of PenA and its derivatives as chiral selector for enentioresolution of certain other compounds has also been discussed. Copyright © 2009 John Wiley & Sons, Ltd. [source] A fully automated amino acid analyzer using NBD-F as a ,uorescent derivatization reagentBIOMEDICAL CHROMATOGRAPHY, Issue 9 2004Chiaki Aoyama Abstract A fully automated amino acid analyzer using NBD-F (4-,uoro-7-nitro-2,1,3-benzoxadiazole) as a ,uorescent derivatization reagent was developed. The whole analytical process was fully automated from derivatization, injection to HPLC separation and quantitation. The derivatization reaction conditions were re-evaluated and optimized. Amino acids were derivatized by NBD-F for 40 min at room temperature in the borate buffer (pH 9.5). The derivatives were separated within 100 min and ,uorometrically detected at 540 nm with excitation at 470 nm. The detection limits for amino acids were in the range of 2.8,20 fmol. The calibration curves were linear over the range of 20 fmol to 20 pmol on column with the correlation coef,cients of 0.999. The coef,cients of variation were less than 5% at 3 pmol injection for all amino acids. Amino acids in rat plasma were determined by the proposed HPLC method. Copyright © 2004 John Wiley & Sons, Ltd. [source] Determination of the ,avone tricin in human plasma by high-performance liquid chromatographyBIOMEDICAL CHROMATOGRAPHY, Issue 7 2003Hong Cai Abstract Tricin is a ,avone constituent of brown rice and rice bran, which interferes potently with the survival of human-derived breast and colon cancer cells in vitro. A speci,c and simple high-performance liquid chromatographic (HPLC) method was developed for the determination of tricin in human plasma with UV,visible detection. HPLC separation on Hypersil-BDS C18 (4.6 × 250 mm) was carried out with an isocratic mobile phase of 52% methanol in 0.1 m ammonium acetate, pH 5.10, containing 0.27 mm disodium ethylenediamine tetraacetic acid and detection at 355 nm. The retention times of tricin and quercetin (internal standard) were 14.2 and 7.8 min, respectively. The assay was linear in the range 1,100 µg/mL (r2 , 0.995). Tricin in plasma was ef,ciently extracted with 0.1 m acetic acid in acetone, and the recoveries were in the range 92.6,102.8% (n = 6) with relative standard deviation below 10% for three concentrations of tricin, 5, 10 and 100 µg/mL. The lower limit of quantitation (relative standard deviation <20%) was 1 µg/mL. Copyright © 2003 John Wiley & Sons, Ltd. [source] Simultaneous determination of nicotinamide, pyridoxine hydrochloride, thiamine mononitrate and riboflavin in multivitamin with minerals tablets by reversed-phase ion-pair high performance liquid chromatographyBIOMEDICAL CHROMATOGRAPHY, Issue 8 2002Ke Li A reversed-phase ion-pair high performance liquid chromatographic method (HPLC) has been developed and validated for the routine analysis of nicotinamide, pyridoxine hydrochloride, thiamine mononitrate and riboflavin in multivitamin with minerals tablets. HPLC separation of the vitamins was performed on a Hypersil C18 column and detected by ultraviolet absorbance at 280,nm. The use of methanol-aqueous 0.5% acetic acid solution (18:82, v/v; containing 2.5,mM sodium hexanesulfonate, pH,=,2.8) as the mobile phase at a flow-rate of 1.2,mL/min enables the baseline separation of the four analytes free from interferences with isocratic elution at 30°C. The analysis time was 17,min per injection. The method was linear in the ranges of 5,90, 2.5,90, 5,95 and 25,450,µg/mL for thiamine mononitrate, riboflavin, pyridoxine hydrochloride and nicotinamide, respectively. The average coefficients of variation of within- and between-day assays were 2.2 and 3.6% for thiamine mononitrate, 1.8 and 2.4% for riboflavin, 1.3 and 1.7% for pyridoxine hydrochloride and 1.0 and 1.5% for nicotinamide, respectively. The average recoveries of thiamine mononitrate, riboflavin, pyridoxine hydrochloride and nicotinamide were 97.0, 97.2, 98.9 and 100.4% for the tablets, respectively. The method has been successfully applied to the simultaneous determination of nicotinamide, pyridoxine hydrochloride, thiamine mononitrate and riboflavin in multivitamin with minerals tablets. Copyright © 2002 John Wiley & Sons, Ltd. [source] How to Spoil the Taste of Insect Prey?CHEMBIOCHEM, Issue 12 2010A Novel Feeding Deterrent against Ants Released by Larvae of the Alder Leaf Beetle, Agelastica alni Abstract Chemical defense of leaf beetle larvae (Chrysomelidae) against enemies is provided by secretions containing a wide range of deterrent compounds or by unpalatable hemolymph constituents. Here we report a new, very strong feeding deterrent against ants released by larvae of the alder leaf beetle Agelastica alni when attacked. The larvae release a defensive fluid from openings of pairwise, dorsolaterally located tubercles on the first to the eighth abdominal segments. The fluid, consisting of hemolymph and probably a glandular cell secretion, has previously been shown to contain a very stable, non-volatile feeding deterrent. The major deterrent component was isolated by repeated HPLC separation and analyzed by NMR and MS. The compound proved to be ,- L -glutamyl- L -2-furylalanine (1), a novel dipeptide containing the unusual amino acid L -2-furylalanine. This amino acid, although synthetically well known, has not previously been reported from natural sources. The absolute configuration of the natural compound was elucidated by enantioselective gas chromatography after derivatization. The structure of the dipeptide was verified by the synthesis of several isomeric dipeptides. In bioassays a concentration of 1 ,g,,L,1 was sufficient to deter polyphagous Myrmica rubra ants from feeding. [source] Qualitative and Quantitative HPLC/MS Determination of Proanthocyanidins in Areca Nut (Areca catechu)CHEMISTRY & BIODIVERSITY, Issue 12 2007Qingli Wu Abstract Proanthocyanidins (PACs) in areca nut (Areca catechu L.) were analyzed by high-performance liquid chromatography (HPLC) combined with electrospray-ionization mass spectrometry (ESI-MS) and compared to grape seed extract. Under optimized conditions, the separated PACs were individually analyzed and identified on the basis of their [M+H]+ peaks. The PAC distribution in areca nut was found to be very similar to that in grape seed, but lacking any gallate conjugates. Based on reverse-phase HPLC separation, the PAC monomers (+)-catechin (CA, 1) and (,)-epicatechin (EC; 2) were successfully quantified by ESI-MS in the selected-ion-monitoring (SIM) mode, (,)-epigallocatechin (EGC; 3) being used as internal standard. Detailed quality and validation assays showed that the accuracy and repeatability (n=8) were within 10% for each analyte. [source] Determination of absolute configurations by X-ray crystallography and 1H NMR anisotropyCHIRALITY, Issue 5 2008Nobuyuki Harada Abstract To determine the absolute configurations of chiral compounds, many spectroscopic and diffraction methods have been developed. Among them, X-ray crystallographic Bijvoet method, CD exciton chirality method, and the combination of vibrational circular dichroism and quantum mechanical calculations are of nonempirical nature. On the other hand, X-ray crystallography using a chiral internal reference, and 1H NMR spectroscopy using chiral anisotropy reagents are relative and/or empirical methods. In addition to absolute configurational determinations, preparations of enantiopure compounds are strongly desired. As chiral reagents useful for both the preparation of enantiopure compounds by HPLC separation and the simultaneous determination of their absolute configurations, we have developed camphorsultam dichlorophthalic acid (CSDP acid) for X-ray crystallography and 2-methoxy-2-(1-naphthyl)propionic acid (M,NP acid) for 1H NMR spectroscopy. In this review, the principles and applications of these X-ray and NMR methods are explained using mostly our own data. Chirality, 2008. © 2007 Wiley-Liss, Inc. [source] Chiral HPLC separation and CD spectra of the enantiomers of the alkaloid tacamonine and related compoundsCHIRALITY, Issue 10 2001Salvatore Caccamese Abstract The HPLC enantiomeric separation of racemic indole alkaloids tacamonine, 17,-hydroxytacamonine, deethyleburnamonine, and vindeburnol was accomplished using Chiralpak AD and Chiralcel OD as chiral stationary phases. Small structural differences affect the enantioselectivity ability of these phases. Single enantiomers of tacamonine and vindeburnol were isolated by semipreparative HPLC and their CD spectra and optical rotations were measured. Chirality 13:691,693, 2001. © 2001 Wiley-Liss, Inc. [source] Alkylated poly(styrene-divinylbenzene) monolithic columns for ,-HPLC and CEC separation of phenolic acidsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 17 2007Zdenka Ku, erová Abstract Macroporous poly(styrene-divinylbenzene) monolithic columns were prepared in fused silica capillaries of 100 ,m id by in-situ copolymerization of styrene with divinylbenzene in the presence of propan-1-ol and formamide as the porogen system. The monoliths were subsequently alkylated with linear alkyl C-18 groups via Friedel-Crafts reaction to improve the retention and chromatographic resolution of strongly polar phenolic acids. A new thermally initiated grafting procedure was developed in order to shorten the time of the alkylation process. The grafting procedure was optimized with respect to the reaction temperature, time, the grafting reactant concentration, and the solvent used. The type of solvent and the grafting temperature are the most significant factors affecting the hydrodynamic properties, porosity, and efficiency of the columns. While the equivalent particle diameter of the grafted column increased, the capillary-like flow-through pore diameter decreased in comparison to non-alkylated monoliths. The hydrodynamic permeability of the monolith decreased, but the monolithic column still permitted fast ,-HPLC separations. [source] An initial assessment of the use of gradient elution in microemulsion and micellar liquid chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 17-18 2004Simon M. Bryant Abstract Novel microemulsion and micellar HPLC separations have been achieved using gradient elution and columns packed with reverse phase material. Initial attempts at gradient microemulsion liquid chromatography proved impossible on use of a microemulsion successfully used in capillary electrophoresis. Optimisation of the microemulsion composition allowed the generation of stable microemulsions to achieve separations in HPLC. The novel use of organic-solvent micellar chromatography in gradient elution mode was shown to give efficient separations. A range of efficient separations of pharmaceuticals and related impurities were obtained. Acidic, basic, and neutral solutes were resolved covering a wide range of water solubilities and polarities. Elution times were in the order of 4,15 minutes. Separations were briefly compared to those accomplished with a micellar HPLC system. It is proposed that gradient elution in both microemulsion and micellar HPLC can be regarded as a highly successful means of achieving resolution of complex mixtures and should be considered for routine analysis and further investigation. [source] Mixed-mode chromatography/isotope ratio mass spectrometry,RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 5 2010James S. O. McCullagh Liquid chromatography coupled to molecular mass spectrometry (LC/MS) has been a standard technique since the early 1970s but liquid chromatography coupled to high-precision isotope ratio mass spectrometry (LC/IRMS) has only been available commercially since 2004. This development has, for the first time, enabled natural abundance and low enrichment ,13C measurements to be applied to individual analytes in aqueous mixtures creating new opportunities for IRMS applications, particularly for the isotopic study of biological molecules. A growing number of applications have been published in a range of areas including amino acid metabolism, carbohydrates studies, quantification of cellular and plasma metabolites, dietary tracer and nucleic acid studies. There is strong potential to extend these to new compounds and complex matrices but several challenges face the development of LC/IRMS methods. To achieve accurate isotopic measurements, HPLC separations must provide baseline-resolution between analyte peaks; however, the design of current liquid interfaces places severe restrictions on compatible flow rates and in particular mobile phase compositions. These create a significant challenge on which reports associated with LC/IRMS have not previously focused. Accordingly, this paper will address aspects of chromatography in the context of LC/IRMS, in particular focusing on mixed-mode separations and their benefits in light of these restrictions. It aims to provide an overview of mixed-mode stationary phases and of ways to improve high aqueous separations through manipulation of parameters such as column length, temperature and mobile phase pH. The results of several practical experiments are given using proteogenic amino acids and nucleosides both of which are of noted importance in the LC/IRMS literature. This communication aims to demonstrate that mixed-mode stationary phases provide a flexible approach given the constraints of LC/IRMS interface design and acts as a practical guide for the development of new chromatographic methods compatible with LC/IRMS applications. Copyright © 2010 John Wiley & Sons, Ltd. [source] High-throughput determination of atrasentan in human plasma by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometryBIOMEDICAL CHROMATOGRAPHY, Issue 9 2005Perry G. Wang Abstract Atrasentan (A-147627) is an endothelin antagonist receptor being developed at Abbott Laboratories for the treatment of prostate cancer. A quick and sensitive method for the determination of atrasentan in human plasma has been developed and validated using high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry. A dual-column, single mass spectrometer system is used to provide a reliable and routine means to increase sample throughput. The analytical method involves liquid,liquid extraction and internal standard (A-166790). The plasma samples and internal standard are acidified with 0.3 m hydrochloric acid prior to being extracted into 1:1 (v[sol ]v) hexanes,methyl t -butyl ether. The organic extract was evaporated to dryness using heated nitrogen stream and reconstituted with mobile phase. Atrasentan and internal standard were separated with no interference in a Zorbax SB-C18 analytical column with 2.1 × 50 mm, 5 µm, and a Zorbax C8 guard column using a mobile phase consisting of 50:50 (v:v) acetonitrile,0.05 m ammonium acetate, pH 4.5, at a flow rate of 0.30 mL[sol ]min to provide 4 min chromatograms. For a 250 µL plasma sample volume, the limit of quantitation was approximately 0.3 ng[sol ]mL. The calibration was linear from 0.30 to 98.0 ng[sol ]mL (r2 > 0.995). A significant advantage of the method is the ability to employ parallel HPLC separations with detection by a single MS[sol ]MS system to provide sensitivity and selectivity sufficient to achieve robust analytical results with a lower limit of quantitation of 0.30 ng[sol ]mL and high throughput. Copyright © 2005 John Wiley & Sons, Ltd. [source] | |||||||||||||